HIV Resistance Prediction to Reverse Transcriptase Inhibitors: Focus on Open Data.

PubMed

Tarasova, Olga; Poroikov, Vladimir

2018-04-19

Research and development of new antiretroviral agents are in great demand due to issues with safety and efficacy of the antiretroviral drugs. HIV reverse transcriptase (RT) is an important target for HIV treatment. RT inhibitors targeting early stages of the virus-host interaction are of great interest for researchers. There are a lot of clinical and biochemical data on relationships between the occurring of the single point mutations and their combinations in the pol gene of HIV and resistance of the particular variants of HIV to nucleoside and non-nucleoside reverse transcriptase inhibitors. The experimental data stored in the databases of HIV sequences can be used for development of methods that are able to predict HIV resistance based on amino acid or nucleotide sequences. The data on HIV sequences resistance can be further used for (1) development of new antiretroviral agents with high potential for HIV inhibition and elimination and (2) optimization of antiretroviral therapy. In our communication, we focus on the data on the RT sequences and HIV resistance, which are available on the Internet. The experimental methods, which are applied to produce the data on HIV-1 resistance, the known data on their concordance, are also discussed.

Molecular docking and 3D-QSAR studies on triazolinone and pyridazinone, non-nucleoside inhibitor of HIV-1 reverse transcriptase.

PubMed

Sivan, Sree Kanth; Manga, Vijjulatha

2010-06-01

Nonnucleoside reverse transcriptase inhibitors (NNRTIs) are allosteric inhibitors of the HIV-1 reverse transcriptase. Recently a series of Triazolinone and Pyridazinone were reported as potent inhibitors of HIV-1 wild type reverse transcriptase. In the present study, docking and 3D quantitative structure activity relationship (3D QSAR) studies involving comparative molecular field analysis (CoMFA) and comparative molecular similarity indices analysis (CoMSIA) were performed on 31 molecules. Ligands were built and minimized using Tripos force field and applying Gasteiger-Hückel charges. These ligands were docked into protein active site using GLIDE 4.0. The docked poses were analyzed; the best docked poses were selected and aligned. CoMFA and CoMSIA fields were calculated using SYBYL6.9. The molecules were divided into training set and test set, a PLS analysis was performed and QSAR models were generated. The model showed good statistical reliability which is evident from the r2 nv, q2 loo and r2 pred values. The CoMFA model provides the most significant correlation of steric and electrostatic fields with biological activities. The CoMSIA model provides a correlation of steric, electrostatic, acceptor and hydrophobic fields with biological activities. The information rendered by 3D QSAR model initiated us to optimize the lead and design new potential inhibitors.

Combination nucleoside/nucleotide reverse transcriptase inhibitors for treatment of HIV infection.

PubMed

Akanbi, Maxwell O; Scarsi, Kimberly K; Scarci, Kimberly; Taiwo, Babafemi; Murphy, Robert L

2012-01-01

The combination of two nucleoside/nucleotide reverse transcriptase inhibitors (N(t)RTIs) and a third agent from another antiretroviral class is currently recommended for initial antiretroviral therapy. In general, N(t)RTIs remain relevant in subsequent regimens. There are currently six nucleoside reverse transcriptase inhibitors and one nucleotide reverse transcriptase inhibitor drug entities available, and several formulations that include two or more N(t)RTIs in a fixed-dose combination. These entities have heterogeneous pharmacological and clinical properties. Accordingly, toxicity, pill burden, dosing frequency, potential drug-drug interaction, preexisting antiretroviral drug resistance and comorbid conditions should be considered when constructing a regimen. This approach is critical in order to optimize virologic efficacy and clinical outcomes. This article reviews N(t)RTI combinations used in the treatment of HIV-infected adults. The pharmacological properties of each N(t)RTI, and the clinical trials that have influenced treatment guidelines are discussed. It is likely that N(t)RTIs will continue to dominate the global landscape of HIV treatment and prevention, despite emerging interest in N(t)RTI-free combination therapy. Clinical domains where only few alternatives to N(t)RTIs exist include treatment of HIV/HBV coinfection and HIV-2. There is a need for novel N(t)RTIs with enhanced safety and resistance profiles compared with current N(t)RTIs.

Nonnucleoside reverse transcriptase inhibitor phenotypic hypersusceptibility can be demonstrated in different assays.

PubMed

Shulman, Nancy S; Delgado, Jamael; Bosch, Ronald J; Winters, Mark A; Johnston, Elizabeth; Shafer, Robert W; Katzenstein, David A; Merigan, Thomas C

2005-05-01

HIV-1 isolates harboring multiple nucleoside reverse transcriptase inhibitor (NRTI) resistance mutations are more susceptible ("hypersusceptible") to the nonnucleoside reverse transcriptase inhibitors (NNRTIs) than isolates lacking NRTI resistance mutations, but this has only been reported with a single-cycle replication phenotypic assay. In fact, there was a report that a commercial multicycle assay did not readily detect hypersusceptibility. To see whether NNRTI hypersusceptibility can be demonstrated in other types of phenotypic assays, including multicycle assays and enzyme inhibition assays. The susceptibility of HIV-1 clones derived from different patients in multicycle assays was tested in peripheral blood mononuclear cells (PBMCs) and in an established cell line. In addition, the reverse transcriptase (RT) of many of these clones was expressed and their susceptibility tested in an RT inhibition assay. Nevirapine and efavirenz susceptibilities were tested and compared with a control wild-type virus or RT. Hypersusceptibility to nevirapine and efavirenz was detected using each of the methods described above. R values correlating the other methods with single-cycle assay values were between 0.66 and 0.96. In addition to the high correlations, the different methods gave similar numeric results. NNRTI hypersusceptibility is readily seen in multicycle susceptibility assays and in enzyme inhibition assays.

HIV type 1 genotypic variation in an antiretroviral treatment-naive population in southern India.

PubMed

Balakrishnan, Pachamuthu; Kumarasamy, Nagalingeswaran; Kantor, Rami; Solomon, Suniti; Vidya, Sundararajan; Mayer, Kenneth H; Newstein, Michael; Thyagarajan, Sadras P; Katzenstein, David; Ramratnam, Bharat

2005-04-01

Most studies of HIV-1 drug resistance have examined subtype B viruses; fewer data are available from developing countries, where non-B subtypes predominate. We determined the prevalence of mutations at protease and reverse transcriptase drug resistance positions in antiretroviral drug-naive individuals in southern India. The pol region of the genome was amplified from plasma HIV-1 RNA in 50 patients. All sequences clustered with HIV-1 subtype C. All patients had at least one protease and/or RT mutation at a known subtype B drug resistance position. Twenty percent of patients had mutations at major protease inhibitor resistance positions and 100% had mutations at minor protease inhibitor resistance positions. Six percent and 14% of patients had mutations at nucleoside reverse transcriptase inhibitor and/or nonnucleoside reverse transcriptase inhibitor resistance positions, respectively. Larger scale studies need to be undertaken to better define the genotypic variation of circulating Indian subtype C viruses and their potential impact on drug susceptibility and clinical outcome in treated individuals.

Unconventional plasticity of HIV-1 reverse transcriptase: how inhibitors could open a connection "gate" between allosteric and catalytic sites.

PubMed

Bellucci, Luca; Angeli, Lucilla; Tafi, Andrea; Radi, Marco; Botta, Maurizio

2013-12-23

Targeted molecular dynamics (TMD) simulations allowed for identifying the chemical/structural features of the nucleotide-competitive HIV-1 inhibitor DAVP-1, which is responsible for the disruption of the T-shape motif between Try183 and Trp229 of the reverse transcriptase (RT). DAVP-1 promoted the opening of a connection "gate" between allosteric and catalytic sites of HIV-1 RT, thus explaining its peculiar mechanism of action and providing useful insights to develop novel nucleotide-competitive RT inhibitors.

The public health approach to identify antiretroviral therapy failure: high-level nucleoside reverse transcriptase inhibitor resistance among Malawians failing first-line antiretroviral therapy

PubMed Central

Hosseinipour, Mina C.; van Oosterhout, Joep J.G.; Weigel, Ralf; Phiri, Sam; Kamwendo, Debbie; Parkin, Neil; Fiscus, Susan A.; Nelson, Julie A.E.; Eron, Joseph J.; Kumwenda, Johnstone

2010-01-01

Background Over 150 000 Malawians have started antiretroviral therapy (ART), in which first-line therapy is stavudine/lamivudine/nevirapine. We evaluated drug resistance patterns among patients failing first-line ART on the basis of clinical or immunological criteria in Lilongwe and Blantyre, Malawi. Methods Patients meeting the definition of ART failure (new or progressive stage 4 condition, CD4 cell count decline more than 30%, CD4 cell count less than that before treatment) from January 2006 to July 2007 were evaluated. Among those with HIV RNA of more than 1000 copies/ml, genotyping was performed. For complex genotype patterns, phenotyping was performed. Results Ninety-six confirmed ART failure patients were identified. Median (interquartile range) CD4 cell count, log10 HIV-1 RNA, and duration on ART were 68 cells/μl (23–174), 4.72 copies/ml (4.26–5.16), and 36.5 months (26.6–49.8), respectively. Ninety-three percent of samples had nonnucleoside reverse transcriptase inhibitor mutations, and 81% had the M184V mutation. The most frequent pattern included M184V and nonnucleoside reverse transcriptase inhibitor mutations along with at least one thymidine analog mutation (56%). Twenty-three percent of patients acquired the K70E or K65R mutations associated with tenofovir resistance; 17% of the patients had pan-nucleoside resistance that corresponded to K65R or K70E and additional resistance mutations, most commonly the 151 complex. Emergence of the K65R and K70E mutations was associated with CD4 cell count of less than 100 cells/μl (odds ratio 6.1) and inversely with the use of zidovudine (odds ratio 0.18). Phenotypic susceptibility data indicated that the nucleoside reverse transcriptase inhibitor backbone with the highest activity for subsequent therapy was zidovudine/lamivudine/tenofovir, followed by lamivudine/tenofovir, and then abacavir/didanosine. Conclusion When clinical and CD4 cell count criteria are used to monitor first-line ART failure, extensive nucleoside reverse transcriptase inhibitor and nonnucleoside reverse transcriptase inhibitor resistance emerges, with most patients having resistance profiles that markedly compromise the activity of second-line ART. PMID:19417582

Synthesis, Activity and Structural Analysis of Novel α-Hydroxytropolone Inhibitors of Human Immunodeficiency Virus Reverse Transcriptase-Associated Ribonuclease H

PubMed Central

Chung, Suhman; Himmel, Daniel M.; Jiang, Jian-Kang; Wojtak, Krzysztof; Bauman, Joseph D.; Rausch, Jason W.; Wilson, Jennifer A.; Beutler, John A.; Thomas, Craig J.; Arnold, Eddy; Le Grice, Stuart F.J.

2011-01-01

The α-hydroxytroplone, manicol (5,7-dihydroxy-2-isopropenyl-9-methyl-1,2,3,4-tetrahydro-benzocyclohepten-6-one) potently and specifically inhibits ribonuclease H (RNase H) activity of human immunodeficiency virus reverse transcriptase (HIV RT) in vitro. However, manicol was ineffective in reducing virus replication in culture. Ongoing efforts to improve the potency and specificity over the lead compound led us to synthesize 14 manicol derivatives that retain the divalent metal-chelating α-hydroxytropolone pharmacophore. These efforts were augmented by a high resolution structure of p66/p51 HIV-1 RT containing the nonnucleoside reverse transcriptase inhibitor (NNRTI), TMC278 and manicol in the DNA polymerase and RNase H active sites, respectively. We demonstrate here that several modified α-hydroxytropolones exhibit antiviral activity at non-cytotoxic concentrations. Inclusion of RNase H active site mutants indicated that manicol analogs can occupy an additional site in or around the DNA polymerase catalytic center. Collectively, our studies will promote future structure-based design of improved α-hydroxytropolones to complement the NRTI and NNRTI currently in clinical use. PMID:21568335

Recent advances in the development of next generation non-nucleoside reverse transcriptase inhibitors.

PubMed

Tarby, Christine M

2004-01-01

Since their discovery, non-nucleoside reverse transcriptase inhibitors (NNRTIs) have become one of the cornerstones of highly active anti-retroviral therapy (HAART). Currently, three NNRTI agents, efavirenz, nevirapine and delavirdine are commercially available. Efavirenz and nevirapine, used in combination with nucleoside reverse transcriptase inhibitors (NRTIs), provide durable regimens with efficacy comparable to protease inhibitor (PI) containing therapies. When virological failure occurs following treatment with an NNRTI, the resistance mutations can confer reduced sensitivity to the entire agent class. Therefore, the strategy for the development of next generation NNRTIs has been to focus on compounds which have improved potencies against the clinically relevant viral mutants. Agents with improved virological profiles and which maintain the ease of administration and favorable safety profiles of the current agents should find use in anti-retroviral naïve patients as well as in components of salvage regimens in the anti-retroviral experienced patient. This review summarizes the recent developments with compounds in clinical trials as of January 2002 as well as to summarize information on new agents appearing in the primary and patent literature between January 2001 and December 2002.

Update on HIV-1 acquired and transmitted drug resistance in Africa.

PubMed

Ssemwanga, Deogratius; Lihana, Raphael W; Ugoji, Chinenye; Abimiku, Alash'le; Nkengasong, John; Dakum, Patrick; Ndembi, Nicaise

2015-01-01

The last ten years have witnessed a significant scale-up and access to antiretroviral therapy in Africa, which has improved patient quality of life and survival. One major challenge associated with increased access to antiretroviral therapy is the development of antiretroviral resistance due to inconsistent drug supply and/or poor patient adherence. We review the current state of both acquired and transmitted drug resistance in Africa over the past ten years (2001-2011) to identify drug resistance associated with the different drug regimens used on the continent and to help guide affordable strategies for drug resistance surveillance. A total of 161 references (153 articles, six reports and two conference abstracts) were reviewed. Antiretroviral resistance data was available for 40 of 53 African countries. A total of 5,541 adult patients from 99 studies in Africa were included in this analysis. The pooled prevalence of drug resistance mutations in Africa was 10.6%, and Central Africa had the highest prevalence of 54.9%. The highest prevalence of nucleoside reverse transcriptase inhibitor mutations was in the west (55.3%) and central (54.8%) areas; nonnucleoside reverse transcriptase inhibitor mutations were highest in East Africa (57.0%) and protease inhibitors mutations highest in Southern Africa (16.3%). The major nucleoside reverse transcriptase inhibitor mutation in all four African regions was M184V. Major nonnucleoside reverse transcriptase inhibitor as well as protease inhibitor mutations varied by region. The prevalence of drug resistance has remained low in several African countries although the emergence of drug resistance mutations varied across countries. Continued surveillance of antiretroviral therapy resistance remains crucial in gauging the effectiveness of country antiretroviral therapy programs and strategizing on effective and affordable strategies for successful treatment.

In Vitro Evaluation of Nonnucleoside Reverse Transcriptase Inhibitors UC-781 and TMC120-R147681 as Human Immunodeficiency Virus Microbicides†

PubMed Central

Van Herrewege, Yven; Michiels, Jo; Van Roey, Jens; Fransen, Katrien; Kestens, Luc; Balzarini, Jan; Lewi, Paul; Vanham, Guido; Janssen, Paul

2004-01-01

The nonnucleoside reverse transcriptase inhibitors UC-781 and TMC120-R147681 (Dapivirine) effectively prevented human immunodeficiency virus (HIV) infection in cocultures of monocyte-derived dendritic cells and T cells, representing primary targets in sexual transmission. Both drugs had a favorable therapeutic index. A 24-h treatment with 1,000 nM UC-781 or 100 nM TMC120-R147681 prevented cell-free HIV infection, whereas 10-fold-higher concentrations blocked cell-associated HIV. PMID:14693562

Antiretroviral Drug Use in a Cross-Sectional Population Survey in Africa: NIMH Project Accept (HPTN 043).

PubMed

Fogel, Jessica M; Clarke, William; Kulich, Michal; Piwowar-Manning, Estelle; Breaud, Autumn; Olson, Matthew T; Marzinke, Mark A; Laeyendecker, Oliver; Fiamma, Agnès; Donnell, Deborah; Mbwambo, Jessie K K; Richter, Linda; Gray, Glenda; Sweat, Michael; Coates, Thomas J; Eshleman, Susan H

2017-02-01

Antiretroviral (ARV) drug treatment benefits the treated individual and can prevent HIV transmission. We assessed ARV drug use in a community-randomized trial that evaluated the impact of behavioral interventions on HIV incidence. Samples were collected in a cross-sectional survey after a 3-year intervention period. ARV drug testing was performed using samples from HIV-infected adults at 4 study sites (Zimbabwe; Tanzania; KwaZulu-Natal and Soweto, South Africa; survey period 2009-2011) using an assay that detects 20 ARV drugs (6 nucleoside/nucleotide reverse transcriptase inhibitors, 3 nonnucleoside reverse transcriptase inhibitors, and 9 protease inhibitors; maraviroc; raltegravir). ARV drugs were detected in 2011 (27.4%) of 7347 samples; 88.1% had 1 nonnucleoside reverse transcriptase inhibitors ± 1-2 nucleoside/nucleotide reverse transcriptase inhibitors. ARV drug detection was associated with sex (women>men), pregnancy, older age (>24 years), and study site (P < 0.0001 for all 4 variables). ARV drugs were also more frequently detected in adults who were widowed (P = 0.006) or unemployed (P = 0.02). ARV drug use was more frequent in intervention versus control communities early in the survey (P = 0.01), with a significant increase in control (P = 0.004) but not in intervention communities during the survey period. In KwaZulu-Natal, a 1% increase in ARV drug use was associated with a 0.14% absolute decrease in HIV incidence (P = 0.018). This study used an objective, biomedical approach to assess ARV drug use on a population level. This analysis identified factors associated with ARV drug use and provided information on ARV drug use over time. ARV drug use was associated with lower HIV incidence at 1 study site.

Nelfinavir and non-nucleoside reverse transcriptase inhibitor-based salvage regimens in heavily HIV pretreated patients

PubMed Central

Baril, Jean-Guy; Lefebvre, Eric A; Lalonde, Richard G; Shafran, Stephen D; Conway, Brian

2003-01-01

OBJECTIVE: To assess the efficacy of nelfinavir mesylate (NFV) in combination with delavirdine mesylate (DLV) or efavirenz (EFV) and other antiretroviral agents following virological failure on other protease inhibitor (PI)-based regimens. DESIGN: Multicentre, retrospective chart review. METHODS: One hundred-one patients who were naive to both NFV and non-nucleoside reverse transcriptase inhibitors (NNRTIs) and who initiated NFV plus DLV or EFV-based salvage regimens were reviewed. Response to treatment was defined as a reduction in HIV ribonucleic acid (RNA) levels to unquantifiable levels (less than 50 copies/mL, less than 400 copies/mL, less than 500 copies/mL) on at least one occasion after the initiation of salvage therapy. Baseline correlates of response, including prior duration of HIV infection, prior number of regimens, viral load and CD4 cell counts were also evaluated. RESULTS: Patients had a mean duration of HIV infection of 10 years, a mean duration of prior therapy of four years, a median of four prior nucleoside reverse transcriptase inhibitors and a median of two prior PIs. At the time of review the mean duration of salvage therapy was 63.4 weeks. Virological suppression was achieved in 59 (58.4%) patients within a mean of eight weeks and maintained for a mean of 44.9 weeks (the mean follow-up was 78 weeks). Of the non-responders, 16 (38%) achieved a less than 1 log10 decrease in HIV RNA levels. Although there was no association between baseline correlates, response rate (75.7%) was significantly higher in patients with HIV RNA levels of 50,000 copies/mL or lower and CD4 counts greater than 200 cells/mm3. CONCLUSION: NFV/NNRTI-based highly active antiretroviral therapy regimens are an effective therapy in many patients who have experienced virological breakthroughs on at least one prior PI-based regimen. PMID:18159457

Antitumor effect of combination of the inhibitors of two new oncotargets: proton pumps and reverse transcriptase.

PubMed

Lugini, Luana; Sciamanna, Ilaria; Federici, Cristina; Iessi, Elisabetta; Spugnini, Enrico Pierluigi; Fais, Stefano

2017-01-17

Tumor therapy needs new approaches in order to improve efficacy and reduce toxicity of the current treatments. The acidic microenvironment and the expression of high levels of endogenous non-telomerase reverse transcriptase (RT) are common features of malignant tumor cells. The anti-acidic proton pump inhibitor Lansoprazole (LAN) and the non-nucleoside RT inhibitor Efavirenz (EFV) have shown independent antitumor efficacy. LAN has shown to counteract drug tumor resistance. We tested the hypothesis that combination of LAN and EFV may improve the overall antitumor effects. We thus pretreated human metastatic melanoma cells with LAN and then with EFV, both in 2D and 3D spheroid models. We evaluated the treatment effect by proliferation and cell death/apoptosis assays in classical and in pulse administration experiments. The action of EFV was negatively affected by the tumor microenvironmental acidity, and LAN pretreatment overcame the problem. LAN potentiated the cytotoxicity of EFV to melanoma cells and, when administered during the drug interruption period, prevented the replacement of tumor cell growth.This study supports the implementation of the current therapies with combination of Proton Pumps and Reverse Transcriptase inhibitors.

Antitumor effect of combination of the inhibitors of two new oncotargets: proton pumps and reverse transcriptase

PubMed Central

Lugini, Luana; Sciamanna, Ilaria; Federici, Cristina; Iessi, Elisabetta; Spugnini, Enrico Pierluigi; Fais, Stefano

2017-01-01

Tumor therapy needs new approaches in order to improve efficacy and reduce toxicity of the current treatments. The acidic microenvironment and the expression of high levels of endogenous non-telomerase reverse transcriptase (RT) are common features of malignant tumor cells. The anti-acidic proton pump inhibitor Lansoprazole (LAN) and the non-nucleoside RT inhibitor Efavirenz (EFV) have shown independent antitumor efficacy. LAN has shown to counteract drug tumor resistance. We tested the hypothesis that combination of LAN and EFV may improve the overall antitumor effects. We thus pretreated human metastatic melanoma cells with LAN and then with EFV, both in 2D and 3D spheroid models. We evaluated the treatment effect by proliferation and cell death/apoptosis assays in classical and in pulse administration experiments. The action of EFV was negatively affected by the tumor microenvironmental acidity, and LAN pretreatment overcame the problem. LAN potentiated the cytotoxicity of EFV to melanoma cells and, when administered during the drug interruption period, prevented the replacement of tumor cell growth. This study supports the implementation of the current therapies with combination of Proton Pumps and Reverse Transcriptase inhibitors. PMID:27926505

Do non-nucleoside reverse transcriptase inhibitors contribute to lipodystrophy?

PubMed

Nolan, David

2005-01-01

Lipodystrophy complications, including lipoatrophy (pathological fat loss) and metabolic complications, have emerged as important long-term toxicities associated with antiretroviral therapy in the current era. The wealth of data that has accumulated over the past 6 years has now clarified the contribution of specific antiretroviral drugs to the risk of these clinical endpoints, with evidence that lipoatrophy is strongly associated with the choice of nucleoside reverse transcriptase inhibitor therapy (specifically, stavudine and to a lesser extent zidovudine). The aetiological basis of metabolic complications of antiretroviral therapy has proven to be complex, in that the risk appears to be modulated by a number of lifestyle factors that have made the metabolic syndrome highly prevalent in the general population, with additional contributions from HIV disease status itself, as well as from individual drugs within the HIV protease inhibitor class. The currently licensed non-nucleoside reverse transcriptase inhibitor (NNRTI) drugs, efavirenz and nevirapine, have been proven to have a favourable safety profile in terms of lipodystrophy complications. However, it must be noted that NNRTI drugs also have individual toxicity profiles that must be accounted for when considering and/or monitoring their use in the treatment of HIV infection.

Pharmacophore Identification, Molecular Docking, Virtual Screening, and In Silico ADME Studies of Non-Nucleoside Reverse Transcriptase Inhibitors.

PubMed

Pirhadi, Somayeh; Ghasemi, Jahan B

2012-12-01

Non-nucleoside reverse transcriptase inhibitors (NNRTIs) have gained a definitive place due to their unique antiviral potency, high specificity and low toxicity in antiretroviral combination therapies used to treat HIV. In this study, chemical feature based pharmacophore models of different classes of NNRT inhibitors of HIV-1 have been developed. The best HypoRefine pharmacophore model, Hypo 1, which has the best correlation coefficient (0.95) and the lowest RMS (0.97), contains two hydrogen bond acceptors, one hydrophobic and one ring aromatic feature, as well as four excluded volumes. Hypo 1 was further validated by test set and Fischer validation method. The best pharmacophore model was then utilized as a 3D search query to perform a virtual screening to retrieve potential inhibitors. The hit compounds were subsequently subjected to filtering by Lipinski's rule of five and docking studies by Libdock and Gold methods to refine the retrieved hits. Finally, 7 top ranked compounds based on Gold score fitness function were subjected to in silico ADME studies to investigate for compliance with the standard ranges. Copyright © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

High Potency of Indolyl Aryl Sulfone Nonnucleoside Inhibitors towards Drug-Resistant Human Immunodeficiency Virus Type 1 Reverse Transcriptase Mutants Is Due to Selective Targeting of Different Mechanistic Forms of the Enzyme

PubMed Central

Cancio, Reynel; Silvestri, Romano; Ragno, Rino; Artico, Marino; De Martino, Gabriella; La Regina, Giuseppe; Crespan, Emmanuele; Zanoli, Samantha; Hübscher, Ulrich; Spadari, Silvio; Maga, Giovanni

2005-01-01

Indolyl aryl sulfone (IAS) nonnucleoside inhibitors have been shown to potently inhibit the growth of wild-type and drug-resistant human immunodeficiency virus type 1 (HIV-1), but their exact mechanism of action has not been elucidated yet. Here, we describe the mechanism of inhibition of HIV-1 reverse transcriptase (RT) by selected IAS derivatives. Our results showed that, depending on the substitutions introduced in the IAS common pharmacophore, these compounds can be made selective for different enzyme-substrate complexes. Moreover, we showed that the molecular basis for this selectivity was a different association rate of the drug to a particular enzymatic form along the reaction pathway. By comparing the activities of the different compounds against wild-type RT and the nonnucleoside reverse transcriptase inhibitor-resistant mutant Lys103Asn, it was possible to hypothesize, on the basis of their mechanism of action, a rationale for the design of drugs which could overcome the steric barrier imposed by the Lys103Asn mutation. PMID:16251294

Polyurethane intravaginal ring for controlled delivery of dapivirine, a nonnucleoside reverse transcriptase inhibitor of HIV-1.

PubMed

Gupta, Kavita M; Pearce, Serena M; Poursaid, Azadeh E; Aliyar, Hyder A; Tresco, Patrick A; Mitchnik, Mark A; Kiser, Patrick F

2008-10-01

Women-controlled methods for prevention of male-to-female sexual transmission of HIV-1 are urgently needed. Providing inhibitory concentrations of HIV-1 reverse transcriptase inhibitors to impede the replication of the virus in the female genital tissue offers a mechanism for prophylaxis of HIV-1. To this end, an intravaginal ring device that can provide long duration delivery of dapivirine, a nonnucleoside reverse transcriptase inhibitor of HIV-1, was developed utilizing a medical-grade polyether urethane. Monolithic intravaginal rings were fabricated and sustained release with cumulative flux linear with time was demonstrated under sink conditions for a period of 30 days. The release rate was directly proportional to the amount of drug loaded. Another release study conducted for a week utilizing liposome dispersions as sink conditions, to mimic the partitioning of dapivirine into vaginal tissue, also demonstrated release rates constant with time. These results qualify polyether urethanes for development of intravaginal rings for sustained delivery of microbicidal agents. (c) 2008 Wiley-Liss, Inc. and the American Pharmacists Association

Tenofovir-related nephrotoxicity: case report and review of the literature.

PubMed

James, Christopher W; Steinhaus, Mary C; Szabo, Susan; Dressier, Robert M

2004-03-01

Tenofovir is a nucleotide reverse transcriptase inhibitor for treatment of human immunodeficiency virus (HIV) infection. Several cases of renal failure associated with tenofovir therapy recently have been reported. A 54-year-old man with HIV experienced decreasing renal function and Fanconi's syndrome secondary to tenofovir therapy. His condition gradually improved after discontinuation of the drug. The available medical literature for reported cases of tenofovir-related nephrotoxicity indicates that this complication is apparently rare. However, our case report and literature review underscore the importance of monitoring renal function when treating patients with any nucleotide reverse transcriptase inhibitor.

Structural investigation of HIV-1 nonnucleoside reverse transcriptase inhibitors: 2-Aryl-substituted benzimidazoles

NASA Astrophysics Data System (ADS)

Ziółkowska, Natasza E.; Michejda, Christopher J.; Bujacz, Grzegorz D.

2009-11-01

Acquired immunodeficiency syndrome (AIDS) caused by the human immunodeficiency virus (HIV) is one of the most destructive epidemics in history. Inhibitors of HIV enzymes are the main targets to develop drugs against that disease. Nonnucleoside reverse transcriptase inhibitors of HIV-1 (NNRTIs) are potentially effective and nontoxic. Structural studies provide information necessary to design more active compounds. The crystal structures of four NNRTI derivatives of 2-aryl-substituted N-benzyl-benzimidazole are presented here. Analysis of the geometrical parameters shows that the structures of the investigated inhibitors are rigid. The important geometrical parameter is the dihedral angle between the planes of the π-electron systems of the benzymidazole and benzyl moieties. The values of these dihedral angles are in a narrow range for all investigated inhibitors. There is no significant difference between the structure of the free inhibitor and the inhibitor in the complex with RT HIV-1. X-ray structures of the investigated inhibitors are a good basis for modeling enzyme-inhibitor interactions in rational drug design.

Pharmacokinetics of antiretroviral drugs in anatomical sanctuary sites: the male and female genital tract.

PubMed

Else, Laura J; Taylor, Stephen; Back, David J; Khoo, Saye H

2011-01-01

HIV resides within anatomical 'sanctuary sites', where local drug exposure and viral dynamics may differ significantly from the systemic compartment. Suboptimal antiretroviral concentrations in the genital tract may result in compartmentalized viral replication, selection of resistant mutations and possible re-entry of wild-type/resistant virus into the systemic circulation. Therefore, achieving adequate antiretroviral exposure in the genital tract has implications for the prevention of sexual and vertical transmission of HIV. Penetration of antiretrovirals in the genital tract is expressed by accumulation ratios derived from the measurement of drug concentrations in time-matched seminal plasma/cervicovaginal fluid and plasma samples. Penetration varies by gender and may be drug (as opposed to class) specific with high interindividual variability. Concentrations in seminal plasma are highest for nucleoside analogues and lowest for protease inhibitors and efavirenz. Seminal accumulation of newer agents, raltegravir and maraviroc, is moderate (rank order of accumulation is nucleoside/nucleotide reverse transcriptase inhibitors [lamivudine/zidovudine/tenofovir/didanosine > stavudine/abacavir] > raltegravir > indinavir/maraviroc/nevirapine > efavirenz/protease inhibitors [amprenavir/atazanavir/darunavir > lopinavir/ritonavir > saquinavir] > enfuvirtide). In the female genital tract, the nucleoside analogues exhibit high accumulation ratios, whereas protease inhibitors have limited penetration; however, substantial variability exists between individuals and study centres. Second generation non-nucleoside reverse transcriptase inhibitor etravirine, and maraviroc and raltegravir, demonstrate effective accumulation in cervicovaginal secretions (rank order of accumulation is nucleoside/nucleotide reverse transcriptase inhibitor [zidovudine/lamivudine/didanosine > emtricitabine/tenofovir] > indinavir > maraviroc/raltegravir/darunavir/etravirine > nevirapine/abacavir > protease inhibitors [amprenavir/atazanavir/ritonavir] > lopinavir/stavudine/efavirenz > saquinavir).

Crystallographic Study of a Novel Sub-Nanomolar Inhibitor Provides Insight on the Binding Interactions of Alkenyldiarylmethanes with Human Immunodeficiency Virus-1 (HIV-1) Reverse Transcriptase†

PubMed Central

Cullen, Matthew D.; Ho, William C.; Bauman, Joseph D.; Das, Kalyan; Arnold, Eddy; Hartman, Tracy L.; Watson, Karen M.; Buckheit, Robert W.; Pannecouque, Christophe; De Clercq, Erik; Cushman, Mark

2009-01-01

Two crystal structures have been solved for separate complexes of alkenyldiarylmethane (ADAM) non-nucleoside reverse transcriptase inhibitors (NNRTI) 3 and 4 with HIV-1 reverse transcriptase (RT). The structures reveal inhibitor binding is exclusively hydrophobic in nature and the shape of the inhibitor-bound NNRTI binding pocket is unique among other reported inhibitor-RT crystal structures. Primarily, ADAMs 3 and 4 protrude from a large gap in the backside of the binding pocket, placing portions of the inhibitors unusually close to the polymerase active site and allowing 3 to form a weak hydrogen bond with Lys223. The lack of additional stabilizing interactions, beyond the observed hydrophobic surface contacts, between 4 and RT is quite perplexing given the extreme potency of the compound (IC50 ≤ nM). ADAM 4 was designed to be hydrolytically stable in blood plasma, and an investigation of its hydrolysis in rat plasma demonstrated it has a significantly prolonged half-life in comparison to ADAM lead compounds 1 and 2. PMID:19775161

Probing the molecular mechanism of action of the HIV-1 reverse transcriptase inhibitor 4'-ethynyl-2-fluoro-2'-deoxyadenosine (EFdA) using pre-steady-state kinetics.

PubMed

Muftuoglu, Yagmur; Sohl, Christal D; Mislak, Andrea C; Mitsuya, Hiroaki; Sarafianos, Stefan G; Anderson, Karen S

2014-06-01

The novel antiretroviral 4'-ethynyl-2-fluoro-2'-deoxyadenosine (EFdA) is a potent nucleoside HIV-1 reverse transcriptase (RT) inhibitor (NRTI). Unlike other FDA-approved NRTIs, EFdA contains a 3'-hydroxyl. Pre-steady-state kinetics showed RT preferred incorporating EFdA-TP over native dATP. Moreover, RT slowly inserted nucleotides past an EFdA-terminated primer, resulting in delayed chain termination with unaffected fidelity. This is distinct from KP1212, another 3'-hydroxyl-containing RT inhibitor considered to promote viral lethal mutagenesis. New mechanistic features of RT inhibition by EFdA are revealed. Copyright © 2014 Elsevier B.V. All rights reserved.

Probing the molecular mechanism of action of the HIV-1 reverse transcriptase inhibitor 4′-ethynyl-2-fluoro-2′-deoxyadenosine (EFdA) using pre-steady-state kinetics

PubMed Central

Muftuoglu, Yagmur; Sohl, Christal D.; Mislak, Andrea C.; Mitsuya, Hiroaki; Sarafianos, Stefan G.; Anderson, Karen S.

2014-01-01

The novel antiretroviral 4′-ethynyl-2-fluoro-2′-deoxyadenosine (EFdA) is a potent nucleoside HIV-1 reverse transcriptase (RT) inhibitor (NRTI). Unlike other FDA-approved NRTIs, EFdA contains a 3′-hydroxyl. Pre-steady-state kinetics showed RT preferred incorporating EFdA-TP over native dATP. Moreover, RT slowly inserted nucleotides past an EFdA-terminated primer, resulting in delayed chain termination with unaffected fidelity. This is distinct from KP1212, another 3′-hydroxyl-containing RT inhibitor considered to promote viral lethal mutagenesis. New mechanistic features of RT inhibition by EFdA are revealed. PMID:24632447

Susceptibility of recombinant porcine endogenous retrovirus reverse transcriptase to nucleoside and non-nucleoside inhibitors.

PubMed

Wilhelm, M; Fishman, J A; Pontikis, R; Aubertin, A M; Wilhelm, F X

2002-12-01

Transplantation of organs, tissues or cells from pigs to humans could be a potential solution to the shortage of human organs for transplantation. Porcine endogenous retroviruses (PERVs) remain a major safety concern for porcine xenotransplantation. Thus, finding drugs that could be used as virological prophylaxis (or therapy) against PERV replication would be desirable. One of the most effective ways to block retroviral multiplication is to inhibit the enzyme reverse transcriptase (RT) which catalyzes the reverse transcription of viral RNA to proviral double-stranded DNA. We report here the cloning and expression of PERV RT and its susceptibility to several inhibitors. Our data demonstrate PERV susceptibility in vitro to the triphosphorylated nucleoside analog of zidovudine (AZT) and to ddGTP and to a lesser extent to ddTTP but almost no susceptibility to the non-nucleoside RT inhibitors tested.

Fragment Based Strategies for Discovery of Novel HIV-1 Reverse Transcriptase and Integrase Inhibitors.

PubMed

Latham, Catherine F; La, Jennifer; Tinetti, Ricky N; Chalmers, David K; Tachedjian, Gilda

2016-01-01

Human immunodeficiency virus (HIV) remains a global health problem. While combined antiretroviral therapy has been successful in controlling the virus in patients, HIV can develop resistance to drugs used for treatment, rendering available drugs less effective and limiting treatment options. Initiatives to find novel drugs for HIV treatment are ongoing, although traditional drug design approaches often focus on known binding sites for inhibition of established drug targets like reverse transcriptase and integrase. These approaches tend towards generating more inhibitors in the same drug classes already used in the clinic. Lack of diversity in antiretroviral drug classes can result in limited treatment options, as cross-resistance can emerge to a whole drug class in patients treated with only one drug from that class. A fresh approach in the search for new HIV-1 drugs is fragment-based drug discovery (FBDD), a validated strategy for drug discovery based on using smaller libraries of low molecular weight molecules (<300 Da) screened using primarily biophysical assays. FBDD is aimed at not only finding novel drug scaffolds, but also probing the target protein to find new, often allosteric, inhibitory binding sites. Several fragment-based strategies have been successful in identifying novel inhibitory sites or scaffolds for two proven drug targets for HIV-1, reverse transcriptase and integrase. While any FBDD-generated HIV-1 drugs have yet to enter the clinic, recent FBDD initiatives against these two well-characterised HIV-1 targets have reinvigorated antiretroviral drug discovery and the search for novel classes of HIV-1 drugs.

Prevalence of HIV-1 Subtypes and Drug Resistance-Associated Mutations in HIV-1-Positive Treatment-Naive Pregnant Women in Pointe Noire, Republic of the Congo (Kento-Mwana Project).

PubMed

Bruzzone, Bianca; Saladini, Francesco; Sticchi, Laura; Mayinda Mboungou, Franc A; Barresi, Renata; Caligiuri, Patrizia; Calzi, Anna; Zazzi, Maurizio; Icardi, Giancarlo; Viscoli, Claudio; Bisio, Francesca

2015-08-01

The Kento-Mwana project was carried out in Pointe Noire, Republic of the Congo, to prevent mother-to-child HIV-1 transmission. To determine the prevalence of different subtypes and transmitted drug resistance-associated mutations, 95 plasma samples were collected at baseline from HIV-1-positive naive pregnant women enrolled in the project during the years 2005-2008. Full protease and partial reverse transcriptase sequencing was performed and 68/95 (71.6%) samples were successfully sequenced. Major mutations to nucleoside reverse transcriptase inhibitors, nonnucleoside reverse transcriptase inhibitors, and protease inhibitors were detected in 4/68 (5.9%), 3/68 (4.4%), and 2/68 (2.9%) samples, respectively. Phylogenetic analysis of HIV-1 isolates showed a high prevalence of unique recombinant forms (24/68, 35%), followed by CRF45_cpx (7/68, 10.3%) and subsubtype A3 and subtype G (6/68 each, 8.8%). Although the prevalence of transmitted drug resistance mutations appears to be currently limited, baseline HIV-1 genotyping is highly advisable in conjunction with antiretroviral therapy scale-up in resource-limited settings to optimize treatment and prevent perinatal transmission.

Dolutegravir Plus Two Nucleoside Reverse Transcriptase Inhibitors versus Efavirenz Plus Two Nucleoside Reverse Transcriptase Inhibitors As Initial Antiretroviral Therapy for People with HIV: A Systematic Review.

PubMed

Rutherford, George W; Horvath, Hacsi

2016-01-01

Dolutegravir (DTG) is a once-daily unboosted second-generation integrase-inhibitor that along with two nucleoside reverse transcriptase inhibitors is one of several regimens recommended by the United States, United Kingdom and European Union for first-line antiretroviral treatment of people with HIV infection. Our objective was to review the evidence for the efficacy and safety of DTG-based first-line regimens compared to efavirenz (EFV)-based regimens. We conducted a systematic review. We comprehensively searched a range of databases as well as conference abstracts and a trials registry. We used Cochrane methods in screening and data collection and assessed each study's risk of bias with the Cochrane tool. We meta-analyzed data using a fixed-effects model. We used GRADE to assess evidence quality. From 492 search results, we identified two randomized controlled trials, reported in five peer-reviewed articles and one conference abstract. One trial tested two DTG-based regimens (DTG + abacavir (ABC) + lamivudine (3TC) or DTG + tenofovir + emtricitabine) against an EFV-based regimen (EFV+ ABC+3TC). The other trial tested DTG+ABC+3TC against EFV+ABC+3TC. In meta-analysis, DTG-containing regimens were superior to EFV-containing regimens at 48 weeks and at 96 weeks (RR = 1.10, 95% CI 1.04-1.16; and RR = 1.12, 95% CI 1.04-1.21, respectively). In one trial, the DTG-containing regimen was superior at 144 weeks (RR = 1.13, 95% CI 1.02-1.24). DTG-containing regimens were superior in reducing treatment discontinuation compared to those containing EFV at 96 weeks and at 144 weeks (RR = 0.27, 95% CI 0.15-0.50; and RR = 0.28, 95% CI 0.16-0.48, respectively). Risk of serious adverse events was similar in each regimen at 96 weeks (RR = 1.15, 95% CI 0.80-1.63) and 144 weeks (RR = 0.93, 95% CI 0.68-1.29). Risk of bias was moderate overall, as was GRADE evidence quality. DTG-based regimens should be considered in future World Health Organization guidelines for initial HIV treatment.

High Rates of Baseline Drug Resistance and Virologic Failure Among ART-naive HIV-infected Children in Mali.

PubMed

Crowell, Claudia S; Maiga, Almoustapha I; Sylla, Mariam; Taiwo, Babafemi; Kone, Niaboula; Oron, Assaf P; Murphy, Robert L; Marcelin, Anne-Geneviève; Traore, Ban; Fofana, Djeneba B; Peytavin, Gilles; Chadwick, Ellen G

2017-11-01

Limited data exist on drug resistance and antiretroviral treatment (ART) outcomes in HIV-1-infected children in West Africa. We determined the prevalence of baseline resistance and correlates of virologic failure (VF) in a cohort of ART-naive HIV-1-infected children <10 years of age initiating ART in Mali. Reverse transcriptase and protease genes were sequenced at baseline (before ART) and at 6 months. Resistance was defined according to the Stanford HIV Genotypic Resistance database. VF was defined as viral load ≥1000 copies/mL after 6 months of ART. Logistic regression was used to evaluate factors associated with VF or death >1 month after enrollment. Post hoc, antiretroviral concentrations were assayed on baseline samples of participants with baseline resistance. One-hundred twenty children with a median age 2.6 years (interquartile range: 1.6-5.0) were included. Eighty-eight percent reported no prevention of mother-to-child transmission exposure. At baseline, 27 (23%), 4 (3%) and none had non-nucleoside reverse transcriptase inhibitor (NNRTI), nucleoside reverse transcriptase inhibitor or protease inhibitor resistance, respectively. Thirty-nine (33%) developed VF and 4 died >1 month post-ART initiation. In multivariable analyses, poor adherence [odds ratio (OR): 6.1, P = 0.001], baseline NNRTI resistance among children receiving NNRTI-based ART (OR: 22.9, P < 0.001) and protease inhibitor-based ART initiation among children without baseline NNRTI resistance (OR: 5.8, P = 0.018) were significantly associated with VF/death. Ten (38%) with baseline resistance had detectable levels of nevirapine or efavirenz at baseline; 7 were currently breastfeeding, but only 2 reported maternal antiretroviral use. Baseline NNRTI resistance was common in children without reported NNRTI exposure and was associated with increased risk of treatment failure. Detectable NNRTI concentrations were present despite few reports of maternal/infant antiretroviral use.

Novel Method for Simultaneous Quantification of Phenotypic Resistance to Maturation, Protease, Reverse Transcriptase, and Integrase HIV Inhibitors Based on 3′Gag(p2/p7/p1/p6)/PR/RT/INT-Recombinant Viruses: a Useful Tool in the Multitarget Era of Antiretroviral Therapy▿†

PubMed Central

Weber, Jan; Vazquez, Ana C.; Winner, Dane; Rose, Justine D.; Wylie, Doug; Rhea, Ariel M.; Henry, Kenneth; Pappas, Jennifer; Wright, Alison; Mohamed, Nizar; Gibson, Richard; Rodriguez, Benigno; Soriano, Vicente; King, Kevin; Arts, Eric J.; Olivo, Paul D.; Quiñones-Mateu, Miguel E.

2011-01-01

Twenty-six antiretroviral drugs (ARVs), targeting five different steps in the life cycle of the human immunodeficiency virus type 1 (HIV-1), have been approved for the treatment of HIV-1 infection. Accordingly, HIV-1 phenotypic assays based on common cloning technology currently employ three, or possibly four, different recombinant viruses. Here, we describe a system to assess HIV-1 resistance to all drugs targeting the three viral enzymes as well as viral assembly using a single patient-derived, chimeric virus. Patient-derived p2-INT (gag-p2/NCp7/p1/p6/pol-PR/RT/IN) products were PCR amplified as a single fragment (3,428 bp) or two overlapping fragments (1,657 bp and 2,002 bp) and then recombined into a vector containing a near-full-length HIV-1 genome with the Saccharomyces cerevisiae uracil biosynthesis gene (URA3) replacing the 3,428 bp p2-INT segment (Dudley et al., Biotechniques 46:458–467, 2009). P2-INT-recombinant viruses were employed in drug susceptibility assays to test the activity of protease (PI), nucleoside/nucleotide reverse transcriptase (NRTI), nonnucleoside reverse transcriptase (NNRTI), and integrase strand-transfer (INSTI) inhibitors. Using a single standardized test (ViralARTS HIV), this new technology permits the rapid and automated quantification of phenotypic resistance for all known and candidate antiretroviral drugs targeting all viral enzymes (PR, RT, including polymerase and RNase H activities, and IN), some of the current and potential assembly inhibitors, and any drug targeting Pol or Gag precursor cleavage sites (relevant for PI and maturation inhibitors) This novel assay may be instrumental (i) in the development and clinical assessment of novel ARV drugs and (ii) to monitor patients failing prior complex treatment regimens. PMID:21628544

Synthesis and evaluation of "AZT-HEPT", "AZT-pyridinone", and "ddC-HEPT" conjugates as inhibitors of HIV reverse transcriptase.

PubMed

Pontikis, R; Dollé, V; Guillaumel, J; Dechaux, E; Note, R; Nguyen, C H; Legraverend, M; Bisagni, E; Aubertin, A M; Grierson, D S; Monneret, C

2000-05-18

To test the concept that HIV reverse transcriptase could be effectively inhibited by "mixed site inhibitors", a series of seven conjugates containing both a nucleoside analogue component (AZT 1, ddC 2) and a nonnucleoside type inhibitor (HEPT analogue 12, pyridinone 27) were synthesized and evaluated for their ability to block HIV replication. The (N-3 and C-5)AZT-HEPT conjugates 15, 22, and 23 displayed 2-5 microM anti-HIV activity, but they had no effect on the replication of HIV-2 or the HIV-1 strain with the Y181C mutation. The (C-5)AZT-pyridinone conjugates 34-37 were found to be inactive. In marked contrast, the ddC-HEPT molecule 26 displayed the same potency (EC(50) = 0.45 microM) against HIV-1 (wild type and the Y181C nevirapine-resistant strain) and HIV-2 in cell culture. No synergistic effect was observed for these bis-substrate inhibitors, suggesting that the two individual inhibitor components in these molecules do not bind simultaneously in their respective sites. Interestingly, however, the results indicate that the AZT-HEPT conjugates and the ddC-HEPT derivative 26 inhibit reverse transcriptase (RT) in an opposite manner. One explanation for this difference is that the former compounds interact preferentially with the hydrophobic pocket in RT, whereas 26 (after supposed triphosphorylation) inhibits RT through binding in the catalytic site.

Simultaneous determination of the HIV nucleoside analogue reverse transcriptase inhibitors lamivudine, didanosine, stavudine, zidovudine and abacavir in human plasma by reversed phase high performance liquid chromatography.

PubMed

Verweij-van Wissen, C P W G M; Aarnoutse, R E; Burger, D M

2005-02-25

A reversed phase high performance liquid chromatography method was developed for the simultaneous quantitative determination of the nucleoside reverse transcriptase inhibitors (NRTIs) lamivudine, didanosine, stavudine, zidovudine and abacavir in plasma. The method involved solid-phase extraction with Oasis MAX cartridges from plasma, followed by high performance liquid chromatography with a SymmetryShield RP 18 column and ultraviolet detection set at a wavelength of 260 nm. The assay was validated over the concentration range of 0.015-5 mg/l for all five NRTIs. The average accuracies for the assay were 92-102%, inter- and intra-day coefficients of variation (CV) were <2.5% and extraction recoveries were higher than 97%. This method proved to be simple, accurate and precise, and is currently in use in our laboratory for the quantitative analysis of NRTIs in plasma.

[Thyroid dysfunction in adults infected by human immunodeficiency virus].

PubMed

Abelleira, Erika; De Cross, Graciela A; Pitoia, Fabián

2014-01-01

Patients infected with human immunodeficiency virus (HIV) have a higher prevalence of thyroid dysfunction when compared with the general population. The most frequently observed manifestations are euthyroid sick syndrome, Graves' disease and subclinical hypothyroidism. The relationship between the use of highly active antiretroviral therapy and the increased prevalence of thyroid dysfunction has been demonstrated in several series of patients. Grave's disease is recognized as a consequence of immune restitution syndrome. Besides, several studies have suggested an association between hypothyroidism and the use of nucleoside reverse transcriptase inhibitors, particularly stavudine and non-nucleoside reverse transcriptase inhibitors such as efavirenz. Further studies could provide additional evidence of the need for routine assessment of thyroid function in HIV-infected patients.

In search of a treatment for HIV--current therapies and the role of non-nucleoside reverse transcriptase inhibitors (NNRTIs).

PubMed

Reynolds, Chevonne; de Koning, Charles B; Pelly, Stephen C; van Otterlo, Willem A L; Bode, Moira L

2012-07-07

The human immunodeficiency virus (HIV) causes AIDS (acquired immune deficiency syndrome), a disease in which the immune system progressively deteriorates, making sufferers vulnerable to all manner of opportunistic infections. Currently, world-wide there are estimated to be 34 million people living with HIV, with the vast majority of these living in sub-Saharan Africa. Therefore, an important research focus is development of new drugs that can be used in the treatment of HIV/AIDS. This review gives an overview of the disease and addresses the drugs currently used for treatment, with specific emphasis on new developments within the class of allosteric non-nucleoside reverse transcriptase inhibitors (NNRTIs).

In Vitro Cross-Resistance Profiles of Rilpivirine, Dapivirine, and MIV-150, Nonnucleoside Reverse Transcriptase Inhibitor Microbicides in Clinical Development for the Prevention of HIV-1 Infection

PubMed Central

Giacobbi, Nicholas S.

2017-01-01

ABSTRACT Rilpivirine (RPV), dapivirine (DPV), and MIV-150 are in development as microbicides. It is not known whether they will block infection of circulating nonnucleoside reverse transcriptase inhibitor (NNRTI)-resistant human immunodeficiency virus type 1 (HIV-1) variants. Here, we demonstrate that the activity of DPV and MIV-150 is compromised by many resistant viruses containing single or double substitutions. High DPV genital tract concentrations from DPV ring use may block replication of resistant viruses. However, MIV-150 genital tract concentrations may be insufficient to inhibit many resistant viruses, including those harboring K103N or Y181C. PMID:28507107

Development and evaluation of a simple and effective RT-qPCR inhibitory assay for detection of the efficacy of compounds towards HIV reverse transcriptase.

PubMed

Marino-Merlo, Francesca; Frezza, Caterina; Papaianni, Emanuela; Valletta, Elena; Mastino, Antonio; Macchi, Beatrice

2017-11-01

Assessing the actual efficacy of compounds to directly inhibit HIV reverse transcriptase (RT) activity is a main goal in preclinical antiretroviral studies. Our previous studies demonstrated that the effects of inhibitor compounds towards HIV-RT could be efficiently assessed through a simple cell-free assay based on conventional reverse transcription PCR. In the present study, we describe a modified variant of our assay, termed RT real-time quantitative PCR inhibitory assay (RT-qPCR-IA), in which the ability of compounds to restrict the complementary DNA (cDNA) generation by HIV-RT using a specific RNA template is performed by the real-time technique, in order to improve both accuracy and sensitivity of the method. As specific RNA template, RNA extracted from stable transfectants ectopically expressing the herpes simplex virus 1 glycoprotein D gene was utilized. HIV-RT, of both commercial or house-made viral lysate origin, was employed for the assay. To assess the reliability of RT-qPCR-IA, we performed a comparative, quantitative analysis of the dose-dependent effect exerted by known nucleotide and non-nucleotide reverse-transcriptase inhibitors, using the SYBR Green dye chemistry as detection system. The results obtained with RT-qPCR-IA were compared to that obtained using a one-step PicoGreen technology-based commercial kit. The outcome of our study indicates that the development of the novel RT-qPCR-IA will provide rapid and accurate evaluation of the inhibitory efficacy of compounds towards HIV-RT activity. This evaluation could be very useful for large-scale screening of potential new anti-HIV drugs.

Indolylarylsulfones carrying a heterocyclic tail as very potent and broad spectrum HIV-1 non-nucleoside reverse transcriptase inhibitors.

PubMed

Famiglini, Valeria; La Regina, Giuseppe; Coluccia, Antonio; Pelliccia, Sveva; Brancale, Andrea; Maga, Giovanni; Crespan, Emmanuele; Badia, Roger; Riveira-Muñoz, Eva; Esté, José A; Ferretti, Rosella; Cirilli, Roberto; Zamperini, Claudio; Botta, Maurizio; Schols, Dominique; Limongelli, Vittorio; Agostino, Bruno; Novellino, Ettore; Silvestri, Romano

2014-12-11

We synthesized new indolylarylsulfone (IAS) derivatives carrying a heterocyclic tail at the indole-2-carboxamide nitrogen as potential anti-HIV/AIDS agents. Several new IASs yielded EC50 values <1.0 nM against HIV-1 WT and mutant strains in MT-4 cells. The (R)-11 enantiomer proved to be exceptionally potent against the whole viral panel; in the reverse transcriptase (RT) screening assay, it was remarkably superior to NVP and EFV and comparable to ETV. The binding poses were consistent with the one previously described for the IAS non-nucleoside reverse transcriptase inhibitors. Docking studies showed that the methyl group of (R)-11 points toward the cleft created by the K103N mutation, different from the corresponding group of (S)-11. By calculating the solvent-accessible surface, we observed that the exposed area of RT in complex with (S)-11 was larger than the area of the (R)-11 complex. Compounds 6 and 16 and enantiomer (R)-11 represent novel robust lead compounds of the IAS class.

Antiviral Activity of MK-4965, a Novel Nonnucleoside Reverse Transcriptase Inhibitor▿

PubMed Central

Lai, Ming-Tain; Munshi, Vandna; Touch, Sinoeun; Tynebor, Robert M.; Tucker, Thomas J.; McKenna, Philip M.; Williams, Theresa M.; DiStefano, Daniel J.; Hazuda, Daria J.; Miller, Michael D.

2009-01-01

Nonnucleoside reverse transcriptase inhibitors (NNRTIs) are the mainstays of therapy for the treatment of human immunodeficiency virus type 1 (HIV-1) infections. However, the effectiveness of NNRTIs can be hampered by the development of resistance mutations which confer cross-resistance to drugs in the same class. Extensive efforts have been made to identify new NNRTIs that can suppress the replication of the prevalent NNRTI-resistant viruses. MK-4965 is a novel NNRTI that possesses both diaryl ether and indazole moieties. The compound displays potency at subnanomolar concentrations against wild-type (WT), K103N, and Y181C reverse transcriptase (RT) in biochemical assays. MK-4965 is also highly potent against the WT virus and two most prevalent NNRTI-resistant viruses (viruses that harbor the K103N or the Y181C mutation), against which it had 95% effective concentrations (EC95s) of <30 nM in the presence of 10% fetal bovine serum. The antiviral EC95 of MK-4965 was reduced approximately four- to sixfold when it was tested in 50% human serum. Moreover, MK-4965 was evaluated with a panel of 15 viruses with NNRTI resistance-associated mutations and showed a superior mutant profile to that of efavirenz but not to that of etravirine. MK-4965 was similarly effective against various HIV-1 subtypes and viruses containing nucleoside reverse transcriptase inhibitor or protease inhibitor resistance-conferring mutations. A two-drug combination study showed that the antiviral activity of MK-4965 was nonantagonistic with each of the 18 FDA-licensed drugs tested vice versa in the present study. Taken together, these in vitro data show that MK-4965 possesses the desired properties for further development as a new NNRTI for the treatment of HIV-1 infection. PMID:19289522

HIV Protease Inhibitor Use During Pregnancy Is Associated With Decreased Progesterone Levels, Suggesting a Potential Mechanism Contributing to Fetal Growth Restriction

PubMed Central

Papp, Eszter; Mohammadi, Hakimeh; Loutfy, Mona R.; Yudin, Mark H.; Murphy, Kellie E.; Walmsley, Sharon L.; Shah, Rajiv; MacGillivray, Jay; Silverman, Michael; Serghides, Lena

2015-01-01

Background. Protease inhibitor (PI)–based combination antiretroviral therapy (cART) is administered during pregnancy to prevent perinatal human immunodeficiency virus (HIV) transmission. However, PI use has been associated with adverse birth outcomes, including preterm delivery and small-for-gestational-age (SGA) births. The mechanisms underlying these outcomes are unknown. We hypothesized that PIs contribute to these adverse events by altering progesterone levels. Methods. PI effects on trophoblast progesterone production were assessed in vitro. A mouse pregnancy model was used to assess the impact of PI-based cART on pregnancy outcomes and progesterone levels in vivo. Progesterone levels were assessed in plasma specimens from 27 HIV-infected and 17 HIV-uninfected pregnant women. Results. PIs (ritonavir, lopinavir, and atazanavir) but not nucleoside reverse transcriptase inhibitors (NRTIs) or nonnucleoside reverse transcriptase inhibitors reduced trophoblast progesterone production in vitro. In pregnant mice, PI-based cART but not dual-NRTI therapy was associated with significantly lower progesterone levels that directly correlated with fetal weight. Progesterone supplementation resulted in a significant improvement in fetal weight. We observed lower progesterone levels and smaller infants in HIV-infected women receiving PI-based cART, compared with the control group. In HIV-infected women, progesterone levels correlated significantly with birth weight percentile. Conclusions. Our data suggest that PI use in pregnancy may lead to lower progesterone levels that could contribute to adverse birth outcomes. PMID:25030058

HIV protease inhibitor use during pregnancy is associated with decreased progesterone levels, suggesting a potential mechanism contributing to fetal growth restriction.

PubMed

Papp, Eszter; Mohammadi, Hakimeh; Loutfy, Mona R; Yudin, Mark H; Murphy, Kellie E; Walmsley, Sharon L; Shah, Rajiv; MacGillivray, Jay; Silverman, Michael; Serghides, Lena

2015-01-01

Protease inhibitor (PI)-based combination antiretroviral therapy (cART) is administered during pregnancy to prevent perinatal human immunodeficiency virus (HIV) transmission. However, PI use has been associated with adverse birth outcomes, including preterm delivery and small-for-gestational-age (SGA) births. The mechanisms underlying these outcomes are unknown. We hypothesized that PIs contribute to these adverse events by altering progesterone levels. PI effects on trophoblast progesterone production were assessed in vitro. A mouse pregnancy model was used to assess the impact of PI-based cART on pregnancy outcomes and progesterone levels in vivo. Progesterone levels were assessed in plasma specimens from 27 HIV-infected and 17 HIV-uninfected pregnant women. PIs (ritonavir, lopinavir, and atazanavir) but not nucleoside reverse transcriptase inhibitors (NRTIs) or nonnucleoside reverse transcriptase inhibitors reduced trophoblast progesterone production in vitro. In pregnant mice, PI-based cART but not dual-NRTI therapy was associated with significantly lower progesterone levels that directly correlated with fetal weight. Progesterone supplementation resulted in a significant improvement in fetal weight. We observed lower progesterone levels and smaller infants in HIV-infected women receiving PI-based cART, compared with the control group. In HIV-infected women, progesterone levels correlated significantly with birth weight percentile. Our data suggest that PI use in pregnancy may lead to lower progesterone levels that could contribute to adverse birth outcomes. © The Author 2014. Published by Oxford University Press on behalf of the Infectious Diseases Society of America.

Discovery of piperidin-4-yl-aminopyrimidine derivatives as potent non-nucleoside HIV-1 reverse transcriptase inhibitors.

PubMed

Wan, Zheng-Yong; Yao, Jin; Tao, Yuan; Mao, Tian-Qi; Wang, Xin-Long; Lu, Yi-Pei; Wang, Hai-Feng; Yin, Hong; Wu, Yan; Chen, Fen-Er; De Clercq, Erik; Daelemans, Dirk; Pannecouque, Christophe

2015-06-05

A novel series of piperidin-4-yl-aminopyrimidine derivatives were designed fusing the pharmacophore templates of etravirine-VRX-480773 hybrids our group previously described and piperidine-linked aminopyrimidines. Most compounds displayed significantly improved activity against wild-type HIV-1 with EC50 values in single-digit nanomolar concentrations compared to etravirine-VRX-480773 hybrids. Selected compounds were also evaluated for activity against reverse transcriptase, and had lower IC50 values than that of nevirapine. The improved potency observed in this in vitro model of HIV RNA replication partly validates the mechanism by which this class of allosteric pyrimidine derivatives inhibits reverse transcriptase, and represents a remarkable step forward in the development of AIDS therapeutics. Copyright © 2015 Elsevier Masson SAS. All rights reserved.

Efficacy of non-nucleoside reverse transcriptase inhibitor-based highly active antiretroviral therapy in Thai HIV-infected children aged two years or less.

PubMed

Puthanakit, Thanyawee; Aurpibul, Linda; Sirisanthana, Thira; Sirisanthana, Virat

2009-03-01

Twenty-six Thai HIV-infected children, aged 2 years or less were prospectively enrolled to receive non-nucleoside reverse transcription inhibitor-based highly active antiretroviral therapy (HAART). Twenty-two children (85%) had World Health Organization clinical stage 3 or 4. The median baseline CD4 cell percentage and plasma HIV RNA were 17% and 5.9 log 10 copies/mL, respectively. The median age at HAART initiation was 9.8 months (range, 1.5-24.0). One child died. The mean CD4 cell percentages at 24, 48, and 96 weeks of treatment were 26%, 31%, and 37%, respectively. The proportions of children with virologic suppression (<400 copies/mL) at week 24 and 48 were 14/26 (54%) and 19/26 (73%), respectively. Non-nucleoside reverse transcription inhibitor-based HAART is safe and effective in HIV-infected young children in a resource-limited setting.

Characteristics of a group of nonnucleoside reverse transcriptase inhibitors with structural diversity and potent anti-human immunodeficiency virus activity.

PubMed

Yang, S S; Fliakas-Boltz, V; Bader, J P; Buckheit, R W

1995-10-01

Current thrust in controlling the Acquired Immune Deficiency Syndrome (AIDS) focuses on antiviral drug development targeting the infection and replication of the human immunodeficiency virus (HIV), the causative agent of AIDS. To date, treatment of AIDS has relied on nucleoside reverse transcriptase inhibitors such as AZT, ddI, and ddC, which eventually become ineffective upon the emergence of resistant mutants bearing specific nucleotide substitutions. The Anti-AIDS Drug Screening Program of the NCI conducts and coordinates a high-capacity semi-robotic in vitro screening of synthetic or natural compounds submitted by academic, research and pharmaceutical institutions world-wide. About 10,000 synthetic compounds are screened annually for anti-HIV activity. Confirmed active agents are subjected to in-depth studies on range and mechanism of action. Emerging from this intense screening activity were a number of potentially promising categories of nonnucleoside reverse transcriptase inhibitors (NNRTI) with structural diversity but strong and reproducible anti-HIV activity. Over 2500 active compounds were evaluated for their inhibitory activity against a panel of both laboratory and clinical virus isolates in the appropriate established cell line or fresh human peripheral blood leukocyte and macrophage preparations. Out of these, 40 agents could be placed structurally in nine categories with an additional 16 unique compounds that share the characteristics of NNRTI. These NNRTIs were shown to inhibit reverse transcriptase enzymatically using homopolymeric or ribosomal RNA as templates. NNRTIs demonstrated similarity in their inhibitory pattern against the HIV-1 laboratory strains IIIB and RF, and an AZT-resistant strain; all were inactive against HIV-2. These compounds were further tested against NNRTI-resistant HIV-1 isolates. NNRTI-resistant HIV-1 isolates were selected and characterized with respect to the change(s) in the viral reverse transcriptase nucleotide sequence. Also, differential cross-resistance or sensitivity patterns to NNRTIs were studied in detail among NNRTI-resistant mutants. When tested in combination with AZT, all of the NNRTI's uniformly exhibited synergistic inhibition of HIV-1, suggesting that combination antiviral therapy of NNRTIs with AZT may be therapeutically promising for AIDS treatment.

Impact of Noncoding Satellite Repeats on Pancreatic Cancer Metastasis

DTIC Science & Technology

2014-09-01

nucleoside reverse transcriptase inhibitor ddC as a small molecule inhibitor of HSATII reverse transcription. Initial data indicates there are anti...proliferative effects of ddC in cancer cell lines. We will evaluate ddC and anti-sense locked nucleic acids as methods for inhibiting this process and...of these hybrids, we tested the effect of the nucleoside analog RT inhibitor (NRTI) 2’,3’-dideoxycytidine ( ddC ) in COLO205 cells (Fig. 2e). Notably

In Vitro Cross-Resistance Profiles of Rilpivirine, Dapivirine, and MIV-150, Nonnucleoside Reverse Transcriptase Inhibitor Microbicides in Clinical Development for the Prevention of HIV-1 Infection.

PubMed

Giacobbi, Nicholas S; Sluis-Cremer, Nicolas

2017-07-01

Rilpivirine (RPV), dapivirine (DPV), and MIV-150 are in development as microbicides. It is not known whether they will block infection of circulating nonnucleoside reverse transcriptase inhibitor (NNRTI)-resistant human immunodeficiency virus type 1 (HIV-1) variants. Here, we demonstrate that the activity of DPV and MIV-150 is compromised by many resistant viruses containing single or double substitutions. High DPV genital tract concentrations from DPV ring use may block replication of resistant viruses. However, MIV-150 genital tract concentrations may be insufficient to inhibit many resistant viruses, including those harboring K103N or Y181C. Copyright © 2017 American Society for Microbiology.

Revealing the drug-resistant mechanism for diarylpyrimidine analogue inhibitors of HIV-1 reverse transcriptase.

PubMed

Zhang, Hao; Qin, Fang; Ye, Wei; Li, Zeng; Ma, Songyao; Xia, Yan; Jiang, Yi; Zhu, Jiayi; Li, Yixue; Zhang, Jian; Chen, Hai-Feng

2011-09-01

Diaryltriazine (DATA) and diarylpyrimidine (DAPY) were two category inhibitors with highly potent activity for wild type (wt) and four principal mutant types (L100I, K103N, Y181C and Y188L) of HIV-1 reverse transcriptase (RT). We had revealed the drug-resistant mechanism of DATA analogue inhibitors with molecular dynamics simulation and three-dimensional quantitative structure-activity relationship (3D-QSAR) methods. In this work, we investigated the drug-resistant mechanism of DAPY analogue inhibitors. It was found that DAPY analogue inhibitors form more hydrogen bonds and hydrophobic contacts with wild type and mutants of HIV-1 RT than DATA inhibitors. This could explain that DAPY analogue inhibitors are more potent than DATA for the wild type and mutants of HIV-1 RT. Then, 3D-QSAR models were constructed for these inhibitors of wild type and four principal mutant types HIV-1 RT and evaluated by test set compounds. These combined models can be used to design new chemical entities and make quantitative prediction of the bioactivities for HIV-1 RT inhibitors before resorting to in vitro and in vivo experiment. © 2011 John Wiley & Sons A/S.

The development of HEPT-type HIV non-nucleoside reverse transcriptase inhibitors and its implications for DABO family.

PubMed

Chen, Wenmin; Zhan, Peng; Wu, Jingde; Li, Zhenyu; Liu, Xinyong

2012-01-01

1-[(2-hydroxyethoxy)methyl]-6-(phenylthio)thymine (HEPT) was discovered as the first HIV-1 non-nucleoside reverse transcriptase inhibitors (NNRTIs) in 1989. The research on HEPT derivatives (HEPTs) has been lasted for more than 20 years and HEPT family is probably the most investigated NNRTI. Extensive molecular modifications on HEPT have led to many highly potent compounds with broad-resistance spectrum and optimal pharmacokinetic profiles. Moreover, X-crystallographic studies of HEPTs/RT complexes revealed the binding mode of HEPTs and the action mechanism of NNRTI, which has greatly facilitated the design of novel NNRTIs. Recently, the development of HEPTs was accelerated by the application of the "follow-on"-based chemical evolution strategies, such as designed multiple ligands (DMLs) and molecular hybridization (MH). Herein, this article will provide an insight into the development of HEPTs, including structural modifications, crystal structure of RT complexed with HEPTs and its structure-activity relationship (SAR). Additionally, this review also covers the emerging HEPT related dual inhibitors and HEPT-pyridinone hybrids, as well as the contributions of HEPTs to the development of dihydro-alkoxy-benzyl-oxopyrimidine (DABO) family, thus highlighting the importance of HEPTs on the development of NNRTIs.

Docking, molecular dynamics and quantitative structure-activity relationship studies for HEPTs and DABOs as HIV-1 reverse transcriptase inhibitors.

PubMed

Mao, Yating; Li, Yan; Hao, Ming; Zhang, Shuwei; Ai, Chunzhi

2012-05-01

As a key component in combination therapy for acquired immunodeficiency syndrome (AIDS), non-nucleoside reverse transcriptase inhibitors (NNRTIs) have been proven to be an essential way in stopping HIV-1 replication. In the present work, in silico studies were conducted on a series of 119 NNRTIs, including 1-(2-hydroxyethoxymethyl)-6-(phenylthio)thymine (HEPT) and dihydroalkoxybenzyloxopyrimidine (DABO) derivatives by using the comparative molecular field analysis (CoMFA), comparative molecular similarity indices analysis (CoMSIA), docking simulations and molecular dynamics (MD). The statistical results of the optimal model, the ligand-based CoMSIA one (Q(2) = 0.48, R(ncv)(2) =0.847, R(pre)(2) = 0.745) validates its satisfactory predictive capacity both internally and externally. The contour maps, docking and MD results correlate well with each other, drawing conclusions as follows: 1) Compounds with bulky substituents in position-6 of ring A, hydrophobic groups around position- 1, 2, 6 are preferable to the biological activities; 2) Two hydrogen bonds between RT inhibitor and the Tyr 318, Lys 101 residues, respectively, and a π-π bond between the inhibitor and Trp 188 are formed and crucial to the orientation of the active conformation of the molecules; 3) The binding pocket is essentially hydrophobic, which are determined by residues such as Trp 229, Tyr 318, Val 179, Tyr 188 and Val 108, and hydrophobic substituents may bring an improvement to the biological activity; 4) DABO and HEPT derivatives have different structures but take a similar mechanism to inhibit RT. The potency difference between two isomers in HEPTs can be explained by the distinct locations of the 6-naphthylmethyl substituent and the reasons are explained in details. All these results could be employed to alter the structural scaffold in order to develop new HIV-1 RT inhibitors that have an improved biological property. To the best of our knowledge, this is the first report on 3D-QSAR modeling of this series of HEPT and DABO NNRTs. The QSAR model and the information derived, we hope, will be of great help in presenting clear guidelines and accurate activity predictions for newly designed HIV-1 reverse transcriptase (RT) inhibitor.

Structure-based design, synthesis, and biological evaluation of novel pyrrolyl aryl sulfones: HIV-1 non-nucleoside reverse transcriptase inhibitors active at nanomolar concentrations.

PubMed

Artico, M; Silvestri, R; Pagnozzi, E; Bruno, B; Novellino, E; Greco, G; Massa, S; Ettorre, A; Loi, A G; Scintu, F; La Colla, P

2000-05-04

Pyrrolyl aryl sulfones (PASs) have been recently reported as a new class of human immunodeficiency virus type 1 (HIV-1) reverse transcriptase (RT) inhibitors acting at the non-nucleoside binding site of this enzyme (Artico, M.; et al. J. Med. Chem. 1996, 39, 522-530). Compound 3, the most potent inhibitor within the series (EC(50) = 0.14 microM, IC(50) = 0.4 microM, and SI > 1429), was then selected as a lead compound for a synthetic project based on molecular modeling studies. Using the three-dimensional structure of RT cocrystallized with the alpha-APA derivative R95845, we derived a model of the RT/3 complex by taking into account previously developed structure-activity relationships. Inspection of this model and docking calculations on virtual compounds prompted the design of novel PAS derivatives and related analogues. Our computational approach proved to be effective in making qualitative predictions, that is in discriminating active versus inactive compounds. Among the compounds synthesized and tested, 20 was the most active one, with EC(50) = 0.045 microM, IC(50) = 0.05 microM, and SI = 5333. Compared with the lead 3, these values represent a 3- and 8-fold improvement in the cell-based and enzyme assays, respectively, together with the highest selectivity achieved so far in the PAS series.

Simple and simultaneous determination of the hiv-protease inhibitors amprenavir, atazanavir, indinavir, lopinavir, nelfinavir, ritonavir and saquinavir plus M8 nelfinavir metabolite and the nonnucleoside reverse transcriptase inhibitors efavirenz and nevirapine in human plasma by reversed-phase liquid chromatography.

PubMed

Poirier, Jean-Marie; Robidou, Pascal; Jaillon, Patrice

2005-04-01

Several studies suggest that therapeutic drug monitoring of protease inhibitors and nonnucleoside reverse transcriptase inhibitors may contribute to the clinical outcome of HIV-infected patients. Because of the growing number of antiretroviral drugs and of drug combinations than can be administered to these patients, an accurate high-performance liquid chromatographic (HPLC) method allowing the simultaneous determination of these drugs may be useful. To date, the authors present the first simultaneous HPLC determination of the new protease inhibitor atazanavir with all the others currently in use (M8 nelfinavir metabolite included) and the 2 widely used nonnucleoside reverse transcriptase inhibitors efavirenz and nevirapine. This simple HPLC method allows the analysis all these drugs at a single ultraviolet wavelength following a 1-step liquid-liquid extraction procedure. A 500-muL plasma sample was spiked with internal standard and subjected to liquid-liquid extraction using by diethyl ether at pH 10. HPLC was performed using a Symmetry Shield RP18 and gradient elution. All the drugs of interest and internal standard were detected with ultraviolet detection at 210 nm. Calibration curves were linear in the range 50-10,000 ng/mL. The observed concentrations of the quality controls at plasma concentrations ranging from 50 to 5000 ng/mL for these drugs showed that the overall accuracy varied from 92% to 104% and 92% to 106% for intraday and day-to-day analysis, respectively. No metabolites of the assayed compounds or other drugs commonly coadministered to HIV-positive patients were found to coelute with the drugs of interest or with the internal standard. This assay was developed for the purpose of therapeutic monitoring (TDM) in HIV-infected patients.

Antiviral interactions of combinations of highly potent 2,4(1H,3H)-pyrimidinedione congeners and other anti-HIV agents.

PubMed

Hartman, Tracy L; Yang, Lu; Buckheit, Robert W

2011-12-01

Structure-activity relationship evaluation of seventy-four 2,4(1H,3H)-pyrimidinedione derivatives identified seven lead compounds based on anti-HIV-1 potency, extended range of action to include HIV-2, virus entry inhibition, reverse transcriptase inhibition, and lack of cytotoxicity to human cells. The selected pyrimidinedione congeners are highly active inhibitors of HIV-1 with EC(50) values ranging from 0.6 to 2 nM in CEM-SS cells infected with laboratory derived viruses, 11-20 nM in fresh human PBMCs infected with subtype B (HT/92/599) virus, and 2-7 nM in PBMCs infected with the clinical subtype C (ZA/97/003) virus. Combination antiviral assays were performed using the laboratory adapted RF strain of HIV-1 in CEM-SS cells and with a clade B and C low passage clinical isolate in fresh human peripheral mononuclear cells and the compound interactions were analyzed using MacSynergy II. The seven pyrimidinedione compounds resulted in additive to synergistic interactions in combination with entry and fusion inhibitors, nonnucleoside and nucleoside reverse transcriptase inhibitors, and the protease inhibitors. No evidence of antagonistic antiviral activity or synergistic cytotoxicity was detected with the combinations of compounds tested. The dual mechanism of action of the pyrimidinediones resulting in inhibition of both virus entry and reverse transcription suggests excellent potential of these lead pyrimidinediones as candidates for combination therapy with other approved HIV inhibitors of varying mechanism of action. Copyright © 2011. Published by Elsevier B.V.

A trypsin inhibitor from rambutan seeds with antitumor, anti-HIV-1 reverse transcriptase, and nitric oxide-inducing properties.

PubMed

Fang, Evandro Fei; Ng, Tzi Bun

2015-04-01

Nephelium lappaceum L., commonly known as "rambutan," is a typical tropical tree and is well known for its juicy and sweet fruit which has an exotic flavor. Chemical studies on rambutan have led to the identification of various components such as monoterpene lactones and volatile compounds. Here, a 22.5-kDa trypsin inhibitor (N . lappaceum trypsin inhibitor (NLTI)) was isolated from fresh rambutan seeds using liquid chromatographical techniques. NLTI reduced the proteolytic activities of both trypsin and α-chymotrypsin. Dithiothreitol reduced the trypsin inhibitory activity of NLTI at a concentration of 1 mM, indicating that an intact disulfide bond is essential to the activity. NLTI inhibited HIV-1 reverse transcriptase with an IC50 of 0.73 μM. In addition, NLTI manifested a time- and dose-dependent inhibitory effect on growth in many tumor cells. NLTI is one of the few trypsin inhibitors with nitric oxide-inducing activity and may find application in tumor therapy.

Approved Antiviral Drugs over the Past 50 Years

PubMed Central

2016-01-01

SUMMARY Since the first antiviral drug, idoxuridine, was approved in 1963, 90 antiviral drugs categorized into 13 functional groups have been formally approved for the treatment of the following 9 human infectious diseases: (i) HIV infections (protease inhibitors, integrase inhibitors, entry inhibitors, nucleoside reverse transcriptase inhibitors, nonnucleoside reverse transcriptase inhibitors, and acyclic nucleoside phosphonate analogues), (ii) hepatitis B virus (HBV) infections (lamivudine, interferons, nucleoside analogues, and acyclic nucleoside phosphonate analogues), (iii) hepatitis C virus (HCV) infections (ribavirin, interferons, NS3/4A protease inhibitors, NS5A inhibitors, and NS5B polymerase inhibitors), (iv) herpesvirus infections (5-substituted 2′-deoxyuridine analogues, entry inhibitors, nucleoside analogues, pyrophosphate analogues, and acyclic guanosine analogues), (v) influenza virus infections (ribavirin, matrix 2 protein inhibitors, RNA polymerase inhibitors, and neuraminidase inhibitors), (vi) human cytomegalovirus infections (acyclic guanosine analogues, acyclic nucleoside phosphonate analogues, pyrophosphate analogues, and oligonucleotides), (vii) varicella-zoster virus infections (acyclic guanosine analogues, nucleoside analogues, 5-substituted 2′-deoxyuridine analogues, and antibodies), (viii) respiratory syncytial virus infections (ribavirin and antibodies), and (ix) external anogenital warts caused by human papillomavirus infections (imiquimod, sinecatechins, and podofilox). Here, we present for the first time a comprehensive overview of antiviral drugs approved over the past 50 years, shedding light on the development of effective antiviral treatments against current and emerging infectious diseases worldwide. PMID:27281742

Design, discovery, modelling, synthesis, and biological evaluation of novel and small, low toxicity s-triazine derivatives as HIV-1 non-nucleoside reverse transcriptase inhibitors.

PubMed

Viira, Birgit; Selyutina, Anastasia; García-Sosa, Alfonso T; Karonen, Maarit; Sinkkonen, Jari; Merits, Andres; Maran, Uko

2016-06-01

A set of top-ranked compounds from a multi-objective in silico screen was experimentally tested for toxicity and the ability to inhibit the activity of HIV-1 reverse transcriptase (RT) in cell-free assay and in cell-based assay using HIV-1 based virus-like particles. Detailed analysis of a commercial sample that indicated specific inhibition of HIV-1 reverse transcription revealed that a minor component that was structurally similar to that of the main compound was responsible for the strongest inhibition. As a result, novel s-triazine derivatives were proposed, modelled, discovered, and synthesised, and their antiviral activity and cellular toxicity were tested. Compounds 18a and 18b were found to be efficient HIV-1 RT inhibitors, with an IC50 of 5.6±1.1μM and 0.16±0.05μM in a cell-based assay using infectious HIV-1, respectively. Compound 18b also had no detectable toxicity for different human cell lines. Their binding mode and interactions with the RT suggest that there was strong and adaptable binding in a tight (NNRTI) hydrophobic pocket. In summary, this iterative study produced structural clues and led to a group of non-toxic, novel compounds to inhibit HIV-RT with up to nanomolar potency. Copyright © 2016 Elsevier Ltd. All rights reserved.

Transmitted drug resistance in patients with acute/recent HIV infection in Brazil.

PubMed

Ferreira, Ana Cristina G; Coelho, Lara E; Grinsztejn, Eduarda; Jesus, Carlos S de; Guimarães, Monick L; Veloso, Valdiléa G; Grinsztejn, Beatriz; Cardoso, Sandra W

The widespread use of antiretroviral therapy increased the transmission of antiretroviral resistant HIV strains. Antiretroviral therapy initiation during acute/recent HIV infection limits HIV reservoirs and improves immune response in HIV infected individuals. Transmitted drug resistance may jeopardize the early goals of early antiretroviral treatment among acute/recent HIV infected patients. Patients with acute/recent HIV infection who underwent resistance test before antiretroviral treatment initiation were included in this analysis. HIV-1 sequences were obtained using an in house protease/reverse transcriptase genotyping assay. Transmitted drug resistance was identified according to the Stanford HIV Database for Transmitted Drug Resistance Mutations, based on WHO 2009 surveillance list, and HIV-1 subtyping according to Rega HIV-1 subtyping tool. Comparison between patients with and without transmitted drug resistance was made using Kruskal-Wallis and Chi-square tests. Forty-three patients were included, 13 with acute HIV infection and 30 with recent HIV infection. The overall transmitted drug resistance prevalence was 16.3% (95% confidence interval [CI]: 8.1-30.0%). The highest prevalence of resistance (11.6%, 95% CI: 8.1-24.5) was against non-nucleoside reverse transcriptase inhibitors, and K103N was the most frequently identified mutation. The high prevalence of nonnucleoside reverse transcriptase inhibitors resistance indicates that efavirenz-based regimen without prior resistance testing is not ideal for acutely/recently HIV-infected individuals in our setting. In this context, the recent proposal of including integrase inhibitors as a first line regimen in Brazil could be an advantage for the treatment of newly HIV infected individuals. However, it also poses a new challenge, since integrase resistance test is not routinely performed for antiretroviral naive individuals. Further studies on transmitted drug resistance among acutely/recently HIV-infected are needed to inform the predictors of transmitted resistance and the antiretroviral therapy outcomes among these population. Copyright © 2017 Sociedade Brasileira de Infectologia. Published by Elsevier Editora Ltda. All rights reserved.

Lower genetic variability of HIV-1 and antiretroviral drug resistance in pregnant women from the state of Pará, Brazil.

PubMed

Machado, Luiz Fernando Almeida; Costa, Iran Barros; Folha, Maria Nazaré; da Luz, Anderson Levy Bessa; Vallinoto, Antonio Carlos Rosário; Ishak, Ricardo; Ishak, Marluisa Oliveira Guimarães

2017-04-12

The present study aimed to describe the genetic diversity of HIV-1, as well as the resistance profile of the viruses identified in HIV-1 infected pregnant women under antiretroviral therapy in the state of Pará, Northern Brazil. Blood samples were collected from 45 HIV-1 infected pregnant to determine the virus subtypes according to the HIV-1 protease (PR) gene and part of the HIV-1 reverse transcriptase (RT) gene by sequencing the nucleotides of these regions. Drug resistance mutations and susceptibility to antiretroviral drugs were analyzed by the Stanford HIV Drug Resistance Database. Out of 45 samples, only 34 could be amplified for PR and 30 for RT. Regarding the PR gene, subtypes B (97.1%) and C (2.9%) were identified; for the RT gene, subtypes B (90.0%), F (6.7%), and C (3.3%) were detected. Resistance to protease inhibitors (PI) was identified in 5.8% of the pregnant, and mutations conferring resistance to nucleoside reverse transcriptase inhibitors were found in 3.3%, while mutations conferring resistance to non-nucleoside reverse transcriptase inhibitors were found in 3.3%. These results showed a low frequency of strains resistant to antiretroviral drugs, the prevalence of subtypes B and F, and the persistent low transmission of subtype C in pregnant of the state of Pará, Brazil.

Chemical system biology based molecular interactions to identify inhibitors against Q151M mutant of HIV-1 reverse transcriptase.

PubMed

Pandey, Rajan Kumar; Sharma, Drista; Ojha, Rupal; Bhatt, Tarun Kumar; Prajapati, Vijay Kumar

2018-05-09

The emergence of mutations leading to drug resistance is the main cause of therapeutic failure in the human HIV infection. Chemical system biology approach has drawn great attention to discover new antiretroviral hits with high efficacy and negligible toxicity, which can be used as a prerequisite for HIV drug resistance global action plan 2017-21. To discover potential hits, we docked 49 antiretroviral analogs (n = 6294) against HIV-1 reverse transcriptase Q151M mutant & its wild-type form and narrow downed their number in three sequential modes of docking using Schrödinger suite. Later on, 80 ligands having better docking score than reference ligands (tenofovir and lamivudine) were screened for ADME, toxicity prediction, and binding energy estimation. Simultaneously, the area under the curve (AUC) was estimated using receiver operating characteristics (ROC) curve analysis to validate docking protocols. Finally, single point energy and molecular dynamics simulation approaches were performed for best two ligands (L3 and L14). This study reveals the antiretroviral efficacy of obtained two best ligands and delivers the hits against HIV-1 reverse transcriptase Q151M mutant. Copyright © 2018 Elsevier B.V. All rights reserved.

Chemical crosslinking of the subunits of HIV-1 reverse transcriptase.

PubMed Central

Debyser, Z.; De Clercq, E.

1996-01-01

The reverse transcriptase (RT) of the human immunodeficiency virus type 1 (HIV-1) is composed of two subunits of 66 and 51 kDa in a 1 to 1 ratio. Because dimerization is a prerequisite for enzymatic activity, interference with the dimerization process could constitute an alternative antiviral strategy for RT inhibition. Here we describe an in vitro assay for the study of the dimerization state of HIV-1 reverse transcriptase based on chemical crosslinking of the subunits with dimethylsuberimidate. Crosslinking results in the formation of covalent bonds between the subunits, so that the crosslinked species can be resolved by denaturing gel electrophoresis. Crosslinked RT species with molecular weight greater than that of the dimeric form accumulate during a 1-15-min time course. Initial evidence suggests that those high molecular weight species represent trimers and tetramers and may be the result of intramolecular crosslinking of the subunits of a higher-order RT oligomer. A peptide that corresponds to part of the tryptophan repeat motif in the connection domain of HIV-1 RT inhibits crosslink formation as well as enzymatic activity. The crosslinking assay thus allows the investigation of the effect of inhibitors on the dimerization of HIV-1 RT. PMID:8745406

Hepatotoxicity of nucleoside reverse transcriptase inhibitors.

PubMed

Montessori, Valentina; Harris, Marianne; Montaner, Julio S G

2003-05-01

Hepatotoxicity is an adverse effect of all available classes of antiretrovirals, including nucleoside reverse transcriptase inhibitors (NRTI). A syndrome of hepatic steatosis and lactic acidosis has been recognized as a rare, potentially fatal complication since the advent of NRTI monotherapy in the early 1990s. Today, NRTI remain the backbone of antiretroviral combination regimens, and, with the success of current treatment strategies, exposure to two or more of these agents may occur over a number of years. Hepatic steatosis and lactic acidosis are accordingly being observed more frequently, along with a more recently recognized syndrome of chronic hyperlactatemia. These as well as other adverse effects of NRTI are mediated by inhibition of human DNA polymerase gamma, resulting in mitochondrial dysfunction in the liver and other tissues. Early recognition and intervention are essential to avert serious outcomes.

Perinatal exposure of patas monkeys to antiretroviral nucleoside reverse-transcriptase inhibitors induces genotoxicity persistent for up to 3 years of age.

PubMed

Olivero, Ofelia A; Torres, Lorangelly Rivera; Gorjifard, Sayeh; Momot, Dariya; Marrogi, Eryney; Divi, Rao L; Liu, Yongmin; Woodward, Ruth A; Sowers, Marsha J; Poirier, Miriam C

2013-07-15

Erythrocebus patas (patas) monkeys were used to model antiretroviral (ARV) drug in human immunodeficiency virus type 1-infected pregnant women. Pregnant patas dams were given human-equivalent doses of ARVs daily during 50% of gestation. Mesenchymal cells, cultured from bone marrow of patas offspring obtained at birth and at 1 and 3 years of age, were examined for genotoxicity, including centrosomal amplification, micronuclei, and micronuclei containing whole chromosomes. Compared with controls, statistically significant increases (P < .05) in centrosomal amplification, micronuclei, and micronuclei containing whole chromosomes were found in mesenchymal cells from most groups of offspring at the 3 time points. Transplacental nucleoside reverse-transcriptase inhibitor exposures induced fetal genotoxicity that was persistent for 3 years.

TNF α is involved in neuropathic pain induced by nucleoside reverse transcriptase inhibitor in rats

PubMed Central

Zheng, Xuexing; Ouyang, Handong; Liu, Shue; Mata, Marina; Fink, David J.; Hao, Shuanglin

2011-01-01

In patients with HIV/AIDS, neuropathic pain is a common neurological complication. Infection with the HIV itself may lead to neuropathic pain, and painful symptoms are enhanced when patients are treated with nucleoside reverse transcriptase inhibitors (NRTI). The mechanisms by which NRTIs contribute to the development of neuropathic pain are not known. In the current studies, we tested the role of TNFα in antiretroviral drug-induced neuropathic pain. We administered 2′,3′-dideoxycytidine (ddC, one of the NRTIs) systemically to induce mechanical allodynia. We found that ddC induced overexpression of both mRNA and proteins of GFAP and TNFα in the spinal dorsal horn. TNFα was colocalized with GFAP in the spinal dorsal horn and with NeuN in the DRG. Knockdown of TNFα with siRNA blocked the mechanical allodynia induced by ddC. Intrathecal administration of glial inhibitor or recombinant TNF soluble receptor, reversed mechanical allodynia induced by ddC. These results suggest that TNFα is involved in NRTI-induced neuropathic pain. PMID:21741472

Characterization of UC781-Tenofovir Combination Gel Products for HIV-1 Infection Prevention in an Ex Vivo Ectocervical Model

PubMed Central

Cost, Marilyn; Dezzutti, Charlene S.; Clark, Meredith R.; Friend, David R.; Akil, Ayman

2012-01-01

HIV continues to be a problem worldwide. Topical vaginal microbicides represent one option being evaluated to stop the spread of HIV. With drug candidates that have a specific action against HIV now being studied, it is important that, when appropriate and based on the mechanism of action, the drug permeates the tissue so that it can be delivered to specific targets which reside there. Novel formulations of the nucleotide reverse transcriptase inhibitor tenofovir (TFV) and the nonnucleoside reverse transcriptase inhibitor UC781 have been developed and evaluated here. Gels with three distinct rheological properties were prepared. The three gels released both UC781 and TFV under in vitro conditions at concentrations equal to or above the reported 50% effective concentrations (EC50s). The drug concentrations in ectocervical tissues were well in excess of the reported EC50s. The gels maintain ectocervical viability and prevent infection of ectocervical explants after a HIV-1 challenge. This study successfully demonstrates the feasibility of using this novel combination of antiretroviral agents in an aqueous gel as an HIV infection preventative. PMID:22430977

Optimal therapies of a virus replication model with pharmacological delays based on reverse transcriptase inhibitors and protease inhibitors

NASA Astrophysics Data System (ADS)

Pei, Yongzhen; Li, Changguo; Liang, Xiyin

2017-11-01

A short delay in the pharmacological effect on account of the time required for drug absorption, distribution, and penetration into target cells after application of any anti-viral drug, is defined by the pharmacological delay (Herz et al 1996 Proc. Natl Acad. Sci. USA 93 7247-51). In this paper, a virus replication model with Beddington-DeAngelis incidence rate and the pharmacological and intracellular delays is presented to describe the treatment to cure the virus infection. The optimal controls represent the efficiency of reverse transcriptase inhibitors and protease inhibitors in suppressing viral production and prohibiting new infections. Due to the fact that both the control and state variables contain delays, we derive a necessary conditions for our optimal problem. Based on these results, numerical simulations are implemented not only to show the optimal therapeutic schedules for different infection and release rates, but also to compare the effective of three treatment programs. Furthermore, comparison of therapeutic effects under different maximum tolerable dosages is shown. Our research indicates that (1) the proper and specific treatment program should be determined according to the infection rates of different virus particles; (2) the optimal combined drug treatment is the most efficient; (3) the appropriate proportion of medicament must be formulated during the therapy due to the non-monotonic relationship between maximum tolerable dosages and therapeutic effects; (4) the therapeutic effect is advantageous when the pharmacological delay is considered.

Impact of human immunodeficiency virus type 1 reverse transcriptase inhibitor drug resistance mutation interactions on phenotypic susceptibility.

PubMed

Trivedi, Vinod; Von Lindern, Jana; Montes-Walters, Miguel; Rojo, Daniel R; Shell, Elisabeth J; Parkin, Neil; O'Brien, William A; Ferguson, Monique R

2008-10-01

The role specific reverse transcriptase (RT) drug resistance mutations play in influencing phenotypic susceptibility to RT inhibitors in virus strains with complex resistance interaction patterns was assessed using recombinant viruses that consisted of RT-PCR-amplified pol fragments derived from plasma HIV-1 RNA from two treatment-experienced patients. Specific modifications of key RT amino acids were performed by site-directed mutagenesis. A panel of viruses with defined genotypic resistance mutations was assessed for phenotypic drug resistance. Introduction of M184V into several different clones expressing various RT resistance mutations uniformly decreased susceptibility to abacavir, lamivudine, and didanosine, and increased susceptibility to zidovudine, stavudine, and tenofovir; replication capacity was decreased. The L74V mutation had similar but slightly different effects, contributing to decreased susceptibility to abacavir, lamivudine, and didanosine and increased susceptibility to zidovudine and tenofovir, but in contrast to M184V, L74V contributed to decreased susceptibility to stavudine. In virus strains with the nonnucleoside reverse transcriptase inhibitor (NNRTI) mutations K101E and G190S, the L74V mutation increased replication capacity, consistent with published observations, but replication capacity was decreased in strains without NNRTI resistance mutations. K101E and G190S together tend to decrease susceptibility to all nucleoside RT inhibitors, but the K103N mutation had little effect on nucleoside RT inhibitor susceptibility. Mutational interactions can have a substantial impact on drug resistance phenotype and replication capacity, and this has been exploited in clinical practice with the development of fixed-dose combination pills. However, we are the first to report these mutational interactions using molecularly cloned recombinant strains derived from viruses that occur naturally in HIV-infected individuals.

Impact of Human Immunodeficiency Virus Type 1 Reverse Transcriptase Inhibitor Drug Resistance Mutation Interactions on Phenotypic Susceptibility

PubMed Central

Trivedi, Vinod; Von Lindern, Jana; Montes-Walters, Miguel; Rojo, Daniel R.; Shell, Elisabeth J.; Parkin, Neil; O'Brien, William A.

2008-01-01

Abstract The role specific reverse transcriptase (RT) drug resistance mutations play in influencing phenotypic susceptibility to RT inhibitors in virus strains with complex resistance interaction patterns was assessed using recombinant viruses that consisted of RT-PCR-amplified pol fragments derived from plasma HIV-1 RNA from two treatment-experienced patients. Specific modifications of key RT amino acids were performed by site-directed mutagenesis. A panel of viruses with defined genotypic resistance mutations was assessed for phenotypic drug resistance. Introduction of M184V into several different clones expressing various RT resistance mutations uniformly decreased susceptibility to abacavir, lamivudine, and didanosine, and increased susceptibility to zidovudine, stavudine, and tenofovir; replication capacity was decreased. The L74V mutation had similar but slightly different effects, contributing to decreased susceptibility to abacavir, lamivudine, and didanosine and increased susceptibility to zidovudine and tenofovir, but in contrast to M184V, L74V contributed to decreased susceptibility to stavudine. In virus strains with the nonnucleoside reverse transcriptase inhibitor (NNRTI) mutations K101E and G190S, the L74V mutation increased replication capacity, consistent with published observations, but replication capacity was decreased in strains without NNRTI resistance mutations. K101E and G190S together tend to decrease susceptibility to all nucleoside RT inhibitors, but the K103N mutation had little effect on nucleoside RT inhibitor susceptibility. Mutational interactions can have a substantial impact on drug resistance phenotype and replication capacity, and this has been exploited in clinical practice with the development of fixed-dose combination pills. However, we are the first to report these mutational interactions using molecularly cloned recombinant strains derived from viruses that occur naturally in HIV-infected individuals. PMID:18844463

Antiretroviral drug susceptibility among drug-naive adults with recent HIV infection in Rakai, Uganda

PubMed Central

Eshleman, Susan H.; Laeyendecker, Oliver; Parkin, Neil; Huang, Wei; Chappey, Colombe; Paquet, Agnes C.; Serwadda, David; Reynolds, Steven J.; Kiwanuka, Noah; Quinn, Thomas C.; Gray, Ronald; Wawer, Maria

2009-01-01

Objective To analyze antiretroviral drug susceptibility in HIV from recently infected adults in Rakai, Uganda, prior to the availability of antiretroviral drug treatment. Methods Samples obtained at the time of HIV seroconversion (1998–2003) were analyzed using the GeneSeq HIV and PhenoSense HIV assays (Monogram Biosciences, Inc., South San Francisco, California, USA). Results Test results were obtained for 104 samples (subtypes: 26A, 1C, 66D, 9A/D, 1C/D, 1 intersubtype recombinant). Mutations used for genotypic surveillance of transmitted antiretroviral drug resistance were identified in six samples: three had nucleoside reverse transcriptase inhibitor (NRTI) surveillance mutations (two had M41L, one had K219R), and three had protease inhibitor surveillance mutations (I47V, F53L, N88D); none had nonnucleoside reverse transcriptase inhibitor (NNRTI) surveillance mutations. Other resistance-associated mutations were identified in some samples. However, none of the samples had a sufficient number of mutations to predict reduced antiretroviral drug susceptibility. Ten (9.6%) of the samples had reduced phenotypic susceptibility to at least one drug (one had partial susceptibility to didanosine, one had nevirapine resistance, and eight had resistance or partial susceptibility to at least one protease inhibitor). Fifty-three (51%) of the samples had hypersusceptibility to at least one drug (seven had zidovudine hypersusceptibility, 28 had NNRTI hypersusceptibility, 34 had protease inhibitor hypersusceptibility). Delavirdine hyper-susceptibility was more frequent in subtype A than D. In subtype D, efavirenz hypersusceptibility was associated with substitutions at codon 11 in HIV-reverse transcriptase. Conclusion Phenotyping detected reduced antiretroviral drug susceptibility and hypersusceptibility in HIV from some antiretroviral-naive Ugandan adults that was not predicted by genotyping. Phenotyping may complement genotyping for analysis of antiretroviral drug susceptibility in populations with nonsubtype B HIV infection. PMID:19276794

Antiretroviral drug susceptibility among drug-naive adults with recent HIV infection in Rakai, Uganda.

PubMed

Eshleman, Susan H; Laeyendecker, Oliver; Parkin, Neil; Huang, Wei; Chappey, Colombe; Paquet, Agnes C; Serwadda, David; Reynolds, Steven J; Kiwanuka, Noah; Quinn, Thomas C; Gray, Ronald; Wawer, Maria

2009-04-27

To analyze antiretroviral drug susceptibility in HIV from recently infected adults in Rakai, Uganda, prior to the availability of antiretroviral drug treatment. Samples obtained at the time of HIV seroconversion (1998-2003) were analyzed using the GeneSeq HIV and PhenoSense HIV assays (Monogram Biosciences, Inc., South San Francisco, California, USA). Test results were obtained for 104 samples (subtypes: 26A, 1C, 66D, 9A/D, 1C/D, 1 intersubtype recombinant). Mutations used for genotypic surveillance of transmitted antiretroviral drug resistance were identified in six samples: three had nucleoside reverse transcriptase inhibitor (NRTI) surveillance mutations (two had M41L, one had K219R), and three had protease inhibitor surveillance mutations (I47V, F53L, N88D); none had nonnucleoside reverse transcriptase inhibitor (NNRTI) surveillance mutations. Other resistance-associated mutations were identified in some samples. However, none of the samples had a sufficient number of mutations to predict reduced antiretroviral drug susceptibility. Ten (9.6%) of the samples had reduced phenotypic susceptibility to at least one drug (one had partial susceptibility to didanosine, one had nevirapine resistance, and eight had resistance or partial susceptibility to at least one protease inhibitor). Fifty-three (51%) of the samples had hypersusceptibility to at least one drug (seven had zidovudine hypersusceptibility, 28 had NNRTI hypersusceptibility, 34 had protease inhibitor hypersusceptibility). Delavirdine hypersusceptibility was more frequent in subtype A than D. In subtype D, efavirenz hypersusceptibility was associated with substitutions at codon 11 in HIV-reverse transcriptase. Phenotyping detected reduced antiretroviral drug susceptibility and hypersusceptibility in HIV from some antiretroviral-naive Ugandan adults that was not predicted by genotyping. Phenotyping may complement genotyping for analysis of antiretroviral drug susceptibility in populations with nonsubtype B HIV infection.

HIV/AIDS Medicines

MedlinePlus

... few years. But today, there are many effective medicines to fight the infection, and people with HIV ... healthier lives. There are five major types of medicines: Reverse transcriptase (RT) inhibitors - interfere with a critical ...

Executive summary of the GESIDA/National AIDS Plan Consensus Document on Antiretroviral Therapy in Adults Infected by the Human Immunodeficiency Virus (Updated January 2016).

PubMed

2016-01-01

In this update, antiretroviral therapy (ART) is recommended for all patients infected by type 1 human immunodeficiency virus (HIV-1). The objective of ART is to achieve an undetectable plasma viral load (PVL). Initial ART should comprise 3 drugs, namely, 2 nucleoside reverse transcriptase inhibitors (NRTI), and 1 drug from another family. Four of the recommended regimens, all of which have an integrase strand transfer inhibitor (INSTI) as the third drug, are considered a preferred regimen; a further 6 regimens, which are based on an INSTI, a non-nucleoside reverse transcriptase inhibitor (NNRTI), or a protease inhibitor boosted with cobicistat or ritonavir (PI/COBI, PI/r), are considered alternatives. The reasons and criteria for switching ART are presented both for patients with an undetectable PVL and for patients who experience virological failure, in which case the rescue regimen should include 3 (or at least 2) drugs that are fully active against HIV. The specific criteria for ART in special situations (acute infection, HIV-2 infection, pregnancy) and comorbid conditions (tuberculosis and other opportunistic infections, kidney disease, liver disease, and cancer) are updated. Copyright © 2016 Elsevier España, S.L.U. and Sociedad Española de Enfermedades Infecciosas y Microbiología Clínica. All rights reserved.

5-Hydroxypyrido[2,3-b]pyrazin-6(5H)-one derivatives as novel dual inhibitors of HIV-1 reverse transcriptase-associated ribonuclease H and integrase.

PubMed

Sun, Lin; Gao, Ping; Dong, Guanyu; Zhang, Xujie; Cheng, Xiqiang; Ding, Xiao; Wang, Xueshun; Daelemans, Dirk; De Clercq, Erik; Pannecouque, Christophe; Menéndez-Arias, Luis; Zhan, Peng; Liu, Xinyong

2018-06-18

We reported herein the design, synthesis and biological evaluation of a series of 5-hydroxypyrido[2,3-b]pyrazin-6(5H)-one derivatives as HIV-1 reverse transcriptase (RT) ribonuclease H (RNase H) inhibitors using a privileged structure-guided scaffold refining strategy. In view of the similarities between the pharmacophore model of RNase H and integrase (IN) inhibitors as well as their catalytic sites, we also performed IN inhibition assays. Notably, the majority of these derivatives inhibited RNase H and IN at micromolar concentrations. Among them, compound 7a exhibited similar inhibitory activity against RNase H and IN (IC 50 RNase H  = 1.77 μM, IC 50 IN  = 1.18 μM, ratio = 1.50). To the best of our knowledge, this is the first reported dual HIV-1 RNase H-IN inhibitor based on a 5-hydroxypyrido[2,3-b]pyrazin-6(5H)-one structure. Molecular modeling has been used to predict the binding mode of 7a in complex with the catalytic cores of HIV-1 RNase H and IN. Taken together these results strongly support the feasibility of developing HIV-1 dual inhibitors from analog-based optimization of divalent metal ion chelators. Recently, the identification of dual inhibitors proved to be a highly effective strategy for novel antivirals discovery. Therefore, these compounds appear to be useful leads that can be further modified to develop more valuable anti-HIV-1 molecules with suitable drug profiles. Copyright © 2018 Elsevier Masson SAS. All rights reserved.

Five Antiretroviral Drug Class-Resistant HIV-1 in a Treatment-Naïve Patient Successfully Suppressed with Optimized Antiretroviral Drug Selection.

PubMed

Volpe, Joseph M; Ward, Douglas J; Napolitano, Laura; Phung, Pham; Toma, Jonathan; Solberg, Owen; Petropoulos, Christos J; Walworth, Charles M

2015-01-01

Transmitted HIV-1 exhibiting reduced susceptibility to protease and reverse transcriptase inhibitors is well documented but limited for integrase inhibitors and enfuvirtide. We describe here a case of transmitted 5 drug class-resistance in an antiretroviral (ARV)-naïve patient who was successfully treated based on the optimized selection of an active ARV drug regimen. The value of baseline resistance testing to determine an optimal ARV treatment regimen is highlighted in this case report. © The Author(s) 2015.

Complete inactivation of HIV-1 using photo-labeled non-nucleoside reverse transcriptase inhibitors.

PubMed

Rios, Adan; Quesada, Jorge; Anderson, Dallas; Goldstein, Allan; Fossum, Theresa; Colby-Germinario, Susan; Wainberg, Mark A

2011-01-01

We demonstrate that a photo-labeled derivative of the non-nucleoside reverse transcriptase inhibitor (NNRTI) dapivirine termed DAPY, when used together with exposure to ultraviolet light, was able to completely and irreversibly inactivate both HIV-1 RT activity as well as infectiousness in each of a T cell line and peripheral blood mononuclear cells. Control experiments using various concentrations of DAPY revealed that a combination of exposure to ultraviolet light together with use of the specific, high affinity photo-labeled compound was necessary for complete inactivation to occur. This method of HIV RT inactivation may have applicability toward preservation of an intact viral structure and warrants further investigation in regard to the potential of this approach to elicit a durable, broad protective immune response. Copyright © 2010 Elsevier B.V. All rights reserved.

Structural optimization of N1-aryl-benzimidazoles for the discovery of new non-nucleoside reverse transcriptase inhibitors active against wild-type and mutant HIV-1 strains.

PubMed

Monforte, Anna Maria; De Luca, Laura; Buemi, Maria Rosa; Agharbaoui, Fatima E; Pannecouque, Christophe; Ferro, Stefania

2018-02-01

Non-nucleoside reverse transcriptase inhibitors (NNRTIs) are recommended components of preferred combination antiretroviral therapies used for the treatment of human immunodeficiency virus (HIV) infection. These regimens are extremely effective in suppressing virus replication. Recently, our research group identified some N 1 -aryl-2-arylthioacetamido-benzimidazoles as a novel class of NNRTIs. In this research work we report the design, the synthesis and the structure-activity relationship studies of new compounds (20-34) in which some structural modifications have been introduced in order to investigate their effects on reverse transcriptase (RT) inhibition and to better define the features needed to increase the antiviral activity. Most of the new compounds proved to be highly effective in inhibiting both RT enzyme at nanomolar concentrations and HIV-1 replication in MT4 cells with minimal cytotoxicity. Among them, the most promising N 1 -aryl-2-arylthioacetamido-benzimidazoles and N 1 -aryl-2-aryloxyacetamido-benzimidazoles were also tested toward a panel of single- and double-mutants strain responsible for resistance to NNRTIs, showing in vitro antiviral activity toward single mutants L100I, K103N, Y181C, Y188L and E138K. The best results were observed for derivatives 29 and 33 active also against the double mutants F227L and V106A. Computational approaches were applied in order to rationalize the potency of the new synthesized inhibitors. Copyright © 2017 Elsevier Ltd. All rights reserved.

The future of pre-exposure prophylaxis (PrEP) for human immunodeficiency virus (HIV) infection.

PubMed

Özdener, Ayşe Elif; Park, Tae Eun; Kalabalik, Julie; Gupta, Rachna

2017-05-01

People at high risk for HIV acquisition should be offered pre-exposure prophylaxis (PrEP). Tenofovir disoproxil fumarate (TDF)/emtricitabine (FTC) is currently the only medication recommended for pre-exposure prophylaxis (PrEP) by the Centers for Disease Control and Prevention (CDC) in people at high risk for HIV acquisition. This article will review medications currently under investigation and the future landscape of PrEP therapy. Areas covered: This article will review clinical trials that have investigated nontraditional regimens of TDF/FTC, antiretroviral agents from different drug classes such as integrase strand transfer inhibitors (INSTI), nucleoside reverse transcriptase inhibitors (NRTI), and non-nucleoside reverse transcriptase inhibitors (NNRTI) as potential PrEP therapies. Expert commentary: Currently, there are several investigational drugs in the pipeline for PrEP against HIV infection. Increased utilization of PrEP therapy depends on provider identification of people at high risk for HIV transmission. Advances in PrEP development will expand options and access for people and reduce the risk of HIV acquisition.

Antiretroviral therapy for human immunodeficiency virus infection in 1997.

PubMed Central

Katzenstein, D A

1997-01-01

It has become clear that the acquired immunodeficiency syndrome follows continuous replication of the human immunodeficiency virus (HIV) and a decrease in immune capability, most obviously a decline in the number of CD4 lymphocytes. An understanding of key elements in the infectious life cycle of HIV has led to the development of potent antiretroviral drugs selectively targeting unique reverse transcriptase and protease enzymes of the virus. Completed clinical trials have shown that antiretroviral therapy for HIV infection, begun early, reduces viral replication and reverses the decline in CD4 lymphocyte numbers. Recent studies of combination therapies have shown that decreases in plasma HIV viremia to low levels and sustained increases in CD4 cell numbers are associated with longer survival. Potent combination regimens including protease inhibitors and non-nucleoside reverse transcriptase inhibitors suppress detectable viral replication and have demonstrated clinical benefits in patients with advanced disease. Progress in antiretroviral therapy and methods to monitor responses to treatment are providing new hope in the treatment of HIV infection. PMID:9217434

Dipyridodiazepinone analogs as human immunodeficiency virus type 1-specific non-nucleoside reverse transcriptase inhibitors: an overview.

PubMed

Lv, M; Xu, H

2010-01-01

According to World Health Organization (WHO)/Joint United Nations Programme on human immunodeficiency virus (HIV)/acquired immune deficiency syndrome (AIDS) (UNAIDS) Report in 2007, 33.2 million people are living with HIV, 2.5 million ones have been newly infected with HIV, and 2.1 million ones died from AIDS, including 330,000 children. Therefore, HIV/AIDS still remains a public health emergency and a leading cause of mortality worldwide. It is believed that reverse transcriptase (RT) is a crucial enzyme in the life cycle of HIV-1, and thereby RT has been the important drug target for antiretroviral (ARV) chemotherapy against AIDS. To our knowledge, dipyridodiazepinone analogs have been considered as one class of potential non-nucleoside reverse transcriptase inhibitors (NNRTIs), especially the structurally and chemically related nevirapine (Viramune(R)), which was the first NNRTI approved by the U. S. Food and Drug Administration (FDA) for the treatment of HIV-1 infection for adults in 1996 and for children in 1998. This review mainly highlights the progress of synthesis and structure-activity relationship (SAR) of dipyridodiazepinone analogs; in the meantime, the mechanism of action is also presented. It will pave the way for the design and development of novel dipyridodiazepinone analogs as NNRTIs in AIDS chemotherapy in the future.

Towards discovering dual functional inhibitors against both wild type and K103N mutant HIV-1 reverse transcriptases: molecular docking and QSAR studies on 4,1-benzoxazepinone analogues

NASA Astrophysics Data System (ADS)

Zhang, Zhenshan; Zheng, Mingyue; Du, Li; Shen, Jianhua; Luo, Xiaomin; Zhu, Weiliang; Jiang, Hualiang

2006-05-01

To find useful information for discovering dual functional inhibitors against both wild type (WT) and K103N mutant reverse transcriptases (RTs) of HIV-1, molecular docking and 3D-QSAR approaches were applied to a set of twenty-five 4,1-benzoxazepinone analogues of efavirenz (SUSTIVA®), some of them are active against the two RTs. 3D-QSAR models were constructed, based on their binding conformations determined by molecular docking, with r 2 cv values ranging from 0.656 to 0.834 for CoMFA and CoMSIA, respectively. The models were then validated to be highly predictive and extrapolative by inhibitors in two test sets with different molecular skeletons. Furthermore, CoMFA models were found to be well matched with the binding sites of both WT and K103N RTs. Finally, a reasonable pharmacophore model of 4,1-benzoxazepinones were established. The application of the model not only successfully differentiated the experimentally determined inhibitors from non-inhibitors, but also discovered two potent inhibitors from the compound database SPECS. On the basis of both the 3D-QSAR and pharmacophore models, new clues for discovering and designing potent dual functional drug leads against HIV-1 were proposed: (i) adopting positively charged aliphatic group at the cis-substituent of C3; (ii) reducing the electronic density at the position of O4; (iii) positioning a small branched aliphatic group at position of C5; (iv) using the negatively charged bulky substituents at position of C7.

Base modifications affecting RNA polymerase and reverse transcriptase fidelity.

PubMed

Potapov, Vladimir; Fu, Xiaoqing; Dai, Nan; Corrêa, Ivan R; Tanner, Nathan A; Ong, Jennifer L

2018-06-20

Ribonucleic acid (RNA) is capable of hosting a variety of chemically diverse modifications, in both naturally-occurring post-transcriptional modifications and artificial chemical modifications used to expand the functionality of RNA. However, few studies have addressed how base modifications affect RNA polymerase and reverse transcriptase activity and fidelity. Here, we describe the fidelity of RNA synthesis and reverse transcription of modified ribonucleotides using an assay based on Pacific Biosciences Single Molecule Real-Time sequencing. Several modified bases, including methylated (m6A, m5C and m5U), hydroxymethylated (hm5U) and isomeric bases (pseudouridine), were examined. By comparing each modified base to the equivalent unmodified RNA base, we can determine how the modification affected cumulative RNA polymerase and reverse transcriptase fidelity. 5-hydroxymethyluridine and N6-methyladenosine both increased the combined error rate of T7 RNA polymerase and reverse transcriptases, while pseudouridine specifically increased the error rate of RNA synthesis by T7 RNA polymerase. In addition, we examined the frequency, mutational spectrum and sequence context of reverse transcription errors on DNA templates from an analysis of second strand DNA synthesis.

Antiretroviral treatment sequencing strategies to overcome HIV type 1 drug resistance in adolescents and adults in low-middle-income countries.

PubMed

De Luca, Andrea; Hamers, Raphael L; Schapiro, Jonathan M

2013-06-15

Antiretroviral treatment (ART) is expanding to human immunodeficiency virus type 1 (HIV-1)-infected persons in low-middle income countries, thanks to a public health approach. With 3 available drug classes, 2 ART sequencing lines are programmatically foreseen. The emergence and transmission of viral drug resistance represents a challenge to the efficacy of ART. Knowledge of HIV-1 drug resistance selection associated with specific drugs and regimens and the consequent activity of residual drug options are essential in programming ART sequencing options aimed at preserving ART efficacy for as long as possible. This article determines optimal ART sequencing options for overcoming HIV-1 drug resistance in resource-limited settings, using currently available drugs and treatment monitoring opportunities. From the perspective of drug resistance and on the basis of limited virologic monitoring data, optimal sequencing seems to involve use of a tenofovir-containing nonnucleoside reverse-transcriptase inhibitor-based first-line regimen, followed by a zidovudine-containing, protease inhibitor (PI)-based second-line regimen. Other options and their consequences are explored by considering within-class and between-class sequencing opportunities, including boosted PI monotherapies and future options with integrase inhibitors. Nucleoside reverse-transcriptase inhibitor resistance pathways in HIV-1 subtype C suggest an additional reason for accelerating stavudine phase out. Viral load monitoring avoids the accumulation of resistance mutations that significantly reduce the activity of next-line options. Rational use of resources, including broader access to viral load monitoring, will help ensure 3 lines of fully active treatment options, thereby increasing the duration of ART success.

Mitochondrial DNA replication, nucleoside reverse-transcriptase inhibitors, and AIDS cardiomyopathy.

PubMed

Lewis, William

2003-01-01

Nucleoside reverse-transcriptase inhibitors (NRTIs) in combination with other antiretrovirals (HAART) are the cornerstones of current AIDS therapy, but extensive use brought mitochondrial side effects to light. Clinical experience, pharmacological, cell, and molecular biological evidence links altered mitochondrial (mt-) DNA replication to the toxicity of NRTIs in many tissues, and conversely, mtDNA replication defects and mtDNA depletion in target tissues are observed. Organ-specific pathological changes or diverse systemic effects result from and are frequently attributed to HAART in which NRTIs are included. The shared features of mtDNA depletion and energy depletion became key observations and related the clinical and in vivo experimental findings to inhibition of mtDNA replication by NRTI triphosphates in vitro. Subsequent to those findings, other observations suggested that mitochondrial energy deprivation is concomitant with or the result of mitochondrial oxidative stress in AIDS (from HIV, for example) or from NRTI therapy itself. Copyright 2003, Elsevier Science (USA)

Synthesis, biological evaluation and molecular modeling of 2-Hydroxyisoquinoline-1,3-dione analogues as inhibitors of HIV reverse transcriptase associated ribonuclease H and polymerase.

PubMed

Tang, Jing; Vernekar, Sanjeev Kumar V; Chen, Yue-Lei; Miller, Lena; Huber, Andrew D; Myshakina, Nataliya; Sarafianos, Stefan G; Parniak, Michael A; Wang, Zhengqiang

2017-06-16

Human immunodeficiency virus (HIV) reverse transcriptase (RT) associated ribonuclease H (RNase H) remains the only virally encoded enzymatic function not clinically validated as an antiviral target. 2-Hydroxyisoquinoline-1,3-dione (HID) is known to confer active site directed inhibition of divalent metal-dependent enzymatic functions, such as HIV RNase H, integrase (IN) and hepatitis C virus (HCV) NS5B polymerase. We report herein the synthesis and biochemical evaluation of a few C-5, C-6 or C-7 substituted HID subtypes as HIV RNase H inhibitors. Our data indicate that while some of these subtypes inhibited both the RNase H and polymerase (pol) functions of RT, potent and selective RNase H inhibition was achieved with subtypes 8-9 as exemplified with compounds 8c and 9c. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

Vaginal Microbicide Film Combinations of Two Reverse Transcriptase Inhibitors, EFdA and CSIC, for the Prevention of HIV-1 Sexual Transmission.

PubMed

Zhang, Wei; Hu, Minlu; Shi, Yuan; Gong, Tiantian; Dezzutti, Charlene S; Moncla, Bernard; Sarafianos, Stefan G; Parniak, Michael A; Rohan, Lisa C

2015-09-01

EFdA is a potent nucleoside reverse transcriptase inhibitor (NRTI) with activity against a wide spectrum of wild-type and drug resistant HIV-1 variants. CSIC is a tight-binding non-nucleoside reverse transcriptase inhibitor (NNRTI) with demonstrated anti-HIV properties important for use in topical prevention of HIV transmission. The objective of this study was to develop and characterize film-formulated EFdA and CSIC for use as a female-controlled vaginal microbicide to prevent sexual transmission of HIV. Assessments of EFdA- and CSIC-loaded films included physicochemical characteristics, in vitro cytotoxicity, epithelia integrity studies, compatibility with the normal vaginal Lactobacillus flora and anti-HIV bioactivity evaluations. No significant change in physicochemical properties or biological activity of the combination films were noted during 3 months storage. In vitro cytotoxicity and bioactivity testing showed that 50% cytotoxic concentration (CC50) of either EFdA or CSIC was several orders of magnitude higher than the 50% effective concentration (EC50) values. Film-formulated EFdA and CSIC combination showed additive inhibitory activity against wild type and drug-resistant variants of HIV. Epithelial integrity studies demonstrated that the combination vaginal film had a much lower toxicity to HEC-1A monolayers compared to that of VCF®, a commercial vaginal film product containing nonoxynol-9. Polarized ectocervical explants showed films with drug alone or in combination were effective at preventing HIV infection. Our data suggest that vaginal microbicide films containing a combination of the NRTI EFdA and the NNRTI CSIC have potential to prevent HIV-1 sexual transmission.

An Intravaginal Ring for the Simultaneous Delivery of an HIV-1 Maturation Inhibitor and Reverse-Transcriptase Inhibitor for Prophylaxis of HIV Transmission.

PubMed

Ugaonkar, Shweta R; Clark, Justin T; English, Lexie B; Johnson, Todd J; Buckheit, Karen W; Bahde, Robert J; Appella, Daniel H; Buckheit, Robert W; Kiser, Patrick F

2015-10-01

Nucleocapsid 7 (NCp7) inhibitors have been investigated extensively for their role in impeding the function of HIV-1 replication machinery and their ability to directly inactivate the virus. A class of NCp7 zinc finger inhibitors, S-acyl-2-mercaptobenzamide thioesters (SAMTs), was investigated for topical drug delivery. SAMTs are inherently unstable because of their hydrolytically labile thioester bond, thus requiring formulation approaches that can lend stability. We describe the delivery of N-[2-(3,4,5-trimethoxybenzoylthio)benzoyl]-β-alaninamide (SAMT-10), as a single agent antiretroviral (ARV) therapeutic and in combination with the HIV-1 reverse-transcriptase inhibitor pyrimidinedione IQP-0528, from a hydrophobic polyether urethane (PEU) intravaginal ring (IVR) for a month. The physicochemical stability of the ARV-loaded IVRs was confirmed after 3 months at 40°C/75% relative humidity. In vitro, 25 ± 3 mg/IVR of SAMT-10 and 86 ± 13 mg/IVR of IQP-0528 were released. No degradation of the hydrolytically labile SAMT-10 was observed within the matrix. The combination of ARVs had synergistic antiviral activity when tested in in vitro cell-based assays. Toxicological evaluations performed on an organotypic EpiVaginal(™) tissue model demonstrated a lack of formulation toxicity. Overall, SAMT-10 and IQP-0528 were formulated in a stable PEU IVR for sustained release. Our findings support the need for further preclinical evaluation. © 2015 Wiley Periodicals, Inc. and the American Pharmacists Association J Pharm Sci 104:3426-3439, 2015. © 2015 Wiley Periodicals, Inc. and the American Pharmacists Association.

2-(Alkyl/aryl)amino-6-benzylpyrimidin-4(3H)-ones as inhibitors of wild-type and mutant HIV-1: enantioselectivity studies.

PubMed

Rotili, Dante; Samuele, Alberta; Tarantino, Domenico; Ragno, Rino; Musmuca, Ira; Ballante, Flavio; Botta, Giorgia; Morera, Ludovica; Pierini, Marco; Cirilli, Roberto; Nawrozkij, Maxim B; Gonzalez, Emmanuel; Clotet, Bonaventura; Artico, Marino; Esté, José A; Maga, Giovanni; Mai, Antonello

2012-04-12

The single enantiomers of two pyrimidine-based HIV-1 non-nucleoside reverse transcriptase inhibitors, 1 (MC1501) and 2 (MC2082), were tested in both cellular and enzyme assays. In general, the R forms were more potent than their S counterparts and racemates and (R)-2 was more efficient than (R)-1 and the reference compounds, with some exceptions. Interestingly, (R)-2 displayed a faster binding to K103N RT with respect to WT RT, while (R)-1 showed the opposite behavior. © 2012 American Chemical Society

Clinical and virologic follow-up in perinatally HIV-1-infected children and adolescents in Madrid with triple-class antiretroviral drug-resistant viruses.

PubMed

Rojas Sánchez, P; de Mulder, M; Fernandez-Cooke, E; Prieto, L; Rojo, P; Jiménez de Ory, S; José Mellado, M; Navarro, M; Tomas Ramos, J; Holguín, Á

2015-06-01

Drug resistance mutations compromise the success of antiretroviral treatment in human immunodeficiency virus type 1 (HIV-1)-infected children. We report the virologic and clinical follow-up of the Madrid cohort of perinatally HIV-infected children and adolescents after the selection of triple-class drug-resistant mutations (TC-DRM). We identified patients from the cohort carrying HIV-1 variants with TC-DRM to nucleoside reverse transcriptase inhibitors, nonnucleoside reverse transcriptase inhibitors and protease inhibitors according to IAS-USA-2013. We recovered pol sequences or resistance profiles from 2000 to 2011 and clinical-immunologic-virologic data from the moment of TC-DRM detection until December 2013. Viruses harbouring TC-DRM were observed in 48 (9%) of the 534 children and adolescents from 2000 to 2011, rising to 24.4% among those 197 with resistance data. Among them, 95.8% were diagnosed before 2003, 91.7% were Spaniards, 89.6% carried HIV-1-subtype B and 75% received mono/dual therapy as first regimen. The most common TC-DRM present in ≥50% of them were D67NME, T215FVY, M41L and K103N (retrotranscriptase) and L90M (protease). The susceptibility to darunavir, tipranavir, etravirine and rilpivirine was 67.7%, 43.7%, 33.3% and 33.3%, respectively, and all reported high resistance to didanosine, abacavir and nelfinavir. Despite the presence of HIV-1 resistance mutations to the three main antiretroviral families in our paediatric cohort, some drugs maintained their susceptibility, mainly the new protease inhibitors (tipranavir and darunavir) and nonnucleoside reverse transcriptase inhibitors (etravirine and rilpivirine). These data will help to improve the clinical management of HIV-infected children with triple resistance in Spain. Copyright © 2015 European Society of Clinical Microbiology and Infectious Diseases. Published by Elsevier Ltd. All rights reserved.

Executive summary of the GeSIDA/National AIDS Plan consensus document on antiretroviral therapy in adults infected by the human immunodeficiency virus (updated January 2014).

PubMed

Berenguer, Juan; Polo, Rosa; Lozano, Fernando; López Aldeguer, José; Antela, Antonio; Arribas, José Ramón; Asensi, Víctor; Blanco, José Ramón; Clotet, Bonaventura; Domingo, Pere; Galindo, María José; Gatell, José María; González-García, Juan; Iribarren, José Antonio; Locutura, Jaime; López, Juan Carlos; Mallolas, Josep; Martínez, Esteban; Miralles, Celia; Miró, José M; Moreno, Santiago; Palacios, Rosario; Pérez Elías, María Jesús; Pineda, Juan Antonio; Podzamczer, Daniel; Portilla, Joaquín; Pulido, Federico; Ribera, Esteban; Riera, Melchor; Rubio, Rafael; Santos, Jesús; Sanz, Jesús; Tuset, Montserrat; Vidal, Francesc; Rivero, Antonio

2014-01-01

In this update, antiretroviral therapy (ART) is recommended for all patients infected by type 1 human immunodeficiency virus (HIV-1). The strength and grade of the recommendation varies with clinical circumstances, number of CD4 cells, comorbid conditions and prevention of transmission of HIV. The objective of ART is to achieve an undetectable plasma viral load. Initial ART should always comprise a combination of 3 drugs, including 2 nucleoside reverse transcriptase inhibitors and a third drug from a different family (non-nucleoside reverse transcriptase inhibitor, protease inhibitor, or integrase inhibitor). This update presents the causes and criteria for switching ART in patients with undetectable plasma viral load and in cases of virological failure. An update is also provided for the specific criteria for ART in special situations (acute infection, HIV-2 infection, and pregnancy) and with comorbid conditions (tuberculosis or other opportunistic infections, kidney disease, liver disease, and cancer). Copyright © 2014 Elsevier España, S.L.U. y Sociedad Española de Enfermedades Infecciosas y Microbiología Clínica. All rights reserved.

[Use of Nadis(®) software to improve adverse drug reaction reporting of antiretroviral drugs: experience in south west of France (midi-pyrénées)].

PubMed

Pochard, Liselotte; Hauviller, Laurent; Cuzin, Lise; Eyvrard, Fréderic; Sommet, Agnès; Montastruc, Jean-Louis; Bagheri, Haleh

2014-01-01

To study the value of the module of pharmacovigilance in Nadis® to improve the antiretroviral (ARV) drugs-induced adverse drug reactions (ADRs) reporting. We collected the ADRs reported for 17 months from November 2010 until April 2012. Following data were recorded: characteristics of patients, ADRs, ARV drugs. The number of ADRs was compared to those collected in the same period (17 months) before use of Nadis®. The 119 ADRs reported (an increase of 183%) for 109 patients ADRs were mainly gastrointestinal (21.8%) followed by renal (20.2%), neuro-psychiatric (16.8%), hepatic (13.5%), cutaneous (8.4%), metabolic (6.7%) and others (12.6%). The repartition of ARV drugs was: nucleoside (31.8%), nucleotide (13.6%) reverse transcriptase inhibitors respectively, non-nucleoside reverse transcriptase inhibitors (13.1%), protease inhibitors (36.4%), and integrase inhibitors (5.1%). Our results show the improvement of ARV-induced ADRs reporting by Nadis® which could be used to reduce the rate of under-reporting in patients exposed to these drugs. © 2014 Société Française de Pharmacologie et de Thérapeutique.

Synthesis, Biological Activity, and Crystal Structure of Potent Nonnucleoside Inhibitors of HIV-1 Reverse Transcriptase That Retain Activity against Mutant Forms of the Enzyme†

PubMed Central

Morningstar, Marshall L.; Roth, Thomas; Farnsworth, David W.; Smith, Marilyn Kroeger; Watson, Karen; Buckheit, Robert W.; Das, Kalyan; Zhang, Wanyi; Arnold, Eddy; Julias, John G.; Hughes, Stephen H.; Michejda, Christopher J.

2010-01-01

In an ongoing effort to develop novel and potent nonnucleoside HIV-1 reverse transcriptase (RT) inhibitors that are effective against the wild type (WT) virus and clinically observed mutants, 1,2-bis-substituted benzimidazoles were synthesized and tested. Optimization of the N1 and C2 positions of benzimidazole led to the development of 1-(2,6-difluorobenzyl)-2-(2,6-difluorophenyl)-4-methylbenzimidazole (1) (IC50 = 0.2 μM, EC50 = 0.44 μM, and TC50 ≥ 100 against WT). This paper describes how substitution on the benzimidazole ring profoundly affects activity. Substituents at the benzimidazole C4 dramatically enhanced potency, while at C5 or C6 substituents were generally detrimental or neutral to activity, respectively. A 7-methyl analogue did not inhibit HIV-1 RT. Determination of the crystal structure of 1 bound to RT provided the basis for accurate modeling of additional analogues, which were synthesized and tested. Several derivatives were nanomolar inhibitors of wild-type virus and were effective against clinically relevant HIV-1 mutants. PMID:17663538

Molecular Characterization of the Human Immunodeficiency Virus Type 1 in Women and Their Vertically Infected Children.

PubMed

Vaz, Sara Nunes; Giovanetti, Marta; Rego, Filipe Ferreira de Almeida; Oliveira, Tulio de; Danaviah, Siva; Gonçalves, Maria Luiza Freire; Alcantara, Luiz Carlos Junior; Brites, Carlos

2015-10-01

Approximately 35 million people worldwide are infected with human immunodeficiency virus (HIV) around 3.2 million of whom are children under 15 years. Mother-to-child-transmission (MTCT) of HIV-1 accounts for 90% of all infections in children. Despite great advances in the prevention of MTCT in Brazil, children are still becoming infected. Samples from 19 HIV-1-infected families were collected. DNA was extracted and fragments from gag, pol, and env were amplified and sequenced directly. Phylogenetic reconstruction was performed. Drug resistance analyses were performed in pol and env sequences. We found 82.1% of subtype B and 17.9% of BF recombinants. A prevalence of 43.9% drug resistance-associated mutations in pol sequences was identified. Of the drug-naive children 33.3% presented at least one mutation related to protease inhibitor/nucleoside reverse transcriptase inhibitor/nonnucleoside reverse transcriptase inhibitor (PI/NRTI/NNRTI) resistance. The prevalence of transmitted drug resistance mutations was 4.9%. On env we found a low prevalence of HR1 (4.9%) and HR2 (14.6%) mutations.

Micronuclei induced by reverse transcriptase inhibitors in mononucleated and binucleated cells as assessed by the cytokinesis-block micronucleus assay

PubMed Central

2010-01-01

This study evaluated the clastogenic and/or aneugenic potential of three nucleoside reverse transcriptase inhibitors (zidovudine - AZT, lamivudine - 3TC and stavudine - d4T) using the cytokinesis-block micronucleus (CBMN) assay in human lymphocyte cultures. All three inhibitors produced a positive response when tested in binucleated cells. The genotoxicity of AZT and 3TC was restricted to binucleated cells since there was no significant increase in the frequency of micronuclei in mononucleated cells. This finding indicated that AZT and 3TC caused chromosomal breakage and that their genotoxicity was related to a clastogenic action. In addition to the positive response observed with d4T in binucleated cells, this drug also increased the frequency of micronuclei in mononucleated cells, indicating clastogenic and aneugenic actions. Since the structural differences between AZT and 3TC and AZT and d4T involve the 3' position in the 2'-deoxyribonucleoside and in an unsaturated 2',3',dideoxyribose, respectively, we suggest that an unsaturated 2', 3', dideoxyribose is responsible for the clastogenic and aneugenic actions of d4T. PMID:21637587

Efficacy and durability of nevirapine in antiretroviral drug näive patients.

PubMed

Lange, Joep M A

2003-09-01

Nevirapine is a non-nucleoside reverse transcriptase inhibitor (NNRTI) that was first reported in the scientific literature in 1990. Varying doses of nevirapine (NVP) and a number of regimens containing this NNRTI have been studied in antiretroviral (ARV) näive patients. Four key studies have compared the efficacy and safety of triple drug regimens containing NVP in ARV näive, HIV-1 infected patients. The INCAS study was the first demonstration of how to use NVP in an effective and durable manner: as a component of a triple drug regimen. The COMBINE Study was a comparison of protease inhibitor (PI)-based and NVP-based triple regimens. The Atlantic Study is comparing the safety and efficacy of three triple drug regimens in ARV näive patients. In this study, treatment consists of a divergent drug regimen (PI and nucleoside reverse transcriptase inhibitors, NRTIs) targeting both HIV-1 protease and reverse transcriptase or a convergent regimen targeting reverse transcriptase alone (three NRTIs or two NRTIs plus a NNRTI). A clinical endpoint study (BI 1090) compared the efficacy and durability of multi-drug regimens in ARV näive patients with high baseline plasma HIV-1 RNA levels (pVLs) and low peripheral blood CD4+ lymphocyte counts. Data from these studies confirm that triple regimens containing NVP suppressed viral replication for up to one year, even when the ARV näive patients had low CD4+ cell counts at baseline. Nevirapine-containing regimens suppressed pVLs to < 50 copies/ mL in approximately 50% of patients in the studies discussed (Intent to Treat analyses). Data from 96 weeks of follow up in the Atlantic Study demonstrates that the regimens containing didanosine and stavudine plus indinavir or NVP were significantly more successful in suppressing pVLs to < 50 copies/mL during this period than a regimen composed of these NRTIs and lamivudine (p < or = 0.001). As with other ARV drugs, NVP should always be used as part of a fully suppressive ARV regimen. When used in this way, it is an effective ARV drug, which contributes to durable virological and immunological responses in approximately half of all treated patients. Nevirapine-containing regimens are effective in patients with advanced HIV-1 infection, i.e., low CD4+ cell counts. Data will soon be available from the 2NN Study that compares the efficacy and safety of four different regimens using NVP once daily, NVP twice daily, efavirenz once daily or a combination of NVP and efavirenz. All four arms of the study include a backbone of stavudine and lamivudine.

Applications of 2D IR spectroscopy to peptides, proteins, and hydrogen-bond dynamics

PubMed Central

Kim, Yung Sam; Hochstrasser, Robin M.

2010-01-01

Following a survey of 2D IR principles this Feature Article describes recent experiments on the hydrogen-bond dynamics of small ions, amide-I modes, nitrile probes, peptides, reverse transcriptase inhibitors, and amyloid fibrils. PMID:19351162

The MONET trial: darunavir/ritonavir with or without nucleoside analogues, for patients with HIV RNA below 50 copies/ml.

PubMed

Arribas, Jose R; Horban, Andrzej; Gerstoft, Jan; Fätkenheuer, Gerdt; Nelson, Mark; Clumeck, Nathan; Pulido, Federico; Hill, Andrew; van Delft, Yvon; Stark, Thomas; Moecklinghoff, Christiane

2010-01-16

In virologically suppressed patients, darunavir-ritonavir (DRV/r) monotherapy could maintain virological suppression similarly to DRV/r and two nucleosides. Two hundred and fifty-six patients with HIV RNA less than 50 copies/ml for over 24 weeks on current antiretrovirals [non-nucleoside reverse transcriptase inhibitor (NNRTI)-based (43%), or protease inhibitor-based (57%)], switched to DRV/r 800/100 mg once daily, either as monotherapy (n = 127) or with two nucleoside reverse transcriptase inhibitors (NRTIs) (n = 129). Treatment failure was defined as two consecutive HIV RNA levels above 50 copies/ml (TLOVR) by week 48, or switches off study treatment. The trial had 80% power to show noninferiority for the monotherapy arm (delta = -12%). Patients were 81% male and 91% Caucasian, with mean age 44 years, and CD4 cell count of 574 cells/microl. In the primary efficacy analysis, HIV RNA less than 50 copies/ml by week 48 (per protocol) was 86.2 versus 87.8% in the monotherapy and triple therapy arms; by intent-to-treat switch equals failure, efficacy was 84.3 versus 85.3%; by a switch-included analysis, efficacy was 93.5 versus 95.1%: all three comparisons showed noninferior efficacy for DRV/r monotherapy. CD4 cell counts remained stable during the trial in both arms. One patient per arm showed at least one protease inhibitor mutation, and one patient in the triple therapy arm showed an NRTI mutation. Nine patients per arm discontinued randomized treatment for either adverse events or other reasons. No new or unexpected safety signals were detected. In this study for patients with HIV RNA less than 50 copies/ml on other antiretrovirals at baseline, switching to DRV/r monotherapy showed noninferior efficacy versus triple antiretroviral therapy.

The molecular basis of resilience to the effect of the Lys103Asn mutation in non-nucleoside HIV-1 reverse transcriptase inhibitors studied by targeted molecular dynamics simulations.

PubMed

Rodríguez-Barrios, Fátima; Balzarini, Jan; Gago, Federico

2005-05-25

A series of targeted molecular dynamics simulations have been carried out in an attempt to assess the effect that the common Lys103Asn mutation in HIV-1 reverse transcriptase (RT) has on the binding of three representative non-nucleoside RT inhibitors (NNRTI), nevirapine, efavirenz, and etravirine. We have shown previously that, in the absence of an incoming inhibitor, creation of the NNRTI binding pocket is hampered due to the existence of a hydrogen bond between the side chains of Asn103 and Tyr188 for which no equivalent exists in the wild-type enzyme. As an extension of this work, we now apply the same methodology to drive the enzyme's conformation from the unbound state to the drug-bound state in the presence of the NNRTI. The location of each drug outside the binding pocket was determined by an automated docking program, and steering into the binding pocket followed a route that is likely to represent the actual entrance pathway. The additional hurdle to inhibitor entry imposed by the extra Asn103-Tyr188 hydrogen bond is seen to affect each NNRTI differently, with the ability to disrupt this interaction increasing in the order etravirine > efavirenz > or = nevirapine, in good accord with the experimental findings. This coherent picture strongly suggests that attempts to overcome resistance through structure-based drug design may be considerably more successful if dynamic structural aspects of the type studied here are considered, particularly in cases where binding energy-based structure-activity relationship methods are unable to provide the required information.

Vaginal microbicide film combinations of two reverse transcriptase inhibitors, EFdA and CSIC, for the prevention of HIV-1 sexual transmission

PubMed Central

Zhang, Wei; Hu, Minlu; Shi, Yuan; Gong, Tiantian; Dezzutti, Charlene S.; Moncla, Bernard; Sarafianos, Stefan G.; Parniak, Michael A.; Rohan, Lisa C.

2015-01-01

Purpose EFdA is a potent nucleoside reverse transcriptase inhibitor (NRTI) with activity against a wide spectrum of wild-type and drug resistant HIV-1 variants. CSIC is a tight-binding non-nucleoside reverse transcriptase inhibitor (NNRTI) with demonstrated anti-HIV properties important for use in topical prevention of HIV transmission. The objective of this study was to develop and characterize film-formulated EFdA and CSIC for use as a female-controlled vaginal microbicide to prevent sexual transmission of HIV. Methods Assessments of EFdA- and CSIC-loaded films included physicochemical characteristics, in vitro cytotoxicity, epithelia integrity studies, compatibility with the normal vaginal Lactobacillus flora and anti-HIV bioactivity evaluations. Results No significant change in physicochemical properties or biological activity of the combination films were noted during 3 months storage. In vitro cytotoxicity and bioactivity testing showed that 50% cytotoxic concentration (CC50) of either EFdA or CSIC was several orders of magnitude higher than the 50% effective concentration (EC50) values. Film-formulated EFdA and CSIC combination showed additive inhibitory activity against wild type and drug-resistant variants of HIV. Epithelial integrity studies demonstrated that the combination vaginal film had a much lower toxicity to HEC-1A monolayers compared to that of VCF®, a commercial vaginal film product containing nonoxynol-9. Polarized ectocervical explants showed films with drug alone or in combination were effective at preventing HIV infection. Conclusions Our data suggest that vaginal microbicide films containing a combination of the NRTI EFdA and the NNRTI CSIC have potential to prevent HIV-1 sexual transmission. PMID:25794967

Transmitted drug resistance is still low in newly diagnosed human immunodeficiency virus type 1 CRF06_cpx-infected patients in Estonia in 2010.

PubMed

Avi, Radko; Huik, Kristi; Pauskar, Merit; Ustina, Valentina; Karki, Tõnis; Kallas, Eveli; Jõgeda, Ene-Ly; Krispin, Tõnu; Lutsar, Irja

2014-03-01

The presence of transmitted drug resistance (TDR) in treatment-naive HIV-1-positive subjects is of concern, especially in the countries of the former Soviet Union in which the number of subjects exposed to antiretrovirals (ARV) has exponentially increased during the past decade. We assessed the rate of TDR among newly diagnosed subjects in Estonia in 2010 and compared it to that in 2008. The study included 325 subjects (87% of all subjects tested HIV positive from January 1 to December 31, 2010). Of the 244 sequenced viral genomic RNA in the reverse transcriptase (RT) region 214 were CRF06_cpx, nine were subtype A1, three (one each) were subtype B and subtype C, CRF02_AG, and CRF03_AB; 15 viruses remained unclassified as putative recombinant forms between CRF06_cpx and subtype A1. HIV-1 TDR mutations in 2010 and 2008 (n=145) occurred at similar frequency in 4.5% (95% CI 2.45; 7.98) and 5.5% (95% CI 1.8; 9.24) of the patients, respectively. In 2010, 2.5% (6/244) of the sequences harbored nonnucleoside reverse transcriptase inhibitor (NNRTI) (K103N and K101E), 1.6% (4/244) nucleoside reverse transcriptase inhibitor (NRTI) (M41L, M184I, and K219E), and 0.4% (1/244) protease inhibitor (PI) (V82A) mutations. Our findings indicate that in spite of the increased consumption of ARVs the rate of TDR in Estonia has remained unchanged over the past 3 years. Similar stabilizing or even decreasing trends have been described in Western Europe and North America albeit at higher levels and in different socioeconomic backgrounds.

A decade of HIV-1 drug resistance in the United States: trends and characteristics in a large protease/reverse transcriptase and co-receptor tropism database from 2003 to 2012.

PubMed

Paquet, Agnes C; Solberg, Owen D; Napolitano, Laura A; Volpe, Joseph M; Walworth, Charles; Whitcomb, Jeannette M; Petropoulos, Christos J; Haddad, Mojgan

2014-01-01

Drug resistance testing and co-receptor tropism determination are key components of the management of antiretroviral therapy for HIV-1-infected individuals. The purpose of this study was to examine trends of HIV-1 resistance and viral evolution in the past decade by surveying a large commercial patient testing database. Temporal trends of drug resistance, viral fitness and co-receptor usage among samples submitted for routine phenotypic and genotypic resistance testing to protease inhibitors (PIs), nucleoside reverse transcriptase inhibitors (NRTIs) and non-nucleoside reverse transcriptase inhibitors (NNRTIs), as well as for tropism determination were investigated. Within 62,397 resistant viruses reported from 2003 to 2012, we observed a decreasing trend in the prevalence of three-class resistance (from 25% to 9%) driven by decreased resistance to PIs (43% to 21%) and NRTIs (79% to 57%), while observing a slight increase in NNRTI resistance (68% to 75%). The prevalence of CXCR4-mediated entry among tropism testing samples (n=52,945) declined over time from 47% in 2007 to 40% in 2012. A higher proportion of CXCR4-tropic viruses was observed within samples with three-class resistance (50%) compared with the group with no resistance (36%). Decreased prevalence of three-class resistance and increased prevalence of one-class resistance was observed within samples reported between 2003 and 2012. The fraction of CXCR4-tropic viruses has decreased over time; however, CXCR4 usage was more prevalent among multi-class-resistant samples, which may be due to the more advanced disease stage of treatment-experienced patients. These trends have important implications for clinical practice and future drug discovery and development.

Potent NLRP3 Inflammasome Activation by the HIV Reverse Transcriptase Inhibitor Abacavir.

PubMed

Toksoy, Atiye; Sennefelder, Helga; Adam, Christian; Hofmann, Sonja; Trautmann, Axel; Goebeler, Matthias; Schmidt, Marc

2017-02-17

There is experimental and clinical evidence that some exanthematous allergic drug hypersensitivity reactions are mediated by drug-specific T cells. We hypothesized that the capacity of certain drugs to directly stimulate the innate immune system may contribute to generate drug-specific T cells. Here we analyzed whether abacavir, an HIV-1 reverse transcriptase inhibitor often inducing severe delayed-type drug hypersensitivity, can trigger innate immune activation that may contribute to its allergic potential. We show that abacavir fails to generate direct innate immune activation in human monocytes but potently triggers IL-1β release upon pro-inflammatory priming with phorbol ester or Toll-like receptor stimulation. IL-1β processing and secretion were sensitive to Caspase-1 inhibition, NLRP3 knockdown, and K + efflux inhibition and were not observed with other non-allergenic nucleoside reverse transcriptase inhibitors, identifying abacavir as a specific inflammasome activator. It further correlated with dose-dependent mitochondrial reactive oxygen species production and cytotoxicity, indicating that inflammasome activation resulted from mitochondrial damage. However, both NLRP3 depletion and inhibition of K + efflux mitigated abacavir-induced mitochondrial reactive oxygen species production and cytotoxicity, suggesting that these processes were secondary to NLRP3 activation. Instead, depletion of cardiolipin synthase 1 abolished abacavir-induced IL-1β secretion, suggesting that mitochondrial cardiolipin release may trigger abacavir-induced inflammasome activation. Our data identify abacavir as a novel inflammasome-stimulating drug allergen. They implicate a potential contribution of innate immune activation to medication-induced delayed-type hypersensitivity, which may stimulate new concepts for treatment and prevention of drug allergies. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

Drug Susceptibility and Resistance Mutations After First-Line Failure in Resource Limited Settings

PubMed Central

Wallis, Carole L.; Aga, Evgenia; Ribaudo, Heather; Saravanan, Shanmugam; Norton, Michael; Stevens, Wendy; Kumarasamy, Nagalingeswaran; Bartlett, John; Katzenstein, David

2014-01-01

Background. The development of drug resistance to nucleoside reverse transcriptase inhibitors (NRTIs) and nonnucleoside reverse transcriptase inhibitors (NNRTIs) has been associated with baseline human immunodeficiency virus (HIV)-1 RNA level (VL), CD4 cell counts (CD4), subtype, and treatment failure duration. This study describes drug resistance and levels of susceptibility after first-line virologic failure in individuals from Thailand, South Africa, India, Malawi, Tanzania. Methods. CD4 and VL were captured at AIDs Clinical Trial Group (ACTG) A5230 study entry, a study of lopinavir/ritonavir (LPV/r) monotherapy after first-line virologic failure on an NNRTI regimen. HIV drug-resistance mutation associations with subtype, site, study entry VL, and CD4 were evaluated using Fisher exact and Kruskall–Wallis tests. Results. Of the 207 individuals who were screened for A5230, sequence data were available for 148 individuals. Subtypes observed: subtype C (n = 97, 66%) AE (n = 27, 18%), A1 (n = 12, 8%), and D (n = 10, 7%). Of the 148 individuals, 93% (n = 138) and 96% (n = 142) had at least 1 reverse transcriptase (RT) mutation associated with NRTI and NNRTI resistance, respectively. The number of NRTI mutations was significantly associated with a higher study screening VL and lower study screening CD4 (P < .001). Differences in drug-resistance patterns in both NRTI and NNRTI were observed by site. Conclusions. The degree of NNRTI and NRTI resistance after first-line virologic failure was associated with higher VL at study entry. Thirty-two percent of individuals remained fully susceptible to etravirine and rilpivirine, protease inhibitor resistance was rare. Some level of susceptibility to NRTI remained; however, VL monitoring and earlier virologic failure detection may result in lower NRTI resistance. PMID:24795328

Structure of the HIV-1 reverse transcriptase Q151M mutant: insights into the inhibitor resistance of HIV-1 reverse transcriptase and the structure of the nucleotide-binding pocket of Hepatitis B virus polymerase

DOE Office of Scientific and Technical Information (OSTI.GOV)

Nakamura, Akiyoshi; Tamura, Noriko; Yasutake, Yoshiaki, E-mail: y-yasutake@aist.go.jp

The structure of the HIV-1 reverse transcriptase Q151M mutant was determined at a resolution of 2.6 Å in space group P321. Hepatitis B virus polymerase (HBV Pol) is an important target for anti-HBV drug development; however, its low solubility and stability in vitro has hindered detailed structural studies. Certain nucleotide reverse transcriptase (RT) inhibitors (NRTIs) such as tenofovir and lamivudine can inhibit both HBV Pol and Human immunodeficiency virus 1 (HIV-1) RT, leading to speculation on structural and mechanistic analogies between the deoxynucleotide triphosphate (dNTP)-binding sites of these enzymes. The Q151M mutation in HIV-1 RT, located at the dNTP-binding site,more » confers resistance to various NRTIs, while maintaining sensitivity to tenofovir and lamivudine. The residue corresponding to Gln151 is strictly conserved as a methionine in HBV Pol. Therefore, the structure of the dNTP-binding pocket of the HIV-1 RT Q151M mutant may reflect that of HBV Pol. Here, the crystal structure of HIV-1 RT Q151M, determined at 2.6 Å resolution, in a new crystal form with space group P321 is presented. Although the structure of HIV-1 RT Q151M superimposes well onto that of HIV-1 RT in a closed conformation, a slight movement of the β-strands (β2–β3) that partially create the dNTP-binding pocket was observed. This movement might be caused by the introduction of the bulky thioether group of Met151. The structure also highlighted the possibility that the hydrogen-bonding network among amino acids and NRTIs is rearranged by the Q151M mutation, leading to a difference in the affinity of NRTIs for HIV-1 RT and HBV Pol.« less

Role of the K101E Substitution in HIV-1 Reverse Transcriptase in Resistance to Rilpivirine and Other Nonnucleoside Reverse Transcriptase Inhibitors

PubMed Central

Xu, Hong-Tao; Colby-Germinario, Susan P.; Huang, Wei; Oliveira, Maureen; Han, Yingshan; Quan, Yudong; Petropoulos, Christos J.

2013-01-01

Resistance to the recently approved nonnucleoside reverse transcriptase inhibitor (NNRTI) rilpivirine (RPV) commonly involves substitutions at positions E138K and K101E in HIV-1 reverse transcriptase (RT), together with an M184I substitution that is associated with resistance to coutilized emtricitabine (FTC). Previous biochemical and virological studies have shown that compensatory interactions between substitutions E138K and M184I can restore enzyme processivity and the viral replication capacity. Structural modeling studies have also shown that disruption of the salt bridge between K101 and E138 can affect RPV binding. The current study was designed to investigate the impact of K101E, alone or in combination with E138K and/or M184I, on drug susceptibility, viral replication capacity, and enzyme function. We show here that K101E can be selected in cell culture by the NNRTIs etravirine (ETR), efavirenz (EFV), and dapivirine (DPV) as well as by RPV. Recombinant RT enzymes and viruses containing K101E, but not E138K, were highly resistant to nevirapine (NVP) and delavirdine (DLV) as well as ETR and RPV, but not EFV. The addition of K101E to E138K slightly enhanced ETR and RPV resistance compared to that obtained with E138K alone but restored susceptibility to NVP and DLV. The K101E substitution can compensate for deficits in viral replication capacity and enzyme processivity associated with M184I, while M184I can compensate for the diminished efficiency of DNA polymerization associated with K101E. The coexistence of K101E and E138K does not impair either viral replication or enzyme fitness. We conclude that K101E can play a significant role in resistance to RPV. PMID:24002090

Role of the K101E substitution in HIV-1 reverse transcriptase in resistance to rilpivirine and other nonnucleoside reverse transcriptase inhibitors.

PubMed

Xu, Hong-Tao; Colby-Germinario, Susan P; Huang, Wei; Oliveira, Maureen; Han, Yingshan; Quan, Yudong; Petropoulos, Christos J; Wainberg, Mark A

2013-11-01

Resistance to the recently approved nonnucleoside reverse transcriptase inhibitor (NNRTI) rilpivirine (RPV) commonly involves substitutions at positions E138K and K101E in HIV-1 reverse transcriptase (RT), together with an M184I substitution that is associated with resistance to coutilized emtricitabine (FTC). Previous biochemical and virological studies have shown that compensatory interactions between substitutions E138K and M184I can restore enzyme processivity and the viral replication capacity. Structural modeling studies have also shown that disruption of the salt bridge between K101 and E138 can affect RPV binding. The current study was designed to investigate the impact of K101E, alone or in combination with E138K and/or M184I, on drug susceptibility, viral replication capacity, and enzyme function. We show here that K101E can be selected in cell culture by the NNRTIs etravirine (ETR), efavirenz (EFV), and dapivirine (DPV) as well as by RPV. Recombinant RT enzymes and viruses containing K101E, but not E138K, were highly resistant to nevirapine (NVP) and delavirdine (DLV) as well as ETR and RPV, but not EFV. The addition of K101E to E138K slightly enhanced ETR and RPV resistance compared to that obtained with E138K alone but restored susceptibility to NVP and DLV. The K101E substitution can compensate for deficits in viral replication capacity and enzyme processivity associated with M184I, while M184I can compensate for the diminished efficiency of DNA polymerization associated with K101E. The coexistence of K101E and E138K does not impair either viral replication or enzyme fitness. We conclude that K101E can play a significant role in resistance to RPV.

Allosteric mechanism of quinoline inhibitors for HIV RT-associated RNase with MD simulation and dynamics fluctuation network.

PubMed

Cai, Yi; Liu, Hao; Chen, Haifeng

2018-03-01

The human immunodeficiency virus (HIV) is a retrovirus which infects T lymphocyte of human body and causes immunodeficiency. Reverse transcriptase inhibitors (RTIs) can inhibit some functions of RT, preventing virus synthesis (double-stranded DNA), so that HIV virus replication can be reduced. Experimental results indicate a series of benzimidazole-based inhibitors which target HIV RT-associated RNase to inhibit the reverse transcription of HIV virus. However, the allosteric mechanism is still unclear. Here, molecular dynamics simulations and dynamics fluctuation network analysis were used to reveal the binding mode between the inhibitors and RT-associated RNase. The most active molecule has more hydrophobic and electrostatic interactions than the less active inhibitor. Dynamics correlation network analysis indicates that the most active inhibitor perturbs the network of RT-associated RNase and decreases the correlation of nodes. 3D-QSAR model suggests that two robust and reliable models were constructed and validated by independent test set. 3D-QSAR model also shows that bulky negatively charged or hydrophilic substituent is favorable to bioactivity. These results reveal the allosteric mechanism of quinoline inhibitors and help to improve the bioactivity. © 2017 John Wiley & Sons A/S.

Executive summary of the GESIDA/National AIDS Plan Consensus Document on antiretroviral therapy in adults infected by the human immunodeficiency virus (updated January 2015).

PubMed

Berenguer, Juan; Polo, Rosa; Aldeguer, José López; Lozano, Fernando; Aguirrebengoa, Koldo; Arribas, José Ramón; Blanco, José Ramón; Boix, Vicente; Casado, José Luis; Clotet, Bonaventura; Crespo, Manuel; Domingo, Pere; Estrada, Vicente; García, Federico; Gatell, José María; González-García, Juan; Gutiérrez, Félix; Iribarren, José Antonio; Knobel, Hernando; Llibre, Josep María; Locutura, Jaime; López, Juan Carlos; Miró, José M; Moreno, Santiago; Podzamczer, Daniel; Portilla, Joaquín; Pulido, Federico; Ribera, Esteban; Riera, Melchor; Rubio, Rafael; Santos, Jesús; Sanz-Moreno, José; Sanz, Jesús; Téllez, María Jesús; Tuset, Montserrat; Rivero, Antonio

2015-10-01

In this update, antiretroviral therapy (ART) is recommended for all patients infected by type 1 human immunodeficiency virus (HIV-1). The strength and grade of the recommendation vary depending on the CD4+ T-lymphocyte count, the presence of opportunistic infections or comorbid conditions, age, and the efforts to prevent the transmission of HIV. The objective of ART is to achieve an undetectable plasma viral load (PVL). Initial ART should comprise three drugs, namely, two nucleoside reverse transcriptase inhibitors (NRTI) and one drug from another family. Three of the recommended regimens, all of which have an integrase strand transfer inhibitor (INSTI) as the third drug, are considered a preferred regimen; a further seven regimens, which are based on an INSTI, an non-nucleoside reverse transcriptase inhibitor (NNRTI), or a protease inhibitor boosted with ritonavir (PI/r), are considered alternatives. The reasons and criteria for switching ART are presented both for patients with an undetectable PVL and for patients who experience virological failure, in which case the rescue regimen should include three (or at least two) drugs that are fully active against HIV. The specific criteria for ART in special situations (acute infection, HIV-2 infection, pregnancy) and comorbid conditions (tuberculosis and other opportunistic infections, kidney disease, liver disease, and cancer) are updated. Copyright © 2015 Elsevier España, S.L.U. y Sociedad Española de Enfermedades Infecciosas y Microbiología Clínica. All rights reserved.

Activities of the human immunodeficiency virus type 1 (HIV-1) protease inhibitor nelfinavir mesylate in combination with reverse transcriptase and protease inhibitors against acute HIV-1 infection in vitro.

PubMed

Patick, A K; Boritzki, T J; Bloom, L A

1997-10-01

Nelfinavir mesylate (formerly AG1343) is a potent and selective, nonpeptidic inhibitor of human immunodeficiency virus type 1 (HIV-1) protease that was discovered by protein structure-based design methodologies. We evaluated the antiviral and cytotoxic effects of two-drug combinations of nelfinavir with the clinically approved antiretroviral therapeutics zidovudine (ZDV), lamivudine (3TC), dideoxycytidine (ddC; zalcitabine), stavudine (d4T), didanosine (ddI), indinavir, saquinavir, and ritonavir and a three-drug combination of nelfinavir with ZDV and 3TC against an acute HIV-1 strain RF infection of CEM-SS cells in vitro. Quantitative assessment of drug interaction was evaluated by a universal response surface approach (W. R. Greco, G. Bravo, and J. C. Parsons, Pharm. Rev. 47:331-385, 1995) and by the method of M. N. Prichard and C. Shipman (Antiviral Res. 14:181-206, 1990). Both analytical methods yielded similar results and showed that the two-drug combinations of nelfinavir with the reverse transcriptase inhibitors ZDV, 3TC, ddI, d4T, and ddC and the three-drug combination with ZDV and 3TC resulted in additive to statistically significant synergistic interactions. In a similar manner, the combination of nelfinavir with the three protease inhibitors resulted in additive (ritonavir and saquinavir) to slightly antagonistic (indinavir) interactions. In all combinations, minimal cellular cytotoxicity was observed with any drug alone and in combination. These results suggest that administration of combinations of the appropriate doses of nelfinavir with other currently approved antiretroviral therapeutic agents in vivo may result in enhanced antiviral activity with no associated increase in cellular cytotoxicity.

Activities of the human immunodeficiency virus type 1 (HIV-1) protease inhibitor nelfinavir mesylate in combination with reverse transcriptase and protease inhibitors against acute HIV-1 infection in vitro.

PubMed Central

Patick, A K; Boritzki, T J; Bloom, L A

1997-01-01

Nelfinavir mesylate (formerly AG1343) is a potent and selective, nonpeptidic inhibitor of human immunodeficiency virus type 1 (HIV-1) protease that was discovered by protein structure-based design methodologies. We evaluated the antiviral and cytotoxic effects of two-drug combinations of nelfinavir with the clinically approved antiretroviral therapeutics zidovudine (ZDV), lamivudine (3TC), dideoxycytidine (ddC; zalcitabine), stavudine (d4T), didanosine (ddI), indinavir, saquinavir, and ritonavir and a three-drug combination of nelfinavir with ZDV and 3TC against an acute HIV-1 strain RF infection of CEM-SS cells in vitro. Quantitative assessment of drug interaction was evaluated by a universal response surface approach (W. R. Greco, G. Bravo, and J. C. Parsons, Pharm. Rev. 47:331-385, 1995) and by the method of M. N. Prichard and C. Shipman (Antiviral Res. 14:181-206, 1990). Both analytical methods yielded similar results and showed that the two-drug combinations of nelfinavir with the reverse transcriptase inhibitors ZDV, 3TC, ddI, d4T, and ddC and the three-drug combination with ZDV and 3TC resulted in additive to statistically significant synergistic interactions. In a similar manner, the combination of nelfinavir with the three protease inhibitors resulted in additive (ritonavir and saquinavir) to slightly antagonistic (indinavir) interactions. In all combinations, minimal cellular cytotoxicity was observed with any drug alone and in combination. These results suggest that administration of combinations of the appropriate doses of nelfinavir with other currently approved antiretroviral therapeutic agents in vivo may result in enhanced antiviral activity with no associated increase in cellular cytotoxicity. PMID:9333041

Intracellular studies of the nucleoside reverse transcriptase inhibitor active metabolites: a review.

PubMed

Rodriguez Orengo, J F; Santana, J; Febo, I; Diaz, C; Rodriguez, J L; Garcia, R; Font, E; Rosario, O

2000-03-01

Nucleoside reverse transcriptase inhibitors (NRTIs) plasma concentrations do not correlate with clinical efficacy or toxicity. These agents need to be phosphorylated to become active against HIV-infection. Thus, the characterization of the NRTIs intracellular metabolite pharmacological parameters will provide a better understanding that could lead to the development of more rational dose regimens in the HIV-infected population. Furthermore, intracellular measurements of NRTIs may provide a better marker with respect to clinical efficacy and toxicity than plasma concentrations. Thus, in this article we review the latest information regarding the intracellular pharmacological parameters of zidovudine (ZDV) and lamivudine (3TC) active metabolites in HIV-infected patients including the results from our recent clinical studies. We will start the discussion with ZDV and 3TC clinical efficacy, followed by systemic pharmacokinetics studies. We will then discuss the in vitro and in vivo intracellular studies with particular emphasis in the method development to measure these metabolites and we will conclude with the most current data from our clinical trials.

Reverse transcriptase inhibitors as microbicides.

PubMed

Lewi, Paul; Heeres, Jan; Ariën, Kevin; Venkatraj, Muthusamy; Joossens, Jurgen; Van der Veken, Pieter; Augustyns, Koen; Vanham, Guido

2012-01-01

The CAPRISA 004 study in South Africa has accelerated the development of vaginal and rectal microbicides containing antiretrovirals that target specific enzymes in the reproduction cycle of HIV, especially reverse transcriptase inhibitors (RTI). In this review we discuss the potential relevance of HIV-1 RTIs as microbicides, focusing in the nucleotide RTI tenofovir and six classes of nonnucleoside RTIs (including dapivirine, UC781, urea and thiourea PETTs, DABOs and a pyrimidinedione). Although tenofovir and dapivirine appear to be most advanced in clinical trials as potential microbicides, several issues remain unresolved, e.g., the importance of nonhuman primates as a "gatekeeper" for clinical trials, the emergence and spread of drug-resistant mutants, the combination of microbicides that target different phases of viral reproduction and the accessibility to microbicides in low-income countries. Thus, here we discuss the latest research on RTI as microbicides in the light of the continuing spread of the HIV pandemic from the point of view of medicinal chemistry, virological, and pharmaceutical studies.

Expanded-spectrum nonnucleoside reverse transcriptase inhibitors inhibit clinically relevant mutant variants of human immunodeficiency virus type 1.

PubMed

Corbett, J W; Ko, S S; Rodgers, J D; Jeffrey, S; Bacheler, L T; Klabe, R M; Diamond, S; Lai, C M; Rabel, S R; Saye, J A; Adams, S P; Trainor, G L; Anderson, P S; Erickson-Viitanen, S K

1999-12-01

A research program targeted toward the identification of expanded-spectrum nonnucleoside reverse transcriptase inhibitors which possess increased potency toward K103N-containing mutant human immunodeficiency virus (HIV) and which maintain pharmacokinetics consistent with once-a-day dosing has resulted in the identification of the 4-cyclopropylalkynyl-4-trifluoromethyl-3, 4-dihydro-2(1H)quinazolinones DPC 961 and DPC 963 and the 4-cyclopropylalkenyl-4-trifluoromethyl-3, 4-dihydro-2(1H)quinazolinones DPC 082 and DPC 083 for clinical development. DPC 961, DPC 963, DPC 082, and DPC 083 all exhibit low-nanomolar potency toward wild-type virus, K103N and L100I single-mutation variants, and many multiply amino acid-substituted HIV type 1 mutants. This high degree of potency is combined with a high degree of oral bioavailability, as demonstrated in rhesus monkeys and chimpanzees, and with plasma serum protein binding that can result in significant free levels of drug.

Dual HIV-1 reverse transcriptase and integrase inhibitors from Limonium morisianum Arrigoni, an endemic species of Sardinia (Italy).

PubMed

Sanna, Cinzia; Rigano, Daniela; Corona, Angela; Piano, Dario; Formisano, Carmen; Farci, Domenica; Franzini, Genni; Ballero, Mauro; Chianese, Giuseppina; Tramontano, Enzo; Taglialatela-Scafati, Orazio; Esposito, Francesca

2018-02-04

During our search for potential templates of HIV-1 reverse transcriptase (RT) and integrase (IN) dual inhibitors, the methanolic extract obtained from aerial parts of Limonium morisianum was investigated. Repeated bioassay-guided chromatographic purifications led to the isolation of the following secondary metabolites: myricetin, myricetin 3-O-rutinoside, myricetin-3-O-(6″-O-galloyl)-β-d-galactopyranoside, (-)-epigallocatechin 3-O-gallate, tryptamine, ferulic and phloretic acids. The isolated compounds were tested on both HIV-1 RT-associated RNase H and IN activities. Interestingly, (-)-epigallocatechin-3-O-gallate and myricetin-3-O-(6″-O-galloyl)-β-d-galactopyranoside potently inhibited both enzyme activities with IC 50 values ranging from 0.21 to 10.9 μM. Differently, tryptamine and ferulic acid exhibited a significant inhibition only on the IN strand transfer reaction, showing a selectivity for this viral enzyme. Taken together these results strongly support the potential of this plant as a valuable anti HIV-1 drugs source worthy of further investigations.

High Levels of Transmitted HIV Drug Resistance in a Study in Papua New Guinea.

PubMed

Lavu, Evelyn; Kave, Ellan; Mosoro, Euodia; Markby, Jessica; Aleksic, Eman; Gare, Janet; Elsum, Imogen A; Nano, Gideon; Kaima, Petronia; Dala, Nick; Gurung, Anup; Bertagnolio, Silvia; Crowe, Suzanne M; Myatt, Mark; Hearps, Anna C; Jordan, Michael R

2017-01-01

Papua New Guinea is a Pacific Island nation of 7.3 million people with an estimated HIV prevalence of 0.8%. ART initiation and monitoring are guided by clinical staging and CD4 cell counts, when available. Little is known about levels of transmitted HIV drug resistance in recently infected individuals in Papua New Guinea. Surveillance of transmitted HIV drug resistance in a total of 123 individuals recently infected with HIV and aged less than 30 years was implemented in Port Moresby (n = 62) and Mount Hagen (n = 61) during the period May 2013-April 2014. HIV drug resistance testing was performed using dried blood spots. Transmitted HIV drug resistance was defined by the presence of one or more drug resistance mutations as defined by the World Health Organization surveillance drug resistance mutations list. The prevalence of non-nucleoside reverse transcriptase inhibitor transmitted HIV drug resistance was 16.1% (95% CI 8.8%-27.4%) and 8.2% (95% CI 3.2%-18.2%) in Port Moresby and Mount Hagen, respectively. The prevalence of nucleoside reverse transcriptase inhibitor transmitted HIV drug resistance was 3.2% (95% CI 0.2%-11.7%) and 3.3% (95% CI 0.2%-11.8%) in Port Moresby and Mount Hagen, respectively. No protease inhibitor transmitted HIV drug resistance was observed. The level of non-nucleoside reverse transcriptase inhibitor drug resistance in antiretroviral drug naïve individuals recently infected with HIV in Port Moresby is amongst the highest reported globally. This alarming level of transmitted HIV drug resistance in a young sexually active population threatens to limit the on-going effective use of NNRTIs as a component of first-line ART in Papua New Guinea. To support the choice of nationally recommended first-line antiretroviral therapy, representative surveillance of HIV drug resistance among antiretroviral therapy initiators in Papua New Guinea should be urgently implemented.

HIV drug resistance in infants increases with changing prevention of mother-to-child transmission regimens.

PubMed

Poppe, Lisa K; Chunda-Liyoka, Catherine; Kwon, Eun H; Gondwe, Clement; West, John T; Kankasa, Chipepo; Ndongmo, Clement B; Wood, Charles

2017-08-24

The objectives of this study were to determine HIV drug resistance (HIVDR) prevalence in Zambian infants upon diagnosis, and to determine how changing prevention of mother-to-child transmission (PMTCT) drug regimens affect drug resistance. Dried blood spot (DBS) samples from infants in the Lusaka District of Zambia, obtained during routine diagnostic screening, were collected during four different years representing three different PMTCT drug treatment regimens. DNA extracted from dried blood spot samples was used to sequence a 1493 bp region of the reverse transcriptase gene. Sequences were analyzed via the Stanford HIVDRdatabase (http://hivdb.standford.edu) to screen for resistance mutations. HIVDR in infants increased from 21.5 in 2007/2009 to 40.2% in 2014. Nonnucleoside reverse transcriptase inhibitor resistance increased steadily over the sampling period, whereas nucleoside reverse transcriptase inhibitor resistance and dual class resistance both increased more than threefold in 2014. Analysis of drug resistance scores in each group revealed increasing strength of resistance over time. In 2014, children with reported PMTCT exposure, defined as infant prophylaxis and/or maternal treatment, showed a higher prevalence and strength of resistance compared to those with no reported exposure. HIVDR is on the rise in Zambia and presents a serious problem for the successful lifelong treatment of HIV-infected children. PMTCT affects both the prevalence and strength of resistance and further research is needed to determine how to mitigate its role leading to resistance.

Role of Ligand Reorganization and Conformational Restraints on the Binding Free Energies of DAPY Non-Nucleoside Inhibitors to HIV Reverse Transcriptase

PubMed Central

Gallicchio, Emilio

2012-01-01

The results of computer simulations of the binding of etravirine (TMC125) and rilpivirine (TMC278) to HIV reverse transcriptase are reported. It is confirmed that consistent binding free energy estimates are obtained with or without the application of torsional restraints when the free energies of imposing the restraints are taken into account. The restraints have a smaller influence on the thermodynamics and apparent kinetics of binding of TMC125 compared to the more flexible TMC278 inhibitor. The concept of the reorganization free energy of binding is useful to understand and categorize these effects. Contrary to expectations, the use of conformational restraints did not consistently enhance convergence of binding free energy estimates due to suppression of binding/unbinding pathways and due to the influence of rotational degrees of freedom not directly controlled by the restraints. Physical insights concerning the thermodynamic driving forces for binding and the role of “jiggling” and “wiggling” motion of the ligands are discussed. Based on these insights we conclude that an ideal inhibitor, if chemically realizable, would possess the electrostatic charge distribution of TMC125, so as to form strong interactions with the receptor, and the larger and more flexible substituents of TMC278, so as to minimize reorganization free energy penalties and the effects of resistance mutations, suitably modified, as in TMC125, so as to disfavor the formation of non-binding competent extended conformations when free in solution. PMID:22708073

Human immunodeficiency virus types 1 and 2 exhibit comparable sensitivities to Zidovudine and other nucleoside analog inhibitors in vitro.

PubMed

Smith, Robert A; Gottlieb, Geoffrey S; Anderson, Donovan J; Pyrak, Crystal L; Preston, Bradley D

2008-01-01

Using an indicator cell assay that directly quantifies viral replication, we show that human immunodeficiency virus types 1 and 2 (HIV-1 and HIV-2, respectively) exhibit similar sensitivities to 3'-azido-3'-deoxythymidine (zidovudine) as well as other nucleoside analog inhibitors of reverse transcriptase. These data support the use of nucleoside analogs for antiviral therapy of HIV-2 infection.

Drug resistance mutations in HIV type 1 isolates from naive patients eligible for first line antiretroviral therapy in JJ Hospital, Mumbai, India.

PubMed

Deshpande, Alake; Karki, Surendra; Recordon-Pinson, Patricia; Fleury, Herve J

2011-12-01

More than 50 HIV-1-infected patients, naive of antiretroviral therapy (ART) but eligible for first line ART in JJ Hospital, Mumbai, India were investigated for surveillance drug resistance mutations (SDRMs); all but one virus belonged to subtype C; we could observe SDRMs to nonnucleoside reverse transcriptase inhibitors and protease inhibitors in 9.6% of the patients.

The Reverse Transcription Inhibitor Abacavir Shows Anticancer Activity in Prostate Cancer Cell Lines

PubMed Central

Molinari, Agnese; Parisi, Chiara; Bozzuto, Giuseppina; Toccacieli, Laura; Formisano, Giuseppe; De Orsi, Daniela; Paradisi, Silvia; Grober, OlÌ Maria Victoria; Ravo, Maria; Weisz, Alessandro; Arcieri, Romano; Vella, Stefano; Gaudi, Simona

2010-01-01

Background Transposable Elements (TEs) comprise nearly 45% of the entire genome and are part of sophisticated regulatory network systems that control developmental processes in normal and pathological conditions. The retroviral/retrotransposon gene machinery consists mainly of Long Interspersed Nuclear Elements (LINEs-1) and Human Endogenous Retroviruses (HERVs) that code for their own endogenous reverse transcriptase (RT). Interestingly, RT is typically expressed at high levels in cancer cells. Recent studies report that RT inhibition by non-nucleoside reverse transcriptase inhibitors (NNRTIs) induces growth arrest and cell differentiation in vitro and antagonizes growth of human tumors in animal model. In the present study we analyze the anticancer activity of Abacavir (ABC), a nucleoside reverse transcription inhibitor (NRTI), on PC3 and LNCaP prostate cancer cell lines. Principal Findings ABC significantly reduces cell growth, migration and invasion processes, considerably slows S phase progression, induces senescence and cell death in prostate cancer cells. Consistent with these observations, microarray analysis on PC3 cells shows that ABC induces specific and dose-dependent changes in gene expression, involving multiple cellular pathways. Notably, by quantitative Real-Time PCR we found that LINE-1 ORF1 and ORF2 mRNA levels were significantly up-regulated by ABC treatment. Conclusions Our results demonstrate the potential of ABC as anticancer agent able to induce antiproliferative activity and trigger senescence in prostate cancer cells. Noteworthy, we show that ABC elicits up-regulation of LINE-1 expression, suggesting the involvement of these elements in the observed cellular modifications. PMID:21151977

Switching regimens in virologically suppressed HIV-1-infected patients: evidence base and rationale for integrase strand transfer inhibitor (INSTI)-containing regimens.

PubMed

Raffi, F; Esser, S; Nunnari, G; Pérez-Valero, I; Waters, L

2016-10-01

In an era when most individuals with treated HIV infection can expect to live into old age, clinicians should proactively review their patients' current and future treatment needs and challenges. Clinical guidelines acknowledge that, in the setting of virological suppression, treatment switch may yield benefits in terms of tolerability, regimen simplification, adherence, convenience and long-term health considerations, particularly in the context of ageing. In this paper, we review evidence from six key clinical studies on switching virologically suppressed patients to regimens based on integrase strand transfer inhibitors (INSTIs), the antiretroviral class increasingly preferred as initial therapy in clinical guidelines. We review these studies and focus on the virological efficacy, safety, and tolerability of switching to INSTI-based regimens in suppressed HIV-positive individuals. We review the early switch studies SWITCHMRK and SPIRAL [assessing a switch from a ritonavir-boosted protease inhibitor (PI/r) to raltegravir (RAL)-containing regimens], together with data from STRATEGY-PI [assessing a switch to elvitegravir (EVG)-containing regimens; EVG/cobicistat (COBI)/emtricitabine (FTC)/tenofovir disoproxil fumarate (TDF) vs. remaining on a PI/r-containing regimen], STRATEGY-NNRTI [assessing a switch to EVG/COBI/FTC/TDF vs. continuation of a nonnucleoside reverse transcriptase inhibitor (NNRTI) and two nucleoside reverse transcriptase inhibitors (NRTIs)], STRIIVING [assessing a switch to a dolutegravir (DTG)-containing regimen (abacavir (ABC)/lamivudine (3TC)/DTG) vs. staying on the background regimen], and GS study 109 [assessing a switch to EVG/COBI/FTC/tenofovir alafenamide fumarate (TAF) vs. continuation of FTC/TDF-based regimens]. Switching to INSTI-containing regimens has been shown to support good virological efficacy, with evidence from two studies demonstrating superior virological efficacy for a switch to EVG-containing regimens. In addition, switching to INSTI regimens was associated with improved tolerability and greater reported patient satisfaction and outcomes in some studies. INSTI-based regimens offer an important contemporary switch option that may be tailored to meet and optimize the needs of many patients. © 2016 British HIV Association.

Investigational Antiretroviral Drugs: What is Coming Down the Pipeline.

PubMed

Gulick, Roy M

2018-04-01

Over the past 30 years, antiretroviral drug regimens for treating HIV infection have become more effective, safer, and more convenient. Despite 31 currently approved drugs, the pipeline of investigational HIV drugs remains full. Investigational antiretroviral drugs include the nucleoside analogue reverse transcriptase translocation inhibitor (NRTTI) MK-8591, a long-acting compound that could be dosed once weekly. Investigational nonnucleoside analogue reverse transcriptase inhibitors (NNRTIs) include doravirine, which is active in vitro against NNRTI-resistant HIV and was potent and well-tolerated when used in combination with a dual-nucleoside analogue RTI (nRTI) backbone in treatment-naive individuals.New integrase strand transfer inhibitors (InSTIs) include recently approved bictegravir, which is active against InSTI-resistant viral strains in vitro and was potent and well-tolerated in combination regimens in treatment-naive individuals, and investigational cabotegravir, which is being studied with monthly parenteral dosing for HIV maintenance treatment and with bimonthly dosing for HIV preexposure prophylaxis (PrEP). Investigational HIV entry inhibitors include the new CD4 attachment inhibitor fostemsavir, which targets HIV envelope glycoprotein 120, and recently approved ibalizumab, which binds the CD4 receptor. This article summarizes presentations by Roy M. Gulick, MD, MPH, at the IAS-USA continuing education program, Improving the Management of HIV Disease, held in Los Angeles, California, in April 2017, and at the 2017 Ryan White HIV/AIDS Program Clinical Conference, held in San Antonio, Texas, in August 2017.

Etravirine and rilpivirine resistance in HIV-1 subtype CRF01_AE-infected adults failing non-nucleoside reverse transcriptase inhibitor-based regimens.

PubMed

Bunupuradah, Torsak; Ananworanich, Jintanat; Chetchotisakd, Ploenchan; Kantipong, Pacharee; Jirajariyavej, Supunnee; Sirivichayakul, Sunee; Munsakul, Warangkana; Prasithsirikul, Wisit; Sungkanuparph, Somnuek; Bowonwattanuwong, Chureeratana; Klinbuayaem, Virat; Petoumenos, Kathy; Hirschel, Bernard; Bhakeecheep, Sorakij; Ruxrungtham, Kiat

2011-01-01

We studied prevalence of etravirine (ETR) and rilpivirine (RPV) resistance in HIV-1 subtype CRF01_AE infection with first-line non-nucleoside reverse transcriptase inhibitor (NNRTI) failure. A total of 225 adults failing two nucleoside reverse transcriptase inhibitors (NRTIs) plus 1 NNRTI in Thailand with HIV RNA>1,000 copies/ml were included. Genotypic resistance results and HIV-1 subtype were interpreted by Stanford DR database. ETR resistance was calculated by the new Monogram weighted score (Monogram WS; ≥ 4 indicating high-level ETR resistance) and by DUET weighted score (DUET WS; 2.5-3.5 and ≥ 4 resulted in intermediate and reduce ETR response, respectively). RPV resistance interpretation was based on previous reports. Median (IQR) age was 38 (34-42) years, 41% were female and CDC A:B:C were 22%:21%:57%. HIV subtypes were 96% CRF01_AE and 4% B. Antiretrovirals at failure were lamivudine (100%), stavudine (93%), nevirapine (90%) and efavirenz (10%) with a median (IQR) duration of 3.4 (1.8-4.5) years. Median (IQR) CD4(+) T-cell count and HIV RNA were 194 (121-280) cells/mm³ and 4.1 (3.6-4.6) log₁₀ copies/ml, respectively. The common NNRTI mutations were Y181C (41%), G190A (22%) and K103N (19%). The proportion of patients with Monogram WS score ≥ 4 was 61.3%. By DUET WS, 49.8% and 7.5% of patients were scored 2.5-3.5 and ≥4, respectively. Only HIV RNA ≥ 4 log₁₀ copies/ml at failure was associated with both Monogram WS ≥ 4 (OR 2.3, 95% CI 1.3-3.9; P=0.003) and DUET WS ≥ 2.5 (OR 1.9, 95% CI 1.1-3.3; P=0.02). The RVP resistance-associated mutations (RAMs) detected were K101P (1.8%), Y181I (2.7%) and Y181V (3.6%). All patients with RPV mutation had ETR resistance. No E138R/E138K mutations were detected. Approximately 60% of patients had high-level ETR resistance. The role of ETR in second-line therapy is limited in late NNRTI failure settings. RVP RAMs were uncommon, but cross-resistance between ETR and RVP was high.

Adipocytes impair efficacy of antiretroviral therapy.

PubMed

Couturier, Jacob; Winchester, Lee C; Suliburk, James W; Wilkerson, Gregory K; Podany, Anthony T; Agarwal, Neeti; Xuan Chua, Corrine Ying; Nehete, Pramod N; Nehete, Bharti P; Grattoni, Alessandro; Sastry, K Jagannadha; Fletcher, Courtney V; Lake, Jordan E; Balasubramanyam, Ashok; Lewis, Dorothy E

2018-06-01

Adequate distribution of antiretroviral drugs to infected cells in HIV patients is critical for viral suppression. In humans and primates, HIV- and SIV-infected CD4 T cells in adipose tissues have recently been identified as reservoirs for infectious virus. To better characterize adipose tissue as a pharmacological sanctuary for HIV-infected cells, in vitro experiments were conducted to assess antiretroviral drug efficacy in the presence of adipocytes, and drug penetration in adipose tissue cells (stromal-vascular-fraction cells and mature adipocytes) was examined in treated humans and monkeys. Co-culture experiments between HIV-1-infected CD4 T cells and primary human adipocytes showed that adipocytes consistently reduced the antiviral efficacy of the nucleotide reverse transcriptase inhibitor tenofovir and its prodrug forms tenofovir disoproxil fumarate (TDF) and tenofovir alafenamide (TAF). In HIV-infected persons, LC-MS/MS analysis of intracellular lysates derived from adipose tissue stromal-vascular-fraction cells or mature adipocytes suggested that integrase inhibitors penetrate adipose tissue, whereas penetration of nucleoside/nucleotide reverse transcriptase inhibitors such as TDF, emtricitabine, abacavir, and lamivudine is restricted. The limited distribution and functions of key antiretroviral drugs within fat depots may contribute to viral persistence in adipose tissue. Copyright © 2018 Elsevier B.V. All rights reserved.

Single Active Site Mutation Causes Serious Resistance of HIV Reverse Transcriptase to Lamivudine: Insight from Multiple Molecular Dynamics Simulations.

PubMed

Moonsamy, Suri; Bhakat, Soumendranath; Walker, Ross C; Soliman, Mahmoud E S

2016-03-01

Molecular dynamics simulations, binding free energy calculations, principle component analysis (PCA), and residue interaction network analysis were employed in order to investigate the molecular mechanism of M184I single mutation which played pivotal role in making the HIV-1 reverse transcriptase (RT) totally resistant to lamivudine. Results showed that single mutations at residue 184 of RT caused (1) distortion of the orientation of lamivudine in the active site due to the steric conflict between the oxathiolane ring of lamivudine and the side chain of beta-branched amino acids Ile at position 184 which, in turn, perturbs inhibitor binding, (2) decrease in the binding affinity by (~8 kcal/mol) when compared to the wild-type, (3) variation in the overall enzyme motion as evident from the PCA for both systems, and (4) distortion of the hydrogen bonding network and atomic interactions with the inhibitor. The comprehensive analysis presented in this report can provide useful information for understanding the drug resistance mechanism against lamivudine. The results can also provide some potential clues for further design of novel inhibitors that are less susceptible to drug resistance.

Adipocytes Impair Efficacy of Antiretroviral Therapy

PubMed Central

Couturier, Jacob; Winchester, Lee C.; Suliburk, James W.; Wilkerson, Gregory K.; Podany, Anthony T.; Agarwal, Neeti; Chua, Corrine Ying Xuan; Nehete, Pramod N.; Nehete, Bharti P.; Grattoni, Alessandro; Sastry, K. Jagannadha; Fletcher, Courtney V.; Lake, Jordan E.; Balasubramanyan, Ashok; Lewis, Dorothy E.

2018-01-01

Adequate distribution of antiretroviral drugs to infected cells in HIV patients is critical for viral suppression. In humans and primates, HIV- and SIV-infected CD4 T cells in adipose tissues have recently been identified as reservoirs for infectious virus. To better characterize adipose tissue as a pharmacological sanctuary for HIV-infected cells, in vitro experiments were conducted to assess antiretroviral drug efficacy in the presence of adipocytes, and drug penetration in adipose tissue cells (stromal-vascular-fraction cells and mature adipocytes) was examined in treated humans and monkeys. Co-culture experiments between HIV-1-infected CD4 T cells and primary human adipocytes showed that adipocytes consistently reduced the antiviral efficacy of the nucleotide reverse transcriptase inhibitor tenofovir and its prodrug forms tenofovir disoproxil fumarate (TDF) and tenofovir alafenamide (TAF). In HIV-infected persons, LC-MS/MS analysis of intracellular lysates derived from adipose tissue stromal-vascular-fraction cells or mature adipocytes suggested that integrase inhibitors penetrate adipose tissue, whereas penetration of nucleoside/nucleotide reverse transcriptase inhibitors such as TDF, emtricitabine, abacavir, and lamivudine is restricted. The limited distribution and functions of key antiretroviral drugs within fat depots may contribute to viral persistence in adipose tissue. PMID:29630975

In Vitro Resistance Profile of the Candidate HIV-1 Microbicide Drug Dapivirine

PubMed Central

Schader, Susan M.; Oliveira, Maureen; Ibanescu, Ruxandra-Ilinca; Moisi, Daniela; Colby-Germinario, Susan P.

2012-01-01

Antiretroviral-based microbicides may offer a means to reduce the sexual transmission of HIV-1. Suboptimal use of a microbicide may, however, lead to the development of drug resistance in users that are already, or become, infected with HIV-1. In such cases, the efficacy of treatments may be compromised since the same (or similar) antiretrovirals used in treatments are being developed as microbicides. To help predict which drug resistance mutations may develop in the context of suboptimal use, HIV-1 primary isolates of different subtypes and different baseline resistance profiles were used to infect primary cells in vitro in the presence of increasing suboptimal concentrations of the two candidate microbicide antiretrovirals dapivirine (DAP) and tenofovir (TFV) alone or in combination. Infections were ongoing for 25 weeks, after which reverse transcriptase genotypes were determined and scrutinized for the presence of any clinically recognized reverse transcriptase drug resistance mutations. Results indicated that suboptimal concentrations of DAP alone facilitated the emergence of common nonnucleoside reverse transcriptase inhibitor resistance mutations, while suboptimal concentrations of DAP plus TFV gave rise to fewer mutations. Suboptimal concentrations of TFV alone did not frequently result in the development of resistance mutations. Sensitivity evaluations for stavudine (d4T), nevirapine (NVP), and lamivudine (3TC) revealed that the selection of resistance as a consequence of suboptimal concentrations of DAP may compromise the potential for NVP to be used in treatment, a finding of potential relevance in developing countries. PMID:22123692

In vitro resistance profile of the candidate HIV-1 microbicide drug dapivirine.

PubMed

Schader, Susan M; Oliveira, Maureen; Ibanescu, Ruxandra-Ilinca; Moisi, Daniela; Colby-Germinario, Susan P; Wainberg, Mark A

2012-02-01

Antiretroviral-based microbicides may offer a means to reduce the sexual transmission of HIV-1. Suboptimal use of a microbicide may, however, lead to the development of drug resistance in users that are already, or become, infected with HIV-1. In such cases, the efficacy of treatments may be compromised since the same (or similar) antiretrovirals used in treatments are being developed as microbicides. To help predict which drug resistance mutations may develop in the context of suboptimal use, HIV-1 primary isolates of different subtypes and different baseline resistance profiles were used to infect primary cells in vitro in the presence of increasing suboptimal concentrations of the two candidate microbicide antiretrovirals dapivirine (DAP) and tenofovir (TFV) alone or in combination. Infections were ongoing for 25 weeks, after which reverse transcriptase genotypes were determined and scrutinized for the presence of any clinically recognized reverse transcriptase drug resistance mutations. Results indicated that suboptimal concentrations of DAP alone facilitated the emergence of common nonnucleoside reverse transcriptase inhibitor resistance mutations, while suboptimal concentrations of DAP plus TFV gave rise to fewer mutations. Suboptimal concentrations of TFV alone did not frequently result in the development of resistance mutations. Sensitivity evaluations for stavudine (d4T), nevirapine (NVP), and lamivudine (3TC) revealed that the selection of resistance as a consequence of suboptimal concentrations of DAP may compromise the potential for NVP to be used in treatment, a finding of potential relevance in developing countries.

Novel nonnucleoside inhibitors that select nucleoside inhibitor resistance mutations in human immunodeficiency virus type 1 reverse transcriptase.

PubMed

Zhang, Zhijun; Walker, Michelle; Xu, Wen; Shim, Jae Hoon; Girardet, Jean-Luc; Hamatake, Robert K; Hong, Zhi

2006-08-01

Mutations in and around the catalytic site of the reverse transcriptase (RT) of human immunodeficiency virus type 1 (HIV-1) are associated with resistance to nucleoside RT inhibitors (NRTIs), whereas changes in the hydrophobic pocket of the RT are attributed to nonnucleoside RT inhibitor (NNRTI) resistance. In this study, we report a novel series of nonnucleoside inhibitors of HIV-1, exemplified by VRX-329747 and VRX-413638, which inhibit both NNRTI- and NRTI-resistant HIV-1 isolates. Enzymatic studies indicated that these compounds are HIV-1 RT inhibitors. Surprisingly, however, following prolonged (6 months) tissue culture selection, this series of nonnucleoside inhibitors did not select NNRTI-resistant mutations in HIV-1 RT. Rather, four mutations (M41L, A62T/V, V118I, and M184V) known to cause resistance to NRTIs and two additional novel mutations (S68N and G112S) adjacent to the catalytic site of the enzyme were selected. Although the M184V mutation appears to be the initial mutation to establish resistance, this mutation alone confers only a two- to fourfold decrease in susceptibility to VRX-329747 and VRX-413638. At least two additional mutations must accumulate for significant resistance. Moreover, while VRX-329747-selected viruses are resistant to lamivudine and emtricitabine due to the M184V mutation, they remain susceptible to zidovudine, stavudine, dideoxyinosine, abacavir, tenofovir, and efavirenz. These results directly demonstrate that VRX-329747 and VRX-413638 are novel nonnucleoside inhibitors of HIV-1 RT with the potential to augment current therapies.

Novel Nonnucleoside Inhibitors That Select Nucleoside Inhibitor Resistance Mutations in Human Immunodeficiency Virus Type 1 Reverse Transcriptase

PubMed Central

Zhang, Zhijun; Walker, Michelle; Xu, Wen; Shim, Jae Hoon; Girardet, Jean-Luc; Hamatake, Robert K.; Hong, Zhi

2006-01-01

Mutations in and around the catalytic site of the reverse transcriptase (RT) of human immunodeficiency virus type 1 (HIV-1) are associated with resistance to nucleoside RT inhibitors (NRTIs), whereas changes in the hydrophobic pocket of the RT are attributed to nonnucleoside RT inhibitor (NNRTI) resistance. In this study, we report a novel series of nonnucleoside inhibitors of HIV-1, exemplified by VRX-329747 and VRX-413638, which inhibit both NNRTI- and NRTI-resistant HIV-1 isolates. Enzymatic studies indicated that these compounds are HIV-1 RT inhibitors. Surprisingly, however, following prolonged (6 months) tissue culture selection, this series of nonnucleoside inhibitors did not select NNRTI-resistant mutations in HIV-1 RT. Rather, four mutations (M41L, A62T/V, V118I, and M184V) known to cause resistance to NRTIs and two additional novel mutations (S68N and G112S) adjacent to the catalytic site of the enzyme were selected. Although the M184V mutation appears to be the initial mutation to establish resistance, this mutation alone confers only a two- to fourfold decrease in susceptibility to VRX-329747 and VRX-413638. At least two additional mutations must accumulate for significant resistance. Moreover, while VRX-329747-selected viruses are resistant to lamivudine and emtricitabine due to the M184V mutation, they remain susceptible to zidovudine, stavudine, dideoxyinosine, abacavir, tenofovir, and efavirenz. These results directly demonstrate that VRX-329747 and VRX-413638 are novel nonnucleoside inhibitors of HIV-1 RT with the potential to augment current therapies. PMID:16870771

Modified telomeric repeat amplification protocol: a quantitative radioactive assay for telomerase without using electrophoresis.

PubMed

Szatmari, I; Tókés, S; Dunn, C B; Bardos, T J; Aradi, J

2000-06-15

A polymerase chain reaction (PCR)-based radioactive telomerase assay was developed in our laboratory which is quantitative and does not require electrophoretic evaluation (designated as TP-TRAP; it utilizes two reverse primers). The main steps of the assay include (1) extension of a 20-mer oligonucleotide substrate (MTS) by telomerase, (2) amplification of the telomerase products in the presence of [(3)H]dTTP using the substrate oligonucleotide and two reverse primers (RPC3, 38 mer; RP, 20 mer), (3) isolation of the amplified radioactive dsDNA by precipitation and filtration, (4) determination of the radioactivity of the acid-insoluble DNA. The length of the telomerase products does not increase on amplification. This valuable feature of the assay is achieved by utilization of the two reverse primers and a highly specific PCR protocol. The assay is linear, accurate, and suitable for cell-biological studies where slight quantitative differences in telomerase activity must be detected. The assay is also suitable for screening and characterization of telomerase inhibitors, as shown with a chemically modified oligonucleotide reverse transcriptase inhibitor [(s(4)dU)(35)]. Copyright 2000 Academic Press.

Effect of dolutegravir in combination with Nucleoside Reverse Transcriptase Inhibitors (NRTIs) on people living with HIV who have pre-existing NRTI mutations.

PubMed

Sörstedt, Erik; Carlander, Christina; Flamholc, Leo; Hejdeman, Bo; Svedhem, Veronica; Sönnerborg, Anders; Gisslén, Magnus; Yilmaz, Aylin

2018-05-01

Until the introduction of dolutegravir (DTG), people living with HIV (PLWH) who have developed nucleoside reverse transcriptase inhibitor (NRTI) mutations have had few other treatment options outside of regimens based on ritonavir-boosted protease inhibitors (PI/r). Here we report treatment results among PLWH in Sweden with pre-existing NRTI mutations on antiretroviral treatment (ART) with DTG and one to two NRTIs. All PLWH on ART with DTG and one to two NRTIs with pre-existing NRTI mutations were retrospectively identified from the National InfCare HIV database. As controls, PLWH on PI/r and one to two NRTIs, matched according to Genotypic Susceptibility Score and observation time, were included. Data were collected as long as the study population was on treatment with DTG; controls were monitored for the same interval. Outcome was classified as either treatment success or failure. In total, 244 participants (122 individuals treated with DTG and 122 individuals treated with PI/r) were included. Median observation time was 78 weeks (interquartile range 50-98 weeks) for participants on DTG and 75 weeks (50-101 weeks) for individuals on PI/r. Viral failure was detected in four individuals treated with DTG and three individuals treated with PI/r, resulting in similar success rates of 96.7% and 97.5%, respectively. No new mutations were found among participants with treatment failure. DTG in combination with one to two NRTIs was as efficient as PI/r in individuals with pre-existing NRTI mutations in this setting. It may be considered an alternative to PI/r-based ART even in the presence of NRTI resistance. Copyright © 2018 Elsevier B.V. and International Society of Chemotherapy. All rights reserved.

Differential Regulation of Telomerase Reverse Transcriptase Promoter Activation and Protein Degradation by Histone Deacetylase Inhibition.

PubMed

Qing, Hua; Aono, Jun; Findeisen, Hannes M; Jones, Karrie L; Heywood, Elizabeth B; Bruemmer, Dennis

2016-06-01

Telomerase reverse transcriptase (TERT) maintains telomeres and is rate limiting for replicative life span. While most somatic tissues silence TERT transcription resulting in telomere shortening, cells derived from cancer or cardiovascular diseases express TERT and activate telomerase. In the present study, we demonstrate that histone deacetylase (HDAC) inhibition induces TERT transcription and promoter activation. At the protein level in contrast, HDAC inhibition decreases TERT protein abundance through enhanced degradation, which decreases telomerase activity and induces senescence. Finally, we demonstrate that HDAC inhibition decreases TERT expression during vascular remodeling in vivo. These data illustrate a differential regulation of TERT transcription and protein stability by HDAC inhibition and suggest that TERT may constitute an important target for the anti-proliferative efficacy of HDAC inhibitors. © 2015 Wiley Periodicals, Inc.

Molecular docking of (5E)-3-(2-aminoethyl)-5-(2- thienylmethylene)-1, 3-thiazolidine-2, 4-dione on HIV-1 reverse transcriptase: novel drug acting on enzyme.

PubMed

Seniya, Chandrabhan; Yadav, Ajay; Uchadia, Kuldeep; Kumar, Sanjay; Sagar, Nitin; Shrivastava, Priyanka; Shrivastava, Shilpi; Wadhwa, Gulshan

2012-01-01

The study of Human immunodeficiency virus (HIV) in humans and animal models in last 31 years suggested that it is a causative agent of AIDS. This causes serious pandemic public health concern globally. It was reported that the HIV-1 reverse transcriptase (RT) played a critical role in the life cycle of HIV. Therefore, inhibition of HIV-1RT enzyme is one of the major and potential targets in the treatment of AIDS. The enzyme (HIV-1RT) was successfully targeted by non nucleotide reverse transcriptase inhibitors (NNRTIs). But frequent application of NNRTIs led drug resistance mutation on HIV infections. Therefore, there is a need to search new NNRTIs with appropriate pharmacophores. For the purpose, a virtually screened 3D model of unliganded HIV-1RT (1DLO) was explored. The unliganded HIV-1RT (1DLO) was docked with 4-thiazolidinone and its derivatives (ChemBank Database) by using AutoDock4. The best seven docking solutions complex were selected and analyzed by Ligplot. The analysis showed that derivative (5E)-3-(2- aminoethyl)-5-(2- thienylmethylene)-1, 3-thiazolidine-2, 4-dione (CID 3087795) has maximum potential against unliganded HIV-1RT (1DLO). The analysis was done on the basis of scoring and binding ability. The derivative (5E)-3-(2- aminoethyl)-5-(2- thienylmethylene)-1, 3-thiazolidine-2, 4-dione (CID 3087795) indicated minimum energy score and highest number of interactions with active site residue and could be a promising inhibitor for HIV-1 RT as Drug target.

Time to Switch to Second-line Antiretroviral Therapy in Children With Human Immunodeficiency Virus in Europe and Thailand.

PubMed

2018-02-01

Data on durability of first-line antiretroviral therapy (ART) in children with human immunodeficiency virus (HIV) are limited. We assessed time to switch to second-line therapy in 16 European countries and Thailand. Children aged <18 years initiating combination ART (≥2 nucleoside reverse transcriptase inhibitors [NRTIs] plus nonnucleoside reverse transcriptase inhibitor [NNRTI] or boosted protease inhibitor [PI]) were included. Switch to second-line was defined as (i) change across drug class (PI to NNRTI or vice versa) or within PI class plus change of ≥1 NRTI; (ii) change from single to dual PI; or (iii) addition of a new drug class. Cumulative incidence of switch was calculated with death and loss to follow-up as competing risks. Of 3668 children included, median age at ART initiation was 6.1 (interquartile range (IQR), 1.7-10.5) years. Initial regimens were 32% PI based, 34% nevirapine (NVP) based, and 33% efavirenz based. Median duration of follow-up was 5.4 (IQR, 2.9-8.3) years. Cumulative incidence of switch at 5 years was 21% (95% confidence interval, 20%-23%), with significant regional variations. Median time to switch was 30 (IQR, 16-58) months; two-thirds of switches were related to treatment failure. In multivariable analysis, older age, severe immunosuppression and higher viral load (VL) at ART start, and NVP-based initial regimens were associated with increased risk of switch. One in 5 children switched to a second-line regimen by 5 years of ART, with two-thirds failure related. Advanced HIV, older age, and NVP-based regimens were associated with increased risk of switch. © The Author(s) 2017. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail: journals.permissions@oup.com.

Design and synthesis of N₁-aryl-benzimidazoles 2-substituted as novel HIV-1 non-nucleoside reverse transcriptase inhibitors.

PubMed

Monforte, Anna-Maria; Ferro, Stefania; De Luca, Laura; Lo Surdo, Giuseppa; Morreale, Francesca; Pannecouque, Christophe; Balzarini, Jan; Chimirri, Alba

2014-02-15

A series of novel N1-aryl-2-arylthioacetamido-benzimidazoles were synthesized and evaluated as inhibitors of human immunodeficiency virus type-1 (HIV-1). Some of them proved to be effective in inhibiting HIV-1 replication at submicromolar and nanomolar concentration acting as HIV-1 non-nucleoside RT inhibitors (NNRTIs), with low cytotoxicity. The preliminary structure-activity relationship (SAR) of these new derivatives was discussed and rationalized by docking studies. Copyright © 2014 Elsevier Ltd. All rights reserved.

Metabolic syndrome in patients on first-line antiretroviral therapy containing zidovudine or tenofovir in rural Lesotho, Southern Africa.

PubMed

Labhardt, Niklaus Daniel; Müller, Urs Franz; Ringera, Isaac; Ehmer, Jochen; Motlatsi, Mokete M; Pfeiffer, Karolin; Hobbins, Michael A; Muhairwe, Josephine A; Muser, Juergen; Hatz, Christoph

2017-06-01

To assess the prevalence of metabolic syndrome (MetS) among patients in rural Lesotho who are taking first-line antiretroviral therapy (ART) containing either zidovudine or tenofovir disoproxil. Cross-sectional survey in 10 facilities in Lesotho among adult (≥16 years) patients on non-nucleoside reverse transcriptase inhibitor (NNRTI)-based first-line ART for ≥6 months. MetS was defined according to the International Diabetes Federation criteria. Among 1166 patients (65.8% female), 22.2% (95% CI: 19.3-25.3) of women and 6.3% (4.1-9.1) of men met the IDF definition of MetS (P < 0.001). In both sexes, there was no significant difference in MetS prevalence between NNRTIs. However, in women taking zidovudine as nucleoside reverse transcriptase inhibitor (NRTI), MetS prevalence was 27.9%, vs. 18.8% in those taking tenofovir. In the multivariate logistic regression allowing for socio-demographic and clinical covariates, ART containing zidovudine was associated with MetS in women (aOR 2.17 (1.46-3.22), P < 0.001) but not in men. In this study, taking ART containing zidovudine instead of tenofovir disoproxil was an independent predictor of MetS in women but not in men. This finding endorses WHO's recommendation of tenofovir as preferred NRTI. © 2017 John Wiley & Sons Ltd.

Prevalence of HIV Antiretroviral Drug Resistance and Its Impacts on HIV-1 Virological Failures in Jiangsu, China: A Cross-Sectional Study.

PubMed

Zhou, Ying; Lu, Jing; Wang, Jinge; Yan, Hongjing; Li, Jianjun; Xu, Xiaoqin; Zhang, Zhi; Qiu, Tao; Ding, Ping; Fu, Gengfeng; Huan, Xiping; Hu, Haiyang

2016-01-01

Antiretroviral therapy (ART) has been shown to improve survival of patients with Human Immunodeficiency Virus (HIV) infection and to reduce HIV-1 transmission. Therefore, the Chinese central government initiated a national program to provide ART free of charge to HIV-1 patients. We conducted a cross-sectional survey in Jiangsu province to determine the level of drug resistance (DR) in HIV-1 infected patients and the correlates of DR in virological failures in 2012. Approximately 10.4% of the HIV-1 patients in the study experienced virological failure after one year of ART and were divided into drug sensitive and drug resistant groups based on genotype determination. The viral loads (VLs) in the drug resistant group were significantly lower than the drug sensitive group. There were two independent predictors of virological failure: male gender and increasing duration of treatment. The primary mutations observed in the study were against nucleoside reverse transcriptase inhibitors (NRTIs) which were M184V (79.45%) and K103N (33.70%) in nonnucleoside reverse transcriptase inhibitors (NNRTIs). The overall rate of DR in Jiangsu province is still relatively low among treated patients. However, close monitoring of drug resistance in male patients in the early stages of treatment is vital to maintaining and increasing the benefits of HIV ART achieved to date.

Parallel screening of drug-like natural compounds using Caco-2 cell permeability QSAR model with applicability domain, lipophilic ligand efficiency index and shape property: A case study of HIV-1 reverse transcriptase inhibitors

NASA Astrophysics Data System (ADS)

Patel, Rikin D.; Kumar, Sivakumar Prasanth; Patel, Chirag N.; Shankar, Shetty Shilpa; Pandya, Himanshu A.; Solanki, Hitesh A.

2017-10-01

The traditional drug design strategy centrally focuses on optimizing binding affinity with the receptor target and evaluates pharmacokinetic properties at a later stage which causes high rate of attrition in clinical trials. Alternatively, parallel screening allows evaluation of these properties and affinity simultaneously. In a case study to identify leads from natural compounds with experimental HIV-1 reverse transcriptase (RT) inhibition, we integrated various computational approaches including Caco-2 cell permeability QSAR model with applicability domain (AD) to recognize drug-like natural compounds, molecular docking to study HIV-1 RT interactions and shape similarity analysis with known crystal inhibitors having characteristic butterfly-like model. Further, the lipophilic properties of the compounds refined from the process with best scores were examined using lipophilic ligand efficiency (LLE) index. Seven natural compound hits viz. baicalien, (+)-calanolide A, mniopetal F, fagaronine chloride, 3,5,8-trihydroxy-4-quinolone methyl ether derivative, nitidine chloride and palmatine, were prioritized based on LLE score which demonstrated Caco-2 well absorption labeling, encompassment in AD structural coverage, better receptor affinity, shape adaptation and permissible AlogP value. We showed that this integrative approach is successful in lead exploration of natural compounds targeted against HIV-1 RT enzyme.

High Levels of Dual-Class Drug Resistance in HIV-Infected Children Failing First-Line Antiretroviral Therapy in Southern Ethiopia

PubMed Central

Kinloch, Natalie N.; Lapointe, Hope R.; Cobarrubias, Kyle D.; Foster, Byron A.; Jerene, Degu; Makonnen, Eyasu; Brumme, Zabrina L.

2018-01-01

Clinical monitoring of pediatric HIV treatment remains a major challenge in settings where drug resistance genotyping is not routinely available. As a result, our understanding of drug resistance, and its impact on subsequent therapeutic regimens available in these settings, remains limited. We investigate the prevalence and correlates of HIV-1 drug resistance among 94 participants of the Ethiopia Pediatric HIV Cohort failing first-line combination antiretroviral therapy (cART) using dried blood spot-based genotyping. Overall, 81% (73/90) of successfully genotyped participants harbored resistance mutations, including 69% (62/90) who harbored resistance to both Nucleoside Reverse Transcriptase Inhibitors (NRTIs) and Non-nucleoside Reverse Transcriptase Inhibitors (NNRTIs). Strikingly, 42% of resistant participants harbored resistance to all four NRTIs recommended for second-line use in this setting, meaning that there are effectively no remaining cART options for these children. Longer cART duration and prior regimen changes were significantly associated with detection of drug resistance mutations. Replicate genotyping increased the breadth of drug resistance detected in 34% of cases, and thus is recommended for consideration when typing from blood spots. Implementation of timely drug resistance testing and access to newer antiretrovirals and drug classes are urgently needed to guide clinical decision-making and improve outcomes for HIV-infected children on first-line cART in Ethiopia. PMID:29389912

Crystal structures of the reverse transcriptase-associated ribonuclease H domain of xenotropic murine leukemia-virus related virus

DOE Office of Scientific and Technical Information (OSTI.GOV)

Zhou, Dongwen; Chung, Suhman; Miller, Maria

2012-06-19

The ribonuclease H (RNase H) domain of retroviral reverse transcriptase (RT) plays a critical role in the life cycle by degrading the RNA strands of DNA/RNA hybrids. In addition, RNase H activity is required to precisely remove the RNA primers from nascent (-) and (+) strand DNA. We report here three crystal structures of the RNase H domain of xenotropic murine leukemia virus-related virus (XMRV) RT, namely (i) the previously identified construct from which helix C was deleted, (ii) the intact domain, and (iii) the intact domain complexed with an active site {alpha}-hydroxytropolone inhibitor. Enzymatic assays showed that the intactmore » RNase H domain retained catalytic activity, whereas the variant lacking helix C was only marginally active, corroborating the importance of this helix for enzymatic activity. Modeling of the enzyme-substrate complex elucidated the essential role of helix C in binding a DNA/RNA hybrid and its likely mode of recognition. The crystal structure of the RNase H domain complexed with {beta}-thujaplicinol clearly showed that coordination by two divalent cations mediates recognition of the inhibitor.« less

New Antiretroviral Therapies for Pediatric HIV Infection

PubMed Central

Morris, Jennifer L.; Kraus, Donna M.

2005-01-01

Human immunodeficiency virus (HIV) infection and acquired immunodeficiency syndrome affect millions of children worldwide. The development of antiretroviral therapy has significantly improved the morbidity and mortality of pediatric patients infected with HIV. Currently, 4 classes of antiretroviral agents exist: nucleoside / nucleotide reverse transcriptase inhibitors, non-nucleoside reverse transcriptase inhibitors, protease inhibitors, and entry inhibitors. A total of 21 single-entity antiretroviral agents and 4 co-formulated antiretroviral products hold Food and Drug Administration (FDA) approval for treatment of HIV-1 infection. However, not all of these agents are indicated for use in patients less than 18 years of age. Since the year 2000, 7 new antiretroviral agents (atazanavir, emtricitabine, enfuvirtide, fosamprenavir, lopinavir/ritonavir, tenofovir, and tipranavir) have been approved by the FDA for use in adult patients as part of combination therapy for the treatment of HIV-1 infection. Although only 3 of these newer agents (emtricitabine, enfuvirtide, and lopinavir/ritonavir) are currently FDA approved for use in pediatric patients, pediatric clinical studies of the other 4 new agents are currently underway. The purpose of this article is to review these 7 new antiretroviral agents and describe their roles in the treatment of pediatric HIV infection. For each drug, the following information will be addressed: FDA-approved indication and age groups, clinical efficacy, pharmacokinetics, adverse drug reactions, clinically relevant drug interactions, pediatric and adult dosing, dosage forms, administration, and place in the treatment of pediatric HIV infection. PMID:23118639

Tenofovir-based regimens associated with less drug resistance in HIV-1-infected Nigerians failing first-line antiretroviral therapy.

PubMed

Etiebet, Mary-Ann A; Shepherd, James; Nowak, Rebecca G; Charurat, Man; Chang, Harry; Ajayi, Samuel; Elegba, Olufunmilayo; Ndembi, Nicaise; Abimiku, Alashle; Carr, Jean K; Eyzaguirre, Lindsay M; Blattner, William A

2013-02-20

In resource-limited settings, HIV-1 drug resistance testing to guide antiretroviral therapy (ART) selection is unavailable. We retrospectively conducted genotypic analysis on archived samples from Nigerian patients who received targeted viral load testing to confirm treatment failure and report their drug resistance mutation patterns. Stored plasma from 349 adult patients on non-nucleoside reverse transcriptase inhibitor (NNRTI) regimens was assayed for HIV-1 RNA viral load, and samples with more than 1000 copies/ml were sequenced in the pol gene. Analysis for resistance mutations utilized the IAS-US 2011 Drug Resistance Mutation list. One hundred and seventy-five samples were genotyped; the majority of the subtypes were G (42.9%) and CRF02_AG (33.7%). Patients were on ART for a median of 27 months. 90% had the M184V/I mutation, 62% had at least one thymidine analog mutation, and 14% had the K65R mutation. 97% had an NNRTI resistance mutation and 47% had at least two etravirine-associated mutations. In multivariate analysis tenofovir-based regimens were less likely to have at least three nucleoside reverse transcriptase inhibitor (NRTI) mutations after adjusting for subtype, previous ART, CD4, and HIV viral load [P < 0.001, odds ratio (OR) 0.04]. 70% of patients on tenofovir-based regimens had at least two susceptible NRTIs to include in a second-line regimen compared with 40% on zidovudine-based regimens (P = 0.04, OR = 3.4). At recognition of treatment failure, patients on tenofovir-based first-line regimens had fewer NRTI drug-resistant mutations and more active NRTI drugs available for second-line regimens. These findings can inform strategies for ART regimen sequencing to optimize long-term HIV treatment outcomes in low-resource settings.

The reverse transcriptase encoded by LINE-1 retrotransposons in the genesis, progression and therapy of cancer

NASA Astrophysics Data System (ADS)

Sciamanna, Ilaria; De Luca, Chiara; Spadafora, Corrado

2016-02-01

In higher eukaryotic genomes, Long Interspersed Nuclear Element 1 (LINE-1) retrotransposons represent a large family of repeated genomic elements. They transpose using a reverse transcriptase (RT), which they encode as part of the ORF2p product. RT inhibition in cancer cells, either via RNA interference-dependent silencing of active LINE-1 elements, or using RT inhibitory drugs, reduces cancer cell proliferation, promotes their differentiation and antagonizes tumor progression in animal models. Indeed, the nonnucleoside RT inhibitor efavirenz has recently been tested in a phase II clinical trial with metastatic prostate cancer patients. An in-depth analysis of ORF2p in a mouse model of breast cancer showed ORF2p to be precociously expressed in precancerous lesions and highly abundant in advanced cancer stages, while being barely detectable in normal breast tissue, providing a rationale for the finding that RT-expressing tumours are therapeutically sensitive to RT inhibitors. We summarise mechanistic and gene profiling studies indicating that highly abundant LINE-1-derived RT can “sequester” RNA substrates for reverse transcription in tumor cells, entailing the formation of RNA:DNA hybrid molecules and impairing the overall production of regulatory miRNAs, with a global impact on the cell transcriptome. Based on these data, LINE-1-ORF2 encoded RT has a tumor-promoting potential that is exerted at an epigenetic level. We propose a model whereby LINE1-RT drives a previously unrecognized global regulatory process, the deregulation of which drives cell transformation and tumorigenesis and possibly implicated in cancer cell heterogeneity.

A molecular-field-based similarity study of non-nucleoside HIV-1 reverse transcriptase inhibitors

NASA Astrophysics Data System (ADS)

Mestres, Jordi; Rohrer, Douglas C.; Maggiora, Gerald M.

1999-01-01

This article describes a molecular-field-based similarity method for aligning molecules by matching their steric and electrostatic fields and an application of the method to the alignment of three structurally diverse non-nucleoside HIV-1 reverse transcriptase inhibitors. A brief description of the method, as implemented in the program MIMIC, is presented, including a discussion of pairwise and multi-molecule similarity-based matching. The application provides an example that illustrates how relative binding orientations of molecules can be determined in the absence of detailed structural information on their target protein. In the particular system studied here, availability of the X-ray crystal structures of the respective ligand-protein complexes provides a means for constructing an 'experimental model' of the relative binding orientations of the three inhibitors. The experimental model is derived by using MIMIC to align the steric fields of the three protein P66 subunit main chains, producing an overlay with a 1.41 Å average rms distance between the corresponding Cα's in the three chains. The inter-chain residue similarities for the backbone structures show that the main-chain conformations are conserved in the region of the inhibitor-binding site, with the major deviations located primarily in the 'finger' and RNase H regions. The resulting inhibitor structure overlay provides an experimental-based model that can be used to evaluate the quality of the direct a priori inhibitor alignment obtained using MIMIC. It is found that the 'best' pairwise alignments do not always correspond to the experimental model alignments. Therefore, simply combining the best pairwise alignments will not necessarily produce the optimal multi-molecule alignment. However, the best simultaneous three-molecule alignment was found to reproduce the experimental inhibitor alignment model. A pairwise consistency index has been derived which gauges the quality of combining the pairwise alignments and aids in efficiently forming the optimal multi-molecule alignment analysis. Two post-alignment procedures are described that provide information on feature-based and field-based pharmacophoric patterns. The former corresponds to traditional pharmacophore models and is derived from the contribution of individual atoms to the total similarity. The latter is based on molecular regions rather than atoms and is constructed by computing the percent contribution to the similarity of individual points in a regular lattice surrounding the molecules, which when contoured and colored visually depict regions of highly conserved similarity. A discussion of how the information provided by each of the procedures is useful in drug design is also presented.

Efficacy and safety of TMC278 in antiretroviral-naive HIV-1 patients: week 96 results of a phase IIb randomized trial.

PubMed

Pozniak, Anton L; Morales-Ramirez, Javier; Katabira, Elly; Steyn, Dewald; Lupo, Sergio H; Santoscoy, Mario; Grinsztejn, Beatriz; Ruxrungtham, Kiat; Rimsky, Laurence T; Vanveggel, Simon; Boven, Katia

2010-01-02

TMC278 is a next-generation nonnucleoside reverse transcriptase inhibitor highly active against wild-type and nonnucleoside reverse transcriptase inhibitor-resistant HIV-1 in vitro. The week 96 analysis of TMC278-C204, a large dose-ranging study of TMC278 in treatment-naive HIV-1-infected patients, is presented. Phase IIb randomized trial. Three hundred sixty-eight patients were randomized and treated with three blinded once-daily TMC278 doses 25, 75 or 150 mg, or an open-label, active control, efavirenz 600 mg once daily, all with two nucleoside reverse transcriptase inhibitors. The primary analysis was at week 48. No TMC278 dose-response relationship for efficacy and safety was observed. TMC278 demonstrated potent antiviral efficacy comparable with efavirenz over 48 weeks that was sustained to week 96 (76.9-80.0% and 71.4-76.3% of TMC278-treated patients with confirmed viral load <50 copies/ml, respectively; time-to-loss of virological-response algorithm). Median increases from baseline in CD4 cell count with TMC278 at week 96 (138.0-149.0 cells/microl) were higher than at week 48 (108.0-123.0 cells/microl). All TMC278 doses were well tolerated. The incidences of the most commonly reported grade 2-4 adverse events at least possibly related to study medication, including nausea, dizziness, abnormal dreams/nightmare, dyspepsia, asthenia, rash, somnolence and vertigo, were low and lower with TMC278 than with efavirenz. Incidences of serious adverse events, grade 3 or 4 adverse events and discontinuations due to adverse events were similar among groups. All TMC278 doses demonstrated potent and sustained efficacy comparable with efavirenz in treatment-naive patients over 96 weeks. TMC278 was well tolerated with lower incidences of neurological and psychiatric adverse events, rash and lower lipid elevations than those with efavirenz. TMC278 25 mg once daily was selected for further clinical development.

Protein-mediated antagonism between HIV reverse transcriptase ligands nevirapine and MgATP.

PubMed

Zheng, Xunhai; Mueller, Geoffrey A; DeRose, Eugene F; London, Robert E

2013-06-18

Nonnucleoside reverse transcriptase inhibitors (NNRTIs) play a central role in the treatment of AIDS, but their mechanisms of action are incompletely understood. The interaction of the NNRTI nevirapine (NVP) with HIV-1 reverse transcriptase (RT) is characterized by a preference for the open conformation of the fingers/thumb subdomains, and a reported variation of three orders of magnitude between the binding affinity of NVP for RT in the presence or absence of primer/template DNA. To investigate the relationship between conformation and ligand binding, we evaluated the use of methionine NMR probes positioned near the tip of the fingers or thumb subdomains. Such probes would be expected to be sensitive to changes in the local environment depending on the fractions of open and closed RT. Comparisons of the NMR spectra of three conservative mutations, I63M, L74M, and L289M, indicated that M63 showed the greatest shift sensitivity to the addition of NVP. The exchange kinetics of the M63 resonance are fast on the chemical shift timescale, but become slow in the presence of NVP due to the slow binding of RT with the inhibitor. The simplest model consistent with this behavior involves a rapid open/closed equilibrium coupled with a slow interaction of the inhibitor with the open conformation. Studies of RT in the presence of both NVP and MgATP indicate a strong negative cooperativity. Binding of MgATP reduces the fraction of RT bound to NVP, as indicated by the intensity of the NVP-perturbed M230 resonance, and enhances the dissociation rate constant of the NVP, resulting in an increase of the open/closed interconversion rate, so that the M63 resonance moves into the fast/intermediate-exchange regime. Protein-mediated interactions appear to explain most of the affinity variation of NVP for RT. Copyright © 2013 Biophysical Society. Published by Elsevier Inc. All rights reserved.

HIV-1 transmitted drug resistance and genetic diversity among patients from Piauí State, Northeast Brazil.

PubMed

Moura, Maria Edileuza Soares; da Guarda Reis, Mônica Nogueira; Lima, Yanna Andressa Ramos; Eulálio, Kelsen Dantas; Cardoso, Ludimila Paula Vaz; Stefani, Mariane Martins Araújo

2015-05-01

HIV-1 transmitted-drug-resistance and genetic diversity are dynamic and may differ in distinct locations/risk groups. In Brazil, increased AIDS incidence and related mortality have been detected in the Northeast region, differently from the epicenter in the Southeast. This cross-sectional study describes transmitted-dru- resistance and HIV-1 subtypes in protease/PR and reverse transcriptase/RT regions among antiretroviral naïve patients from Piauí State, Northeast Brazil. Among 96 patients recruited 89 (92.7%) had HIV-1 PR/RT regions sequenced: 44 females and 45 males, 22 self-declared as men who have sex with men. Transmitted-drug-resistance was investigated by CPR tool (Stanford HIV-1 Drug Resistance/SDRM). HIV-1 subtypes were assigned by REGA and phylogenetic inference. Overall, transmitted-drug-resistance rate was 11.2% (10/89; CI 95%: 5.8-19.1%); 22.7% among men who have sex with men (5/22; CI 95%: 8.8-43.4%), 10% in heterosexual men (2/20; CI 95%: 1.7-29.3%) and 6.8% in women (3/44; CI 95%: 1.8-17.4%). Singleton mutations to protease-inhibitor/PI, nucleoside-reverse-transcriptase-inhibitor/NRTI or non-nucleoside-reverse-transcriptase-inhibitor/NNRTI predominated (8/10): PI mutations (M46L, V82F, L90M); NRTI mutations (M41L, D67N) and NNRTI mutations (K103N/S). Dual class resistance mutations to NRTI and NNRTI were observed: T215L (NRTI), Y188L (NNRTI) and T215N (NRTI), F227L (NNRTI). Subtype B prevailed (86.6%; 77/89), followed by subtype F1 (1.1%, 1/89) and subtype C (1.1%, 1/89). B/F1 and B/C intersubtype recombinants represented 11.2% (10/89). In Piauí State extensive testing of incidence and transmitted-drug-resistance in all populations with risk behaviors may help control AIDS epidemic locally. © 2015 Wiley Periodicals, Inc.

Exploiting the anti-HIV 6-desfluoroquinolones to design multiple ligands.

PubMed

Sancineto, Luca; Iraci, Nunzio; Barreca, Maria Letizia; Massari, Serena; Manfroni, Giuseppe; Corazza, Gianmarco; Cecchetti, Violetta; Marcello, Alessandro; Daelemans, Dirk; Pannecouque, Christophe; Tabarrini, Oriana

2014-09-01

It is getting clearer that many drugs effective in different therapeutic areas act on multiple rather than single targets. The application of polypharmacology concepts might have numerous advantages especially for disease such as HIV/AIDS, where the rapid emergence of resistance requires a complex combination of more than one drug. In this paper, we have designed three hybrid molecules combining WM5, a quinolone derivative we previously identified as HIV Tat-mediated transcription (TMT) inhibitor, with the tricyclic core of nevirapine and BILR 355BS (BILR) non-nucleoside reverse transcriptase inhibitors (NNRTIs) to investigate whether it could be possible to obtain molecules acting on both transcription steps of the HIV replicative cycle. One among the three designed multiple ligands, reached this goal. Indeed, compound 1 inhibited both TMT and reverse transcriptase (RT) activity. Unexpectedly, while the anti-TMT activity exerted by compound 1 resulted into a selective inhibition of HIV-1 reactivation from latently infected OM10.1 cells, the anti-RT properties shown by all of the synthesized compounds did not translate into an anti-HIV activity in acutely infected cells. Thus, we have herein produced the proof of concept that the design of dual TMT-RT inhibitors is indeed possible, but optimization efforts are needed to obtain more potent derivatives. Copyright © 2014 Elsevier Ltd. All rights reserved.

Active methamphetamine use is associated with transmitted drug resistance to non-nucleoside reverse transcriptase inhibitors in individuals with HIV infection of unknown duration.

PubMed

Cachay, Edward R; Moini, Niousha; Kosakovsky Pond, Sergei L; Pesano, Rick; Lie, Yolanda S; Aiem, Heidi; Butler, David M; Letendre, Scott; Mathews, Wm Christopher; Smith, Davey M

2007-01-01

Frequent methamphetamine use among recently HIV infected individuals is associated with transmitted drug resistance (TDR) to non-nucleoside reverse transcriptase inhibitors (NNRTI); however, the reversion time of TDR to drug susceptible HIV may exceed 3 years. We assessed whether recreational substance use is associated with detectable TDR among individuals newly diagnosed with HIV infection of unknown duration. Cross-sectional analysis. Subjects were enrolled at the University California, San Diego Early Intervention Program. Demographic, clinical and substance use data were collected using structured interviews. Genotypic resistance testing was performed using GeneSeq, Monogram Biosciences. We analyzed the association between substance use and TDR using bivariate analyses and the corresponding transmission networks using phylogenetic models. Between April 2004 and July 2006, 115 individuals with genotype data were enrolled. The prevalence of alcohol, marijuana and methamphetamine use were 98%, 71% and 64% respectively. Only active methamphetamine use in the 30 days prior to HIV diagnosis was independently associated with TDR to NNRTI (OR: 6.6; p=0.002). Despite not knowing the duration of their HIV infection, individuals reporting active methamphetamine use in the 30 days prior to HIV diagnosis are at an increased risk of having HIV strains that are resistant to NNRTI.

Free Energy-Based Virtual Screening and Optimization of RNase H Inhibitors of HIV-1 Reverse Transcriptase.

PubMed

Zhang, Baofeng; D'Erasmo, Michael P; Murelli, Ryan P; Gallicchio, Emilio

2016-09-30

We report the results of a binding free energy-based virtual screening campaign of a library of 77 α-hydroxytropolone derivatives against the challenging RNase H active site of the reverse transcriptase (RT) enzyme of human immunodeficiency virus-1. Multiple protonation states, rotamer states, and binding modalities of each compound were individually evaluated. The work involved more than 300 individual absolute alchemical binding free energy parallel molecular dynamics calculations and over 1 million CPU hours on national computing clusters and a local campus computational grid. The thermodynamic and structural measures obtained in this work rationalize a series of characteristics of this system useful for guiding future synthetic and biochemical efforts. The free energy model identified key ligand-dependent entropic and conformational reorganization processes difficult to capture using standard docking and scoring approaches. Binding free energy-based optimization of the lead compounds emerging from the virtual screen has yielded four compounds with very favorable binding properties, which will be the subject of further experimental investigations. This work is one of the few reported applications of advanced-binding free energy models to large-scale virtual screening and optimization projects. It further demonstrates that, with suitable algorithms and automation, advanced-binding free energy models can have a useful role in early-stage drug-discovery programs.

Synthesis and biological evaluation of 2-thioxopyrimidin-4(1H)-one derivatives as potential non-nucleoside HIV-1 reverse transcriptase inhibitors.

PubMed

Khalifa, Nagy M; Al-Omar, Mohamed A

2014-11-12

A series of new 5-allyl-6-benzylpyrimidin-4(3H)-ones bearing different substituents at the C-2 position of the pyrimidine core have been synthesized and evaluated for their in vitro activities against human immunodeficiency virus type 1 (HIV-1) in the human T-lymphotropic type (MT-4 cell cultures). The majority of the title compounds showed moderate to good activities against HIV-1. Amongst them, 5-allyl-6-benzyl-2-(3-hydroxypropylthio)pyrimidin-4(3H)-one analogue 11c exhibited the most potent anti-HIV-1 activity (IC50 0.32 µM). The biological testing results clearly indicated that the substitution at C-2 position of the pyrimidine ring could increase the anti-HIV-1 reverse transcriptase (RT) activity.

Synthesis and Biological Evaluation of 2-Thioxopyrimidin-4(1H)-one Derivatives as Potential Non-Nucleoside HIV-1 Reverse Transcriptase Inhibitors

PubMed Central

Khalifa, Nagy M.; Al-Omar, Mohamed A.

2014-01-01

A series of new 5-allyl-6-benzylpyrimidin-4(3H)-ones bearing different substituents at the C-2 position of the pyrimidine core have been synthesized and evaluated for their in vitro activities against human immunodeficiency virus type 1 (HIV-1) in the human T-lymphotropic type (MT-4 cell cultures). The majority of the title compounds showed moderate to good activities against HIV-1. Amongst them, 5-allyl-6-benzyl-2-(3-hydroxypropylthio)pyrimidin-4(3H)-one analogue 11c exhibited the most potent anti-HIV-1 activity (IC50 0.32 µM). The biological testing results clearly indicated that the substitution at C-2 position of the pyrimidine ring could increase the anti-HIV-1 reverse transcriptase (RT) activity. PMID:25397597

Special coverage: 9th Conference on Retroviruses. New drugs, new data hold promise for next decade of HIV treatment.

PubMed

2002-05-01

Antiretroviral research presented recently at the 9th Conference on Retroviruses and Opportunistic Infections demonstrates that investigators and pharmaceutical companies continue to strive for the next highly potent and easily tolerated anti-HIV drug. Among the new approaches are entry inhibitor drug and second-generation non-nucleoside reverse transcriptase inhibitors. New studies also looked into potency against multidrug-resistant virus and medication regimens that are simpler to take and have fewer side effects.

1-Benzyl-2-(1H-indol-3-yl)-5-oxo-pyrrolidine-2-carbonitrile.

PubMed

Tamazyan, Rafael; Armen, Ayvazyan; Ashot, Martirosyan; Sahak, Gasparyan; Schinazi, Raymond

2008-01-04

In the title compound, C(20)H(17)N(3)O, a potential anti-human immunodeficiency virus type 1 (HIV-1) non-nucleoside reverse-transcriptase inhibitor, the pyrrolidine ring has an envelope conformation. In the crystal structure, adjacent mol-ecules are connected into infinite chains via an N-H⋯O hydrogen bond.

Suicide Inhibitors of Reverse Transcriptase in the Therapy of AIDS and Other Retroviruses

DTIC Science & Technology

1990-07-01

I and 10 nanonolar) and compared to the E . Coli recombinant HIV-RT (Kindly donated by Dr. Steven Hughes Fort Detrick M.D.) and the wild type HIV-RT...Both the wild type and E . Coli HIV-RT’s were resistant to PFA showing essentially no inhibition at the lOnM level. Previous studies have shown that...10 nanomolar PFA. j, Sentivitv of Recombinant HIV-Reverse Transcriotase to Foscarnet. RECOMBINANT HIV RT ( E . COLI) + FOSCARNET 350001R 300000 PFA .001

In vitro cross-resistance profile of nucleoside reverse transcriptase inhibitor (NRTI) BMS-986001 against known NRTI resistance mutations.

PubMed

Li, Zhufang; Terry, Brian; Olds, William; Protack, Tricia; Deminie, Carol; Minassian, Beatrice; Nowicka-Sans, Beata; Sun, Yongnian; Dicker, Ira; Hwang, Carey; Lataillade, Max; Hanna, George J; Krystal, Mark

2013-11-01

BMS-986001 is a novel HIV nucleoside reverse transcriptase inhibitor (NRTI). To date, little is known about its resistance profile. In order to examine the cross-resistance profile of BMS-986001 to NRTI mutations, a replicating virus system was used to examine specific amino acid mutations known to confer resistance to various NRTIs. In addition, reverse transcriptases from 19 clinical isolates with various NRTI mutations were examined in the Monogram PhenoSense HIV assay. In the site-directed mutagenesis studies, a virus containing a K65R substitution exhibited a 0.4-fold change in 50% effective concentration (EC50) versus the wild type, while the majority of viruses with the Q151M constellation (without M184V) exhibited changes in EC50 versus wild type of 0.23- to 0.48-fold. Susceptibility to BMS-986001 was also maintained in an L74V-containing virus (0.7-fold change), while an M184V-only-containing virus induced a 2- to 3-fold decrease in susceptibility. Increasing numbers of thymidine analog mutation pattern 1 (TAM-1) pathway mutations correlated with decreases in susceptibility to BMS-986001, while viruses with TAM-2 pathway mutations exhibited a 5- to 8-fold decrease in susceptibility, regardless of the number of TAMs. A 22-fold decrease in susceptibility to BMS-986001 was observed in a site-directed mutant containing the T69 insertion complex. Common non-NRTI (NNRTI) mutations had little impact on susceptibility to BMS-986001. The results from the site-directed mutants correlated well with the more complicated genotypes found in NRTI-resistant clinical isolates. Data from clinical studies are needed to determine the clinically relevant resistance cutoff values for BMS-986001.

An integrated molecular dynamics, principal component analysis and residue interaction network approach reveals the impact of M184V mutation on HIV reverse transcriptase resistance to lamivudine.

PubMed

Bhakat, Soumendranath; Martin, Alberto J M; Soliman, Mahmoud E S

2014-08-01

The emergence of different drug resistant strains of HIV-1 reverse transcriptase (HIV RT) remains of prime interest in relation to viral pathogenesis as well as drug development. Amongst those mutations, M184V was found to cause a complete loss of ligand fitness. In this study, we report the first account of the molecular impact of M184V mutation on HIV RT resistance to 3TC (lamivudine) using an integrated computational approach. This involved molecular dynamics simulation, binding free energy analysis, principle component analysis (PCA) and residue interaction networks (RINs). Results clearly confirmed that M184V mutation leads to steric conflict between 3TC and the beta branched side chain of valine, decreases the ligand (3TC) binding affinity by ∼7 kcal mol(-1) when compared to the wild type, changes the overall conformational landscape of the protein and distorts the native enzyme residue-residue interaction network. The comprehensive molecular insight gained from this study should be of great importance in understanding drug resistance against HIV RT as well as assisting in the design of novel reverse transcriptase inhibitors with high ligand efficacy on resistant strains.

The mechano-chemistry of a monomeric reverse transcriptase

PubMed Central

Malik, Omri; Khamis, Hadeel; Rudnizky, Sergei

2017-01-01

Abstract Retroviral reverse transcriptase catalyses the synthesis of an integration-competent dsDNA molecule, using as a substrate the viral RNA. Using optical tweezers, we follow the Murine Leukemia Virus reverse transcriptase as it performs strand-displacement polymerization on a template under mechanical force. Our results indicate that reverse transcriptase functions as a Brownian ratchet, with dNTP binding as the rectifying reaction of the ratchet. We also found that reverse transcriptase is a relatively passive enzyme, able to polymerize on structured templates by exploiting their thermal breathing. Finally, our results indicate that the enzyme enters the recently characterized backtracking state from the pre-translocation complex. PMID:29165701

The role of the glycosyl moiety of myricetin derivatives in anti-HIV-1 activity in vitro.

PubMed

Ortega, Joseph T; Suárez, Alirica I; Serrano, Maria L; Baptista, Jani; Pujol, Flor H; Rangel, Hector R

2017-10-12

Plant extracts are sources of valuable compounds with biological activity, especially for the anti-proliferative activity against pathogens or tumor cells. Myricetin is a flavonoid found in several plants that has been described as an inhibitor of Human immunodeficiency virus type 1 (HIV-1) through its action against the HIV reverse transcriptase, but myricetin derivatives have not been fully studied. The aim of this study was to evaluate the anti-HIV-1 activity of glycosylated metabolites obtained from Marcetia taxifolia and derived from myricetin: myricetin rhamnoside and myricetin 3-(6-rhamnosylgalactoside). Compounds were obtained from organic extracts by maceration of aerial parts of M. taxifolia. All biological assays were performed in the MT4 cell line. Antiviral activity was measured as inhibition of p24 and reverse transcriptase with a fluorescent assay. Both flavonoids have antiviral activity in vitro, with an EC50 of 120 µM for myricetin 3-rhamnoside (MR) and 45 µM for myricetin 3-(6-rhamnosylgalactoside) (MRG), both significantly lower than the EC50 of myricetin (230 µM). Although both compounds inhibited the reverse transcriptase activity, with an IC50 of 10.6 µM for MR and 13.8 µM for MRG, myricetin was the most potent, with an IC50 of 7.6 µM, and an inhibition greater than 80%. Molecular docking approach showed correlation between the free energy of binding with the assays of enzyme inhibition. The results suggest that glycosylated moiety might enhance the anti-HIV-1 activity of myricetin, probably by favoring the internalization of the flavonoid into the cell. The inhibition of the HIV-1 reverse transcriptase is likely responsible for the antiviral activity.

Impact of Human Immunodeficiency Virus Type 1 Minority Variants on the Virus Response to a Rilpivirine-Based First-line Regimen.

PubMed

Raymond, Stéphanie; Nicot, Florence; Pallier, Coralie; Bellecave, Pantxika; Maillard, Anne; Trabaud, Mary Anne; Morand-Joubert, Laurence; Rodallec, Audrey; Amiel, Corinne; Mourez, Thomas; Bocket, Laurence; Beby-Defaux, Agnès; Bouvier-Alias, Magali; Lambert-Niclot, Sidonie; Charpentier, Charlotte; Malve, Brice; Mirand, Audrey; Dina, Julia; Le Guillou-Guillemette, Hélène; Marque-Juillet, Stéphanie; Signori-Schmuck, Anne; Barin, Francis; Si-Mohamed, Ali; Avettand Fenoel, Véronique; Roussel, Catherine; Calvez, Vincent; Saune, Karine; Marcelin, Anne Geneviève; Rodriguez, Christophe; Descamps, Diane; Izopet, Jacques

2018-05-02

Minority resistant variants of human immunodeficiency virus type 1 (HIV-1) could influence the virological response to treatment based on nonnucleoside reverse transcriptase inhibitors (NNRTIs). Data on minority rilpivirine-resistant variants are scarce. This study used next-generation sequencing (NGS) to identify patients harboring minority resistant variants to nucleos(t)ide reverse transcriptase inhibitors and NNRTIs and to assess their influence on the virological response (VR). All the subjects, 541 HIV-1-infected patients started a first-line regimen containing rilpivirine. VR was defined as a HIV-1 RNA load <50 copies/mL at month 6 with continued suppression at month 12. NGS was performed at baseline (retrospectively) on the 454 GS-FLX platform (Roche). NGS revealed resistance-associated mutations accounting for 1% to <5% of variants in 17.2% of samples, for 5%-20% in 5.7% of samples, and for >20% in 29% of samples. We identified 43 (8.8%) and 36 (7.4%) patients who harbored rilpivirine-resistant variants with a 1% sensitivity threshold according to the French National Agency for Research on AIDS and Viral Hepatitis and Stanford algorithms, respectively. The VR was 96.9% at month 12. Detection of minority rilpivirine resistant variants was not associated with virological failure (VF). Multivariate analysis indicated that VF at month 12 was associated with a CD4 count <250 cells/µL at baseline, a slower decrease in viral load at month 3, and rilpivirine resistance at baseline using the Stanford algorithm with a 20% threshold. Minority resistant variants had no impact on the VR of treatment-naive patients to a rilpivirine-based regimen.

Transmission of HIV Drug Resistance and the Predicted Effect on Current First-line Regimens in Europe.

PubMed

Hofstra, L Marije; Sauvageot, Nicolas; Albert, Jan; Alexiev, Ivailo; Garcia, Federico; Struck, Daniel; Van de Vijver, David A M C; Åsjö, Birgitta; Beshkov, Danail; Coughlan, Suzie; Descamps, Diane; Griskevicius, Algirdas; Hamouda, Osamah; Horban, Andrzej; Van Kasteren, Marjo; Kolupajeva, Tatjana; Kostrikis, Leondios G; Liitsola, Kirsi; Linka, Marek; Mor, Orna; Nielsen, Claus; Otelea, Dan; Paraskevis, Dimitrios; Paredes, Roger; Poljak, Mario; Puchhammer-Stöckl, Elisabeth; Sönnerborg, Anders; Staneková, Danica; Stanojevic, Maja; Van Laethem, Kristel; Zazzi, Maurizio; Zidovec Lepej, Snjezana; Boucher, Charles A B; Schmit, Jean-Claude; Wensing, Annemarie M J; Puchhammer-Stockl, E; Sarcletti, M; Schmied, B; Geit, M; Balluch, G; Vandamme, A-M; Vercauteren, J; Derdelinckx, I; Sasse, A; Bogaert, M; Ceunen, H; De Roo, A; De Wit, S; Echahidi, F; Fransen, K; Goffard, J-C; Goubau, P; Goudeseune, E; Yombi, J-C; Lacor, P; Liesnard, C; Moutschen, M; Pierard, D; Rens, R; Schrooten, Y; Vaira, D; Vandekerckhove, L P R; Van den Heuvel, A; Van Der Gucht, B; Van Ranst, M; Van Wijngaerden, E; Vandercam, B; Vekemans, M; Verhofstede, C; Clumeck, N; Van Laethem, K; Beshkov, D; Alexiev, I; Lepej, S Zidovec; Begovac, J; Kostrikis, L; Demetriades, I; Kousiappa, I; Demetriou, V; Hezka, J; Linka, M; Maly, M; Machala, L; Nielsen, C; Jørgensen, L B; Gerstoft, J; Mathiesen, L; Pedersen, C; Nielsen, H; Laursen, A; Kvinesdal, B; Liitsola, K; Ristola, M; Suni, J; Sutinen, J; Descamps, D; Assoumou, L; Castor, G; Grude, M; Flandre, P; Storto, A; Hamouda, O; Kücherer, C; Berg, T; Braun, P; Poggensee, G; Däumer, M; Eberle, J; Heiken, H; Kaiser, R; Knechten, H; Korn, K; Müller, H; Neifer, S; Schmidt, B; Walter, H; Gunsenheimer-Bartmeyer, B; Harrer, T; Paraskevis, D; Hatzakis, A; Zavitsanou, A; Vassilakis, A; Lazanas, M; Chini, M; Lioni, A; Sakka, V; Kourkounti, S; Paparizos, V; Antoniadou, A; Papadopoulos, A; Poulakou, G; Katsarolis, I; Protopapas, K; Chryssos, G; Drimis, S; Gargalianos, P; Xylomenos, G; Lourida, G; Psichogiou, M; Daikos, G L; Sipsas, N V; Kontos, A; Gamaletsou, M N; Koratzanis, G; Sambatakou, H; Mariolis, H; Skoutelis, A; Papastamopoulos, V; Georgiou, O; Panagopoulos, P; Maltezos, E; Coughlan, S; De Gascun, C; Byrne, C; Duffy, M; Bergin, C; Reidy, D; Farrell, G; Lambert, J; O'Connor, E; Rochford, A; Low, J; Coakely, P; O'Dea, S; Hall, W; Mor, O; Levi, I; Chemtob, D; Grossman, Z; Zazzi, M; de Luca, A; Balotta, C; Riva, C; Mussini, C; Caramma, I; Capetti, A; Colombo, M C; Rossi, C; Prati, F; Tramuto, F; Vitale, F; Ciccozzi, M; Angarano, G; Rezza, G; Kolupajeva, T; Vasins, O; Griskevicius, A; Lipnickiene, V; Schmit, J C; Struck, D; Sauvageot, N; Hemmer, R; Arendt, V; Michaux, C; Staub, T; Sequin-Devaux, C; Wensing, A M J; Boucher, C A B; van de Vijver, D A M C; van Kessel, A; van Bentum, P H M; Brinkman, K; Connell, B J; van der Ende, M E; Hoepelman, I M; van Kasteren, M; Kuipers, M; Langebeek, N; Richter, C; Santegoets, R M W J; Schrijnders-Gudde, L; Schuurman, R; van de Ven, B J M; Åsjö, B; Kran, A-M Bakken; Ormaasen, V; Aavitsland, P; Horban, A; Stanczak, J J; Stanczak, G P; Firlag-Burkacka, E; Wiercinska-Drapalo, A; Jablonowska, E; Maolepsza, E; Leszczyszyn-Pynka, M; Szata, W; Camacho, R; Palma, C; Borges, F; Paixão, T; Duque, V; Araújo, F; Otelea, D; Paraschiv, S; Tudor, A M; Cernat, R; Chiriac, C; Dumitrescu, F; Prisecariu, L J; Stanojevic, M; Jevtovic, Dj; Salemovic, D; Stanekova, D; Habekova, M; Chabadová, Z; Drobkova, T; Bukovinova, P; Shunnar, A; Truska, P; Poljak, M; Lunar, M; Babic, D; Tomazic, J; Vidmar, L; Vovko, T; Karner, P; Garcia, F; Paredes, R; Monge, S; Moreno, S; Del Amo, J; Asensi, V; Sirvent, J L; de Mendoza, C; Delgado, R; Gutiérrez, F; Berenguer, J; Garcia-Bujalance, S; Stella, N; de Los Santos, I; Blanco, J R; Dalmau, D; Rivero, M; Segura, F; Elías, M J Pérez; Alvarez, M; Chueca, N; Rodríguez-Martín, C; Vidal, C; Palomares, J C; Viciana, I; Viciana, P; Cordoba, J; Aguilera, A; Domingo, P; Galindo, M J; Miralles, C; Del Pozo, M A; Ribera, E; Iribarren, J A; Ruiz, L; de la Torre, J; Vidal, F; Clotet, B; Albert, J; Heidarian, A; Aperia-Peipke, K; Axelsson, M; Mild, M; Karlsson, A; Sönnerborg, A; Thalme, A; Navér, L; Bratt, G; Karlsson, A; Blaxhult, A; Gisslén, M; Svennerholm, B; Bergbrant, I; Björkman, P; Säll, C; Mellgren, Å; Lindholm, A; Kuylenstierna, N; Montelius, R; Azimi, F; Johansson, B; Carlsson, M; Johansson, E; Ljungberg, B; Ekvall, H; Strand, A; Mäkitalo, S; Öberg, S; Holmblad, P; Höfer, M; Holmberg, H; Josefson, P; Ryding, U

2016-03-01

Numerous studies have shown that baseline drug resistance patterns may influence the outcome of antiretroviral therapy. Therefore, guidelines recommend drug resistance testing to guide the choice of initial regimen. In addition to optimizing individual patient management, these baseline resistance data enable transmitted drug resistance (TDR) to be surveyed for public health purposes. The SPREAD program systematically collects data to gain insight into TDR occurring in Europe since 2001. Demographic, clinical, and virological data from 4140 antiretroviral-naive human immunodeficiency virus (HIV)-infected individuals from 26 countries who were newly diagnosed between 2008 and 2010 were analyzed. Evidence of TDR was defined using the WHO list for surveillance of drug resistance mutations. Prevalence of TDR was assessed over time by comparing the results to SPREAD data from 2002 to 2007. Baseline susceptibility to antiretroviral drugs was predicted using the Stanford HIVdb program version 7.0. The overall prevalence of TDR did not change significantly over time and was 8.3% (95% confidence interval, 7.2%-9.5%) in 2008-2010. The most frequent indicators of TDR were nucleoside reverse transcriptase inhibitor (NRTI) mutations (4.5%), followed by nonnucleoside reverse transcriptase inhibitor (NNRTI) mutations (2.9%) and protease inhibitor mutations (2.0%). Baseline mutations were most predictive of reduced susceptibility to initial NNRTI-based regimens: 4.5% and 6.5% of patient isolates were predicted to have resistance to regimens containing efavirenz or rilpivirine, respectively, independent of current NRTI backbones. Although TDR was highest for NRTIs, the impact of baseline drug resistance patterns on susceptibility was largest for NNRTIs. The prevalence of TDR assessed by epidemiological surveys does not clearly indicate to what degree susceptibility to different drug classes is affected. © The Author 2015. Published by Oxford University Press for the Infectious Diseases Society of America.

Nonnucleoside Reverse-transcriptase Inhibitor- vs Ritonavir-boosted Protease Inhibitor-based Regimens for Initial Treatment of HIV Infection: A Systematic Review and Metaanalysis of Randomized Trials.

PubMed

Borges, Álvaro H; Lundh, Andreas; Tendal, Britta; Bartlett, John A; Clumeck, Nathan; Costagliola, Dominique; Daar, Eric S; Echeverría, Patrícia; Gisslén, Magnus; Huedo-Medina, Tania B; Hughes, Michael D; Huppler Hullsiek, Katherine; Khabo, Paul; Komati, Stephanus; Kumar, Princy; Lockman, Shahin; MacArthur, Rodger D; Maggiolo, Franco; Matteelli, Alberto; Miro, Jose M; Oka, Shinichi; Petoumenos, Kathy; Puls, Rebekah L; Riddler, Sharon A; Sax, Paul E; Sierra-Madero, Juan; Torti, Carlo; Lundgren, Jens D

2016-07-15

Previous studies suggest that nonnucleoside reverse-transcriptase inhibitors (NNRTIs) cause faster virologic suppression, while ritonavir-boosted protease inhibitors (PI/r) recover more CD4 cells. However, individual trials have not been powered to compare clinical outcomes. We searched databases to identify randomized trials that compared NNRTI- vs PI/r-based initial therapy. A metaanalysis calculated risk ratios (RRs) or mean differences (MDs), as appropriate. Primary outcome was death or progression to AIDS. Secondary outcomes were death, progression to AIDS, and treatment discontinuation. We calculated RR of virologic suppression and MD for an increase in CD4 cells at week 48. We included 29 trials with 9047 participants. Death or progression to AIDS occurred in 226 participants in the NNRTI arm and in 221 in the PI/r arm (RR, 1.03; 95% confidence interval, .87-1.22; 12 trials; n = 3825), death in 205 participants in the NNRTI arm vs 198 in the PI/r arm (1.04; 0.86-1.25; 22 trials; n = 8311), and progression to AIDS in 140 participants in the NNRTI arm vs 144 in the PI/r arm (1.00; 0.80-1.25; 13 trials; n = 4740). Overall treatment discontinuation (1.12; 0.93-1.35; 24 trials; n = 8249) and from toxicity (1.21; 0.87-1.68; 21 trials; n = 6195) were comparable, but discontinuation due to virologic failure was more common with NNRTI (1.58; 0.91-2.74; 17 trials; n = 5371). At week 48, there was no difference between NNRTI and PI/r in virologic suppression (RR, 1.03; 0.98-1.09) or CD4(+) recovery (MD, -4.7 cells; -14.2 to 4.8). We found no difference in clinical and viro-immunologic outcomes between NNRTI- and PI/r-based therapy. © The Author 2016. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail journals.permissions@oup.com.

Current and future management of treatment failure in low- and middle-income countries.

PubMed

Boyd, Mark A

2010-01-01

Access to second-line therapy in low- and middle-income countries has been limited to date. The WHO predicts that between 500 000 and 800 000 HIV-infected people on first-line combination antiretroviral therapy will require switch to second-line therapy by 2010. This paper aims to describe and review access to second-line therapy in low- and middle-income countries at present and examine future possibilities. The majority of HIV-infected patients failing first-line combination antiretroviral therapy is identified by way of routine monitoring of clinical and immunological status as a surrogate for virological monitoring. Evidence suggests that immunological and clinical monitoring lack both sensitivity and specificity for virological failure. Consequently, at treatment failure, patients have often selected a degree of resistance within the nucleoside/nucleotide reverse transcriptase inhibitor class that questions the efficacy of using nucleoside/nucleotide reverse transcriptase inhibitors in a second-line regimen. There is a paucity of good-quality evidence on which to base guidelines and policy. Optimally, a second-line regimen would be simple, potent, tolerable and lend itself to provision according to the successful 'public health' approach. Provision of second-line therapy to HIV-infected individuals failing first-line therapy is a major challenge to the ongoing success of access to HIV care programmes in low- and middle-income countries. The optimal second-line combination antiretroviral therapies are unknown. Research trials to help define best practice are in advanced stages of development and implementation.

Naringin Reverses Hepatocyte Apoptosis and Oxidative Stress Associated with HIV-1 Nucleotide Reverse Transcriptase Inhibitors-Induced Metabolic Complications

PubMed Central

Adebiyi, Oluwafeyisetan O.; Adebiyi, Olubunmi A.; Owira, Peter M. O.

2015-01-01

Nucleoside Reverse Transcriptase Inhibitors (NRTIs) have not only improved therapeutic outcomes in the treatment of HIV infection but have also led to an increase in associated metabolic complications of NRTIs. Naringin’s effects in mitigating NRTI-induced complications were investigated in this study. Wistar rats, randomly allotted into seven groups (n = 7) were orally treated daily for 56 days with 100 mg/kg zidovudine (AZT) (groups I, II III), 50 mg/kg stavudine (d4T) (groups IV, V, VI) and 3 mL/kg of distilled water (group VII). Additionally, rats in groups II and V were similarly treated with 50 mg/kg naringin, while groups III and VI were treated with 45 mg/kg vitamin E. AZT or d4T treatment significantly reduced body weight and plasma high density lipoprotein concentrations but increased liver weights, plasma triglycerides and total cholesterol compared to controls, respectively. Furthermore, AZT or d4T treatment significantly increased oxidative stress, adiposity index and expression of Bax protein, but reduced Bcl-2 protein expression compared to controls, respectively. However, either naringin or vitamin E significantly mitigated AZT- or d4T-induced weight loss, dyslipidemia, oxidative stress and hepatocyte apoptosis compared to AZT- or d4T-only treated rats. Our results suggest that naringin reverses metabolic complications associated with NRTIs by ameliorating oxidative stress and apoptosis. This implies that naringin supplements could mitigate lipodystrophy and dyslipidemia associated with NRTI therapy. PMID:26690471

The discovery of novel diarylpyri(mi)dine derivatives with high level activity against a wide variety of HIV-1 strains as well as against HIV-2.

PubMed

Lu, Xueyi; Yang, Jiapei; Kang, Dongwei; Gao, Ping; Daelemans, Dirk; De Clercq, Erik; Pannecouque, Christophe; Zhan, Peng; Liu, Xinyong

2018-05-01

By means of structure-based molecular hybridization strategy, a series of novel diarylpyri(mi)dine derivatives targeting the entrance channel of HIV-1 reverse transcriptase (RT) were designed, synthesized and evaluated as potent non-nucleoside reverse transcriptase inhibitors (NNRTIs). Encouragingly, all the tested compounds showed good activities against wild-type (WT) HIV-1 (IIIB) with EC 50 in the range of 1.36 nM-29 nM, which is much better than those of nevirapine (NVP, EC 50  = 125.42 nM) and azidothymidine (AZT, EC 50  = 11.36 nM). Remarkably, these compounds also displayed effective activity against the most of the single and double-mutated HIV-1 strains with low EC 50 values, which is comparable to the control drugs. Besides, these compounds were also exhibited favorable enzymatic inhibitory activity. Moreover, preliminary structure-activity relationships (SARs) and molecular modeling study were investigated and discussed in detail. Unexpectedly, four diarylpyrimidines yielded moderate anti-HIV-2 activities. To our knowledge, this is rarely reported that diarylpyrimidine-based NNRTIs have potent activity against both HIV-1 and HIV-2 in cell culture. Copyright © 2018 Elsevier Ltd. All rights reserved.

Maraviroc, as a Switch Option, in HIV-1–infected Individuals With Stable, Well-controlled HIV Replication and R5-tropic Virus on Their First Nucleoside/Nucleotide Reverse Transcriptase Inhibitor Plus Ritonavir-boosted Protease Inhibitor Regimen: Week 48 Results of the Randomized, Multicenter MARCH Study

PubMed Central

Pett, Sarah Lilian; Amin, Janaki; Horban, Andrejz; Andrade-Villanueva, Jaime; Losso, Marcelo; Porteiro, Norma; Sierra Madero, Juan; Belloso, Waldo; Tu, Elise; Silk, David; Kelleher, Anthony; Harrigan, Richard; Clark, Andrew; Sugiura, Wataru; Wolff, Marcelo; Gill, John; Gatell, Jose; Fisher, Martin; Clarke, Amanda; Ruxrungtham, Kiat; Prazuck, Thierry; Kaiser, Rolf; Woolley, Ian; Arnaiz, Juan Alberto; Cooper, David; Rockstroh, Jürgen K.; Mallon, Patrick; Emery, Sean; Kelleher, Anthony; Merlin, Kate; Yeung, Julie; Fsadni, Bertha; Marks, Kat; Suzuki, Kazuo; Rismanto, Nick; Salomon, Horacio; Rubio, Andrea E.; Chibo, Doris; Birch, Chris; Harrigan, Richard; Swenson, Luke; Chan, Dennison; Berg, Thomas; Obermeier, Martin; Kaiser, Rolf; Schuelter, Eugen; Sierra Aragon, Saleta; Luebke, Nadine; Coughlan, Suzie; Dean, Jonathan; Sugiura, Wataru; Iwatani, Yasumasa; Reyes Teran, Gustavo; Avila, Santiago; Ruxrungtham, Kiat; Sirivichayakul, Sunee; Naphassanant, May; Ubolyam, Sasiwimol; Kaye, Steve; Land, Sally; Walker, Sarah; Haubrich, Richard; DeJesus, Edwin; Emery, Sean; Pett, Sarah L.; Tu, Elise; Silk, David; Berthon-Jones, Nisha; Amin, Janaki; Espinosa, Natalie; Courtney-Vega, Kymme; Absar, Noorul; Haskelberg, Hila; Robson, Rose; Donaldson, Anna; Losso, Marcelo; Belloso, Waldo; Guelman, Daniel; Gambardella, Luciana; Valdovinos, Mariana; Gatell, Jose; Arnaiz, Juan; Beleta, Helena; Ramos, Nuria; Targa, Marta; Rockstroh, Jurgen; Späth, Brigitta; Boesecke, Christoph; Engelhardt, Angelika; Fisher, Martin; Perry, Nicky; Clarke, Amanda; Gill, John; Beckthold, Brenda; Clark, Andrew; Drummond, Fraser; Lefevre, Eric; Corr, Sharon; Grant, Carol; Lupo, Sergio; Peroni, Luciana; Italiano, Hospital; Sanchez, Marisa; De Paz Sierra, Mariana; Mejia, Ramos; Losso, Marcelo; Viloria, Guillermo; Parlante, Angel; Bissio, Emiliano; Luchetti, Pablo; Warley, Eduardo; Vieni, Ines; Porteiro, Norma; Vilas, Cecilia; Zarate, Abel; Mayer, Gabriela; Elliot, Julian; Hagenauer, Michelle; Kelley, Mark; Rowling, Diane; Gibson, Abby; Latch, Ngaire; Tabrett, Chantal; Warzywoda, Elizabeth; Cooper, David; Pett, Sarah; MacRae, Karen; Sinclair, Brett; Sinn, Kate; Bloch, Mark; Franic, Teo; Vincent, Trina; Stewart, Natasha; Jayewardene, Avindra; Dwyer, Dominic; Kok, Jennifer; Assam, Delene; Taylor, Janette; King, Patricia; Orth, David; Youds, David; Sowden, David; Johnston, Colleen; Murray, Suzanne; Hehir, Jennifer; Wadham, Samantha; Donohue, William; Thompson, Jill; Garsia, Roger; Turnham, Geoffrey; Madden, Tracey; Woolley, Ian; Gillies, Ainsley; Bryant, Mellissa; Gill, John; Beckthold, Brenda; Walmsley, Sharon; Chan, Warmond; LeBlanc, Roger; Lanteigne, Francois; Mouawad, Rima; Rahal, Ines; Guber, Sergio; Ozturk, Sefika; Smith, Graham; Halpenny, Roberta; Reko, Tatjana; Robinette Hills, Jennifer; Wolff, Marcelo; Prazuck, Thierry; Laurent Hocqueloux, Francois; Wolfgang, Johann; Stephan, Christoph; Ebeling, Franziska; Rockstroh, Juergen; Boesecke, Christoph; Spath, Brigitta; Engelhardt, Angelika; Ole Jensen, Bjorn-Erik; Feind, Cecilie; Meyer-Olson, Dirk; Stoll, Matthias; Hoeper, Kirsten; Beider, Renata; Faetkenheur, Gerd; Thomas Baumgarten, Ellen; Baumgarten, Axel; Ingiliz, Patrick; Wienbreyer, Andreas; Behrendt, Daniela; Nienkarken, Tanja; Stein, Jessen; Jessen, Heiko; Zedlack, Carmen; Mallon, Paddy; Simelane, Sibongile; Assmann, Jennifer; Ghavami-Kia, Bijan; Sugiura, Wataru; Imahashi, Mayumi; Tanabe, Kazue; Yokomaku, Yoshiyuki; Imamura, Junji; Andrade-Villanueva, Jaime; Montes de Oca, Melva; Gonzalez, Lucero; Ponce, David; Mendoza, Andrea; Sierra-Madero, Juan; Sanchez Hernandez, Jesus Eduardo; Jaime Ruiz Ballesteros, Eduardo; del Moral Ponce, Sergio; Mosqueda, Luis; Lopez, Monica; Horban, Andrzej; Ignatowska, Anna; Bakowska, Elzbieta; Pulik, Piotr; Sanz-Moreno, Jose; Paredes, Roger; Puig, Jordi; Domingo, Pere; Gutierrez, Mar; Gatell, Jose; González-Cordón, Ana; Callau, Pili; Lopez Aldeguer, Jose; Cuellar Tovar, Sandra; Leal Noval, Manuel; Rivas, Inmaculada; Delgado-Fernandez, Marcial; Ramon Arribas, Jose; Miguel Castro, Juan; Ruxrungtham, Kiat; Avihingsanon, Anchalee; Maek-a-nantawat, Wirach; Intasan, Jintana; Charoenporn, Walairat; Cuprasitrut, Thidarat; Jaisomkom, Pachuen; Pruksakaew, Kanchana; Winston, Alan; Mullaney, Scott; Fisher, Martin; Clarke, Amanda; Barbour, Lisa; Perry, Nicky; Richardson, Celia; Fox, Julie; Murray, Tammy; Leen, Clifford; Morris, Shelia; Satyajit, Das; Sandhu, Rumun; Tucker, James

2016-01-01

Abstract Background. Alternative combination antiretroviral therapies in virologically suppressed human immunodeficiency virus (HIV)–infected patients experiencing side effects and/or at ongoing risk of important comorbidities from current therapy are needed. Maraviroc (MVC), a chemokine receptor 5 antagonist, is a potential alternative component of therapy in those with R5-tropic virus. Methods. The Maraviroc Switch Study is a randomized, multicenter, 96-week, open-label switch study in HIV type 1–infected adults with R5-tropic virus, virologically suppressed on a ritonavir-boosted protease inhibitor (PI/r) plus double nucleoside/nucleotide reverse transcriptase inhibitor (2 N(t)RTI) backbone. Participants were randomized 1:2:2 to current combination antiretroviral therapy (control), or replacing the protease inhibitor (MVC + 2 N(t)RTI arm) or the nucleoside reverse transcriptase inhibitor backbone (MVC + PI/r arm) with twice-daily MVC. The primary endpoint was the difference (switch minus control) in proportion with plasma viral load (VL) <200 copies/mL at 48 weeks. The switch arms were judged noninferior if the lower limit of the 95% confidence interval (CI) for the difference in the primary endpoint was < −12% in the intention-to-treat (ITT) population. Results. The ITT population comprised 395 participants (control, n = 82; MVC + 2 N(t)RTI, n = 156; MVC + PI/r, n = 157). Baseline characteristics were well matched. At week 48, noninferior rates of virological suppression were observed in those switching away from a PI/r (93.6% [95% CI, −9.0% to 2.2%] and 91.7% [95% CI, −9.6% to 3.8%] with VL <200 and <50 copies/mL, respectively) compared to the control arm (97.6% and 95.1% with VL <200 and <50 copies/mL, respectively). In contrast, MVC + PI/r did not meet noninferiority bounds and was significantly inferior (84.1% [95% CI, −19.8% to −5.8%] and 77.7% [95% CI, −24.9% to −8.4%] with VL <200 and <50 copies/mL, respectively) to the control arm in the ITT analysis. Conclusions. These data support MVC as a switch option for ritonavir-boosted PIs when partnered with a 2-N(t)RTI backbone, but not as part of N(t)RTI-sparing regimens comprising MVC with PI/r. Clinical Trials Registration. NCT01384682. PMID:27048747

An Intravaginal Ring That Releases the NNRTI MIV-150 Reduces SHIV Transmission in Macaques

PubMed Central

Rodriguez, Aixa; Kizima, Larisa; Menon, Radhika; Goldman, Daniel; Kenney, Jessica; Aravantinou, Meropi; Seidor, Samantha; Gettie, Agegnehu; Blanchard, James; Piatak, Michael; Lifson, Jeffrey D.; Fernández-Romero, José A.; Robbiani, Melissa; Zydowsky, Thomas M.

2015-01-01

Microbicides may prevent HIV and sexually transmitted infections (STIs) in women; however, determining the optimal means of delivery of active pharmaceutical ingredients remains a major challenge. We previously demonstrated that a vaginal gel containing the non-nucleoside reverse transcriptase inhibitor MIV-150 partially protected macaques from SHIV-RT (simian/HIV reverse transcriptase) infection, and the addition of zinc acetate rendered the gel significantly protective. We test the activity of MIV-150 without the addition of zinc acetate when delivered from either ethylene vinyl acetate (EVA) or silicone intravaginal rings (IVRs). MIV-150 was successfully delivered, because it was detected in vaginal fluids and tissues by radioimmunoassay in pharmacokinetic studies. Moreover, EVA IVRs significantly protected macaques from SHIV-RT infection. Our results demonstrate that MIV-150–containing IVRs have the potential to prevent HIV infection and highlight the possible use of IVRs for delivering drugs that block HIV and other STIs. PMID:22956201

An intravaginal ring that releases the NNRTI MIV-150 reduces SHIV transmission in macaques.

PubMed

Singer, Rachel; Mawson, Paul; Derby, Nina; Rodriguez, Aixa; Kizima, Larisa; Menon, Radhika; Goldman, Daniel; Kenney, Jessica; Aravantinou, Meropi; Seidor, Samantha; Gettie, Agegnehu; Blanchard, James; Piatak, Michael; Lifson, Jeffrey D; Fernández-Romero, José A; Robbiani, Melissa; Zydowsky, Thomas M

2012-09-05

Microbicides may prevent HIV and sexually transmitted infections (STIs) in women; however, determining the optimal means of delivery of active pharmaceutical ingredients remains a major challenge. We previously demonstrated that a vaginal gel containing the non-nucleoside reverse transcriptase inhibitor MIV-150 partially protected macaques from SHIV-RT (simian/HIV reverse transcriptase) infection, and the addition of zinc acetate rendered the gel significantly protective. We test the activity of MIV-150 without the addition of zinc acetate when delivered from either ethylene vinyl acetate (EVA) or silicone intravaginal rings (IVRs). MIV-150 was successfully delivered, because it was detected in vaginal fluids and tissues by radioimmunoassay in pharmacokinetic studies. Moreover, EVA IVRs significantly protected macaques from SHIV-RT infection. Our results demonstrate that MIV-150-containing IVRs have the potential to prevent HIV infection and highlight the possible use of IVRs for delivering drugs that block HIV and other STIs.

Occurrence of etravirine/rilpivirine-specific resistance mutations selected by efavirenz and nevirapine in Kenyan patients with non-B HIV-1 subtypes failing antiretroviral therapy.

PubMed

Crawford, Keith W; Njeru, Dorothy; Maswai, Jonah; Omondi, Milton; Apollo, Duncan; Kimetto, Jane; Gitonga, Lawrence; Munyao, James; Langat, Raphael; Aoko, Appolonia; Tarus, Jemutai; Khamadi, Samoel; Hamm, Tiffany E

2014-01-28

Resistance to efavirenz and nevirapine has not been associated with mutations at position 138 of reverse transcriptase. In an evaluation of virologic suppression rates in PEPFAR (President's Emergency Plan For AIDS Relief) clinics in Kenya among patients on first-line therapy (RV288), 63% (617/975) of randomly selected patients on antiretroviral therapy were suppressed (HIV RNA<400 copies/ml). Among those with non-nucleoside reverse transcriptase inhibitor resistance (n = 101), 14 (13.8%) had substitutions at 138 (A, G, K or Q), mutations selected only by etravirine and rilpivirine in subtype B viruses. All 14 patients received efavirenz or nevirapine, not etravirine or rilpivirine, and were predominantly subtype A1. This may be the first report of efavirenz and nevirapine selecting these mutations in these subtypes.

HIV type 1 diversity in the Seychelles.

PubMed

Razafindratsimandresy, Richter; Hollanda, Justina; Soares, Jean-Louis; Rousset, Dominique; Chetty, Agnes P; Reynes, Jean-Marc

2007-06-01

Subtype determination and drug resistance-associated mutations (DRM) detection were performed on 40 HIV-1 Western blot-positive sera detected, obtained from consecutive patients resident in the Seychelles and consulting the Communicable Disease Control Unit, HIV reference center, in Victoria Hospital (Mahe) from October 2005 to June 2006. Amplification and sequencing of at least two of the partial reverse transcriptase, protease, and partial envelope genes were successful for all strains. All three genes sequences were obtained for 39 strains. A high degree of subtype or circulating recombinant forms (CRF) was observed for these 39 strains: A-A1 (17 cases), C (10 cases), B (8 cases), CRF02_AG (2 cases), D (1 case) and CRF01_AE (1 case). According to the ANRS 2006 DRM list and algorithm, none of the 40 isolates was found to be resistant to any protease or reverse transcriptase inhibitors.

1-Benzyl-2-(1H-indol-3-yl)-5-oxo­pyrrolidine-2-carbonitrile

PubMed Central

Tamazyan, Rafael; Armen, Ayvazyan; Ashot, Martirosyan; Sahak, Gasparyan; Schinazi, Raymond

2008-01-01

In the title compound, C20H17N3O, a potential anti-human immunodeficiency virus type 1 (HIV-1) non-nucleoside reverse-transcriptase inhibitor, the pyrrolidine ring has an envelope conformation. In the crystal structure, adjacent mol­ecules are connected into infinite chains via an N—H⋯O hydrogen bond. PMID:21201400

Creation of a Long-Acting Nanoformulated 2′,3′-Dideoxy-3′-Thiacytidine

PubMed Central

Guo, Dongwei; Zhou, Tian; Araínga, Mariluz; Palandri, Diana; Gautam, Nagsen; Bronich, Tatiana; Alnouti, Yazen; McMillan, JoEllyn; Edagwa, Benson

2017-01-01

Background: Antiretroviral drug discovery and formulation design will facilitate viral clearance in infectious reservoirs. Although progress has been realized for selected hydrophobic integrase and nonnucleoside reverse transcriptase inhibitors, limited success has been seen to date with hydrophilic nucleosides. To overcome these limitations, hydrophobic long-acting drug nanoparticles were created for the commonly used nucleoside reverse transcriptase inhibitor, lamivudine (2′,3′-dideoxy-3′-thiacytidine, 3TC). Methods: A 2-step synthesis created a slow-release long-acting hydrophobic 3TC. Conjugation of 3TC to a fatty acid created a myristoylated prodrug which was encased into a folate-decorated poloxamer 407. Both in vitro antiretroviral efficacy in human monocyte-derived macrophages and pharmacokinetic profiles in mice were evaluated for the decorated nanoformulated drug. Results: A stable drug formulation was produced by poloxamer encasement that improved monocyte–macrophage uptake, antiretroviral activities, and drug pharmacokinetic profiles over native drug formulations. Conclusions: Sustained release of long-acting antiretroviral therapy is a new therapeutic frontier for HIV/AIDS. 3TC depot formation in monocyte-derived macrophages can be facilitated through stable subcellular internalization and slow drug release. PMID:27559685

The emerging profile of cross-resistance among the nonnucleoside HIV-1 reverse transcriptase inhibitors.

PubMed

Sluis-Cremer, Nicolas

2014-07-31

Nonnucleoside reverse transcriptase inhibitors (NNRTIs) are widely used to treat HIV-1-infected individuals; indeed most first-line antiretroviral therapies typically include one NNRTI in combination with two nucleoside analogs. In 2008, the next-generation NNRTI etravirine was approved for the treatment of HIV-infected antiretroviral therapy-experienced individuals, including those with prior NNRTI exposure. NNRTIs are also increasingly being included in strategies to prevent HIV-1 infection. For example: (1) nevirapine is used to prevent mother-to-child transmission; (2) the ASPIRE (MTN 020) study will test whether a vaginal ring containing dapivirine can prevent HIV-1 infection in women; (3) a microbicide gel formulation containing the urea-PETT derivative MIV-150 is in a phase I study to evaluate safety, pharmacokinetics, pharmacodynamics and acceptability; and (4) a long acting rilpivirine formulation is under-development for pre-exposure prophylaxis. Given their widespread use, particularly in resource-limited settings, as well as their low genetic barriers to resistance, there are concerns about overlapping resistance between the different NNRTIs. Consequently, a better understanding of the resistance and cross-resistance profiles among the NNRTI class is important for predicting response to treatment, and surveillance of transmitted drug-resistance.

Exploiting Drug-Resistant Enzymes as Tools to Identify Thienopyrimidinone Inhibitors of Human Immunodeficiency Virus Reverse Transcriptase-Associated Ribonuclease H

PubMed Central

Masaoka, Takashi; Chung, Suhman; Caboni, Pierluigi; Rausch, Jason W.; Wilson, Jennifer A.; Taskent-Sezgin, Humeyra; Beutler, John A.; Tocco, Graziella; Le Grice, Stuart F. J.

2013-01-01

The thienopyrimidinone 5,6-dimethyl-2-(4-nitrophenyl)thieno[2,3-d]pyrimidin-4(3H)-one (DNTP) occupies the interface between the p66 ribonuclease H (RNase H) domain and p51 thumb of human immunodeficiency virus reverse transcriptase (HIV RT), thereby inducing a conformational change incompatible with catalysis. Here, we combined biochemical characterization of 39 DNTP derivatives with antiviral testing of selected compounds. In addition to wild-type HIV-1 RT, derivatives were evaluated with rationally-designed, p66/p51 heterodimers exhibiting high-level DNTP sensitivity or resistance. This strategy identified 3′,4′-dihydroxyphenyl (catechol)-substituted thienopyrimidinones with sub-micromolar in vitro activity against both wild type HIV-1 RT and drug-resistant variants. Thermal shift analysis indicates that, in contrast to active site RNase H inhibitors, these thienopyrimidinones destabilize the enzyme, in some instances reducing the Tm by 5°C. Importantly, catechol-containing thienopyrimidinones also inhibit HIV-1 replication in cells. Our data strengthens the case for allosteric inhibition of HIV RNase H activity, providing a platform for designing improved antagonists for use in combination antiviral therapy. PMID:23631411

Discovery of 3-{5-[(6-Amino-1H-pyrazolo[3,4-b]pyridine-3-yl)methoxy]-2-chlorophenoxy}-5-chlorobenzonitrile (MK-4965): A Potent, Orally Bioavailable HIV-1 Non-Nucleoside Reverse Transcriptase Inhibitor with Improved Potency against Key Mutant Viruses

DOE Office of Scientific and Technical Information (OSTI.GOV)

Tucker, Thomas J.; Sisko, John T.; Tynebor, Robert M.

2009-07-10

Non-nucleoside reverse transcriptase inhibitors (NNRTIs) have been shown to be a key component of highly active antiretroviral therapy (HAART). The use of NNRTIs has become part of standard combination antiviral therapies producing clinical outcomes with efficacy comparable to other antiviral regimens. There is, however, a critical issue with the emergence of clinical resistance, and a need has arisen for novel NNRTIs with a broad spectrum of activity against key HIV-1 RT mutations. Using a combination of traditional medicinal chemistry/SAR analyses, crystallography, and molecular modeling, we have designed and synthesized a series of novel, highly potent NNRTIs that possess broad spectrummore » antiviral activity and good pharmacokinetic profiles. Further refinement of key compounds in this series to optimize physical properties and pharmacokinetics has resulted in the identification of 8e (MK-4965), which has high levels of potency against wild-type and key mutant viruses, excellent oral bioavailability and overall pharmacokinetics, and a clean ancillary profile.« less

Reverse Transcriptase Inhibitors as Potential Colorectal Microbicides▿ †

PubMed Central

Herrera, Carolina; Cranage, Martin; McGowan, Ian; Anton, Peter; Shattock, Robin J.

2009-01-01

We investigated whether reverse transcriptase (RT) inhibitors (RTI) can be combined to inhibit human immunodeficiency virus type 1 (HIV-1) infection of colorectal tissue ex vivo as part of a strategy to develop an effective rectal microbicide. The nucleotide RTI (NRTI) PMPA (tenofovir) and two nonnucleoside RTI (NNRTI), UC-781 and TMC120 (dapivirine), were evaluated. Each compound inhibited the replication of the HIV isolates tested in TZM-bl cells, peripheral blood mononuclear cells, and colorectal explants. Dual combinations of the three compounds, either NRTI-NNRTI or NNRTI-NNRTI combinations, were more active than any of the individual compounds in both cellular and tissue models. Combinations were key to inhibiting infection by NRTI- and NNRTI-resistant isolates in all models tested. Moreover, we found that the replication capacities of HIV-1 isolates in colorectal explants were affected by single point mutations in RT that confer resistance to RTI. These data demonstrate that colorectal explants can be used to screen compounds for potential efficacy as part of a combination microbicide and to determine the mucosal fitness of RTI-resistant isolates. These findings may have important implications for the rational design of effective rectal microbicides. PMID:19258271

Reverse transcriptase inhibitors as potential colorectal microbicides.

PubMed

Herrera, Carolina; Cranage, Martin; McGowan, Ian; Anton, Peter; Shattock, Robin J

2009-05-01

We investigated whether reverse transcriptase (RT) inhibitors (RTI) can be combined to inhibit human immunodeficiency virus type 1 (HIV-1) infection of colorectal tissue ex vivo as part of a strategy to develop an effective rectal microbicide. The nucleotide RTI (NRTI) PMPA (tenofovir) and two nonnucleoside RTI (NNRTI), UC-781 and TMC120 (dapivirine), were evaluated. Each compound inhibited the replication of the HIV isolates tested in TZM-bl cells, peripheral blood mononuclear cells, and colorectal explants. Dual combinations of the three compounds, either NRTI-NNRTI or NNRTI-NNRTI combinations, were more active than any of the individual compounds in both cellular and tissue models. Combinations were key to inhibiting infection by NRTI- and NNRTI-resistant isolates in all models tested. Moreover, we found that the replication capacities of HIV-1 isolates in colorectal explants were affected by single point mutations in RT that confer resistance to RTI. These data demonstrate that colorectal explants can be used to screen compounds for potential efficacy as part of a combination microbicide and to determine the mucosal fitness of RTI-resistant isolates. These findings may have important implications for the rational design of effective rectal microbicides.

Discovery of dapivirine, a nonnucleoside HIV-1 reverse transcriptase inhibitor, as a broad-spectrum antiviral against both influenza A and B viruses.

PubMed

Hu, Yanmei; Zhang, Jiantao; Musharrafieh, Rami Ghassan; Ma, Chunlong; Hau, Raymond; Wang, Jun

2017-09-01

The emergence of multidrug-resistant influenza viruses poses a persistent threat to public health. The current prophylaxis and therapeutic interventions for influenza virus infection have limited efficacy due to the continuous antigenic drift and antigenic shift of influenza viruses. As part of our ongoing effort to develop the next generation of influenza antivirals with broad-spectrum antiviral activity and a high genetic barrier to drug resistance, in this study we report the discovery of dapivirine, an FDA-approved HIV nonnucleoside reverse transcriptase inhibitor, as a broad-spectrum antiviral against multiple strains of influenza A and B viruses with low micromolar efficacy. Mechanistic studies revealed that dapivirine inhibits the nuclear entry of viral ribonucleoproteins at the early stage of viral replication. As a result, viral RNA and protein synthesis were inhibited. Furthermore, dapivirine has a high in vitro genetic barrier to drug resistance, and its antiviral activity is synergistic with oseltamivir carboxylate. In summary, the in vitro antiviral results of dapivirine suggest it is a promising candidate for the development of the next generation of dual influenza and HIV antivirals. Copyright © 2017 Elsevier B.V. All rights reserved.

Diaryltriazine non-nucleoside reverse transcriptase inhibitors are potent candidates for pre-exposure prophylaxis in the prevention of sexual HIV transmission.

PubMed

Ariën, Kevin K; Venkatraj, Muthusamy; Michiels, Johan; Joossens, Jurgen; Vereecken, Katleen; Van der Veken, Pieter; Abdellati, Saïd; Cuylaerts, Vicky; Crucitti, Tania; Heyndrickx, Leo; Heeres, Jan; Augustyns, Koen; Lewi, Paul J; Vanham, Guido

2013-09-01

Pre-exposure prophylaxis and topical microbicides are important strategies in the prevention of sexual HIV transmission, especially since partial protection has been shown in proof-of-concept studies. In search of new candidate drugs with an improved toxicity profile and with activity against common non-nucleoside reverse transcriptase inhibitor (NNRTI)-resistant HIV, we have synthesized and investigated a library of 60 new diaryltriazine analogues. From this library, 15 compounds were evaluated in depth using a broad armamentarium of in vitro assays that are part of a preclinical testing algorithm for microbicide development. Antiviral activity was assessed in a cell line, and in primary human cells, against both subtype B and subtype C HIV-1 and against viruses resistant to therapeutic NNRTIs and the candidate NNRTI microbicide dapivirine. Toxicity towards primary blood-derived cells, cell lines originating from the female reproductive tract and female genital microflora was also studied. We identified several compounds with highly potent antiviral activity and toxicity profiles that are superior to that of dapivirine. In particular, compound UAMC01398 is an interesting new candidate that warrants further investigation because of its superior toxicity profile and potent activity against dapivirine-resistant viruses.

The Emerging Profile of Cross-Resistance among the Nonnucleoside HIV-1 Reverse Transcriptase Inhibitors

PubMed Central

Sluis-Cremer, Nicolas

2014-01-01

Nonnucleoside reverse transcriptase inhibitors (NNRTIs) are widely used to treat HIV-1-infected individuals; indeed most first-line antiretroviral therapies typically include one NNRTI in combination with two nucleoside analogs. In 2008, the next-generation NNRTI etravirine was approved for the treatment of HIV-infected antiretroviral therapy-experienced individuals, including those with prior NNRTI exposure. NNRTIs are also increasingly being included in strategies to prevent HIV-1 infection. For example: (1) nevirapine is used to prevent mother-to-child transmission; (2) the ASPIRE (MTN 020) study will test whether a vaginal ring containing dapivirine can prevent HIV-1 infection in women; (3) a microbicide gel formulation containing the urea-PETT derivative MIV-150 is in a phase I study to evaluate safety, pharmacokinetics, pharmacodynamics and acceptability; and (4) a long acting rilpivirine formulation is under-development for pre-exposure prophylaxis. Given their widespread use, particularly in resource-limited settings, as well as their low genetic barriers to resistance, there are concerns about overlapping resistance between the different NNRTIs. Consequently, a better understanding of the resistance and cross-resistance profiles among the NNRTI class is important for predicting response to treatment, and surveillance of transmitted drug-resistance. PMID:25089538

Emtricitabine: a once-daily nucleoside reverse transcriptase inhibitor.

PubMed

Modrzejewski, Krysten A; Herman, Ronald A

2004-06-01

To review the pharmacology, virology, pharmacokinetics, safety, and efficacy of the nucleoside reverse transcriptase inhibitor (NRTI) emtricitabine. English-language reports were accessed using MEDLINE (1966-June 2003) and the Iowa Drug Information Service database (1966-June 2003) using emtricitabine and Coviracil as key words. (Coviracil was the proposed trade name for the product prior to approval.) The Internet was also searched using the terms HIV/AIDS conferences, then emtricitabine within the conference proceedings. Abstracts, posters, and oral presentations from scientific conferences, both published and unpublished, were included. Preference was given to published controlled trials. Studies providing a description of the pharmacology, virology, effectiveness, safety, or pharmacokinetics of emtricitabine were used in this review. Emtricitabine is an NRTI used to treat HIV-1 infection. Once-daily administration can decrease pill burden and potentially increase adherence to multidrug HIV therapy. Further, emtricitabine has shown equivalent or improved outcomes compared with lamivudine and stavudine. Emtricitabine is a safe and effective option for HIV-1 infection in adults as part of a multidrug regimen. It may be a better alternative than lamivudine for once-daily therapy because of its extended intracellular half-life and better than lamivudine and stavudine because of a possibly decreased potential for drug resistance.

Design, synthesis and biological evaluations of N-Hydroxy thienopyrimidine-2,4-diones as inhibitors of HIV reverse transcriptase-associated RNase H.

PubMed

Kankanala, Jayakanth; Kirby, Karen A; Huber, Andrew D; Casey, Mary C; Wilson, Daniel J; Sarafianos, Stefan G; Wang, Zhengqiang

2017-12-01

Human immunodeficiency virus (HIV) reverse transcriptase (RT) associated ribonuclease H (RNase H) is the only HIV enzymatic function not targeted by current antiviral drugs. Although various chemotypes have been reported to inhibit HIV RNase H, few have shown significant antiviral activities. We report herein the design, synthesis and biological evaluation of a novel N-hydroxy thienopyrimidine-2,3-dione chemotype (11) which potently and selectively inhibited RNase H with considerable potency against HIV-1 in cell culture. Current structure-activity-relationship (SAR) identified analogue 11d as a nanomolar inhibitor of RNase H (IC 50  = 0.04 μM) with decent antiviral potency (EC 50  = 7.4 μM) and no cytotoxicity (CC 50  > 100 μM). In extended biochemical assays compound 11d did not inhibit RT polymerase (pol) while inhibiting integrase strand transfer (INST) with 53 fold lower potency (IC 50  = 2.1 μM) than RNase H inhibition. Crystallographic and molecular modeling studies confirmed the RNase H active site binding mode. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

Genetic characterization and antiretroviral resistance mutations among treatment-naive HIV-infected individuals in Jiaxing, China.

PubMed

Guo, Jinlei; Yan, Yong; Zhang, Jiafeng; Ji, Jimei; Ge, Zhijian; Ge, Rui; Zhang, Xiaofei; Wang, Henghui; Chen, Zhongwen; Luo, Jianyong

2017-03-14

The aim of this study was to characterize HIV-1 genotypes and antiretroviral resistance mutations among treatment-naive HIV-infected individuals in Jiaxing, China. The HIV-1 partial polymerase (pol) genes in 93 of the 99 plasma samples were successfully amplified and analyzed. Phylogenetic analysis revealed the existence of five HIV-1 genotypes, of which the most prevalent genotype was CRF01_AE (38.7%), followed by CRF07_BC (34.4%), CRF08_BC (16.1%), subtype B/B' (5.4%), and CRF55_01B (2.1%). Besides, three types of unique recombination forms (URFs) were also observed, including C/F2/A1, CRF01_AE/B, and CRF08_BC/CRF07_BC. Among 93 amplicons, 46.2% had drug resistance-associated mutations, including 23.7% for protease inhibitors (PIs) mutations, 1.1% for nucleoside reverse transcriptase inhibitors (NRTIs) mutations, and 20.4% for non-nucleoside reverse transcriptase inhibitors (NNRTIs) mutations. Six (6.5%) out of 93 treatment-naive subjects were identified to be resistant to one or more NNRTIs, while resistance to NRTIs or PIs was not observed. Our study showed the genetic diversity of HIV-1 strains circulating in Jiaxing and a relative high proportion of antiretroviral resistance mutations among treatment-naive patients, indicating a serious challenge for HIV prevention and treatment program.

Prevalence of Drug-Resistance Mutations and Non–Subtype B Strains Among HIV-Infected Infants From New York State

PubMed Central

Karchava, Marine; Pulver, Wendy; Smith, Lou; Philpott, Sean; Sullivan, Timothy J.; Wethers, Judith; Parker, Monica M.

2010-01-01

Summary Prevalence studies indicate that transmission of drug-resistant HIV has been rising in the adult population, but data from the perinatally infected pediatric population are limited. In this retrospective study, we sequenced the pol region of HIV from perinatally infected infants diagnosed in New York State in 2001–2002. Analyses of drug resistance, subtype diversity, and perinatal antiretroviral exposure were conducted, and the results were compared with those from a previous study of HIV-infected infants identified in 1998–1999. Eight of 42 infants (19.1%) had provirus carrying at least 1 drug-resistance mutation, an increase of 58% over the 1998–1999 results. Mutations conferring resistance to nucleoside reverse transcriptase inhibitors, nonnucleoside reverse transcriptase inhibitors, and protease inhibitors were detected in 7.1%, 11.9%, and 2.4% of specimens, respectively. Consistent with previous results, perinatal antiretroviral exposure was not associated with drug resistance (P = 0.70). Phylogenetic analysis indicated that 16.7% of infants were infected with a non–subtype B strain of HIV. It seems that drug-resistant and non–subtype B strains of HIV are becoming increasingly common in the perinatally infected population. Our results highlight the value of resistance testing for all HIV-infected infants upon diagnosis and the need to consider subtype diversity in diagnostic and treatment strategies. PMID:16868498

Low prevalence of primary HIV resistance in western Massachusetts.

PubMed

Iarikov, Dmitri E; Irizarry-Acosta, Melina; Martorell, Claudia; Hoffman, Robert P; Skiest, Daniel J

2010-01-01

Most studies of primary antiretroviral (ARV) resistance have been conducted in large metropolitan areas with reported rates of 8% to 25%. We collected data on 99 HIV-1-infected antiretroviral-naive patients from several sites in Springfield, MA, who underwent genotypic resistance assay between 2004 and 2008. Only major resistance mutations per International AIDS Society-USA (IAS-USA) drug resistance mutations list were considered. The prevalence of resistance was 5% (5 of 99). Three patients had one nonnucleoside reverse transcriptase inhibitor (NNRTI) mutation: 103N, 103N, and 190A, 1 patient had a protease inhibitor (PI) mutation: 90M; and 1 patient had 3-class resistance with NNRTI: 181C, 190A, PI: 90M, and nucleoside analogue reverse transcriptase inhibitor (NRTI): 41L, 210W. Mean time from HIV diagnosis to resistance testing was shorter in patients with resistance versus those without: 9 (range 0.3-42 months) versus 27 (range 0.1-418 months), P = .11. There was a trend to lower mean CD4 count in those with resistance, 170 versus 318 cells/mm(3), P = .06. No differences were noted in gender, age, HIV risk category, or HIV RNA level. The low prevalence of primary resistance may be explained by differences in demographic and risk factors or may reflect the time from infection to resistance testing. Our findings emphasize the importance of continued resistance surveillance.

Active Methamphetamine Use is Associated with Transmitted Drug Resis-tance to Non-Nucleoside Reverse Transcriptase Inhibitors in Individuals with HIV Infection of Unknown Duration

PubMed Central

Cachay, Edward R; Moini, Niousha; Kosakovsky Pond, Sergei L; Pesano, Rick; Lie, Yolanda S; Aiem, Heidi; Butler, David M; Letendre, Scott; Mathews, Wm. Christopher; Smith, Davey M

2007-01-01

Background: Frequent methamphetamine use among recently HIV infected individuals is associated with transmitted drug resistance (TDR) to non-nucleoside reverse transcriptase inhibitors (NNRTI); however, the reversion time of TDR to drug susceptible HIV may exceed 3 years. We assessed whether recreational substance use is associated with detectable TDR among individuals newly diagnosed with HIV infection of unknown duration. Design: Cross-sectional analysis. Methods: Subjects were enrolled at the University California, San Diego Early Intervention Program. Demographic, clinical and substance use data were collected using structured interviews. Genotypic resistance testing was performed using GeneSeq™, Monogram Biosciences. We analyzed the association between substance use and TDR using bivariate analyses and the corresponding transmission networks using phylogenetic models. Results: Between April 2004 and July 2006, 115 individuals with genotype data were enrolled. The prevalence of alcohol, marijuana and methamphetamine use were 98%, 71% and 64% respectively. Only active methamphetamine use in the 30 days prior to HIV diagnosis was independently associated with TDR to NNRTI (OR: 6.6; p=0.002). Conclusion: Despite not knowing the duration of their HIV infection, individuals reporting active methamphetamine use in the 30 days prior to HIV diagnosis are at an increased risk of having HIV strains that are resistant to NNRTI. PMID:18923691

Scaffold hopping: exploration of acetanilide-containing uracil analogues as potential NNRTIs.

PubMed

Babkov, Denis A; Valuev-Elliston, Vladimir T; Paramonova, Maria P; Ozerov, Alexander A; Ivanov, Alexander V; Chizhov, Alexander O; Khandazhinskaya, Anastasia L; Kochetkov, Sergey N; Balzarini, Jan; Daelemans, Dirk; Pannecouque, Christophe; Seley-Radtke, Katherine L; Novikov, Mikhail S

2015-03-01

In order to identify novel nonnucleoside inhibitors of HIV-1 reverse transcriptase two series of amide-containing uracil derivatives were designed as hybrids of two scaffolds of previously reported inhibitors. Subsequent biological evaluation confirmed acetamide uracil derivatives 15a-k as selective micromolar NNRTIs with a first generation-like resistance profile. Molecular modeling of the most active compounds 15c and 15i was employed to provide insight on their inhibitory properties and direct future design efforts. Copyright © 2015 Elsevier Ltd. All rights reserved.

Alteration of Substrate and Inhibitor Specificity of Feline Immunodeficiency Virus Protease

PubMed Central

Lin, Ying-Chuan; Beck, Zachary; Lee, Taekyu; Le, Van-Duc; Morris, Garrett M.; Olson, Arthur J.; Wong, Chi-Huey; Elder, John H.

2000-01-01

Feline immunodeficiency virus (FIV) protease is structurally very similar to human immunodeficiency virus (HIV) protease but exhibits distinct substrate and inhibitor specificities. We performed mutagenesis of subsite residues of FIV protease in order to define interactions that dictate this specificity. The I37V, N55M, M56I, V59I, and Q99V mutants yielded full activity. The I37V, N55M, V59I, and Q99V mutants showed a significant increase in activity against the HIV-1 reverse transcriptase/integrase and P2/nucleocapsid junction peptides compared with wild-type (wt) FIV protease. The I37V, V59I, and Q99V mutants also showed an increase in activity against two rapidly cleaved peptides selected by cleavage of a phage display library with HIV-1 protease. Mutations at Q54K, I98P, and L101I dramatically reduced activity. Mutants containing a I35D or I57G substitution showed no activity against either FIV or HIV substrates. FIV proteases all failed to cut HIV-1 matrix/capsid, P1/P6, P6/protease, and protease/reverse transcriptase junctions, indicating that none of the substitutions were sufficient to change the specificity completely. The I37V, N55M, M56I, V59I, and Q99V mutants, compared with wt FIV protease, all showed inhibitor specificity more similar to that of HIV-1 protease. The data also suggest that FIV protease prefers a hydrophobic P2/P2′ residue like Val over Asn or Glu, which are utilized by HIV-1 protease, and that S2/S2′ might play a critical role in distinguishing FIV and HIV-1 protease by specificity. The findings extend our observations regarding the interactions involved in substrate binding and aid in the development of broad-based inhibitors. PMID:10775609

Viro-immunological response of drug-naive HIV-1-infected patients starting a first-line regimen with viraemia >500,000 copies/ml in clinical practice.

PubMed

Santoro, Maria Mercedes; Di Carlo, Domenico; Armenia, Daniele; Zaccarelli, Mauro; Pinnetti, Carmela; Colafigli, Manuela; Prati, Francesca; Boschi, Andrea; Antoni, Anna Maria Degli; Lagi, Filippo; Sighinolfi, Laura; Gervasoni, Cristina; Andreoni, Massimo; Antinori, Andrea; Mussini, Cristina; Perno, Carlo Federico; Borghi, Vanni; Sterrantino, Gaetana

2017-09-22

Virological success (VS) and immunological reconstitution (IR) of antiretroviral-naive HIV-1-infected patients with pre-therapy viral load (VL) >500,000 copies/ml was assessed after 12 months of treatment according to initial drug-class regimens. An observational multicentre retrospective study was performed. VS was defined as the first VL <50 copies/ml from treatment start. IR was defined as an increase of at least 150 CD4 + T-lymphocytes from treatment start. Survival analysis was used to estimate the probability and predictors of VS and IR by 12 months of therapy. 428 HIV-1-infected patients were analysed. Patients were grouped according to the different first-line drug-classes used: a non-nucleoside reverse transcriptase inhibitor (NNRTI) plus two nucleoside reverse transcriptase inhibitors (NRTIs; NNRTI-group; n=105 [24.5%]); a protease inhibitor (PI) plus two NRTIs (PI-group; n=260 [60.8%]); a four-drug regimen containing a PI-regimen plus an integrase inhibitor (PI+INI-group; n=63 [14.7%]). Patients in the PI-group showed the lowest probability of VS (PI-group: 72.4%; NNRTI-group: 75.5%; PI+INI-group: 81.0%; P<0.0001). By Cox regression, patients in PI+INI and NNRTI-groups showed a higher adjusted hazard ratio (95% CI) of VS compared to those in the PI-group (PI+INI-group: 1.48 [1.08, 2.03]; P=0.014; NNRTI-group: 1.37 [1.06-1.78]; P=0.015). The probability of IR was 76.2%, and was similar among groups. Patients with AIDS showed a lower adjusted hazard ratio (95% CI) of IR compared to non-AIDS presenters (0.70 [0.54, 0.90]; P=0.005). In this multicentre retrospective study, patients with viraemia >500,000 copies/ml who start a first-line regimen containing PI+INI or NNRTI yield a better VS compared to those receiving a PI-based regimen.

One-, two-, and three-class resistance among HIV-infected patients on antiretroviral therapy in private care clinics: Mumbai, India.

PubMed

Gupta, Amita; Saple, Dattaray G; Nadkarni, Girish; Shah, Bijal; Vaidya, Satish; Hingankar, Nitin; Chaturbhuj, Devidas; Deshmukh, Praveen; Walshe, Louise; Hudelson, Sarah E; James, Maria; Paranjape, Ramesh S; Eshleman, Susan H; Tripathy, Srikanth

2010-01-01

HIV-infected patients receiving antiretroviral (ARV) therapy (ART) in India are not all adequately virally suppressed. We analyzed ARV drug resistance in adults receiving ART in three private clinics in Mumbai, India. HIV viral load was measured in 200 patients with the Roche AMPLICOR HIV-1 Monitor Test, v1.5. HIV genotyping was performed with the ViroSeq HIV-1 Genotyping System for 61 participants who had HIV-1 RNA >1000 copies/ml. Genotyping results were obtained for 51 samples. The participants with resistance results were on ART for a median of 24 months and were on their current regimen for a median of 12 months (median CD4 cell count: 217 cells/mm(3); median HIV viral load: 28,200 copies/ml). ARV regimens included nonnucleoside reverse transcriptase inhibitor (NNRTI)-based regimens (n = 27), dual nucleoside reverse transcriptase inhibitors (NRTIs, n = 19), protease inhibitor (PI)-based regimens (n = 3), and other regimens (n = 2). Twenty-six participants (51.0%) were on their first ARV regimen and 24 (47%) reported >95% adherence. Forty-nine participants (96.1%) had resistance to at least one ARV drug; 47 (92.2%) had NRTI resistance, 32 (62.7%) had NNRTI resistance, and four (7.8%) had PI resistance. Thirty (58.8%) had two-class resistance and three (5.9%) had three-class resistance. Four (8%) had three or more resistance mutations associated with etravirine resistance and two (4%) had two mutations associated with reduced darunavir susceptibility. Almost all patients with HIV-1 RNA >1000 copies/ml had NRTI resistance and nearly two-thirds had NNRTI resistance; PI resistance was uncommon. Nearly 60% and 6% had two- and three-class resistance, respectively. This emphasizes the need for greater viral load and resistance monitoring, use of optimal ART combinations, and increased availability of second- and third-line agents for patients with ARV resistance.

Trends of drug-resistance-associated mutations in the reverse transcriptase gene of HIV type 1 isolates from North India.

PubMed

Azam, Mohd; Malik, Abida; Rizvi, Meher; Rai, Arvind

2014-04-01

A major cause of failure of antiretroviral therapy (ART) is the presence of drug-resistance-associated mutations in the polymerase gene of HIV-1. The paucity of data regarding potential drug resistance to reverse transcriptase inhibitors (RTIs) prompted us to carry out this study. This information will shed light on the extent of drug resistance already present in HIV strains and will give future directions in patient treatment and in drug design. Drug resistance genotyping of a partial reverse transcriptase gene was done in 103 HIV-1-infected patients, including the ART-naive and ART-experienced population. The drug resistance pattern was analyzed using the Stanford HIV-DR database, the IAS-USA mutation list and the REGA algorithm-v8.0. Subtyping was done using the REGA HIV-1 subtyping tool-v2.01. The majority of our sequences (96 %) were found to be subtype C, and four (3.8 %) were subtype A1. Significant prevalence of DR mutations (28 %) was observed in the RT gene. Major amino acid substitutions were seen at positions 41, 90, 98, 103, 106, 108, 138, 181, 184, 190, 215, and 219, which confer high/intermediate levels of resistance to most RTIs, independently or together. Our results show that there is an urgent need to tailor ART drug regimens to the individual to achieve optimum therapeutic outcome in North India.

Telomerase reverse transcriptase (TERT) - enhancer of zeste homolog 2 (EZH2) network regulates lipid metabolism and DNA damage responses in glioblastoma.

PubMed

Ahmad, Fahim; Patrick, Shruti; Sheikh, Touseef; Sharma, Vikas; Pathak, Pankaj; Malgulwar, Prit Benny; Kumar, Anupam; Joshi, Shanker Datt; Sarkar, Chitra; Sen, Ellora

2017-12-01

Elevated expression of enhancer of zeste homolog 2 (EZH2), a histone H3K27 methyltransferase, was observed in gliomas harboring telomerase reverse transcriptase (TERT) promoter mutations. Given the known involvement of TERT and EZH2 in glioma progression, the correlation between the two and subsequently its involvement in metabolic programming was investigated. Inhibition of human telomerase reverse transcriptase either pharmacologically or through genetic manipulation not only decreased EZH2 expression, but also (i) abrogated FASN levels, (ii) decreased de novo fatty acid accumulation, and (iii) increased ataxia-telangiectasia-mutated (ATM) phosphorylation levels. Conversely, diminished TERT and FASN levels upon siRNA-mediated EZH2 knockdown indicated a positive correlation between TERT and EZH2. Interestingly, ATM kinase inhibitor rescued TERT inhibition-mediated decrease in FASN and EZH2 levels. Importantly, TERT promoter mutant tumors exhibited greater microsatellite instability, heightened FASN levels and lipid accumulation. Coherent with in vitro findings, pharmacological inhibition of TERT by costunolide decreased lipid accumulation and elevated ATM expression in heterotypic xenograft glioma mouse model. By bringing TERT-EZH2 network at the forefront as driver of dysregulated metabolism, our findings highlight the non-canonical but distinct role of TERT in metabolic reprogramming and DNA damage responses in glioblastoma. © 2017 International Society for Neurochemistry.

HIV-1 drug resistance genotyping from antiretroviral therapy (ART) naïve and first-line treatment failures in Djiboutian patients

PubMed Central

2012-01-01

Abstract In this study we report the prevalence of antiretroviral drug resistant HIV-1 genotypes of virus isolated from Djiboutian patients who failed first-line antiretroviral therapy (ART) and from ART naïve patients. Patients and methods A total of 35 blood samples from 16 patients who showed first-line ART failure (>1000 viral genome copies/ml) and 19 ART-naïve patients were collected in Djibouti from October 2009 to December 2009. Both the protease (PR) and reverse transcriptase (RT) genes were amplified and sequenced using National Agency for AIDS Research (ANRS) protocols. The Stanford HIV database algorithm was used for interpretation of resistance data and genotyping. Results Among the 16 patients with first-line ART failure, nine (56.2%) showed reverse transcriptase inhibitor-resistant HIV-1 strains: two (12.5%) were resistant to nucleoside (NRTI), one (6.25%) to non-nucleoside (NNRTI) reverse transcriptase inhibitors, and six (37.5%) to both. Analysis of the DNA sequencing data indicated that the most common mutations conferring drug resistance were M184V (38%) for NRTI and K103N (25%) for NNRTI. Only NRTI primary mutations K101Q, K103N and the PI minor mutation L10V were found in ART naïve individuals. No protease inhibitor resistant strains were detected. In our study, we found no detectable resistance in ∼ 44% of all patients who experienced therapeutic failure which was explained by low compliance, co-infection with tuberculosis and malnutrition. Genotyping revealed that 65.7% of samples were infected with subtype C, 20% with CRF02_AG, 8.5% with B, 2.9% with CRF02_AG/C and 2.9% with K/C. Conclusion The results of this first study about drug resistance mutations in first-line ART failures show the importance of performing drug resistance mutation test which guides the choice of a second-line regimen. This will improve the management of HIV-infected Djiboutian patients. Virtual slides The virtual slide(s) for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/2051206212753973 PMID:23044036

HIV-1 drug resistance genotyping from antiretroviral therapy (ART) naïve and first-line treatment failures in Djiboutian patients.

PubMed

Elmi Abar, Aden; Jlizi, Asma; Darar, Houssein Youssouf; Kacem, Mohamed Ali Ben Hadj; Slim, Amine

2012-10-08

In this study we report the prevalence of antiretroviral drug resistant HIV-1 genotypes of virus isolated from Djiboutian patients who failed first-line antiretroviral therapy (ART) and from ART naïve patients. A total of 35 blood samples from 16 patients who showed first-line ART failure (>1000 viral genome copies/ml) and 19 ART-naïve patients were collected in Djibouti from October 2009 to December 2009. Both the protease (PR) and reverse transcriptase (RT) genes were amplified and sequenced using National Agency for AIDS Research (ANRS) protocols. The Stanford HIV database algorithm was used for interpretation of resistance data and genotyping. Among the 16 patients with first-line ART failure, nine (56.2%) showed reverse transcriptase inhibitor-resistant HIV-1 strains: two (12.5%) were resistant to nucleoside (NRTI), one (6.25%) to non-nucleoside (NNRTI) reverse transcriptase inhibitors, and six (37.5%) to both. Analysis of the DNA sequencing data indicated that the most common mutations conferring drug resistance were M184V (38%) for NRTI and K103N (25%) for NNRTI. Only NRTI primary mutations K101Q, K103N and the PI minor mutation L10V were found in ART naïve individuals. No protease inhibitor resistant strains were detected. In our study, we found no detectable resistance in ∼ 44% of all patients who experienced therapeutic failure which was explained by low compliance, co-infection with tuberculosis and malnutrition. Genotyping revealed that 65.7% of samples were infected with subtype C, 20% with CRF02_AG, 8.5% with B, 2.9% with CRF02_AG/C and 2.9% with K/C. The results of this first study about drug resistance mutations in first-line ART failures show the importance of performing drug resistance mutation test which guides the choice of a second-line regimen. This will improve the management of HIV-infected Djiboutian patients. The virtual slide(s) for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/2051206212753973.

Selection and characterization of a mutant of feline immunodeficiency virus resistant to 2',3'-dideoxycytidine.

PubMed Central

Medlin, H K; Zhu, Y Q; Remington, K M; Phillips, T R; North, T W

1996-01-01

We have selected and plaque purified a mutant of feline immunodeficiency virus (FIV) that is resistant to 2',3'-dideoxycytidine (ddC). This mutant was selected in cultured cells in the continuous presence of 25 microM ddC. The mutant, designated DCR-5c, was fourfold resistant to ddC, threefold resistant to 2',3'-dideoxyinosine, and more than fourfold resistant to phosphonoformic acid. DCR-5c displayed little or no resistance to (-)-beta-2',3'-dideoxy-3'-thiacytidine, 3'-azido-3'-deoxythymidine, or 9-(2-phosphonylmethoxyethyl) adenine. Reverse transcriptase purified from DCR-5c was less susceptible to inhibition by ddCTP, phosphonoformic acid, ddATP, or azido-dTTP than the wild-type FIV reverse transcriptase. Sequence analysis of DCR-5c revealed a single base change (G to C at nucleotide 2342) in the reverse transcriptase-encoding region of FIV. This mutation results in substitution of His for Asp at codon 3 of FIV reverse transcriptase. The role of this mutation in ddC resistance was confirmed by site-directed mutagenesis. PMID:8849258

Osteoporosis and osteopenia are not associated with T-cell activation in older cART-treated HIV-infected patients.

PubMed

Krikke, M; Klomberg, R C W; van der Veer, E; Tesselaar, K; Verhaar, H J J; Hoepelman, A I M; Arends, J E

2017-05-01

A higher risk of developing osteopenia/ osteoporosis has been seen in HIV-infected patients. We compared HIV-infected patients, all treated with combination antiretroviral therapy (cART), with a low bone mineral density (BMD) (T-score < -1) to those with a normal BMD (T-score > -1), examining the relation with T-cell activation and bone turnover markers (c-terminal telopeptide (CTX) and procollagen type 1 amino-terminal propeptide (P1NP)). In this single visit pilot study, bone turnover markers, T-cell activation (CD38 + HLA - DR +) and senescence (CD57+) of T cells were measured in patients who had previously undergone dual energy X-ray absorptiometry scanning. All study participants (n = 16) were male, on cART, with a median age of 61 years (IQR 56-66). Nine patients had osteopenia/osteoporosis. When comparing the patients with osteopenia/osteoporosis with those with a normal BMD, no differences in activation and senescence were found. A relation was seen between higher bone formation (P1NP) and patients who were on cART for longer. The median length of cART use was 5.5 years (IQR 4.5-7.8), with all patients on nucleoside reverse transcriptase inhibitors, 88% on tenofovir, 63% on non-nucleoside reverse transcriptase inhibitors (NNRTIs) and 38% on protease inhibitors. Osteopenia/osteoporosis was seen in 100% of the patients on protease inhibitors versus 30% of those on NNRTIs. This study did not find an association between activated T cells and BMD, thus did not explain the higher prevalence of osteopenia/osteoporosis in HIV-infected patients. Interestingly, this small pilot showed that cART might influence BMD, with a possible negative effect for protease inhibitors and a possible protective effect for NNRTIs. These results warrant further investigation.

Design, synthesis and antiviral evaluation of novel heteroarylcarbothioamide derivatives as dual inhibitors of HIV-1 reverse transcriptase-associated RNase H and RDDP functions.

PubMed

Corona, Angela; Onnis, Valentina; Deplano, Alessandro; Bianco, Giulia; Demurtas, Monica; Distinto, Simona; Cheng, Yung-Chi; Alcaro, Stefano; Esposito, Francesca; Tramontano, Enzo

2017-08-31

In the continuous effort to identify new HIV-1 inhibitors endowed with innovative mechanisms, the dual inhibition of different viral functions would provide a significant advantage against drug-resistant variants. The HIV-1 reverse transcriptase (RT)-associated ribonuclease H (RNase H) is the only viral-encoded enzymatic activity that still lacks an efficient inhibitor. We synthesized a library of 3,5-diamino-N-aryl-1H-pyrazole-4-carbothioamide and 4-amino-5-benzoyl-N-phenyl-2-(substituted-amino)-1H-pyrrole-3-carbothioamide derivatives and tested them against RNase H activity. We identified the pyrazolecarbothioamide derivative A15, able to inhibit viral replication and both RNase H and RNA-dependent DNA polymerase (RDDP) RT-associated activities in the low micromolar range. Docking simulations hypothesized its binding to two RT pockets. Site-directed mutagenesis experiments showed that, with respect to wt RT, V108A substitution strongly reduced A15 IC50 values (12.6-fold for RNase H inhibition and 4.7-fold for RDDP), while substitution A502F caused a 9.0-fold increase in its IC50 value for RNase H, not affecting the RDDP inhibition, reinforcing the hypothesis of a dual-site inhibition. Moreover, A15 retained good inhibition potency against three non-nucleoside RT inhibitor (NNRTI)-resistant enzymes, confirming a mode of action unrelated to NNRTIs and suggesting its potential as a lead compound for development of new HIV-1 RT dual inhibitors active against drug-resistant viruses. © FEMS 2017. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Solution characterization of [methyl-13C]methionine HIV-1 reverse transcriptase by NMR spectroscopy☆

PubMed Central

Zheng, Xunhai; Mueller, Geoffrey A.; DeRose, Eugene F.; London, Robert E.

2013-01-01

HIV reverse transcriptase (RT) is a primary target for drug intervention in the treatment of AIDS. Wereport the first solution NMR studies of [methyl-13 C]methionine HIV-1 RT, aimed at better understanding the conformational and dynamic characteristics of RT, both in the presence and absence of the non-nucleoside RT inhibitor (NNRTI) nevirapine. The selection of methionine as a structural probe was based both on its favorable NMR characteristics, and on the presence of two important active site methionine residues, M18466 and M23066. Observation of the M184 resonance is subunit dependent; in the p66 subunit the solvent-exposed residue produces a readily observed signal with a characteristic resonance shift, while in the globular p51 subunit, the M18451 resonance is shifted and broadened as M184 becomes buried in the protein interior. In contrast, although structural data indicates that the environment of M230 is also strongly subunit dependent, the M230 resonances from both subunits have very similar shift and relaxation characteristics. A comparison of chemical shift and intensity data with model-based predictions gives reasonable agreement for M18466, while M23066, located on the β-hairpin “primer grip”, is more mobile and solvent-exposed than suggested by crystal structures of the apo enzyme which have a “closed” fingers-thumb conformation. This mobility of the primer grip is presumably important for binding of non-nucleoside RT inhibitors (NNRTIs), since the NNRTI binding pocket is not observed in the absence of the inhibitors, requiring instead that the binding pocket be dynamically accessible. In the presence of the nevirapine, both the M18466 and M23066 resonances are significantly perturbed, while none of the methionine resonances in the p51 subunit is sensitive to this inhibitor. Site-directed mutagenesis indicates that both M16 and M357 produce two resonances in each subunit, and for both residues, the intensity ratio of the component peaks is strongly subunit dependent. Conformational features that might explain the multiple peaks are discussed. PMID:19665484

Diketo acids derivatives as integrase inhibitors: the war against the acquired immunodeficiency syndrome.

PubMed

Henao-Mejia, Jorge; Góez, Yenny; Patiño, Pablo; Rugeles, Maria T

2006-06-01

Since the human immunodeficiency virus was identified as etiological agent of the acquired immunodeficiency syndrome, great advances have been accomplished in the therapeutic field leading to reduced morbidity and mortality among infected patients. However, the high mutation rate of the viral genome generates strains resistant to multiple drugs, pointing to the importance of finding new therapeutic targets. Among the HIV structural genes, the POL gene codes for three essential enzymes: reverse transcriptase, protease, and integrase; nineteen of the twenty drugs currently approved by the Food and Drug Administration to treat this viral infection, inhibit the reverse transcriptase and the protease. Although intense research has been carried out in this area during the last 10 years, HIV integrase inhibitors are not yet approved for clinical use; however the fact that presence of this enzyme is a sine qua non for a productive HIV life cycle joined to its unique properties makes it a promissory target for anti-HIV therapy. Many compounds have been claimed to inhibit integrase in vitro; however, few of them have proven to have antiviral activity and low cytotoxicity in cell systems. Diketoacid derivatives are the most promising integrase inhibitors so far reported. Initially discovered independently by Shionogi & Co. and the Merck Research Laboratories, these compounds are highly specific for the integrase with potent antiviral activity in vitro and in vivo, and low cytotoxicity in cell cultures. Some of these compounds have recently entered clinical trials. Due to the high relevance of integrase inhibitors, and specifically of diketoacid derivatives, we review the latest findings and patents in this important field of research.

Development and validation of the first liquid chromatography-tandem mass spectrometry assay for simultaneous quantification of multiple antiretrovirals in meconium.

PubMed

Himes, Sarah K; Scheidweiler, Karl B; Tassiopoulos, Katherine; Kacanek, Deborah; Hazra, Rohan; Rich, Kenneth; Huestis, Marilyn A

2013-02-05

A novel method for the simultaneous quantification of 16 antiretroviral (ARV) drugs and 4 metabolites in meconium was developed and validated. Quantification of 6 nucleoside/nucleotide reverse transcriptase inhibitors, 2 non-nucleoside reverse transcriptase inhibitors, 7 protease inhibitors, and 1 integrase inhibitor was achieved in 0.25 g of meconium. Specimen preparation included methanol homogenization and solid-phase extraction. Separate positive and negative polarity multiple reaction monitoring mode injections were required to achieve sufficient sensitivity. Linearity ranged from 10 to 75 ng/g up to 2500 ng/g for most analytes and 100-500 ng/g up to 25,000 ng/g for some; all correlation coefficients were ≥0.99. Extraction efficiencies from meconium were 32.8-119.5% with analytical recovery of 80.3-108.3% and total imprecision of 2.2-11.0% for all quantitative analytes. Two analytes with analytical recovery (70.0-138.5%) falling outside the 80-120% criteria range were considered semiquantitative. Matrix effects were -98.3-47.0% and -98.0-67.2% for analytes and internal standards, respectively. Analytes were stable (>75%) at room temperature for 24 h, 4 °C for 3 days, -20 °C for 3 freeze-thaw cycles over 3 days, and on the autosampler. Method applicability was demonstrated by analyzing meconium from HIV-uninfected infants born to HIV-positive mothers on ARV therapy. This method can be used as a tool to investigate the potential effects of in utero ARV exposure on childhood health and neurodevelopmental outcomes.

Protecting the fetus against HIV infection: a systematic review of placental transfer of antiretrovirals.

PubMed

McCormack, Shelley A; Best, Brookie M

2014-11-01

Maternal-to-fetal transfer of antiretroviral drugs contributes to prevention of vertical transmission of HIV. This systematic review discusses published studies containing data pertaining to the pharmacokinetics of placental transfer of antiretrovirals in humans, including paired cord and maternal plasma samples collected at the time of delivery as well as ex vivo placental perfusion models. Articles pertaining to placental transfer of antiretrovirals were identified from PubMed, from references of included articles, and from US Department of Health and Human Services Panel on Treatment of HIV-infected Pregnant Women and Prevention of Perinatal Transmission guidelines. Articles from non-human animal models or that had no original maternal-to-fetal transfer data were excluded. PRISMA guidelines were followed. A total of 103 published studies were identified. Data across studies appeared relatively consistent for the nucleoside reverse transcriptase inhibitors (NRTIs) and the non-nucleotide reverse transcriptase inhibitors (NNRTIs), with cord to maternal ratios approaching 1 for many of these agents. The protease inhibitors atazanavir and lopinavir exhibited consistent maternal-to-fetal transfer across studies, although the transfer may be influenced by variations in drug-binding proteins. The protease inhibitors indinavir, nelfinavir, and saquinavir exhibited unreliable placental transport, with cord blood concentrations that were frequently undetectable. Limited data, primarily from case reports, indicate that darunavir and raltegravir provide detectable placental transfer. These findings appear consistent with current guidelines of using two NRTIs plus an NNRTI, atazanavir/ritonavir, or lopinavir/ritonavir to maximize placental transfer as well as to optimally suppress maternal viral load. Darunavir/ritonavir and raltegravir may reasonably serve as second-line agents.

Differences in resistance mutations among HIV-1 non-subtype B infections: a systematic review of evidence (1996–2008)

PubMed Central

2009-01-01

Ninety percent of HIV-1-infected people worldwide harbour non-subtype B variants of HIV-1. Yet knowledge of resistance mutations in non-B HIV-1 and their clinical relevance is limited. Although a few reviews, editorials and perspectives have been published alluding to this lack of data among non-B subtypes, no systematic review has been performed to date. With this in mind, we conducted a systematic review (1996–2008) of all published studies performed on the basis of non-subtype B HIV-1 infections treated with antiretroviral drugs that reported genotype resistance tests. Using an established search string, 50 studies were deemed relevant for this review. These studies reported genotyping data from non-B HIV-1 infections that had been treated with either reverse transcriptase inhibitors or protease inhibitors. While most major resistance mutations in subtype B were also found in non-B subtypes, a few novel mutations in non-B subtypes were recognized. The main differences are reflected in the discoveries that: (i) the non-nucleoside reverse transcriptase inhibitor resistance mutation, V106M, has been seen in subtype C and CRF01_AE, but not in subtype B, (ii) the protease inhibitor mutations L89I/V have been reported in C, F and G subtypes, but not in B, (iii) a nelfinavir selected non-D30N containing pathway predominated in CRF01_AE and CRF02_AG, while the emergence of D30N is favoured in subtypes B and D, (iv) studies on thymidine analog-treated subtype C infections from South Africa, Botswana and Malawi have reported a higher frequency of the K65R resistance mutation than that typically seen with subtype B. Additionally, some substitutions that seem to impact non-B viruses differentially are: reverse transcriptase mutations G196E, A98G/S, and V75M; and protease mutations M89I/V and I93L. Polymorphisms that were common in non-B subtypes and that may contribute to resistance tended to persist or become more frequent after drug exposure. Some, but not all, are recognized as minor resistance mutations in B subtypes. These observed differences in resistance pathways may impact cross-resistance and the selection of second-line regimens with protease inhibitors. Attention to newer drug combinations, as well as baseline genotyping of non-B isolates, in well-designed longitudinal studies with long duration of follow up are needed. PMID:19566959

High rates of virological failure and drug resistance in perinatally HIV-1-infected children and adolescents receiving lifelong antiretroviral therapy in routine clinics in Togo

PubMed Central

Salou, Mounerou; Dagnra, Anoumou Y; Butel, Christelle; Vidal, Nicole; Serrano, Laetitia; Takassi, Elom; Konou, Abla A; Houndenou, Spero; Dapam, Nina; Singo-Tokofaï, Assetina; Pitche, Palokinam; Atakouma, Yao; Prince-David, Mireille; Delaporte, Eric; Peeters, Martine

2016-01-01

Introduction Antiretroviral treatment (ART) has been scaled up over the last decade but compared to adults, children living with HIV are less likely to receive ART. Moreover, children and adolescents are more vulnerable than adults to virological failure (VF) and emergence of drug resistance. In this study we determined virological outcome in perinatally HIV-1-infected children and adolescents receiving ART in Togo. Methods HIV viral load (VL) testing was consecutively proposed to all children and adolescents who were on ART for at least 12 months when attending HIV healthcare services for their routine follow-up visit (June to September 2014). Plasma HIV-1 VL was measured using the m2000 RealTime HIV-1 assay (Abbott Molecular, Des Plaines, IL, USA). Genotypic drug resistance was done for all samples with VL>1000 copies/ml. Results and discussion Among 283 perinatally HIV-1-infected children and adolescents included, 167 (59%) were adolescents and 116 (41%) were children. The median duration on ART was 48 months (interquartile range: 28 to 68 months). For 228 (80.6%), the current ART combination consisted of two nucleoside reverse transcriptase inhibitors (NRTIs) (zidovudine and lamivudine) and one non-nucleoside reverse transcriptase inhibitor (NNRTI) (nevirapine or efavirenz). Only 28 (9.9%) were on a protease inhibitor (PI)-based regimen. VL was below the detection limit (i.e. 40 copies/ml) for 102 (36%), between 40 and 1000 copies/ml for 35 (12.4%) and above 1000 copies/ml for 146 (51.6%). Genotypic drug-resistance testing was successful for 125/146 (85.6%); 110/125 (88.0%) were resistant to both NRTIs and NNRTIs, 1/125 (0.8%) to NRTIs only, 4/125 (3.2%) to NNRTIs only and three harboured viruses resistant to reverse transcriptase and PIs. Overall, 86% (108/125) of children and adolescents experiencing VF and successfully genotyped, corresponding thus to at least 38% of the study population, had either no effective ART or had only a single effective drug in their current ART regimen. Conclusions Our study provided important information on virological outcome on lifelong ART in perinatally HIV-1-infected children and adolescents who were still on ART and continued to attend antiretroviral (ARV) clinics for follow-up visits. Actual conditions for scaling up and monitoring lifelong ART in children in resource-limited countries can have dramatic long-term outcomes and illustrate that paediatric ART receives inadequate attention. PMID:27125320

Differential modulation of P-glycoprotein expression and activity by non-nucleoside HIV-1 reverse transcriptase inhibitors in cell culture.

PubMed

Störmer, Elke; von Moltke, Lisa L; Perloff, Michael D; Greenblatt, David J

2002-07-01

This study investigated the effects of the non-nucleoside HIV-1 reverse transcriptase inhibitors (NNRTI) nevirapine (NVR), efavirenz (EFV), and delavirdine (DLV) on P-glycoprotein (P-gp) activity and expression to anticipate P-gp related drug-drug interactions associated with combination therapy. NNRTIs were evaluated as P-gp substrates by measuring differential transport across Caco-2 cell monolayers. Inhibition of P-gp mediated rhodaminel23 (Rh123) transport in Caco-2 cells was used to assess P-gp inhibition by NNRTIs. Induction of P-gp expression and activity in LS180V cells following 3-day exposure to NNRTIs was measured by western blot analysis and cellular Rh123 uptake, respectively. The NNRTIs showed no differential transport between the basolateral to apical and apical to basolateral direction. NNRTI transport in either direction was not affected by the P-gp inhibitor verapamil. DLV inhibited Rh123 transport, causing a reduction to 15% of control at 100 microM (IC50 = 30 microM). NVR caused a concentration-dependent induction of P-gp expression in LS180V cells resulting in a 3.5-fold increase in immunoreactive P-gp at 100 microM NVR. Induction attributable to EFV and DLV was quantitatively smaller. NVR significantly reduced cellular uptake of Rh123 into LS180V cells, indicating increased drug efflux due to induced P-gp activity; effects of EFV and DLV were smaller. Acute DLV treatment of LS180V cells previously induced with NVR or ritonavir did not reverse the decreased Rh123 cell accumulation. NNRTIs show differential effects on P-gp activity and expression in vitro. Clinical studies are required to elucidate the clinical importance of potential drug interactions.

Antiretroviral therapy modifies the genetic effect of known type 2 diabetes-associated risk variants in HIV-infected women.

PubMed

Frasco, Melissa A; Karim, Roksana; Van Den Berg, David; Watanabe, Richard M; Anastos, Kathryn; Cohen, Mardge; Gange, Stephen J; Gustafson, Deborah R; Liu, Chenglong; Tien, Phyllis C; Mack, Wendy J; Pearce, Celeste L

2014-07-31

Type 2 diabetes mellitus incidence is increased in HIV-infected persons. We examined the associations of diabetes mellitus with known diabetes mellitus-risk alleles from the general population in the context of HIV infection, and explored effect modification by combination antiretroviral therapy (cART). The Women's Interagency HIV Study is a prospective cohort of HIV-infected women. Seventeen European-derived diabetes mellitus-risk polymorphisms were genotyped in the eligible participants of the Women's Interagency HIV Study. Analyses were run separately for non-African Americans (Whites, Hispanics, Asians, and other; n = 378, 49 with incident diabetes mellitus) and African Americans (n = 591, 49 with incident diabetes mellitus). Cox proportional-hazards models were fit to estimate hazard ratios for diabetes mellitus overall and within strata of cART. In non-African Americans, heterogeneity across cART regimen was observed for nine of the 14 polymorphisms (phet < 0.05). One polymorphism was statistically significantly inversely associated with diabetes mellitus risk among women taking two nucleotide reverse transcriptase inhibitors (NRTIs) + non-nucleotide reverse transcriptase inhibitor (NNRTI). Five polymorphisms were statistically significantly associated with diabetes mellitus among women treated with at least two NRTIs + at least one protease inhibitor and one polymorphism was associated with diabetes mellitus among those treated with at least three NRTIs ± NNRTI. The hazard ratio per risk allele for IGF2BP2 rs1470579 was 2.67 (95% confidence interval 1.67-4.31) for women taking cART with at least two NRTIs + at least one protease inhibitor and 2.45 (95% confidence interval 1.08-5.53) in women taking at least three NRTIs ± NNRTI (phet = 2.50 × 10⁻³). No such associations were observed in the African Americans. Genetic susceptibility to diabetes mellitus, based on the variants studied, is substantially elevated among HIV-infected women using cART containing three or more NRTI/protease inhibitor components. A personalized medicine approach to cART selection may be indicated for HIV-infected persons carrying these diabetes mellitus-risk variants.

HIV-1 RT Inhibitors with a Novel Mechanism of Action: NNRTIs that Compete with the Nucleotide Substrate

PubMed Central

Maga, Giovanni; Radi, Marco; Gerard, Marie-Aline; Botta, Maurizio; Ennifar, Eric

2010-01-01

HIV-1 reverse transcriptase (RT) inhibitors currently used in antiretroviral therapy can be divided into two classes: (i) nucleoside analog RT inhibitors (NRTIs), which compete with natural nucleoside substrates and act as terminators of proviral DNA synthesis, and (ii) non-nucleoside RT inhibitors (NNRTIs), which bind to a hydrophobic pocket close to the RT active site. In spite of the efficiency of NRTIs and NNRTIs, the rapid emergence of multidrug-resistant mutations requires the development of new RT inhibitors with an alternative mechanism of action. Recently, several studies reported the discovery of novel non-nucleoside inhibitors with a distinct mechanism of action. Unlike classical NNRTIs, they compete with the nucleotide substrate, thus forming a new class of RT inhibitors: nucleotide-competing RT inhibitors (NcRTIs). In this review, we discuss current progress in the understanding of the peculiar behavior of these compounds. PMID:21994659

A Novel Leu92 Mutant of HIV-1 Reverse Transcriptase with a Selective Deficiency in Strand Transfer Causes a Loss of Viral Replication.

PubMed

Herzig, Eytan; Voronin, Nickolay; Kucherenko, Nataly; Hizi, Amnon

2015-08-01

The process of reverse transcription (RTN) in retroviruses is essential to the viral life cycle. This key process is catalyzed exclusively by the viral reverse transcriptase (RT) that copies the viral RNA into DNA by its DNA polymerase activity, while concomitantly removing the original RNA template by its RNase H activity. During RTN, the combination between DNA synthesis and RNA hydrolysis leads to strand transfers (or template switches) that are critical for the completion of RTN. The balance between these RT-driven activities was considered to be the sole reason for strand transfers. Nevertheless, we show here that a specific mutation in HIV-1 RT (L92P) that does not affect the DNA polymerase and RNase H activities abolishes strand transfer. There is also a good correlation between this complete loss of the RT's strand transfer to the loss of the DNA clamp activity of the RT, discovered recently by us. This finding indicates a mechanistic linkage between these two functions and that they are both direct and unique functions of the RT (apart from DNA synthesis and RNA degradation). Furthermore, when the RT's L92P mutant was introduced into an infectious HIV-1 clone, it lost viral replication, due to inefficient intracellular strand transfers during RTN, thus supporting the in vitro data. As far as we know, this is the first report on RT mutants that specifically and directly impair RT-associated strand transfers. Therefore, targeting residue Leu92 may be helpful in selectively blocking this RT activity and consequently HIV-1 infectivity and pathogenesis. Reverse transcription in retroviruses is essential for the viral life cycle. This multistep process is catalyzed by viral reverse transcriptase, which copies the viral RNA into DNA by its DNA polymerase activity (while concomitantly removing the RNA template by its RNase H activity). The combination and balance between synthesis and hydrolysis lead to strand transfers that are critical for reverse transcription completion. We show here for the first time that a single mutation in HIV-1 reverse transcriptase (L92P) selectively abolishes strand transfers without affecting the enzyme's DNA polymerase and RNase H functions. When this mutation was introduced into an infectious HIV-1 clone, viral replication was lost due to an impaired intracellular strand transfer, thus supporting the in vitro data. Therefore, finding novel drugs that target HIV-1 reverse transcriptase Leu92 may be beneficial for developing new potent and selective inhibitors of retroviral reverse transcription that will obstruct HIV-1 infectivity. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

SJ-3366 Sam Jin Pharmaceutical.

PubMed

Baba, Masanori

2002-08-01

Sam Jin is investigating SJ-3366, a non-nucleoside reverse transcriptase inhibitor (NNRTI), for the potential treatment of HIV infection [302450]. As well as acting as an NNRTI, SJ-3366 also interferes with HIV-1 entry via an intermediate target formed after virus-cell attachment [341146], [363900]. As of June 1998, Sam Jin had been awarded a patent for SJ-3366 in South Africa, with applications pending in 22 other countries [302450].

Inhibition of human immunodeficiency virus type 1 infection by the candidate microbicide dapivirine, a nonnucleoside reverse transcriptase inhibitor.

PubMed

Fletcher, P; Harman, S; Azijn, H; Armanasco, N; Manlow, P; Perumal, D; de Bethune, M-P; Nuttall, J; Romano, J; Shattock, R

2009-02-01

Heterosexual transmission of human immunodeficiency virus (HIV) remains the major route of infection worldwide; thus, there is an urgent need for additional prevention strategies, particularly strategies that could be controlled by women, such as topical microbicides. Potential microbicide candidates must be both safe and effective. Using cellular and tissue explant models, we have evaluated the activity of the nonnucleoside reverse transcriptase inhibitor (NNRTI) dapivirine as a vaginal microbicide. In tissue compatibility studies, dapivirine was well tolerated by epithelial cells, T cells, macrophages, and cervical tissue explants. Dapivirine demonstrated potent dose-dependent inhibitory effects against a broad panel of HIV type 1 isolates from different clades. Furthermore, dapivirine demonstrated potent activity against a wide range of NNRTI-resistant isolates. In human cervical explant cultures, dapivirine was able not only to inhibit direct infection of mucosal tissue but also to prevent the dissemination of the virus by migratory cells. Activity was retained in the presence of semen or a cervical mucus simulant. Furthermore, dapivirine demonstrated prolonged inhibitory effects: it was able to prevent both localized and disseminated infection for as long as 6 days posttreatment. The prolonged protection observed following pretreatment of genital tissue and the lack of observable toxicity suggest that dapivirine has considerable promise as a potential microbicide candidate.

4'-Ethynyl-2-fluoro-2'-deoxyadenosine, MK-8591: a novel HIV-1 reverse transcriptase translocation inhibitor.

PubMed

Markowitz, Martin; Sarafianos, Stefan G

2018-07-01

4'-Ethynyl-2-fluoro-2'-deoxyadenosine (EFdA) is a nucleoside reverse transcriptase inhibitor (NRTI) with a novel mechanism of action, unique structure, and amongst NRTIs, unparalleled anti-HIV-1 activity. We will summarize its structure and function, antiviral activity, resistance profile, and potential as an antiretroviral for use in the treatment and preexposure prophylaxis of HIV-1 infection. EFdA is active against wild-type (EC50 as low as 50 pmol/l) and most highly NRTI-resistant viruses. The active metabolite, EFdA-triphosphate, has been shown to have a prolonged intracellular half-life in human and rhesus (Rh) blood cells. As a result, single drug doses tested in simian immunodeficiency virus mac251-infected Rh macaques and HIV-1-infected individuals exhibited robust antiviral activity of 7-10 days duration. Preclinical studies of EFdA as preexposure prophylaxis in the Rh macaque/simian/human immunodeficiency virus low-dose intrarectal challenge model have shown complete protection when given in clinically relevant doses. EFdA is a novel antiretroviral with activity against both wild-type and NRTI-resistant viruses. As a result of the prolonged intracellular half-life of its active moiety, it is amenable to flexibility in dosing of at least daily to weekly and perhaps longer.

Short communication: high prevalence of drug resistance in HIV type 1-infected children born in Honduras and Belize 2001 to 2004.

PubMed

Parham, Leda; de Rivera, Ivette Lorenzana; Murillo, Wendy; Naver, Lars; Largaespada, Natalia; Albert, Jan; Karlsson, Annika C

2011-10-01

Antiretroviral therapy has had a great impact on the prevention of mother-to-child transmission (MTCT) of HIV-1. However, development of drug resistance, which could be subsequently transmitted to the child, is a major concern. In Honduras and Belize the prevalence of drug resistance among HIV-1-infected children remains unknown. A total of 95 dried blood spot samples was obtained from HIV-1-infected, untreated children in Honduras and Belize born during 2001 to 2004, when preventive antiretroviral therapy was often suboptimal and consisted of monotherapy with nevirapine or zidovudine. Partial HIV-1 pol gene sequences were successfully obtained from 66 children (Honduras n=55; Belize n=11). Mutations associated with drug resistance were detected in 13% of the Honduran and 27% of the Belizean children. Most of the mutations detected in Honduras (43%) and all mutations detected in Belize were associated with resistance to nonnucleoside reverse transcriptase inhibitors, which was expected from the wide use of nevirapine to prevent MTCT during the study period. In addition, although several mothers reported that they had not received antiretroviral therapy, mutations associated with resistance to nucleoside reverse transcriptase inhibitors and protease inhibitors were found in Honduras. This suggests prior and unreported use of these drugs, or that these women had been infected with resistant virus. The present study demonstrates, for the first time, the presence of drug resistance-associated mutations in HIV-1-infected Honduran and Belizean children.

An overview of the molecular and epidemiological features of HIV-1 infection in two major cities of Bahia state, Brazil.

PubMed

Amaral, Amanda Gm; Oliveira, Isabele B; Carneiro, Diego C; Alcantara, Luiz Cj; Monteiro-Cunha, Joana P

2017-06-01

The high mutation rate of the human immunodeficiency virus (HIV) has created a public health challenge because the use of antiretroviral drugs can generate selective pressure that drives resistance in these viruses. The aim of this work was to characterise the molecular and epidemiological profile of HIV in Bahia, Brazil. DNA sequences from regions of HIV gag, pol, and env genes were obtained from previous studies performed in this area between 2002 and 2012. Their genotype and drug-resistance mutations were identified using bioinformatics tools. Clinical and epidemiological data were analysed. Among 263 individuals (46.4% male), 97.5% were asymptomatic and 49.1% were receiving treatment. Most of the individuals were 31 to 40 years old (36.9%) and infected through heterosexual contact (40.7%). The predominant genotype was B (68.1%) followed by BF recombinants (18.6%). Among the individuals infected with either F or BF genotypes, 68.4% were women and 76.8% were infected through heterosexual transmission. The prevalence of associated mutations conferring antiretroviral resistance was 14.2%, with 3.8% of all mutations conferring resistance to protease inhibitors, 9.43% to nucleoside reverse transcriptase inhibitors, and 8.5% to non-nucleoside reverse transcriptase inhibitors. Drug resistance was higher in individuals receiving treatment (26.1%) than in the drug-naïve (4.3%) individuals. This study will contribute to the understanding and monitoring of HIV epidemic in this Brazilian region.

Probing the structural and molecular basis of nucleotide selectivity by human mitochondrial DNA polymerase γ

PubMed Central

Sohl, Christal D.; Szymanski, Michal R.; Mislak, Andrea C.; Shumate, Christie K.; Amiralaei, Sheida; Schinazi, Raymond F.; Anderson, Karen S.; Yin, Y. Whitney

2015-01-01

Nucleoside analog reverse transcriptase inhibitors (NRTIs) are the essential components of highly active antiretroviral (HAART) therapy targeting HIV reverse transcriptase (RT). NRTI triphosphates (NRTI-TP), the biologically active forms, act as chain terminators of viral DNA synthesis. Unfortunately, NRTIs also inhibit human mitochondrial DNA polymerase (Pol γ), causing unwanted mitochondrial toxicity. Understanding the structural and mechanistic differences between Pol γ and RT in response to NRTIs will provide invaluable insight to aid in designing more effective drugs with lower toxicity. The NRTIs emtricitabine [(-)-2,3′-dideoxy-5-fluoro-3′-thiacytidine, (-)-FTC] and lamivudine, [(-)-2,3′-dideoxy-3′-thiacytidine, (-)-3TC] are both potent RT inhibitors, but Pol γ discriminates against (-)-FTC-TP by two orders of magnitude better than (-)-3TC-TP. Furthermore, although (-)-FTC-TP is only slightly more potent against HIV RT than its enantiomer (+)-FTC-TP, it is discriminated by human Pol γ four orders of magnitude more efficiently than (+)-FTC-TP. As a result, (-)-FTC is a much less toxic NRTI. Here, we present the structural and kinetic basis for this striking difference by identifying the discriminator residues of drug selectivity in both viral and human enzymes responsible for substrate selection and inhibitor specificity. For the first time, to our knowledge, this work illuminates the mechanism of (-)-FTC-TP differential selectivity and provides a structural scaffold for development of novel NRTIs with lower toxicity. PMID:26124101

FDA approves efavirenz. Food and Drug Administration.

PubMed

Highleyman, L

1998-10-01

The Food and Drug Administration (FDA) approved DuPont Pharma's new non-nucleoside reverse transcriptase inhibitor (NNRTI) efavirenz (Sustiva, DMP-266). Efavirenz has shown promise in trials with over 2000 participants for up to 24 weeks, and early data suggests it may be as effective as protease inhibitors when used in a combination regimen. It is the first anti-HIV drug approved for once-daily dosing. Efavirenz is well tolerated, and the main side effects reported are dizziness, insomnia, abnormal dreams, and skin rash. Efavirenz has been approved for adults and children, but should not be used by pregnant women. Contact information is provided.

Current concepts of metabolic abnormalities in HIV patients: focus on lipodystrophy.

PubMed

Kolter, Donald P

2003-12-01

HIV infection is associated with a number of metabolic abnormalities, including lipodystrophy, a difficult-to-define disorder whose characteristics include hyperlipidemia, insulin resistance, and fat redistribution. Current data suggest that lipodystrophy is caused by multiple factors. Dual-nucleoside reverse transcriptase inhibitor therapy combined with protease inhibitor therapy has been shown to increase the risk of metabolic abnormalities, but susceptibility independent of drug effects has also been shown. While many of the treatments for the broad range of signs and symptoms of lipodystrophy bring about improvements in patient status, none have been demonstrated to bring about a return to baseline levels.

Maraviroc, as a Switch Option, in HIV-1-infected Individuals With Stable, Well-controlled HIV Replication and R5-tropic Virus on Their First Nucleoside/Nucleotide Reverse Transcriptase Inhibitor Plus Ritonavir-boosted Protease Inhibitor Regimen: Week 48 Results of the Randomized, Multicenter MARCH Study.

PubMed

Pett, Sarah Lilian; Amin, Janaki; Horban, Andrejz; Andrade-Villanueva, Jaime; Losso, Marcelo; Porteiro, Norma; Sierra Madero, Juan; Belloso, Waldo; Tu, Elise; Silk, David; Kelleher, Anthony; Harrigan, Richard; Clark, Andrew; Sugiura, Wataru; Wolff, Marcelo; Gill, John; Gatell, Jose; Fisher, Martin; Clarke, Amanda; Ruxrungtham, Kiat; Prazuck, Thierry; Kaiser, Rolf; Woolley, Ian; Arnaiz, Juan Alberto; Cooper, David; Rockstroh, Jürgen K; Mallon, Patrick; Emery, Sean

2016-07-01

Alternative combination antiretroviral therapies in virologically suppressed human immunodeficiency virus (HIV)-infected patients experiencing side effects and/or at ongoing risk of important comorbidities from current therapy are needed. Maraviroc (MVC), a chemokine receptor 5 antagonist, is a potential alternative component of therapy in those with R5-tropic virus. The Maraviroc Switch Study is a randomized, multicenter, 96-week, open-label switch study in HIV type 1-infected adults with R5-tropic virus, virologically suppressed on a ritonavir-boosted protease inhibitor (PI/r) plus double nucleoside/nucleotide reverse transcriptase inhibitor (2 N(t)RTI) backbone. Participants were randomized 1:2:2 to current combination antiretroviral therapy (control), or replacing the protease inhibitor (MVC + 2 N(t)RTI arm) or the nucleoside reverse transcriptase inhibitor backbone (MVC + PI/r arm) with twice-daily MVC. The primary endpoint was the difference (switch minus control) in proportion with plasma viral load (VL) <200 copies/mL at 48 weeks. The switch arms were judged noninferior if the lower limit of the 95% confidence interval (CI) for the difference in the primary endpoint was < -12% in the intention-to-treat (ITT) population. The ITT population comprised 395 participants (control, n = 82; MVC + 2 N(t)RTI, n = 156; MVC + PI/r, n = 157). Baseline characteristics were well matched. At week 48, noninferior rates of virological suppression were observed in those switching away from a PI/r (93.6% [95% CI, -9.0% to 2.2%] and 91.7% [95% CI, -9.6% to 3.8%] with VL <200 and <50 copies/mL, respectively) compared to the control arm (97.6% and 95.1% with VL <200 and <50 copies/mL, respectively). In contrast, MVC + PI/r did not meet noninferiority bounds and was significantly inferior (84.1% [95% CI, -19.8% to -5.8%] and 77.7% [95% CI, -24.9% to -8.4%] with VL <200 and <50 copies/mL, respectively) to the control arm in the ITT analysis. These data support MVC as a switch option for ritonavir-boosted PIs when partnered with a 2-N(t)RTI backbone, but not as part of N(t)RTI-sparing regimens comprising MVC with PI/r. NCT01384682. © The Author 2016. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail journals.permissions@oup.com.

Deep sequencing analysis of HIV-1 reverse transcriptase at baseline and time of failure in patients receiving rilpivirine in the phase III studies ECHO and THRIVE.

PubMed

Van Eygen, Veerle; Thys, Kim; Van Hove, Carl; Rimsky, Laurence T; De Meyer, Sandra; Aerssens, Jeroen; Picchio, Gaston; Vingerhoets, Johan

2016-05-01

Minority variants (1.0-25.0%) were evaluated by deep sequencing (DS) at baseline and virological failure (VF) in a selection of antiretroviral treatment-naïve, HIV-1-infected patients from the rilpivirine ECHO/THRIVE phase III studies. Linkage between frequently emerging resistance-associated mutations (RAMs) was determined. DS (llIumina®) and population sequencing (PS) results were available at baseline for 47 VFs and time of failure for 48 VFs; and at baseline for 49 responders matched for baseline characteristics. Minority mutations were accurately detected at frequencies down to 1.2% of the HIV-1 quasispecies. No baseline minority rilpivirine RAMs were detected in VFs; one responder carried 1.9% F227C. Baseline minority mutations associated with resistance to other non-nucleoside reverse transcriptase inhibitors (NNRTIs) were detected in 8/47 VFs (17.0%) and 7/49 responders (14.3%). Baseline minority nucleoside/nucleotide reverse transcriptase inhibitor (NRTI) RAMs M184V and L210W were each detected in one VF (none in responders). At failure, two patients without NNRTI RAMs by PS carried minority rilpivirine RAMs K101E and/or E138K; and five additional patients carried other minority NNRTI RAMs V90I, V106I, V179I, V189I, and Y188H. Overall at failure, minority NNRTI RAMs and NRTI RAMs were found in 29/48 (60.4%) and 16/48 VFs (33.3%), respectively. Linkage analysis showed that E138K and K101E were usually not observed on the same viral genome. In conclusion, baseline minority rilpivirine RAMs and other NNRTI/NRTI RAMs were uncommon in the rilpivirine arm of the ECHO and THRIVE studies. DS at failure showed emerging NNRTI resistant minority variants in seven rilpivirine VFs who had no detectable NNRTI RAMs by PS. © 2015 Wiley Periodicals, Inc.

The HEPT Analogue WPR-6 Is Active against a Broad Spectrum of Nonnucleoside Reverse Transcriptase Drug-Resistant HIV-1 Strains of Different Serotypes.

PubMed

Xu, Weisi; Zhao, Jianxiong; Sun, Jianping; Yin, Qianqian; Wang, Yan; Jiao, Yang; Liu, Junyi; Jiang, Shibo; Shao, Yiming; Wang, Xiaowei; Ma, Liying

2015-08-01

Nonnucleoside reverse transcriptase inhibitors (NNRTIs) are important components of the highly active antiretroviral therapy (HAART) used to treat human immunodeficiency type 1 virus (HIV-1). However, because of the emergence of drug resistance and the adverse effects of current anti-HIV drugs, it is essential to develop novel NNRTIs with an excellent safety profile, improved activity against NNRTI-resistant viruses, and enhanced activity against clinical isolates of different subtypes. Here, we have identified 1-[(benzyloxy)methyl]-6-(3,5-dimethylbenzyl)-5-iodopyrimidine-2,4(1H,3H)-dione (WPR-6), a novel NNRTI with a 50% effective concentration (EC50) of 2 to 4 nM against laboratory-adapted HIV-1 strain SF33 and an EC50 of 7 to 14 nM against nucleoside reverse transcriptase inhibitor-resistant HIV-1 strain 7391 with a therapeutic index of >1 × 10(4). A panel of five representative clinical virus isolates of different subtypes circulating predominantly in China was highly sensitive to WPR-6, with EC50s ranging from 1 to 6 nM. In addition, WPR-6 showed excellent antiviral potency against the most prevalent NNRTI-resistant viruses containing the K103N and Y181C mutations. To determine whether WPR-6 selects for novel resistant mutants, in vitro resistance selection was conducted with laboratory-adapted HIV-1 strain SF33 on MT-4 cells. The results demonstrated that V106I and Y188L were the two dominant NNRTI-associated resistance mutations detected in the breakthrough viruses. Taken together, these in vitro data indicate that WPR-6 has greater efficacy than the reference HEPT analogue TNK651 and the marketed drug nevirapine against HIV-1. However, to develop it as a new NNRTI, further improvement of its pharmacological properties is warranted. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

HIV type-1 genotypic resistance profiles in vertically infected patients from Argentina reveal an association between K103N+L100I and L74V mutations.

PubMed

Aulicino, Paula C; Rocco, Carlos A; Mecikovsky, Debora; Bologna, Rosa; Mangano, Andrea; Sen, Luisa

2010-01-01

Patterns and pathways of HIV type-1 (HIV-1) antiretroviral (ARV) drug resistance-associated mutations in clinical isolates are conditioned by ARV history and factors such as viral subtype and fitness. Our aim was to analyse the frequency and association of ARV drug resistance mutations in a group of long-term vertically infected patients from Argentina. Plasma samples from 71 patients (38 children and 33 adolescents) were collected for genotypic HIV-1 ARV resistance testing during the period between February 2006 and October 2008. Statistically significant pairwise associations between ARV resistance mutations in pol, as well as associations between mutations and drug exposure, were identified using Fisher's exact tests with Bonferroni and false discovery rate corrections. Phylogenetic analyses were performed for subtype assignment. In protease (PR), resistance-associated mutations M46I/L, I54M/L/V/A/S and V82A/F/T/S/M/I were associated with each other and with minor mutations at codons 10, 24 and 71. Mutations V82A/F/T/S/M/I were primarily selected by the administration of ritonavir (RTV) in an historical ARV regimen. In reverse transcriptase, thymidine analogue mutation (TAM)1 profile was more common than TAM2. The non-nucleoside K103N+L100I mutations were observed at high frequency (15.5%) and were significantly associated with the nucleoside mutation L74V in BF recombinants. Associations of mutations at PR sites reflect the frequent use of RTV at an early time in this group of patients and convergent resistance mechanisms driven by the high exposure to protease inhibitors, as well as local HIV-1 diversity. The results provide clinical evidence of a molecular interaction between K103N+L100I and L74V mutations at the reverse transcriptase gene in vivo, limiting the future use of second-generation non-nucleoside reverse transcriptase inhibitors such as etravirine.

In Vitro Characterization of MK-1439, a Novel HIV-1 Nonnucleoside Reverse Transcriptase Inhibitor

PubMed Central

Feng, Meizhen; Falgueyret, Jean-Pierre; Tawa, Paul; Witmer, Marc; DiStefano, Daniel; Li, Yuan; Burch, Jason; Sachs, Nancy; Lu, Meiqing; Cauchon, Elizabeth; Campeau, Louis-Charles; Grobler, Jay; Yan, Youwei; Ducharme, Yves; Côté, Bernard; Asante-Appiah, Ernest; Hazuda, Daria J.; Miller, Michael D.

2014-01-01

Nonnucleoside reverse transcriptase inhibitors (NNRTIs) are a mainstay of therapy for treating human immunodeficiency type 1 virus (HIV-1)-infected patients. MK-1439 is a novel NNRTI with a 50% inhibitory concentration (IC50) of 12, 9.7, and 9.7 nM against the wild type (WT) and K103N and Y181C reverse transcriptase (RT) mutants, respectively, in a biochemical assay. Selectivity and cytotoxicity studies confirmed that MK-1439 is a highly specific NNRTI with minimum off-target activities. In the presence of 50% normal human serum (NHS), MK-1439 showed excellent potency in suppressing the replication of WT virus, with a 95% effective concentration (EC95) of 20 nM, as well as K103N, Y181C, and K103N/Y181C mutant viruses with EC95 of 43, 27, and 55 nM, respectively. MK-1439 exhibited similar antiviral activities against 10 different HIV-1 subtype viruses (a total of 93 viruses). In addition, the susceptibility of a broader array of clinical NNRTI-associated mutant viruses (a total of 96 viruses) to MK-1439 and other benchmark NNRTIs was investigated. The results showed that the mutant profile of MK-1439 was superior overall to that of efavirenz (EFV) and comparable to that of etravirine (ETR) and rilpivirine (RPV). Furthermore, E138K, Y181C, and K101E mutant viruses that are associated with ETR and RPV were susceptible to MK-1439 with a fold change (FC) of <3. A two-drug in vitro combination study indicated that MK-1439 acts nonantagonistically in the antiviral activity with each of 18 FDA-licensed drugs for HIV infection. Taken together, these in vitro data suggest that MK-1439 possesses the desired properties for further development as a new antiviral agent. PMID:24379202

Silent mutations at codons 65 and 66 in reverse transcriptase alleviate indel formation and restore fitness in subtype B HIV-1 containing D67N and K70R drug resistance mutations

PubMed Central

Telwatte, Sushama; Hearps, Anna C.; Johnson, Adam; Latham, Catherine F.; Moore, Katie; Agius, Paul; Tachedjian, Mary; Sonza, Secondo; Sluis-Cremer, Nicolas; Harrigan, P. Richard; Tachedjian, Gilda

2015-01-01

Resistance to combined antiretroviral therapy (cART) in HIV-1-infected individuals is typically due to nonsynonymous mutations that change the protein sequence; however, the selection of synonymous or ‘silent’ mutations in the HIV-1 genome with cART has been reported. These silent K65K and K66K mutations in the HIV-1 reverse transcriptase (RT) occur in over 35% of drug-experienced individuals and are highly associated with the thymidine analog mutations D67N and K70R, which confer decreased susceptibility to most nucleoside and nucleotide RT inhibitors. However, the basis for selection of these silent mutations under selective drug pressure is unknown. Using Illumina next-generation sequencing, we demonstrate that the D67N/K70R substitutions in HIV-1 RT increase indel frequency by 100-fold at RT codons 65–67, consequently impairing viral fitness. Introduction of either K65K or K66K into HIV-1 containing D67N/K70R reversed the error-prone DNA synthesis at codons 65–67 in RT and improved viral replication fitness, but did not impact RT inhibitor drug susceptibility. These data provide new mechanistic insights into the role of silent mutations selected during antiretroviral therapy and have broader implications for the relevance of silent mutations in the evolution and fitness of RNA viruses. PMID:25765644

Sargassum fusiforme fraction is a potent and specific inhibitor of HIV-1 fusion and reverse transcriptase

PubMed Central

Paskaleva, Elena E; Lin, Xudong; Duus, Karen; McSharry, James J; Veille, Jean-Claude L; Thornber, Carol; Liu, Yanze; Lee, David Yu-Wei; Canki, Mario

2008-01-01

Sargassum fusiforme (Harvey) Setchell has been shown to be a highly effective inhibitor of HIV-1 infection. To identify its mechanism of action, we performed bioactivity-guided fractionation on Sargassum fusiforme mixture. Here, we report isolation of a bioactive fraction SP4-2 (S. fusiforme), which at 8 μg/ml inhibited HIV-1 infection by 86.9%, with IC50 value of 3.7 μg. That represents 230-fold enhancement of antiretroviral potency as compared to the whole extract. Inhibition was mediated against both CXCR4 (X4) and CCR5 (R5) tropic HIV-1. Specifically, 10 μg/ml SP4-2 blocked HIV-1 fusion and entry by 53%. This effect was reversed by interaction of SP4-2 with sCD4, suggesting that S. fusiforme inhibits HIV-1 infection by blocking CD4 receptor, which also explained observed inhibition of both X4 and R5-tropic HIV-1. SP4-2 also inhibited HIV-1 replication after virus entry, by directly inhibiting HIV-1 reverse transcriptase (RT) in a dose dependent manner by up to 79%. We conclude that the SP4-2 fraction contains at least two distinct and biologically active molecules, one that inhibits HIV-1 fusion by interacting with CD4 receptor, and another that directly inhibits HIV-1 RT. We propose that S. fusiforme is a lead candidate for anti-HIV-1 drug development. PMID:18197976

[GESIDA/National AIDS Plan: Consensus document on antiretroviral therapy in adults infected by the human immunodeficiency virus (Updated January 2015)].

PubMed

2015-10-01

This consensus document is an update of combined antiretroviral therapy (cART) guidelines and recommendations for HIV-1 infected adult patients. To formulate these recommendations, a panel composed of members of the AIDS Study Group and the AIDS National Plan (GeSIDA/Plan Nacional sobre el Sida) reviewed the efficacy and safety advances in clinical trials, and cohort and pharmacokinetic studies published in medical journals (PubMed and Embase) or presented in medical scientific meetings. The strength of the recommendations, and the evidence that supports them, are based on modified criteria of the Infectious Diseases Society of America. In this update, cART is recommended for all patients infected by type 1 human immunodeficiency virus (HIV-1). The strength and level of the recommendation depends on the CD4+T-lymphocyte count, the presence of opportunistic diseases or comorbid conditions, age, and prevention of transmission of HIV. The objective of cART is to achieve an undetectable plasma viral load. Initial cART should always comprise a combination of 3 drugs, including 2 nucleoside reverse transcriptase inhibitors, and a third drug from a different family. Three out of the ten recommended regimes are regarded as preferential (all of them with an integrase inhibitor as the third drug), and the other seven (based on a non-nucleoside reverse transcriptase inhibitor, a ritonavir-boosted protease inhibitor, or an integrase inhibitor) as alternatives. This update presents the causes and criteria for switching cART in patients with undetectable plasma viral load, and in cases of virological failure where rescue cART should comprise 3 (or at least 2) drugs that are fully active against the virus. An update is also provided for the specific criteria for cART in special situations (acute infection, HIV-2 infection, and pregnancy) and with comorbid conditions (tuberculosis or other opportunistic infections, kidney disease, liver disease, and cancer). These new guidelines update previous recommendations related to cART (when to begin and what drugs should be used), how to monitor and what to do in case of viral failure or drug adverse reactions. cART specific criteria in comorbid patients and special situations are equally updated. Copyright © 2015 Elsevier España, S.L.U. y Sociedad Española de Enfermedades Infecciosas y Microbiología Clínica. All rights reserved.

TOPICAL TENOFOVIR, A MICROBICIDE EFFECTIVE AGAINST HIV, INHIBITS HERPES SIMPLEX VIRUS-2 REPLICATION

PubMed Central

Andrei, Graciela; Lisco, Andrea; Vanpouille, Christophe; Introini, Andrea; Balestra, Emanuela; van den Oord, Joost; Cihlar, Tomas; Perno, Carlo-Federico; Snoeck, Robert; Margolis, Leonid; Balzarini, Jan

2011-01-01

SUMMARY The HIV reverse transcriptase inhibitor tenofovir, was recently formulated into a vaginal gel for use as a microbicide. In human trials, a 1% tenofovir gel inhibited HIV sexual transmission by 39% and surprisingly herpes simplex virus-2 (HSV-2) transmission by 51%. We demonstrate that the concentration achieved intravaginally with a 1% tenofovir topical gel has direct anti-herpetic activity. Tenofovir inhibits the replication of HSV clinical isolates in human embryonic fibroblasts, keratinocytes, and organotypic epithelial 3D-rafts, decreases HSV replication in human lymphoid and cervical tissues ex vivo, and delays HSV-induced lesions and death of topically treated HSV-infected mice. The active tenofovir metabolite inhibits HSV DNA-polymerase and HIV reverse transcriptase. Tenofovir must be topically administered to achieve concentrations, which are higher than systemic levels after oral treatment, that exert these dual antiviral effects. These findings indicate that a single topical treatment, like tenofovir, can inhibit the transmission of HIV and its co-pathogens. PMID:22018238

Pausing kinetics dominates strand-displacement polymerization by reverse transcriptase

PubMed Central

Malik, Omri; Khamis, Hadeel; Rudnizky, Sergei; Marx, Ailie

2017-01-01

Abstract Reverse transcriptase (RT) catalyzes the conversion of the viral RNA into an integration-competent double-stranded DNA, with a variety of enzymatic activities that include the ability to displace a non-template strand concomitantly with polymerization. Here, using high-resolution optical tweezers to follow the activity of the murine leukemia Virus RT, we show that strand-displacement polymerization is frequently interrupted. Abundant pauses are modulated by the strength of the DNA duplex ∼8 bp ahead, indicating the existence of uncharacterized RT/DNA interactions, and correspond to backtracking of the enzyme, whose recovery is also modulated by the duplex strength. Dissociation and reinitiation events, which induce long periods of inactivity and are likely the rate-limiting step in the synthesis of the genome in vivo, are modulated by the template structure and the viral nucleocapsid protein. Our results emphasize the potential regulatory role of conserved structural motifs, and may provide useful information for the development of potent and specific inhibitors. PMID:28973474

HIV-1 Drug Resistance and Second-Line Treatment in Children Randomized to Switch at Low Versus Higher RNA Thresholds.

PubMed

Harrison, Linda; Melvin, Ann; Fiscus, Susan; Saidi, Yacine; Nastouli, Eleni; Harper, Lynda; Compagnucci, Alexandra; Babiker, Abdel; McKinney, Ross; Gibb, Diana; Tudor-Williams, Gareth

2015-09-01

The PENPACT-1 trial compared virologic thresholds to determine when to switch to second-line antiretroviral therapy (ART). Using PENPACT-1 data, we aimed to describe HIV-1 drug resistance accumulation on first-line ART by virologic threshold. PENPACT-1 had a 2 × 2 factorial design, randomizing HIV-infected children to start protease inhibitor (PI) versus nonnucleoside reverse transcriptase inhibitor (NNRTI)-based ART, and switch at a 1000 copies/mL versus 30,000 copies/mL threshold. Switch criteria were not achieving the threshold by week 24, confirmed rebound above the threshold thereafter, or Center for Disease Control and Prevention stage C event. Resistance tests were performed on samples ≥1000 copies/mL before switch, resuppression, and at 4-years/trial end. Sixty-seven children started PI-based ART and were randomized to switch at 1000 copies/mL (PI-1000), 64 PIs and 30,000 copies/mL (PI-30,000), 67 NNRTIs and 1000 copies/mL (NNRTI-1000), and 65 NNRTI and 30,000 copies/mL (NNRTI-30,000). Ninety-four (36%) children reached the 1000 copies/mL switch criteria during 5-year follow-up. In 30,000 copies/mL threshold arms, median time from 1000 to 30,000 copies/mL switch criteria was 58 (PI) versus 80 (NNRTI) weeks (P = 0.81). In NNRTI-30,000, more nucleoside reverse transcriptase inhibitor (NRTI) resistance mutations accumulated than other groups. NNRTI mutations were selected before switching at 1000 copies/mL (23% NNRTI-1000, 27% NNRTI-30,000). Sixty-two children started abacavir + lamivudine, 166 lamivudine + zidovudine or stavudine, and 35 other NRTIs. The abacavir + lamivudine group acquired fewest NRTI mutations. Of 60 switched to second-line, 79% PI-1000, 63% PI-30,000, 64% NNRTI-1000, and 100% NNRTI-30,000 were <400 copies/mL 24 weeks later. Children on first-line NNRTI-based ART who were randomized to switch at a higher virologic threshold developed the most resistance, yet resuppressed on second-line. An abacavir + lamivudine NRTI combination seemed protective against development of NRTI resistance.

Site-directed mutagenesis of the conserved Asp-443 and Asp-498 carboxy-terminal residues of HIV-1 reverse transcriptase.

PubMed Central

Mizrahi, V; Usdin, M T; Harington, A; Dudding, L R

1990-01-01

Substitution of the conserved Asp-443 residue of HIV-1 reverse transcriptase by asparagine specifically suppressed the ribonuclease H activity of the enzyme without affecting the reverse transcriptase activity, suggesting involvement of this ionizable residue at the ribonuclease H active site. An analogous asparagine substitution of the Asp-498 residue yielded an unstable enzyme that was difficult to enzymatically characterize. However, the instability caused by the Asn-498 mutation was relieved by the introduction of a second distal Asn-443 substitution, yielding an enzyme with wild type reverse transcriptase activity, but lacking ribonuclease H activity. Images PMID:1699202

A new strategy to inhibit the excision reaction catalysed by HIV-1 reverse transcriptase: compounds that compete with the template–primer

PubMed Central

Cruchaga, Carlos; Anso, Elena; Font, María; Martino, Virginia S.; Rouzaut, Ana; Martinez-Irujo, Juan J.

2007-01-01

Inhibitors of the excision reaction catalysed by HIV-1 RT (reverse transcriptase) represent a promising approach in the fight against HIV, because these molecules would interfere with the main mechanism of resistance of this enzyme towards chain-terminating nucleotides. Only a limited number of compounds have been demonstrated to inhibit this reaction to date, including NNRTIs (non-nucleoside RT inhibitors) and certain pyrophosphate analogues. We have found previously that 2GP (2-O-galloylpunicalin), an antiviral compound extracted from the leaves of Terminalia triflora, was able to inhibit both the RT and the RNase H activities of HIV-1 RT without affecting cell proliferation or viability. In the present study, we show that 2GP also inhibited the ATP- and PPi-dependent phosphorolysis catalysed by wild-type and AZT (3′-azido-3′-deoxythymidine)-resistant enzymes at sub-micromolar concentrations. Kinetic and direct-binding analysis showed that 2GP was a non-competitive inhibitor against the nucleotide substrate, whereas it competed with the binding of RT to the template–primer (Kd=85 nM). As expected from its mechanism of action, 2GP was active against mutations conferring resistance to NNRTIs and AZT. The combination of AZT with 2GP was highly synergistic when tested in the presence of pyrophosphate, indicating that the inhibition of RT-catalysed phosphorolysis was responsible for the synergy found. Although other RT inhibitors that compete with the template–primer have been described, this is the first demonstration that these compounds can be used to block the excision of chain terminating nucleotides, providing a rationale for their combination with nucleoside analogues. PMID:17355225

A new strategy to inhibit the excision reaction catalysed by HIV-1 reverse transcriptase: compounds that compete with the template-primer.

PubMed

Cruchaga, Carlos; Anso, Elena; Font, María; Martino, Virginia S; Rouzaut, Ana; Martinez-Irujo, Juan J

2007-07-01

Inhibitors of the excision reaction catalysed by HIV-1 RT (reverse transcriptase) represent a promising approach in the fight against HIV, because these molecules would interfere with the main mechanism of resistance of this enzyme towards chain-terminating nucleotides. Only a limited number of compounds have been demonstrated to inhibit this reaction to date, including NNRTIs (non-nucleoside RT inhibitors) and certain pyrophosphate analogues. We have found previously that 2GP (2-O-galloylpunicalin), an antiviral compound extracted from the leaves of Terminalia triflora, was able to inhibit both the RT and the RNase H activities of HIV-1 RT without affecting cell proliferation or viability. In the present study, we show that 2GP also inhibited the ATP- and PP(i)-dependent phosphorolysis catalysed by wild-type and AZT (3'-azido-3'-deoxythymidine)-resistant enzymes at sub-micromolar concentrations. Kinetic and direct-binding analysis showed that 2GP was a non-competitive inhibitor against the nucleotide substrate, whereas it competed with the binding of RT to the template-primer (K(d)=85 nM). As expected from its mechanism of action, 2GP was active against mutations conferring resistance to NNRTIs and AZT. The combination of AZT with 2GP was highly synergistic when tested in the presence of pyrophosphate, indicating that the inhibition of RT-catalysed phosphorolysis was responsible for the synergy found. Although other RT inhibitors that compete with the template-primer have been described, this is the first demonstration that these compounds can be used to block the excision of chain terminating nucleotides, providing a rationale for their combination with nucleoside analogues.

Suicide Inhibitors of Reverse Transcriptase in the Therapy of AIDS and Other Retroviruses

DTIC Science & Technology

1991-07-01

of intermediate 3 was detected by 3 P NMR spectroscopy . The observed chemical shifts are comparable to those reported ’in the literature. The spectrum...were characterized by NMR spectroscopy . The presence of the characteristic triphosphate group was confirmed by NMR as indicated in the figure below...Two compounds, 2𔃽’ sulfinyl cytidine hydrochloride and 2,0 2 anhydrocytidine hydrochloride, which have proved to be highly effective against vaccinia

Design, Conformation, and Crystallography of 2-Naphthyl Phenyl Ethers as Potent Anti-HIV Agents

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lee, Won-Gil; Chan, Albert H.; Spasov, Krasimir A.

Catechol diethers that incorporate a 7-cyano-2-naphthyl substituent are reported as non-nucleoside inhibitors of HIV-1 reverse transcriptase (NNRTIs). Many of the compounds have 1–10 nM potencies toward wild-type HIV-1. An interesting conformational effect allows two unique conformers for the naphthyl group in complexes with HIV-RT. X-ray crystal structures for 4a and 4f illustrate the alternatives.

Efficient Single Tobamoviral Vector-Based Bioproduction of Broadly Neutralizing Anti-HIV-1 Monoclonal Antibody VRC01 in Nicotiana benthamiana Plants and Utility of VRC01 in Combination Microbicides

PubMed Central

Hamorsky, Krystal Teasley; Grooms-Williams, Tiffany W.; Husk, Adam S.; Bennett, Lauren J.; Palmer, Kenneth E.

2013-01-01

Broadly neutralizing monoclonal antibodies (bnMAbs) may offer powerful tools for HIV-1 preexposure prophylaxis, such as topical microbicides. However, this option is hampered due to expensive MAb biomanufacturing based on mammalian cell culture. To address this issue, we developed a new production system for bnMAb VRC01 in Nicotiana benthamiana plants using a tobamovirus replicon vector. Unlike conventional two-vector-based expression, this system was designed to overexpress full-length IgG1 from a single polypeptide by means of kex2p-like enzyme recognition sites introduced between the heavy and light chains. An enzyme-linked immunosorbent assay (ELISA) revealed that gp120-binding VRC01 IgG1 was maximally accumulated on 5 to 7 days following vector inoculation, yielding ∼150 mg of the bnMAb per kg of fresh leaf material. The plant-made VRC01 (VRC01p) was efficiently purified by protein A affinity followed by hydrophobic-interaction chromatography. ELISA, surface plasmon resonance, and an HIV-1 neutralization assay demonstrated that VRC01p has gp120-binding affinity and HIV-1-neutralization capacity virtually identical to the human-cell-produced counterpart. To advance VRC01p's use in topical microbicides, we analyzed combinations of the bnMAb with other microbicide candidates holding distinct antiviral mechanisms in an HIV-1 neutralization assay. VRC01p exhibited clear synergy with the antiviral lectin griffithsin, the CCR5 antagonist maraviroc, and the reverse transcriptase inhibitor tenofovir in multiple CCR5-tropic HIV-1 strains from clades A, B, and C. In summary, VRC01p is amenable to robust, rapid, and large-scale production and may be developed as an active component in combination microbicides with other anti-HIV agents such as antiviral lectins, CCR5 antagonists, and reverse transcriptase inhibitors. PMID:23403432

Raltegravir plasma concentrations in treatment-experienced patients receiving salvage regimens based on raltegravir with and without maraviroc coadministration.

PubMed

Baroncelli, Silvia; Villani, Paola; Weimer, Liliana E; Ladisa, Nicoletta; Francisci, Daniela; Tommasi, Chiara; Vullo, Vincenzo; Preziosi, Roberta; Cicalini, Stefania; Cusato, Maria; Galluzzo, Clementina; Floridia, Marco; Regazzi, Mario

2010-05-01

Raltegravir and maraviroc represent new, important resources for HIV-infected patients with intolerance or resistance to other antiretroviral agents. The safety and efficacy of both drugs have been investigated, but there is no information on possible pharmacokinetic interactions between these 2 drugs in clinical practice. To evaluate raltegravir plasma concentrations in heavily treatment-experienced patients receiving salvage regimens and explore, in a preliminary assessment, the potential influence of maraviroc coadministration and other cofactors on raltegravir trough concentrations (C(trough)). Fifty-four HIV-infected patients with triple class (nucleoside reverse transcriptase inhibitor, nonnucleoside reverse transcriptase inhibitor, protease inhibitor) treatment experience starting raltegravir 400 mg twice daily, with (n = 11) or without (n = 43) concomitant maraviroc 300 mg twice daily, were evaluated. All regimens included at least 3 drugs of at least 2 different classes. Raltegravir plasma Ctrough, after at least 1 month of treatment, were analyzed to compare groups of patients taking raltegravir only and raltegravir plus maraviroc. Immunovirological (CD4, HIV-RNA) and clinical data after 6 months of treatment were also collected and described. Raltegravir plasma Ctrough showed a large variability (range <0.020-2.47 microg/mL). Median levels were similar in the 2 groups (raltegravir + maraviroc 0.104 microg/mL, range 0.025-0.826; raltegravir 0.090 microg/mL, range <0.020-2.47, p = 0.400). Detectable (>0.02 microg/mL) raltegravir concentrations were observed in all patients receiving raltegravir + maraviroc and in 74% of patients receiving raltegravir alone (p = 0.060). After 6 months of treatment, the 2 groups had similar clinical, virologic, and immunologic conditions. Coadministration of maraviroc does not seem to have any relevant effects on raltegravir plasma Ctrough in heavily treatment-experienced patients receiving salvage regimens. Further studies should evaluate the potential additional benefits of maraviroc coadministration in terms of virologic and immunologic response.

HIV-1 drug resistance in antiretroviral-naive individuals with HIV-1-associated tuberculous meningitis initiating antiretroviral therapy in Vietnam.

PubMed

Thao, Vu P; Le, Thuy; Török, Estee M; Yen, Nguyen T B; Chau, Tran T H; Jurriaans, Suzanne; van Doorn, H Rogier; van Doorn, Rogier H; de Jong, Menno D; Farrar, Jeremy J; Dunstan, Sarah J

2012-01-01

Access to antiretroviral therapy (ART) for HIV-infected individuals in Vietnam is rapidly expanding, but there are limited data on HIV drug resistance (HIVDR) to guide ART strategies. We retrospectively conducted HIVDR testing in 220 ART-naive individuals recruited to a randomized controlled trial of immediate versus deferred ART in individuals with HIV-associated tuberculous meningitis in Ho Chi Minh City (HCMC) from 2005-2008. HIVDR mutations were identified by population sequencing of the HIV pol gene and were defined based on 2009 WHO surveillance drug resistance mutations (SDRMs). We successfully sequenced 219/220 plasma samples of subjects prior to ART; 218 were subtype CRF01_AE and 1 was subtype B. SDRMs were identified in 14/219 (6.4%) subjects; 8/14 were resistant to nucleoside/nucleotide reverse transcriptase inhibitors (NRTIs; T69D, L74V, V75M, M184V/I and K219R), 5/14 to non-nucleoside reverse transcriptase inhibitors (NNRTIs; K103N, V106M, Y181C, Y188C and G190A), 1/14 to both NRTIs and NNRTIs (D67N and Y181C) and none to protease inhibitors. After 6 months of ART, eight subjects developed protocol-defined virological failure. HIVDR mutations were identified in 5/8 subjects. All five had mutations with high-level resistance to NNRTIs and three had mutations with high-level resistance to NRTIs. Due to a high early mortality rate (58%), the effect of pre-existing HIVDR mutations on treatment outcome could not be accurately assessed. The prevalence of WHO SDRMs in ART-naive individuals with HIV-associated tuberculous meningitis in HCMC from 2005-2008 is 6.4%. The SDRMs identified conferred resistance to NRTIs and/or NNRTIs, reflecting the standard first-line ART regimens in Vietnam.

Arylazolyl(azinyl)thioacetanilides. Part 16: Structure-based bioisosterism design, synthesis and biological evaluation of novel pyrimidinylthioacetanilides as potent HIV-1 inhibitors.

PubMed

Li, Xiao; Lu, Xueyi; Chen, Wenmin; Liu, Huiqing; Zhan, Peng; Pannecouque, Christophe; Balzarini, Jan; De Clercq, Erik; Liu, Xinyong

2014-10-01

A series of novel pyrimidinylthioacetanilides were designed, synthesized, and evaluated for their biological activity as potent HIV-1 non-nucleoside reverse transcriptase inhibitors (NNRTIs). Most of the tested compounds were proved to be effective in inhibiting HIV-1 (IIIB) replication with EC50 ranging from 0.15 μM to 24.2 μM, thereinto compound 15 was the most active lead with favorable inhibitory activity against HIV-1 (IIIB) (EC50=0.15 μM, SI=684). Besides, compound 6 displayed moderate inhibition against the double-mutated HIV-1 strain (K103N/Y181C) (EC50=3.9 μM). Preliminary structure-activity relationships (SARs), structure-cytotoxicity relationships (SCRs) data, and molecular modeling studies were discussed as well, which may provide valuable insights for further optimizations. Copyright © 2014 Elsevier Ltd. All rights reserved.

The Reverse Transcriptase of the Tf1 Retrotransposon Has a Specific Novel Activity for Generating the RNA Self-Primer That Is Functional in cDNA Synthesis▿

PubMed Central

Hizi, Amnon

2008-01-01

The Tf1 retrotransposon of Schizosaccharomyces pombe represents a group of eukaryotic long terminal repeat (LTR) retroelements that, based on their sequences, were predicted to use an RNA self-primer for initiating reverse transcription while synthesizing the negative-sense DNA strand. This feature is substantially different from the one typical to retroviruses and other LTR retrotransposons that all exhibit a tRNA-dependent priming mechanism. Genetic studies have suggested that the self-primer of Tf1 can be generated by a cleavage between the 11th and 12th bases of the Tf1 RNA transcript. The in vitro data presented here show that recombinant Tf1 reverse transcriptase indeed introduces a nick at the end of a duplexed region at the 5′ end of Tf1 genomic RNA, substantiating the prediction that this enzyme is responsible for generating this RNA self-primer. The 3′ end of the primer, generated in this manner, can then be extended upon the addition of deoxynucleoside triphosphates by the DNA polymerase activity of the same enzyme, synthesizing the negative-sense DNA strand. This functional primer must have been generated by the RNase H activity of Tf1 reverse transcriptase, since a mutant enzyme lacking this activity has lost its ability to generate the self-primer. It was also found here that the reverse transcriptases of human immunodeficiency virus type 1 and of murine leukemia virus do not exhibit this specific cleavage activity. In all, it is likely that the observed unique mechanism of self-priming in Tf1 represents an early advantageous form of initiating reverse transcription in LTR retroelements without involving cellular tRNAs. PMID:18753200

The reverse transcriptase of the Tf1 retrotransposon has a specific novel activity for generating the RNA self-primer that is functional in cDNA synthesis.

PubMed

Hizi, Amnon

2008-11-01

The Tf1 retrotransposon of Schizosaccharomyces pombe represents a group of eukaryotic long terminal repeat (LTR) retroelements that, based on their sequences, were predicted to use an RNA self-primer for initiating reverse transcription while synthesizing the negative-sense DNA strand. This feature is substantially different from the one typical to retroviruses and other LTR retrotransposons that all exhibit a tRNA-dependent priming mechanism. Genetic studies have suggested that the self-primer of Tf1 can be generated by a cleavage between the 11th and 12th bases of the Tf1 RNA transcript. The in vitro data presented here show that recombinant Tf1 reverse transcriptase indeed introduces a nick at the end of a duplexed region at the 5' end of Tf1 genomic RNA, substantiating the prediction that this enzyme is responsible for generating this RNA self-primer. The 3' end of the primer, generated in this manner, can then be extended upon the addition of deoxynucleoside triphosphates by the DNA polymerase activity of the same enzyme, synthesizing the negative-sense DNA strand. This functional primer must have been generated by the RNase H activity of Tf1 reverse transcriptase, since a mutant enzyme lacking this activity has lost its ability to generate the self-primer. It was also found here that the reverse transcriptases of human immunodeficiency virus type 1 and of murine leukemia virus do not exhibit this specific cleavage activity. In all, it is likely that the observed unique mechanism of self-priming in Tf1 represents an early advantageous form of initiating reverse transcription in LTR retroelements without involving cellular tRNAs.

Design, synthesis, and anti-HIV-1 activity of 1-aromatic methyl-substituted 3-(3,5-dimethylbenzyl)uracil and N-3,5-dimethylbenzyl-substituted urea derivatives.

PubMed

Sakakibara, Norikazu; Baba, Masanori; Okamoto, Mika; Toyama, Masaaki; Demizu, Yosuke; Misawa, Takashi; Kurihara, Masaaki; Irie, Kohji; Kato, Yoshihisa; Maruyama, Tokumi

2015-02-01

A new series of 1-aromatic methyl-substituted 3-(3,5-dimethylbenzyl)uracil and N-3,5-dimethylbenzyl-substituted urea derivatives were synthesized and evaluated as non-nucleoside HIV-1 reverse transcriptase inhibitors. A series of new 6-azido and 6-amino derivatives of 1-substituted-3-(3,5-dimethylbenzyl)uracils were synthesized using our previously reported method, and three acyclic derivatives were synthesized from urea. The anti-HIV-1 activities of these compounds were determined based on the inhibition of virus-induced cytopathogenicity in MT-4 cells. The cytotoxicities of the compounds were evaluated using the viability of mock-infected cells. Some of these compounds showed good-to-moderate activities against HIV-1 with half maximal effective concentration (EC50) values in the submicromolar or subnanomolar range. Compared with emivirine, compound 6-amino-3-(3,5-dimethylbenzyl)-1-(4-aminobenzyl)uracil showed significant anti-HIV-1 activity with an EC50 value of 10 nM and a high selectivity index of 1923. Preliminary structure-activity relationship studies and molecular modeling analyses were carried out to explore the major interactions between HIV-1 reverse transcriptase and the potent inhibitor 6-amino-3-(3,5-dimethylbenzyl)-1-(4-aminobenzyl)uracil; these results may be important for further development of this class of compounds as anti-HIV-1 agents. The excellent activity of 6-amino-3-(3,5-dimethylbenzyl)-1-(4-aminobenzyl)uracil (EC50: 0.010 ± 0.006 µM, SI: >1923) may serve as the basis for conducting further investigations on the behavior of this class of compounds against drug-resistant mutants. © The Author(s) 2015 Reprints and permissions: sagepub.co.uk/journalsPermissions.nav.

Design, synthesis, and anti-HIV-1 activity of 1-aromatic methyl-substituted 3-(3,5-dimethylbenzyl)uracil and N-3,5-dimethylbenzyl-substituted urea derivatives

PubMed Central

Sakakibara, Norikazu; Baba, Masanori; Okamoto, Mika; Toyama, Masaaki; Demizu, Yosuke; Misawa, Takashi; Kurihara, Masaaki; Irie, Kohji; Kato, Yoshihisa; Maruyama, Tokumi

2015-01-01

Background A new series of 1-aromatic methyl-substituted 3-(3,5-dimethylbenzyl)uracil and N-3,5-dimethylbenzyl-substituted urea derivatives were synthesized and evaluated as non-nucleoside HIV-1 reverse transcriptase inhibitors. Methods A series of new 6-azido and 6-amino derivatives of 1-substituted-3-(3,5-dimethylbenzyl)uracils were synthesized using our previously reported method, and three acyclic derivatives were synthesized from urea. The anti-HIV-1 activities of these compounds were determined based on the inhibition of virus-induced cytopathogenicity in MT-4 cells. The cytotoxicities of the compounds were evaluated using the viability of mock-infected cells. Results Some of these compounds showed good-to-moderate activities against HIV-1 with half maximal effective concentration (EC50) values in the submicromolar or subnanomolar range. Compared with emivirine, compound 6-amino-3-(3,5-dimethylbenzyl)-1-(4-aminobenzyl)uracil showed significant anti-HIV-1 activity with an EC50 value of 10 nM and a high selectivity index of 1923. Preliminary structure–activity relationship studies and molecular modeling analyses were carried out to explore the major interactions between HIV-1 reverse transcriptase and the potent inhibitor 6-amino-3-(3,5-dimethylbenzyl)-1-(4-aminobenzyl)uracil; these results may be important for further development of this class of compounds as anti-HIV-1 agents. Conclusion The excellent activity of 6-amino-3-(3,5-dimethylbenzyl)-1-(4-aminobenzyl)uracil (EC50: 0.010 ± 0.006 µM, SI: >1923) may serve as the basis for conducting further investigations on the behavior of this class of compounds against drug-resistant mutants. PMID:26149262

Transmitted Antiretroviral Drug Resistance in Newly HIV-Infected and Untreated Patients in Ségou and Bamako, Mali

PubMed Central

Fofana, Djeneba Bocar; Maiga, Aichatou Chehy; Diallo, Fodie; Ait-Arkoub, Zaina; Daou, Fatoumata; Cisse, Mamadou; Sarro, Yaya dit Sadio; Oumar, Aboubacar Alassane; Sylla, Aliou; Katlama, Christine; Taiwo, Babafemi; Murphy, Robert; Tounkara, Anatole; Marcelin, Anne-Genevieve; Calvez, Vincent

2013-01-01

Abstract The WHO recommends regular surveillance for transmitted antiretroviral drug-resistant viruses in HIV antiretroviral treatment (ART)-naive patients in resource-limited settings. This study aimed to assess the prevalence of mutations associated with resistance in ART-naive patients newly diagnosed with HIV in Bamako and Ségou in Mali. HIV-positive patients who never received ART were recruited in Bamako and Ségou, Mali. The reverse transcriptase (RT) and protease (PR) genes of these patients were sequenced by the “ViroSeq” method. Analysis and interpretation of the resistance were made according to the WHO 2009 list of drug resistance mutations. In all, 51/54 (94.4%) sample patients were sequenced. The median age (IQR) of our patients was 24 (22–27) years and the median CD4 count was 380 (340–456) cells/mm3. The predominant subtype was recombinant HIV-1 CRF02_AG (66.7%) followed by CRF06_cpx (12%) and CRF09_cpx (4%). Four patients had mutations associated with resistance, giving an overall prevalence of resistance estimated at 7.9%. There were two (4%) patients with nucleoside reverse transcriptase inhibitor (NRTI) mutations (one M184V and one T215Y), two (4%) with non-NRTI mutations (two K103N), and one (2%) with a protease inhibitor mutation (one I54V). The prevalence of primary resistance in newly infected patients in Mali is moderate (7.9%). This indicates that the standard NNRTI-based first-line regimen used in Mali is suboptimal for some patients. This study should be done regularly to inform clinical practice. PMID:22823755

HIV salvage therapy does not require nucleoside reverse transcriptase inhibitors: a randomized trial

PubMed Central

Tashima, Karen T; Smeaton, Laura M; Fichtenbaum, Carl J; Andrade, Adriana; Eron, Joseph J; Gandhi, Rajesh T; Johnson, Victoria A; Klingman, Karin L; Ritz, Justin; Hodder, Sally; Santana, Jorge L; Wilkin, Timothy; Haubrich, Richard H

2015-01-01

Background Nucleoside reverse transcriptase inhibitors (NRTIs) are often included in antiretroviral (ARV) regimens in treatment-experienced patients in the absence of data from randomized trials. Objective To compare treatment success between participants who omit versus Add NRTIs to an optimized ARV regimen of three or more agents. Design Multisite, randomized, controlled trial. Setting Outpatient HIV clinics. Participants HIV-infected patients with three-class ARV experience and/or viral resistance. Intervention Open-label optimized regimens (not including NRTIs) were selected based upon treatment history and susceptibility testing. Participants were randomized to Omit or Add NRTIs. Measurements The primary efficacy outcome was regimen failure through week 48, using a non-inferiority margin of 15%. The primary safety outcome was time to initial episode of severe sign/symptom or laboratory abnormality prior to discontinuation of NRTI assignment. Results 360 participants were randomized and 93% completed a week 48 visit. The cumulative probability of regimen failure was 29.8% in the Omit NRTI arm versus 25.9% in the Add NRTI arm (difference= 3.2%: 95% CI, −6.1 to 12.5). There were no significant differences in the primary safety endpoints or the proportion of participants with HIV RNA <50 copies/mL between arms. No deaths occurred in the Omit NRTIs arm, compared with 7 deaths in the Add NRTIs arm. Limitations Non-blinded study design and may not be applicable to resource poor settings. Conclusion HIV-infected treatment-experienced patients starting a new optimized regimen can safely omit NRTIs without compromising virologic efficacy. Omitting NRTIs will reduce pill burden, cost, and toxicity in this patient population. PMID:26595748

Characterizing Class‐Specific Exposure‐Viral Load Suppression Response of HIV Antiretrovirals Using A Model‐Based Meta‐Analysis

PubMed Central

Xu, Y; Li, YF; Zhang, D; Dockendorf, M; Tetteh, E; Rizk, ML; Grobler, JA; Lai, M‐T; Gobburu, J

2016-01-01

We applied model‐based meta‐analysis of viral suppression as a function of drug exposure and in vitro potency for short‐term monotherapy in human immunodeficiency virus type 1 (HIV‐1)‐infected treatment‐naïve patients to set pharmacokinetic targets for development of nonnucleoside reverse transcriptase inhibitors (NNRTIs) and integrase strand transfer inhibitors (InSTIs). We developed class‐specific models relating viral load kinetics from monotherapy studies to potency normalized steady‐state trough plasma concentrations. These models were integrated with a literature assessment of doses which demonstrated to have long‐term efficacy in combination therapy, in order to set steady‐state trough concentration targets of 6.17‐ and 2.15‐fold above potency for NNRTIs and InSTIs, respectively. Both the models developed and the pharmacokinetic targets derived can be used to guide compound selection during preclinical development and to predict the dose–response of new antiretrovirals to inform early clinical trial design. PMID:27171172

Genotypic tropism testing of proviral DNA to guide maraviroc initiation in aviraemic subjects: 48-week analysis of results from the PROTEST study.

PubMed

Poveda, E; Hernández-Quero, J; Pérez-Elías, M J; Ribas, M A; Martínez-Madrid, O J; Flores, J; Navarro, J; Gutiérrez, F; García-Deltoro, M; Imaz, A; Ocampo, A; Artero, A; Blanco, F; Bernal, E; Pasquau, J; Mínguez-Gallego, C; Pérez, N; Aiestaran, A; García, F; Paredes, R

2017-08-01

Maraviroc (MVC) is a suitable drug for aviraemic subjects on antiretroviral treatment (ART) developing toxicity. Its prescription requires prior tropism testing. It is unknown if proviral DNA genotypic tropism testing is reliable for guiding MVC initiation in aviraemic subjects, so this study was carried out to address this issue. PROTEST was a phase 4, prospective, single-arm clinical trial carried out in 24 HIV care centres in Spain. MVC-naïve HIV-1-infected patients with HIV-1 RNA < 50 copies/mL on stable ART during the previous 6 months who required an ART change because of toxicity and who had R5 HIV, as determined by proviral DNA genotypic tropism testing, initiated MVC with two nucleoside reverse transcriptase inhibitors (NRTIs) and were followed for 48 weeks. Virological failure was defined as two consecutive viral load measurements > 50 copies/mL. Tropism results were available for 141 of 175 (80.6%) subjects screened: 60% had R5 and 85% of these (n = 74) were finally included in the study. Previous ART included protease inhibitors (PIs) in 62% of subjects, nonnucleoside reverse transcriptase inhibitors (NNRTIs) in 36%, and integrase inhibitors (INIs) in 2%. Main reasons for treatment change were dyslipidaemia (42%), gastrointestinal symptoms (22%) and liver toxicity (15%). MVC was given alongside tenofovir (TDF)/emtricitabine (FTC) (54%) and abacavir (ABC)/lamivudine (3TC) (40%) in most patients. Eighty-four per cent of patients maintained a viral load < 50 copies/mL to week 48, whereas 16% discontinued treatment: two withdrew informed consent, one had an R5 to X4 shift between screening and baseline, one was lost to follow-up, one developed an adverse event (rash), two died from non-study-related causes, and five developed protocol-defined virological failure. Initiation of MVC plus two NRTIs in aviraemic subjects based on genotypic tropism testing of proviral HIV-1 DNA is associated with low rates of virological failure for up to 1 year. © 2016 British HIV Association.

Anti-adenoviral effect of anti-HIV agents in vitro in serotypes inducing keratoconjunctivitis.

PubMed

Uchio, Eiichi; Fuchigami, Aki; Kadonosono, Kazuaki; Hayashi, Akio; Ishiko, Hiroaki; Aoki, Koki; Ohno, Shigeaki

2007-09-01

Around one million people are affected by adenoviral keratoconjunctivitis a year in Japan, and it is recognized as one of the major pathogens of ophthalmological nosocomial infection worldwide. Although cidofovir can be used systemically for immunocompromised patients with disseminated adenoviral infection, no specific anti-adenoviral agent has been established for the treatment of adenoviral infection. We evaluated the anti-adenoviral effect of anti-HIV (human immunodeficiency virus) agents in this study. Five anti-HIV agents (zalcitabine, stavudine, nevirapine, indinavir and amprenavir) were subjected to in vitro evaluation. A549 cells were used for viral cell culture, and adenovirus serotypes 3, 4, 8, 19 and 37 were used. After calculating CC(50) (50% cytotoxic concentration) of each agent by MTS (3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium) method, we cultured adenovirus with the agents for seven days and quantitatively measured extracted adenoviral DNA by real-time PCR. Among the anti-HIV drugs, zalcitabine and stavudine, both nucleoside reverse transcriptase inhibitors, showed significant anti-adenoviral activity. In contrast, nevirapine, a non-nucleoside reverse transcriptase inhibitor, and indinavir and amprenavir, which are both protease inhibitors, were ineffective against adenovirus. These results indicate that zalcitabine and stavudine are possible candidates for the local and systemic treatment of adenoviral infection, and the anti-adenoviral effect might depend on the pharmacological properties of anti-HIV agents. The chemical properties on the clinical safety for systemic and local application need to be determined in order to for these drugs to be accepted for the treatment of adenovirus in clinical settings.

The impact of transmission clusters on primary drug resistance in newly diagnosed HIV-1 infection.

PubMed

Yerly, Sabine; Junier, Thomas; Gayet-Ageron, Angèle; Amari, Emmanuelle Boffi El; von Wyl, Viktor; Günthard, Huldrych F; Hirschel, Bernard; Zdobnov, Evgeny; Kaiser, Laurent

2009-07-17

To monitor HIV-1 transmitted drug resistance (TDR) in a well defined urban area with large access to antiretroviral therapy and to assess the potential source of infection of newly diagnosed HIV individuals. All individuals resident in Geneva, Switzerland, with a newly diagnosed HIV infection between 2000 and 2008 were screened for HIV resistance. An infection was considered as recent when the positive test followed a negative screening test within less than 1 year. Phylogenetic analyses were performed by using the maximum likelihood method on pol sequences including 1058 individuals with chronic infection living in Geneva. Of 637 individuals with newly diagnosed HIV infection, 20% had a recent infection. Mutations associated with resistance to at least one drug class were detected in 8.5% [nucleoside reverse transcriptase inhibitors (NRTIs), 6.3%; non-nucleoside reverse transcriptase inhibitors (NNRTIs), 3.5%; protease inhibitors, 1.9%]. TDR (P-trend = 0.015) and, in particular, NNRTI resistance (P = 0.002) increased from 2000 to 2008. Phylogenetic analyses revealed that 34.9% of newly diagnosed individuals, and 52.7% of those with recent infection were linked to transmission clusters. Clusters were more frequent in individuals with TDR than in those with sensitive strains (59.3 vs. 32.6%, respectively; P < 0.0001). Moreover, 84% of newly diagnosed individuals with TDR were part of clusters composed of only newly diagnosed individuals. Reconstruction of the HIV transmission networks using phylogenetic analysis shows that newly diagnosed HIV infections are a significant source of onward transmission, particularly of resistant strains, thus suggesting an important self-fueling mechanism for TDR.

Non-nucleoside reverse transcriptase inhibitor efavirenz increases monolayer permeability of human coronary artery endothelial cells

PubMed Central

Jamaluddin, Md Saha; Lin, Peter H.; Yao, Qizhi; Chen, Changyi

2009-01-01

Highly active antiretroviral therapy (HAART) is often associated with endothelial dysfunction and cardiovascular complications. In this study, we determined whether HIV non-nucleoside reverse transcriptase inhibitor efavirenz (EFV) could increase endothelial permeability. Human coronary artery endothelial cells (HCAECs) were treated with EFV (1, 5 and 10 µg/ml) and endothelial permeability was determined by a transwell system with a fluorescence-labeled dextran tracer. HCAECs treated with EFV showed a significant increase of endothelial permeability in a concentration-dependent manner. With real time PCR analysis, EFV significantly reduced the mRNA levels of tight junction proteins claudin-1, occludin, zonula occluden-1 and junctional adhesion molecule-1 compared with controls (P < 0.05). Protein levels of these tight junction molecules were also reduced substantially in the EFV-treated cells by western blot and flow cytometry analyses. In addition, EFV also increased superoxide anion production with dihydroethidium and cellular glutathione assays, while it decreased mitochondrial membrane potential with JC-staining. Antioxidants (ginkgolide B and MnTBAP) effectively blocked EFV-induced endothelial permeability and mitochondrial dysfunction. Furthermore, EFV increased the phosphorylation of MAPK JNK and IκBα, thereby increasing NFκB translocation to the nucleus. Chemical JNK inhibitor and dominant negative mutant JNK and IkBa adenoviruses effectively blocked the effects of EFV on HCAECs. Thus, EFV increases endothelial permeability which may be due to the decrease of tight junction proteins and the increase of superoxide anion. JNK and NFκB activation may be directly involved in the signal transduction pathway of EFV action in HCAECs. PMID:19674747

Prevalence of antiretroviral drug resistance and resistance-associated mutations in antiretroviral therapy-naïve HIV-infected individuals from 40 United States cities.

PubMed

Ross, Lisa; Lim, Michael L; Liao, Qiming; Wine, Brian; Rodriguez, Allan E; Weinberg, Winkler; Shaefer, Mark

2007-01-01

Transmission of drug-resistant HIV strains to antiretroviral therapy (ART)-naïve subjects can negatively impact therapy response. As treatment strategies and utilization of antiretroviral drugs evolve, patterns of transmitted mutations may shift. Paired genotypic and phenotypic susceptibility data were retrospectively analyzed for 317 ART-naïve, HIV-infected subjects from 40 small and major metropolitan cities in the Northeastern, Midwestern, Southern, Southwestern, and Northwestern United States during 2003. Using current (January 2007) PhenoSense cutoffs, HIV-from 8% of subjects had reduced susceptibility to > or = 1 drug. By class, < 1% had reduced susceptibility to protease inhibitors (PIs), and 1% had reduced susceptibility to nucleoside reverse transcriptase inhibitors (NRTIs); reduced susceptibility to > or = 1 non-nucleoside reverse transcriptase inhibitor (NNRTIs) was seen in 7% of subjects, with 4% of all subjects having reduced susceptibility to all NNRTIs. IAS-USA-defined NRTI, NNRTI, and/or major PI HIV-drug resistance-associated mutations were detected for 0% of the subjects. HIV risk factors included homosexual contact (74%), heterosexual contact (28%), and injectable drug use/transfusion/other (7%). Reduced susceptibility to > or = 1 drug was significantly higher (p = .034) for white subjects than African Americans and Hispanics/others. The high prevalence of drug resistance in these ART-naïve subjects suggests that transmitted resistance is occurring widely within the United States. HIV genotyping and/or phenotyping for antiretroviral-naïve patients seeking treatment should be considered, especially if the therapy will include an NNRTI.

Bayesian network analyses of resistance pathways against efavirenz and nevirapine

PubMed Central

Deforche, Koen; Camacho, Ricardo J.; Grossman, Zehave; Soares, Marcelo A.; Laethem, Kristel Van; Katzenstein, David A.; Harrigan, P. Richard; Kantor, Rami; Shafer, Robert; Vandamme, Anne-Mieke

2016-01-01

Objective To clarify the role of novel mutations selected by treatment with efavirenz or nevirapine, and investigate the influence of HIV-1 subtype on nonnucleoside reverse transcriptase inhibitor (nNRTI) resistance pathways. Design By finding direct dependencies between treatment-selected mutations, the involvement of these mutations as minor or major resistance mutations against efavirenz, nevirapine, or coadministrated nucleoside analogue reverse transcriptase inhibitors (NRTIs) is hypothesized. In addition, direct dependencies were investigated between treatment-selected mutations and polymorphisms, some of which are linked with subtype, and between NRTI and nNRTI resistance pathways. Methods Sequences from a large collaborative database of various subtypes were jointly analyzed to detect mutations selected by treatment. Using Bayesian network learning, direct dependencies were investigated between treatment-selected mutations, NRTI and nNRTI treatment history, and known NRTI resistance mutations. Results Several novel minor resistance mutations were found: 28K and 196R (for resistance against efavirenz), 101H and 138Q (nevirapine), and 31L (lamivudine). Robust interactions between NRTI mutations (65R, 74V, 75I/M, and 184V) and nNRTI resistance mutations (100I, 181C, 190E and 230L) may affect resistance development to particular treatment combinations. For example, an interaction between 65R and 181C predicts that the nevirapine and tenofovir and lamivudine/emtricitabine combination should be more prone to failure than efavirenz and tenofovir and lamivudine/emtricitabine. Conclusion Bayesian networks were helpful in untangling the selection of mutations by NRTI versus nNRTI treatment, and in discovering interactions between resistance mutations within and between these two classes of inhibitors. PMID:18832874

Prevalence of Transmitted HIV Drug Resistance Among Recently Infected Persons in San Diego, CA 1996-2013.

PubMed

Panichsillapakit, Theppharit; Smith, Davey M; Wertheim, Joel O; Richman, Douglas D; Little, Susan J; Mehta, Sanjay R

2016-02-01

Transmitted drug resistance (TDR) remains an important concern when initiating antiretroviral therapy (ART). Here, we describe the prevalence and phylogenetic relationships of TDR among ART-naive, HIV-infected individuals in San Diego from 1996 to 2013. Data were analyzed from 496 participants of the San Diego Primary Infection Cohort who underwent genotypic resistance testing before initiating therapy. Mutations associated with drug resistance were identified according to the WHO-2009 surveillance list. Network and phylogenetic analyses of the HIV-1 pol sequences were used to evaluate the relationships of TDR within the context of the entire cohort. The overall prevalence of TDR was 13.5% (67/496), with an increasing trend over the study period (P = 0.005). TDR was predominantly toward nonnucleoside reverse transcriptase inhibitors (NNRTIs) [8.5% (42/496)], also increasing over the study period (P = 0.005). By contrast, TDR to protease inhibitors and nucleos(t)ide reverse transcriptase inhibitors were 4.4% (22/496) and 3.8% (19/496), respectively, and did not vary with time. TDR prevalence did not differ by age, gender, race/ethnicity, or risk factors. Using phylogenetic analysis, we identified 52 transmission clusters, including 8 with at least 2 individuals sharing the same mutation, accounting for 23.8% (16/67) of the individuals with TDR. Between 1996 and 2013, the prevalence of TDR significantly increased among recently infected ART-naive individuals in San Diego. Around one-fourth of TDR occurred within clusters of recently infected individuals. These findings highlight the importance of baseline resistance testing to guide selection of ART and for public health monitoring.

Anti-human immunodeficiency virus (HIV) activities of halogenated gomisin J derivatives, new nonnucleoside inhibitors of HIV type 1 reverse transcriptase.

PubMed

Fujihashi, T; Hara, H; Sakata, T; Mori, K; Higuchi, H; Tanaka, A; Kaji, H; Kaji, A

1995-09-01

Halogenated gomisin J (a derivative of lignan compound), represented by the bromine derivative 1506 [(6R, 7S, S-biar)-4,9-dibromo-3,10-dihydroxy-1,2,11,12-tetramethoxy-6, 7-dimethyl-5,6,7,8- tetrahydrodibenzo[a,c]cyclo-octene], was found to be a potent inhibitor of the cytopathic effects of human immunodeficiency virus type 1 (HIV-1) on MT-4 human T cells (50% effective dose, 0.1 to 0.5 microM). Gomisin J derivatives were active in preventing p24 production from acutely HIV-1-infected H9 cells. The selective indices (toxic dose/effective dose) of these compounds were as high as > 300 in some systems. 1506 was active against 3'-azido-3'-deoxythymidine-resistant HIV-1 and acted synergistically with AZT and 2',3'-ddC. 1506 inhibited HIV-1 reverse transcriptase (RT) in vitro but not HIV-1 protease. From the time-of-addition experiment, 1506 was found to inhibit the early phase of the HIV life cycle. A 1506-resistant HIV mutant was selected and shown to possess a mutation within the RT-coding region (at position 188 [Tyr to Leu]). The mutant RT expressed in Escherichia coli was resistant to 1506 in the in vitro RT assay. Some of the HIV strains resistant to other nonnucleoside HIV-1 RT inhibitors were also resistant to 1506. Comparison of various gomisin J derivatives with gomisin J showed that iodine, bromine, and chlorine in the fourth and ninth positions increased RT inhibitory activity as well as cytoprotective activity.

Simultaneous Detection of Rift Valley Fever, Bluetongue, Rinderpest, and Peste des Petits Ruminants Viruses by a Single-Tube Multiplex Reverse Transcriptase-PCR Assay Using a Dual-Priming Oligonucleotide System▿

PubMed Central

Yeh, Jung-Yong; Lee, Ji-Hye; Seo, Hyun-Ji; Park, Jee-Yong; Moon, Jin-San; Cho, In-Soo; Choi, In-Soo; Park, Seung-Yong; Song, Chang-Seon; Lee, Joong-Bok

2011-01-01

The aim of this study was to develop a highly sensitive and specific one-step multiplex reverse transcriptase PCR assay for the simultaneous and differential detection of Rift Valley Fever virus (RVFV), bluetongue virus (BTV), rinderpest virus (RPV), and Peste des petits ruminants virus (PPRV). These viruses cause mucosal lesions in cattle, sheep, and goats, and they are difficult to differentiate from one another based solely on their clinical presentation in suspected disease cases. In this study, we developed a multiplex reverse transcriptase PCR to detect these viruses using a novel dual-priming oligonucleotide (DPO). The DPO contains two separate priming regions joined by a polydeoxyinosine linker, which blocks extension of nonspecifically primed templates and consistently allows high PCR specificity even under less-than-optimal PCR conditions. A total of 19 DPO primers were designed to detect and discriminate between RVFV, BTV, RPV, and PPRV by the generation of 205-, 440-, 115-, and 243-bp cDNA products, respectively. The multiplex reverse transcriptase PCR described here enables the early diagnosis of these four viruses and may also be useful as part of a testing regime for cattle, sheep, or goats exhibiting similar clinical signs, including mucosal lesions. PMID:21307219

In vitro resistance development for RO-0335, a novel diphenylether nonnucleoside reverse transcriptase inhibitor.

PubMed

Javanbakht, H; Ptak, R G; Chow, E; Yan, J M; Russell, J D; Mankowski, M K; Hogan, P A; Hogg, J H; Vora, H; Hang, J Q; Li, Y; Su, G; Paul, A; Cammack, N; Klumpp, K; Heilek, G

2010-05-01

Nonnucleoside reverse transcriptase inhibitors (NNRTIs) are important components of current combination therapies for the treatment of human immunodeficiency virus type 1 (HIV-1) infection. However, their low genetic barriers against resistance development, cross-resistance and serious side effects can compromise the benefits of the first generation compounds in this class (efavirenz and nevirapine). To study potential pathways leading to resistance against the novel diphenylether NNRTI, RO-0335, sequential passage experiments at low multiplicity of infection (MOI) were performed to solicit a stepwise selection of resistance mutations. Two pathways to loss of susceptibility to RO-0335 were observed, containing patterns of amino acid changes at either V106I/A plus F227C (with additional contributions from A98G, V108I, E138K, M230L and P236L) or V106I/Y188L (with a potential contribution from L100I, E138K and Y181C). Characterization of the observed mutations by site-directed mutagenesis in the isogenic HXB2D background demonstrated that a minimum of two or more mutations were required for significant loss of susceptibility, with the exception of Y188L, which requires a two-nucleotide change. Patterns containing F227C or quadruple mutations selected by RO-0335 showed a low relative fitness value when compared to wild-type HXB2D.

The design and synthesis of 9-phenylcyclohepta[d]pyrimidine-2,4-dione derivatives as potent non-nucleoside inhibitors of HIV reverse transcriptase.

PubMed

Wang, Xiaowei; Lou, Qinghua; Guo, Ying; Xu, Yang; Zhang, Zhili; Liu, Junyi

2006-09-07

Novel compounds, which can be considered as conformationally restricted analogues of MKC-442, have been synthesized and tested as inhibitors of the reverse transcriptase of human immunodeficiency virus type-1 (HIV-1). Reaction of urea with a beta-ketoester furnished 6,7,8,9-tetrahydro-9-phenyl-1H-cyclohepta[d]pyrimidine-2,4-(3H,5H)-dione (6a) and 6,7,8,9-tetrahydro-9-p-tolyl-1H-cyclohepta[d]pyrimidine-2,4-(3H,5H)-dione (6b) which were then alkylated at the N-1 position with chloromethyl ether, allyl bromide and benzyl bromide to afford the target compounds 7a-b, 8a-b, 9 and 10, respectively. The seven-membered, annelated compounds have a relatively rigid structures and can lock the orientation of the aromatic ring. Chemical modification at N-1 of the pyrinidine ring and the 9-phenyl ring was attempted, with the aim of improving the antiretroviral activity. In particular, replacement of the aliphatic group with the phenyl moiety at the terminus of N-1 side chain can enhance the activity. The most active compounds showed activity in the low micromolar range with IC50 values comparable to that of nevirapine. The biological activity results are in accordance with the docking results.

(PCG) Protein Crystal Growth HIV Reverse Transcriptase

NASA Technical Reports Server (NTRS)

1992-01-01

HIV Reverse Transcriptase crystals grown during the USML-1 (STS-50) mission using Commercial Refrigerator/Incubator Module (CR/IM) at 4 degrees C and the Vapor Diffusion Apparatus (VDA). Reverse transcriptase is an enzyme responsible for copying the nucleic acid genome of the AIDS virus from RNA to DNA. Studies indicated that the space-grown crystals were larger and better ordered (beyond 4 angstroms) than were comparable Earth-grown crystals. Principal Investigators were Charles Bugg and Larry DeLucas.

Risk factors for pre-term birth in a Canadian cohort of HIV-positive women: role of ritonavir boosting?

PubMed Central

Kakkar, Fatima; Boucoiran, Isabelle; Lamarre, Valerie; Ducruet, Thierry; Amre, Devendra; Soudeyns, Hugo; Lapointe, Normand; Boucher, Marc

2015-01-01

Background The risk of pre-term birth (PTB) associated with the use of protease inhibitors (PIs) during pregnancy remains a subject of debate. Recent data suggest that ritonavir boosting of PIs may play a specific role in the initiation of PTB, through an effect on the maternal–fetal adrenal axis. The primary objective of this study is to compare the risk of PTB among women treated with boosted PI versus non-boosted PIs during pregnancy. Methods Between 1988 and 2011, 705 HIV-positive women were enrolled into the Centre Maternel et Infantile sur le SIDA mother–infant cohort at Centre Hospitalier Universitaire Sainte-Justine in Montreal, Canada. Inclusion criteria for the study were: 1) attendance at a minimum of two antenatal obstetric visits and 2) singleton live birth, at 24 weeks gestational or older. The association between PTB (defined as delivery at <37 weeks gestational age), antiretroviral drug exposure and maternal risk factors was assessed retrospectively using logistic regression. Results A total of 525 mother–infant pairs were included in the analysis. Among them, PI-based combination anti-retroviral therapy was used in 37.4%, boosted PI based in 24.4%, non-nucleoside reverse transcriptase inhibitor (NNRTI) or nucleoside reverse transcriptase inhibitor based in 28.1%, and no treatment was given in 10.0% of cases. Overall, 13.5% of women experienced PTB. Among women treated with antiretroviral therapy, the risk of PTB was significantly higher among women who received boosted versus non-boosted PI (OR 2.01, 95% CI 1.02–3.97). This remained significant after adjusting for maternal age, delivery CD4 count, hepatitis C co-infection, history of previous PTB, and parity (aOR 2.17, 95% CI 1.05–4.51). There was no increased risk of PTB with the use of unboosted PIs as compared to NNRTI- or NRTI-based regimens. Conclusion While previous studies on the association between PTB and PI use have generally considered all PIs the same, our results would indicate a possible role of ritonavir boosting as a risk factor for PTB. Further work is needed to understand the pathophysiologic mechanisms involved, and to identify the safest ARV regimens to be used in pregnancy. PMID:26051165

The Discovery of Reverse Transcriptase.

PubMed

Coffin, John M; Fan, Hung

2016-09-29

In 1970 the independent and simultaneous discovery of reverse transcriptase in retroviruses (then RNA tumor viruses) by David Baltimore and Howard Temin revolutionized molecular biology and laid the foundations for retrovirology and cancer biology. In this historical review we describe the formulation of the controversial provirus hypothesis by Temin, which ultimately was proven by his discovery of reverse transcriptase in Rous sarcoma virus virions. Baltimore arrived at the same discovery through his studies on replication of RNA-containing viruses, starting with poliovirus and then moving to vesicular stomatitis virus, where he discovered a virion RNA polymerase. Subsequent studies of reverse transcriptase led to the elucidation of the mechanism of retrovirus replication, the discovery of oncogenes, the advent of molecular cloning, the search for human cancer viruses, and the discovery and treatment of HIV/AIDS.

Long-term effectiveness of initiating non-nucleoside reverse transcriptase inhibitor- versus ritonavir-boosted protease inhibitor-based antiretroviral therapy: implications for first-line therapy choice in resource-limited settings

PubMed Central

Lima, Viviane D; Hull, Mark; McVea, David; Chau, William; Harrigan, P Richard; Montaner, Julio SG

2016-01-01

Introduction In many resource-limited settings, combination antiretroviral therapy (cART) failure is diagnosed clinically or immunologically. As such, there is a high likelihood that patients may stay on a virologically failing regimen for a substantial period of time. Here, we compared the long-term impact of initiating non-nucleoside reverse transcriptase inhibitor (NNRTI)- versus boosted protease inhibitor (bPI)-based cART in British Columbia (BC), Canada. Methods We followed prospectively 3925 ART-naïve patients who started NNRTIs (N=1963, 50%) or bPIs (N=1962; 50%) from 1 January 2000 until 30 June 2013 in BC. At six months, we assessed whether patients virologically failed therapy (a plasma viral load (pVL) >50 copies/mL), and we stratified them based on the pVL at the time of failure ≤500 versus >500 copies/mL. We then followed these patients for another six months and calculated their probability of achieving subsequent viral suppression (pVL <50 copies/mL twice consecutively) and of developing drug resistance. These probabilities were adjusted for fixed and time-varying factors, including cART adherence. Results At six months, virologic failure rates were 9.5 and 14.3 cases per 100 person-months for NNRTI and bPI initiators, respectively. NNRTI initiators who failed with a pVL ≤500 copies/mL had a 16% higher probability of achieving subsequent suppression at 12 months than bPI initiators (0.81 (25th–75th percentile 0.75–0.83) vs. 0.72 (0.61–0.75)). However, if failing NNRTI initiators had a pVL >500 copies/mL, they had a 20% lower probability of suppressing at 12 months than pVL-matched bPI initiators (0.37 (0.29–0.45) vs. 0.46 (0.38–0.54)). In terms of evolving HIV drug resistance, those who failed on NNRTI performed worse than bPI in all scenarios, especially if they failed with a viral load >500 copies/mL. Conclusions Our results show that patients who virologically failed at six months on NNRTI and continued on the same regimen had a lower probability of subsequently achieving viral suppression and a higher chance of evolving HIV drug resistance. These results suggest that improving access to regular virologic monitoring is critically important, especially if NNRTI-based cART is to remain a preferred choice for first-line therapy in resource-limited settings. PMID:27499064

Impact of lopinavir-ritonavir exposure in HIV-1 infected children and adolescents in Madrid, Spain during 2000-2014.

PubMed

Rojas Sánchez, Patricia; Prieto, Luis; Jiménez De Ory, Santiago; Fernández Cooke, Elisa; Navarro, Maria Luisa; Ramos, José Tomas; Holguín, África

2017-01-01

The most-used protease-inhibitor in children is Lopinavir-ritonavir (LPV/r), which provides durable suppression of viral load and increases CD4+T-counts. This study describes the virological outcome of the HIV-1-infected paediatric population exposed to LPV/r during 15 years in Spain. Patients from the Madrid Cohort of HIV-1-infected-children and adolescents exposed to LPV/r as different line therapy during 2000-2014 were selected. The baseline epidemiological-clinical features, viral suppression, changes in CD4+T-CD8+T cell counts and drug susceptibility were recorded before and during LPV/r exposure. Drug resistance mutations (DRM) were identified in viruses from samples collected until 2011. We predicted drug susceptibility to 19 antiretrovirals among those carrying DRM using the Stanford's HIVdb Algorithm. A total of 199 (37.3%) of the 534 patients from the cohort were exposed to LPV/r during 2000-2014 in first (group 1), second (group 2) or more line-therapies (group 3). Patients were mainly Spaniards (81.9%), perinatally infected (96.5%) with subtype-B (65.3%) and HIV-diagnosed before year 2000 (67.8%). The mean age at first LPV/r exposure was 9.7 years. After protease-inhibitor exposure, viral suppression was higher in groups 1 and 2 than in group 3. Viral suppression occurred in 87.5%, 68.6% and 64.8% patients from groups 1, 2 and 3, respectively. Among the 64 patients with available resistance data during LPV/r treatment, 27(42.3%) carried DRM to protease-inhibitor, 28 (58.3%) to reverse-transcriptase-inhibitors and 21 (43.7%) to non-reverse-transcriptase-inhibitors. Darunavir/ritonavir, atazanavir-ritonavir and tipranavir/ritonavir presented the highest susceptibility and nelfinavir the lowest. A better lymphocyte recovering occurred when protease-inhibitor was taken as part of a first-line regimen and a higher number of patients reached viral suppression. The least compromised antiretrovirals for rescue antiretroviral regimens, according to DRM in the LPV/r-exposed-paediatric cohort, were mainly the new protease inhibitors.

Biotechnological applications of mobile group II introns and their reverse transcriptases: gene targeting, RNA-seq, and non-coding RNA analysis.

PubMed

Enyeart, Peter J; Mohr, Georg; Ellington, Andrew D; Lambowitz, Alan M

2014-01-13

Mobile group II introns are bacterial retrotransposons that combine the activities of an autocatalytic intron RNA (a ribozyme) and an intron-encoded reverse transcriptase to insert site-specifically into DNA. They recognize DNA target sites largely by base pairing of sequences within the intron RNA and achieve high DNA target specificity by using the ribozyme active site to couple correct base pairing to RNA-catalyzed intron integration. Algorithms have been developed to program the DNA target site specificity of several mobile group II introns, allowing them to be made into 'targetrons.' Targetrons function for gene targeting in a wide variety of bacteria and typically integrate at efficiencies high enough to be screened easily by colony PCR, without the need for selectable markers. Targetrons have found wide application in microbiological research, enabling gene targeting and genetic engineering of bacteria that had been intractable to other methods. Recently, a thermostable targetron has been developed for use in bacterial thermophiles, and new methods have been developed for using targetrons to position recombinase recognition sites, enabling large-scale genome-editing operations, such as deletions, inversions, insertions, and 'cut-and-pastes' (that is, translocation of large DNA segments), in a wide range of bacteria at high efficiency. Using targetrons in eukaryotes presents challenges due to the difficulties of nuclear localization and sub-optimal magnesium concentrations, although supplementation with magnesium can increase integration efficiency, and directed evolution is being employed to overcome these barriers. Finally, spurred by new methods for expressing group II intron reverse transcriptases that yield large amounts of highly active protein, thermostable group II intron reverse transcriptases from bacterial thermophiles are being used as research tools for a variety of applications, including qRT-PCR and next-generation RNA sequencing (RNA-seq). The high processivity and fidelity of group II intron reverse transcriptases along with their novel template-switching activity, which can directly link RNA-seq adaptor sequences to cDNAs during reverse transcription, open new approaches for RNA-seq and the identification and profiling of non-coding RNAs, with potentially wide applications in research and biotechnology.

Structural basis for the D-stereoselectivity of human DNA polymerase β

PubMed Central

Vyas, Rajan; Reed, Andrew J.; Raper, Austin T.; Zahurancik, Walter J.; Wallenmeyer, Petra C.

2017-01-01

Abstract Nucleoside reverse transcriptase inhibitors (NRTIs) with L-stereochemistry have long been an effective treatment for viral infections because of the strong D-stereoselectivity exhibited by human DNA polymerases relative to viral reverse transcriptases. The D-stereoselectivity of DNA polymerases has only recently been explored structurally and all three DNA polymerases studied to date have demonstrated unique stereochemical selection mechanisms. Here, we have solved structures of human DNA polymerase β (hPolβ), in complex with single-nucleotide gapped DNA and L-nucleotides and performed pre-steady-state kinetic analysis to determine the D-stereoselectivity mechanism of hPolβ. Beyond a similar 180° rotation of the L-nucleotide ribose ring seen in other studies, the pre-catalytic ternary crystal structures of hPolβ, DNA and L-dCTP or the triphosphate forms of antiviral drugs lamivudine ((-)3TC-TP) and emtricitabine ((-)FTC-TP) provide little structural evidence to suggest that hPolβ follows the previously characterized mechanisms of D-stereoselectivity. Instead, hPolβ discriminates against L-stereochemistry through accumulation of several active site rearrangements that lead to a decreased nucleotide binding affinity and incorporation rate. The two NRTIs escape some of the active site selection through the base and sugar modifications but are selected against through the inability of hPolβ to complete thumb domain closure. PMID:28402499

Enzyme engineering through evolution: thermostable recombinant group II intron reverse transcriptases provide new tools for RNA research and biotechnology.

PubMed

Collins, Kathleen; Nilsen, Timothy W

2013-08-01

Current investigation of RNA transcriptomes relies heavily on the use of retroviral reverse transcriptases. It is well known that these enzymes have many limitations because of their intrinsic properties. This commentary highlights the recent biochemical characterization of a new family of reverse transcriptases, those encoded by group II intron retrohoming elements. The novel properties of these enzymes endow them with the potential to revolutionize how we approach RNA analyses.

Molecular modeling studies of HIV-1 reverse transcriptase nonnucleoside inhibitors: total energy of complexation as a predictor of drug placement and activity.

PubMed Central

Kroeger Smith, M. B.; Rouzer, C. A.; Taneyhill, L. A.; Smith, N. A.; Hughes, S. H.; Boyer, P. L.; Janssen, P. A.; Moereels, H.; Koymans, L.; Arnold, E.

1995-01-01

Computer modeling studies have been carried out on three nonnucleoside inhibitors complexed with human immunodeficiency virus type 1 (HIV-1) reverse transcriptase (RT), using crystal coordinate data from a subset of the protein surrounding the binding pocket region. Results from the minimizations of solvated complexes of 2-cyclopropyl-4-methyl-5,11-dihydro-5H-dipyrido[3,2-b :2',3'-e][1,4] diazepin-6-one (nevirapine), alpha-anilino-2, 6-dibromophenylacetamide (alpha-APA), and 8-chloro-tetrahydro-imidazo(4,5,1-jk)(1,4)-benzodiazepin-2(1H)-thi one (TIBO) show that all three inhibitors maintain a very similar conformational shape, roughly overlay each other in the binding pocket, and appear to function as pi-electron donors to aromatic side-chain residues surrounding the pocket. However, side-chain residues adapt to each bound inhibitor in a highly specific manner, closing down around the surface of the drug to make tight van der Waals contacts. Consequently, the results from the calculated minimizations reveal that only when the inhibitors are modeled in a site constructed from coordinate data obtained from their particular RT complex can the calculated binding energies be relied upon to predict the correct orientation of the drug in the pocket. In the correct site, these binding energies correlate with EC50 values determined for all three inhibitors in our laboratory. Analysis of the components of the binding energy reveals that, for all three inhibitors, solvation of the drug is endothermic, but solvation of the protein is exothermic, and the sum favors complex formation. In general, the protein is energetically more stable and the drug less stable in their complexes as compared to the reactant conformations. For all three inhibitors, interaction with the protein in the complex is highly favorable. Interactions of the inhibitors with individual residues correlate with crystallographic and site-specific mutational data. pi-Stacking interactions are important in binding and correlate with drug HOMO RHF/6-31G* energies. Modeling results are discussed with respect to the mechanism of complex formation and the design of nonnucleoside inhibitors that will be more effective against mutants of HIV-1 RT that are resistant to the currently available drugs. PMID:8535257

The effect of transmitted HIV-1 drug resistance on pre-therapy viral load.

PubMed

Harrison, Linda; Castro, Hannah; Cane, Patricia; Pillay, Deenan; Booth, Clare; Phillips, Andrew; Geretti, Anna Maria; Dunn, David

2010-07-31

Reduced replication capacity of viruses expressing drug resistant mutations implies that patients with transmitted drug resistance (TDR) could have lower HIV RNA viral load than those infected with wild-type virus. We performed analysis using data from the UK HIV Drug Resistance Database and the UK CHIC study. Eligible patients had a resistance test performed between 1997 and 2007 while naive to antiretroviral therapy, were 16 years or older, and had a viral load and CD4 cell count measurement within 6 months of this test. Models were adjusted for CD4 cell count, viral subtype, ethnicity, risk group, sex, age, calendar year, clinical centre, and viral load assay. Of a total of 7994 patients included, 709 (9%) had TDR: 604 (85%) had resistance to one drug class only [350 nucleos(t)ide reverse transcriptase inhibitors (NRTIs), 164 non-nucleos(t)ide reverse transcriptase inhibitors (NNRTIs), 90 protease inhibitors (PIs)], 77 (11%) to two classes (42 NRTIs/NNRTIs, 31 NRTIs/PIs, 4 NNRTIs/PIs), and 28 (4%) had resistance to all three classes. The overall mean (SD) viral load at the time of resistance testing was 4.60 (0.82) log(10) copies/ml, and did not differ by class of TDR. However, patients harbouring M184V/I (n = 61) had a significantly lower viral load [adjusted mean difference -0.33 log10 copies/ml (95% CI -0.54 to -0.11), 53% lower (95% CI 22 to 71%), P = 0.002] compared to wild-type virus. Our study provides clear evidence of an in-vivo fitness cost associated with the M184V/I mutation independent of drug effects which select for this mutation. This was not observed for any other mutation, but true effects may have been obscured by reversion of initially resistant viruses to wild-type.

Suicide Inhibitors of Reverse Transcriptase in the Therapy of AIDS and Other Retroviruses

DTIC Science & Technology

1989-07-01

Acquired Immun Deficiency Syndrome (AIDS) (7-8). The most recent evaluation of the AIDS epidemic in the U.S. (4) indicates that the currer total of...are shown below. One of the first, [N-(L-3-tran carboxyxiran-2-carbonyl)-L-leucyl]-amido (4-guanido) butane was isolated from Asperg /II japonicus and...risk of acquired immune deficiency syndrome (AIDS). Science. 2.4: 497- 500 (1983). 9. Kopkrowski H., De Freitas E.C., Harper M.E., Woliheim S.M

Bauhinia variegata var. variegata trypsin inhibitor: From isolation to potential medicinal applications

DOE Office of Scientific and Technical Information (OSTI.GOV)

Fang, Evandro Fei; Wong, Jack Ho; Bah, Clara Shui Fern

Here we report for the first time of a new Kunitz-type trypsin inhibitor (termed BvvTI) from seeds of the Camel's foot tree, Bauhinia variegata var. variegata. BvvTI shares the same reactive site residues (Arg, Ser) and exhibits a homology of N-terminal amino acid sequence to other Bauhinia protease inhibitors. The trypsin inhibitory activity (K{sub i}, 0.1 x 10{sup -9} M) of BvvTI ranks the highest among them. Besides anti-HIV-1 reverse transcriptase activity, BvvTI could significantly inhibit the proliferation of nasopharyngeal cancer CNE-1 cells in a selective way. This may partially be contributed by its induction of cytokines and apoptotic bodies.more » These results unveil potential medicinal applications of BvvTI.« less

Bauhinia variegata var. variegata trypsin inhibitor: from isolation to potential medicinal applications.

PubMed

Fang, Evandro Fei; Wong, Jack Ho; Bah, Clara Shui Fern; Lin, Peng; Tsao, Sai Wah; Ng, Tzi Bun

2010-06-11

Here we report for the first time of a new Kunitz-type trypsin inhibitor (termed BvvTI) from seeds of the Camel's foot tree, Bauhinia variegata var. variegata. BvvTI shares the same reactive site residues (Arg, Ser) and exhibits a homology of N-terminal amino acid sequence to other Bauhinia protease inhibitors. The trypsin inhibitory activity (K(i), 0.1 x 10(-9)M) of BvvTI ranks the highest among them. Besides anti-HIV-1 reverse transcriptase activity, BvvTI could significantly inhibit the proliferation of nasopharyngeal cancer CNE-1 cells in a selective way. This may partially be contributed by its induction of cytokines and apoptotic bodies. These results unveil potential medicinal applications of BvvTI. (c) 2010 Elsevier Inc. All rights reserved.

Balancing Antiviral Potency and Host Toxicity: Identifying a Nucleotide Inhibitor with an Optimal Kinetic Phenotype for HIV-1 Reverse Transcriptase

PubMed Central

Sohl, Christal D.; Kasiviswanathan, Rajesh; Kim, Jiae; Pradere, Ugo; Schinazi, Raymond F.; Copeland, William C.; Mitsuya, Hiroaki; Baba, Masanori

2012-01-01

Two novel thymidine analogs, 3′-fluoro-3′-deoxythymidine (FLT) and 2′,3′-didehydro-3′-deoxy-4′-ethynylthymidine (Ed4T), have been investigated as nucleoside reverse transcriptase inhibitors (NRTIs) for treatment of HIV infection. Ed4T seems very promising in phase II clinical trials, whereas toxicity halted FLT development during this phase. To understand these different molecular mechanisms of toxicity, pre–steady-state kinetic studies were used to examine the interactions of FLT and Ed4T with wild-type (WT) human mitochondrial DNA polymerase γ (pol γ), which is often associated with NRTI toxicity, as well as the viral target protein, WT HIV-1 reverse transcriptase (RT). We report that Ed4T-triphosphate (TP) is the first analog to be preferred over native nucleotides by RT but to experience negligible incorporation by WT pol γ, with an ideal balance between high antiretroviral efficacy and minimal host toxicity. WT pol γ could discriminate Ed4T-TP from dTTP 12,000-fold better than RT, with only an 8.3-fold difference in discrimination being seen for FLT-TP. A structurally related NRTI, 2′,3′-didehydro-2′,3′-dideoxythymidine, is the only other analog favored by RT over native nucleotides, but it exhibits only a 13-fold difference (compared with 12,000-fold for Ed4T) in discrimination between the two enzymes. We propose that the 4′-ethynyl group of Ed4T serves as an enzyme selectivity moiety, critical for discernment between RT and WT pol γ. We also show that the pol γ mutation R964C, which predisposes patients to mitochondrial toxicity when receiving 2′,3′-didehydro-2′,3′-dideoxythymidine to treat HIV, produced some loss of discrimination for FLT-TP and Ed4T-TP. These molecular mechanisms of analog incorporation, which are critical for understanding pol γ-related toxicity, shed light on the unique toxicity profiles observed during clinical trials. PMID:22513406

Homodimerization of the p51 Subunit of HIV-1 Reverse Transcriptase

DOE Office of Scientific and Technical Information (OSTI.GOV)

Zheng, X.; Mueller, G; Cuneo, M

2010-01-01

The dimerization of HIV reverse transcriptase (RT), required to obtain the active form of the enzyme, is influenced by mutations, non-nucleoside reverse transcriptase inhibitors (NNRTIs), nucleotide substrates, Mg ions, temperature, and specifically designed dimerization inhibitors. In this study, we have utilized nuclear magnetic resonance (NMR) spectroscopy of the [methyl-{sup 13}C]methionine-labeled enzyme and small-angle X-ray scattering (SAXS) to investigate how several of these factors influence the dimerization behavior of the p51 subunit. The {sup 1}H-{sup 13}C HSQC spectrum of p51 obtained at micromolar concentrations indicates that a significant fraction of the p51 adopts a 'p66-like' conformation. SAXS data obtained for p51more » samples were used to determine the fractions of monomer and dimer in the sample and to evaluate the conformation of the fingers/thumb subdomain. All of the p51 monomer observed was found to adopt the compact, 'p51C' conformation observed for the p51 subunit in the RT heterodimer. The NMR and SAXS data indicate that the p51 homodimer adopts a structure that is similar to the p66/p51 heterodimer, with one p51C subunit and a second p51 subunit in an extended, 'p51E' conformation that resembles the p66 subunit of the heterodimer. The fractional dimer concentration and the fingers/thumb orientation are found to depend strongly on the experimental conditions and exhibit a qualitative dependence on nevirapine and ionic strength (KCl) that is similar to the behavior reported for the heterodimer and the p66 homodimer. The L289K mutation interferes with p51 homodimer formation as it does with formation of the heterodimer, despite its location far from the dimer interface. This effect is readily interpreted in terms of a conformational selection model, in which p51{sub L289K} has a much greater preference for the compact, p51C conformation. A reduced level of dimer formation then results from the reduced ratio of the p51E{sub L289K} to p51C{sub L289K} monomers.« less

Evidence for retrovirus infections in green turtles Chelonia mydas from the Hawaiian islands

USGS Publications Warehouse

Casey, R.N.; Quackenbush, S.L.; Work, Thierry M.; Balazs, G.H.; Bowser, P.R.; Casey, J.W.

1997-01-01

Apparently normal Hawaiian green turtles Chelonia mydas and those displaying fibropapillomas were analyzed for infection by retroviruses. Strikingly, all samples were positive for polymerase enhanced reverse transcriptase (PERT) with levels high enough to quantitate by the conventional reverse transcriptase (RT) assay. However, samples of skin, even from asymptomatic turtles, were RT positive, although the levels of enzyme activity in healthy turtles hatched and raised in captivity were much lower than those observed in asymptomatic free-ranging turtles. Turtles with fibropapillomas displayed a broad range of reverse transcriptase activity. Skin and eye fibropapillomas and a heart tumor were further analyzed and shown to have reverse transcriptase activity that banded in a sucrose gradient at 1.17 g ml-1. The reverse transcriptase activity purified from the heart tumor displayed a temperature optimum of 37??C and showed a preference for Mn2+ over Mg2+. Sucrose gradient fractions of this sample displaying elevated reverse transcriptase activity contained primarily retrovitalsized particles with prominent envelope spikes, when negatively stained and examined by electron microscopy. Sodium dodecylsulfate-polyacrylamide gel electrophoresis (SDS-PAGE) analysis of gradient-purified virions revealed a conserved profile among 4 independent tumors and showed 7 prominent proteins having molecular weights of 116, 83, 51, 43, 40, 20 and 14 kDa. The data suggest that retroviral infections are widespread in Hawaiian green turtles and a comprehensive investigation is warranted to address the possibility that these agents cause green turtle fibropapillomatosis (GTFP).

Molecular dynamics and Monte Carlo simulations for protein-ligand binding and inhibitor design.

PubMed

Cole, Daniel J; Tirado-Rives, Julian; Jorgensen, William L

2015-05-01

Non-nucleoside inhibitors of HIV reverse transcriptase are an important component of treatment against HIV infection. Novel inhibitors are sought that increase potency against variants that contain the Tyr181Cys mutation. Molecular dynamics based free energy perturbation simulations have been run to study factors that contribute to protein-ligand binding, and the results are compared with those from previous Monte Carlo based simulations and activity data. Predictions of protein-ligand binding modes are very consistent for the two simulation methods; the accord is attributed to the use of an enhanced sampling protocol. The Tyr181Cys binding pocket supports large, hydrophobic substituents, which is in good agreement with experiment. Although some discrepancies exist between the results of the two simulation methods and experiment, free energy perturbation simulations can be used to rapidly test small molecules for gains in binding affinity. Free energy perturbation methods show promise in providing fast, reliable and accurate data that can be used to complement experiment in lead optimization projects. This article is part of a Special Issue entitled "Recent developments of molecular dynamics". Copyright © 2014 Elsevier B.V. All rights reserved.

Base Preferences in Non-Templated Nucleotide Incorporation by MMLV-Derived Reverse Transcriptases

PubMed Central

Zajac, Pawel; Islam, Saiful; Hochgerner, Hannah; Lönnerberg, Peter; Linnarsson, Sten

2013-01-01

Reverse transcriptases derived from Moloney Murine Leukemia Virus (MMLV) have an intrinsic terminal transferase activity, which causes the addition of a few non-templated nucleotides at the 3´ end of cDNA, with a preference for cytosine. This mechanism can be exploited to make the reverse transcriptase switch template from the RNA molecule to a secondary oligonucleotide during first-strand cDNA synthesis, and thereby to introduce arbitrary barcode or adaptor sequences in the cDNA. Because the mechanism is relatively efficient and occurs in a single reaction, it has recently found use in several protocols for single-cell RNA sequencing. However, the base preference of the terminal transferase activity is not known in detail, which may lead to inefficiencies in template switching when starting from tiny amounts of mRNA. Here, we used fully degenerate oligos to determine the exact base preference at the template switching site up to a distance of ten nucleotides. We found a strong preference for guanosine at the first non-templated nucleotide, with a greatly reduced bias at progressively more distant positions. Based on this result, and a number of careful optimizations, we report conditions for efficient template switching for cDNA amplification from single cells. PMID:24392002

Indolyl aryl sulfones as HIV-1 non-nucleoside reverse transcriptase inhibitors: role of two halogen atoms at the indole ring in developing new analogues with improved antiviral activity.

PubMed

Regina, Giuseppe La; Coluccia, Antonio; Piscitelli, Francesco; Bergamini, Alberto; Sinistro, Anna; Cavazza, Antonella; Maga, Giovanni; Samuele, Alberta; Zanoli, Samantha; Novellino, Ettore; Artico, Marino; Silvestri, Romano

2007-10-04

Indolyl aryl sulfones bearing the 4,5-difluoro (10) or 5-chloro-4-fluoro (16) substitution pattern at the indole ring were potent inhibitors of HIV-1 WT and the NNRTI-resistant strains Y181C and K103N-Y181C. These compounds were highly effective against the 112 and the AB1 strains in lymphocytes and inhibited at nanomolar concentration the multiplication of the IIIBBa-L strain in macrophages. Compound 16 was exceptionally potent against RT WT and RTs carrying the K103N, Y181I, and L100I mutations.

Murine Leukemia Virus Reverse Transcriptase: Structural Comparison with HIV-1 Reverse Transcriptase

PubMed Central

Coté, Marie L.; Roth, Monica J.

2008-01-01

Recent X-ray crystal structure determinations of Moloney murine leukemia virus reverse transcriptase (MoMLV RT) have allowed for more accurate structure/function comparisons to HIV-1 RT than were formerly possible. Previous biochemical studies of MoMLV RT in conjunction with knowledge of sequence homologies to HIV-1 RT and overall fold similarities to RTs in general, provided a foundation upon which to build. In addition, numerous crystal structures of the MoMLV RT fingers/palm subdomain had also shed light on one of the critical functions of the enzyme, specifically polymerization. Now in the advent of new structural information, more intricate examination of MoMLV RT in its entirety can be realized, and thus the comparisons with HIV-1 RT may be more critically elucidated. Here, we will review the similarities and differences between MoMLV RT and HIV-1 RT via structural analysis, and propose working models for the MoMLV RT based upon that information. PMID:18294720

Transmitted drug resistance and type of infection in newly diagnosed HIV-1 individuals in Honduras.

PubMed

Murillo, Wendy; Paz-Bailey, Gabriela; Morales, Sonia; Monterroso, Edgar; Paredes, Mayte; Dobbs, Trudy; Parekh, Bharat S; Albert, Jan; Rivera, Ivette Lorenzana de

2010-12-01

Transmitted drug resistance (TDR) reduces the efficacy of antiretroviral treatment and is a public health concern. To gain insight in the epidemiology of TDR in Honduras by evaluating the amount of TDR in a representative sample of newly diagnosed individuals and by determining whether these are recent or established infections. Two hundred treatment-naïve, newly diagnosed HIV-positive individuals representing different population groups (general population, Garifunas ethnic group, female sex workers and men who have sex with men) and different geographic regions were enrolled during April 2004-April 2007. The HIV-1 pol gene was sequenced to identify drug-resistant mutations and TDR was scored as recommended by the WHO. Specimens were classified as recent or established infections using the BED assay. Among 200 samples analyzed from Honduran patients the prevalence of TDR was 7% (95% CI: 3.9-11.5%), 5% for non-nucleoside reverse transcriptase inhibitors (NNRTIs), 3% for nucleoside reverse transcriptase inhibitors (NRTIs) and 0.5% for protease inhibitors (PIs). Testing of these samples with the BED assay revealed that 12% of the specimens were associated with recent infections. TDR was significantly more common in specimens with recent infection (21%) than established infection (5%) (p=0.016). The prevalence of TDR in Honduras was moderate (7%). The percentage of specimens who were recently infected was low (12%), suggesting that late HIV diagnosis is common. The TDR prevalence was higher in recent than in established infections, which may indicate that TDR is increasing over time. The higher prevalence of NNRTI and NRTI mutations as compared to PI mutations is probably due to a broader and longer use of these drugs in Honduras. Copyright © 2010 Elsevier B.V. All rights reserved.

Efficacy and safety of etravirine (TMC125) in patients with highly resistant HIV-1: primary 24-week analysis.

PubMed

Nadler, Jeffrey P; Berger, Daniel S; Blick, Gary; Cimoch, Paul J; Cohen, Calvin J; Greenberg, Richard N; Hicks, Charles B; Hoetelmans, Richard M W; Iveson, Kathy J; Jayaweera, Dushyantha S; Mills, Anthony M; Peeters, Monika P; Ruane, Peter J; Shalit, Peter; Schrader, Shannon R; Smith, Stephen M; Steinhart, Corklin R; Thompson, Melanie; Vingerhoets, Johan H; Voorspoels, Ellen; Ward, Douglas; Woodfall, Brian

2007-03-30

TMC125-C223 is an open-label, partially blinded, randomized clinical trial to evaluate the efficacy and safety of two dosages of etravirine (TMC125), a non-nucleoside reverse transcriptase inhibitor (NNRTI) with activity against wild-type and NNRTI-resistant HIV-1. A total of 199 patients were randomly assigned 2: 2: 1 to twice-daily etravirine 400 mg, 800 mg and control groups, respectively. The primary endpoint was a change in viral load from baseline at week 24 in the intention-to-treat population. Patients had HIV-1 with genotypic resistance to approved NNRTIs and at least three primary protease inhibitor (PI) mutations. Etravirine groups received an optimized background of at least two approved antiretroviral agents [nucleoside reverse transcriptase inhibitors (NRTI) and/or lopinavir/ritonavir and/or enfuvirtide]. Control patients received optimized regimens of at least three antiretroviral agents (NRTIs or PIs and/or enfuvirtide). The mean change from baseline in HIV-1 RNA at week 24 was -1.04, -1.18 and -0.19 log10 copies/ml for etravirine 400 mg twice a day, 800 mg twice a day and the control group, respectively (P < 0.05 for both etravirine groups versus control). Etravirine showed no dose-related effects on safety and tolerability. No consistent pattern of neuropsychiatric symptoms was observed. There were few hepatic adverse events, and rashes were predominantly early onset and mild to moderate in severity. Etravirine plus an optimized background significantly reduced HIV-1-RNA levels from baseline after 24 weeks in patients with substantial NNRTI and PI resistance, and demonstrated a favorable safety profile compared with control.

Safety of Perinatal Exposure to Antiretroviral Medications: Developmental Outcomes in Infants

PubMed Central

Sirois, Patricia A.; Huo, Yanling; Williams, Paige L.; Malee, Kathleen; Garvie, Patricia A.; Kammerer, Betsy; Rich, Kenneth; Van Dyke, Russell B.; Nozyce, Molly L.

2013-01-01

Background This study evaluated effects of perinatal exposure to antiretroviral (ARV) medications on neurodevelopment of HIV-exposed, uninfected infants. Methods HIV-exposed, uninfected infants (age 9-15 months) enrolled in SMARTT, a multisite prospective surveillance study, completed the Bayley Scales of Infant and Toddler Development—Third Edition (Bayley-III), assessing cognition, language, motor skills, social-emotional development, and adaptive behavior. Linear regression models were used to evaluate associations between Bayley-III outcomes in infants with and without perinatal and neonatal ARV exposure, by regimen (combination ARV [cARV] versus non-cARV), type of regimen (defined by drug class), and individual ARVs (for infants with cARV exposure), adjusting for maternal and infant health and demographic covariates. Results As of May 2010, 374 infants had valid Bayley-III evaluations. Median age at testing was 12.7 months; 49% male, 79% black, 16% Hispanic. Seventy-nine percent were exposed to regimens containing protease inhibitors (PIs; 9% of PI-containing regimens also included non-nucleoside reverse transcriptase inhibitors [NNRTIs]), 5% to regimens containing NNRTIs (without PI), and 14% to regimens containing only nucleoside reverse transcriptase inhibitors (NRTIs). Overall, 83% were exposed to cARV. No Bayley-III outcome was significantly associated with overall exposure to cARV, ARV regimen, or neonatal prophylaxis. For individual ARVs, following sensitivity analyses, the adjusted group mean on the Language domain was within age expectations but significantly lower for infants with perinatal exposure to atazanavir (p=0.01). Conclusions These results support the safety of perinatal ARV use. Continued monitoring for adverse neurodevelopmental outcomes in older children is warranted, and the safety of atazanavir merits further study. PMID:23340561

Lipoprotein Changes in HIV-Infected Antiretroviral-Naïve Individuals after Starting Antiretroviral Therapy: ACTG Study A5152s Stein: Lipoprotein Changes on Antiretroviral Therapy

PubMed Central

Stein, James H.; Komarow, Lauren; Cotter, Bruno R.; Currier, Judith S.; Dubé, Michael P.; Fichtenbaum, Carl J.; Gerschenson, Mariana; Mitchell, Carol K.C.; Murphy, Robert L.; Squires, Kathleen; Parker, Robert A.; Torriani, Francesca J.

2008-01-01

Background Dyslipidemia is a frequent complication of antiretroviral therapy (ART) for patients with human immunodeficiency virus infection (HIV). The effects of ART on lipoproteins are less well-understood, and have not been investigated in a prospective study where assignment to ART is randomized. Objective To evaluate the effects of three class-sparing ART regimens on lipids and lipoproteins. Methods This was a substudy of a prospective, multicenter study treatment-naïve HIV-infected individuals randomly assigned to receive a regimen of nucleoside reverse transcriptase inhibitors (NRTIs) + the non-nucleoside reverse transcriptase inhibitor efavirenz, NRTIs + the protease inhibitor lopinavir/ritonavir, or a NRTI-sparing regimen of efavirenz + lopinavir/ritonavir. Lipoproteins were measured by nuclear magnetic resonance spectroscopy. Results Among the 82 participants, total and small low-density lipoprotein concentrations increased (median, interquartile range) by 152 (-49 - +407, p<0.01) and 130 (-98 - +417, p<0.01) nmol/L, respectively, especially in the arms containing lopinavir/ritonavir (pKW<0.04). Very low-density lipoproteins also increased (p<0.01), with a larger increase in the arms that contained lopinavir/ritonavir (p=0.022). High-density lipoproteins increased by 6.0 nmol/L (2.8 - 10.4, p<0.01), but differences between arms were not significant (pKW=0.069). Changes were not related to changes in markers of insulin/glucose metabolism. Conclusions Total and small low-density lipoprotein concentrations increased, especially in the arms containing lopinavir/ritonavir, as did increases in total very low-density lipoproteins. Adverse changes were especially prominent in the arm with efavirenz + lopinavir/ritonavir. PMID:19956354

First trimester maternal uterine artery Doppler examination in HIV-positive women.

PubMed

Savvidou, M D; Samuel, M I; Akolekar, R; Poulton, M; Nicolaides, K H

2011-11-01

The aim of the current study was to assess the effect of maternal HIV infection, treated or untreated, on the degree of placental invasion, as assessed by the pulsatility index of the uterine arteries during a Doppler examination at 11(+0) -13(+6) weeks' gestation. This was a nested case-control study in which a uterine artery Doppler examination was performed in the first trimester in 76 HIV-positive women. Each woman was matched with 30 HIV-negative women. As the pulsatility index of the uterine arteries depends on a number of maternal and fetal characteristics, its values in each case and control were expressed as multiples of the median (MoM) of the unaffected group. Among the 76 HIV-positive women, 33 (43.4%) were on antiretroviral treatment at the time of the Doppler examination, including 14 women (42.4%) on nucleoside reverse transcriptase inhibitors (NRTIs) and a protease inhibitor, 18 women (54.5%) on NRTIs and a nonnucleoside reverse transcriptase inhibitor and one woman (3.1%) on monotherapy. Compared with the HIV-negative women, the HIV-positive women were more likely to be heavier (P<0.01), to be of African origin (P<0.01), to be nonsmokers (P=0.01) and to deliver smaller neonates earlier (P<0.01). The median adjusted pulsatility index of the uterine arteries was not statistically different between the cases and controls [1.07; interquartile range (IQR) 0.85-1.24 MoM vs. 0.99; IQR 0.81-1.20 MoM; P= 0.28] or, in HIV-positive women, between those receiving and not receiving antiretroviral treatment (P=0.12). HIV-positive women with uncomplicated pregnancies have normal placental perfusion in the first trimester of pregnancy. 2011 British HIV Association.

Prevalence of drug resistance and importance of viral load measurements in Honduran HIV-infected patients failing antiretroviral treatment.

PubMed

Murillo, Wendy; de Rivera, I L; Parham, L; Jovel, E; Palou, E; Karlsson, A C; Albert, J

2010-02-01

The Honduran HIV/AIDS Program began to scale up access to HIV therapy in 2002. Up to May 2008, more than 6000 patients received combination antiretroviral therapy (cART). As HIV drug resistance is the major obstacle for effective treatment, the purpose of this study was to assess the prevalence of antiretroviral drug resistance in Honduran HIV-1-infected individuals. We collected samples from 138 individuals (97 adults and 41 children) on cART with virological, immunological or clinical signs of treatment failure. HIV-1 pol sequences were obtained using an in-house method. Resistance mutations were identified according to the 2007 International AIDS Society (IAS)-USA list and predicted susceptibility to cART was scored using the ANRS algorithm. Resistance mutations were detected in 112 patients (81%), 74% in adults and 98% in children. Triple-, dual- and single-class drug resistance was documented in 27%, 43% and 11% of the study subjects, respectively. Multiple logistic regression showed that resistance was independently associated with type of treatment failure [virological failure (odds ratio (OR) = 1) vs. immunological failure (OR = 0.11; 95% confidence interval (CI) 0.030-0.43) vs. clinical failure (OR = 0.037; 95% CI 0.0063-0.22)], route of transmission (OR = 42.8; 95% CI 3.73-491), and years on therapy (OR = 1.81; 95% CI 1.11-2.93). The prevalence of antiretroviral resistance was high in Honduran HIV-infected patients with signs of treatment failure. A majority of study subjects showed dual- or triple-class resistance to nucleoside reverse transcriptase inhibitors, nonnucleoside reverse transcriptase inhibitors and protease inhibitors. Virologically defined treatment failure was a strong predictor of resistance, indicating that viral load testing is needed to correctly identify patients with treatment failure attributable to resistance.

Prevalence of HIV drug resistance mutation in the northern Indian population after failure of the first line antiretroviral therapy.

PubMed

Sinha, S; Shekhar, R C; Ahmad, H; Kumar, N; Samantaray, J C; Sreenivas, V; Khan, N H; Mitsuyasu, R T

2012-09-01

There is limited information available about the prevalence and pattern of human immunodeficiency virus (HIV) drug resistance mutations (DRMs) among antiretroviral therapy (ART) experienced patients from northern India. Results of genotypic drug resistance testing were obtained from plasma samples of 128 patients, who had presented with clinical or immunological failure to treatment after at least six months of ART. Major DRMs associated with any of the three classes of antiretroviral (ARV) drugs, nucleoside reverse transcriptase inhibitors (NRTI), non-nucleoside reverse transcriptase inhibitors (NNRTI) and protease inhibitors (PI), were seen in 120 out of 128 patients (93.8% prevalence). NRTI and NNRTI DRMs were each seen in 115/128 (89.8%) patients, with M184V, M41L, D67N and T215Y being the most frequent among NRTI associated mutations, and K103N, G190A, Y181C and A98G among NNRTI associated ones. PI DRMs were observed in 14/128 (10.9%) patients, with L10I, V82A and L89V being the commonest. These results present a high prevalence of DRMs among ART experienced patients from northern India with clinical or immunological failure of therapy. It emphasizes the need for regular testing of plasma samples of such patients for DRMs in order to detect and replace a failing regimen early, and also the use of HIV drug resistance genotyping of ART naive individuals prior to initiating first line ART for possible transmitted resistance. It is very important to enhance the access of patients to ARV drugs so that their compliance could be improved and hence development of DRMs be minimized.

Antiretroviral drug use in a cross-sectional population survey in Africa: NIMH Project Accept (HPTN 043)

PubMed Central

Fogel, Jessica M.; Clarke, William; Kulich, Michal; Piwowar-Manning, Estelle; Breaud, Autumn; Olson, Matthew T.; Marzinke, Mark A.; Laeyendecker, Oliver; Fiamma, Agnès; Donnell, Deborah; Mbwambo, Jessie K. K.; Richter, Linda; Gray, Glenda; Sweat, Michael; Coates, Thomas J.; Eshleman, Susan H.

2016-01-01

Background Antiretroviral (ARV) drug treatment benefits the treated individual and can prevent HIV transmission. We assessed ARV drug use in a community-randomized trial that evaluated the impact of behavioral interventions on HIV incidence. Methods Samples were collected in a cross-sectional survey after a 3-year intervention period. ARV drug testing was performed using samples from HIV-infected adults at four study sites (Zimbabwe; Tanzania; KwaZulu-Natal and Soweto, South Africa; survey period 2009–2011), using an assay that detects 20 ARV drugs (6 nucleoside/nucleotide reverse transcriptase inhibitors [NRTIs]; 3 non-nucleoside reverse transcriptase inhibitors [NNRTIs]; 9 protease inhibitors; maraviroc; raltegravir). Results ARV drugs were detected in 2,011 (27.4%) of 7,347 samples; 88.1% had 1 NNRTI +/− 1–2 NRTIs. ARV drug detection was associated with sex (women>men), pregnancy, older age (>24 years), and study site (p<0.0001 for all four variables). ARV drugs were also more frequently detected in adults who were widowed (p=0.006) or unemployed (p=0.02). ARV drug use was more frequent in intervention versus control communities early in the survey (p=0.01), with a significant increase in control (p=0.004) but not in intervention communities during the survey period. In KwaZulu-Natal, a 1% increase in ARV drug use was associated with a 0.14% absolute decrease in HIV incidence (p=0.018). Conclusions This study used an objective, biomedical approach to assess ARV drug use on a population level. This analysis identified factors associated with ARV drug use and provided information on ARV drug use over time. ARV drug use was associated with lower HIV incidence at one study site. PMID:27828875

Non-nucleoside reverse transcriptase inhibitors (NNRTIs), their discovery, development, and use in the treatment of HIV-1 infection: a review of the last 20 years (1989-2009).

PubMed

de Béthune, Marie-Pierre

2010-01-01

It is almost 20 years since NNRTIs were identified as a new class of antiretroviral drugs for the treatment of HIV-1 infection. Although they belong to different and diverse chemical families, they share a common and unique mechanism of action: their interaction with HIV-1 reverse transcriptase induces conformational changes that inhibit the catalytic activities of the enzyme. They are characterized by their specificity for HIV-1, which makes them very selective inhibitors of the virus. First generation NNRTIs nevirapine and efavirenz, in combination with other antiretroviral drugs, have become a cornerstone for the treatment of HIV-1 infection, in patients initiating antiretroviral therapy. Further research has led to the discovery and development of next generation NNRTIs with an increased genetic barrier to the development of resistance. Etravirine is the first NNRTI to show sustained virologic efficacy in patients with NNRTI resistant HIV-1. This review covers the NNRTI class of anti-HIV-1 drugs, from the initial discovery of the class in 1990 to the current compounds in clinical development, i.e. around 20 years of research and development efforts. It describes the characteristics of the NNRTIs, their mechanisms of action, HIV-1 resistance to the inhibitors, and the drugs that have been approved for the treatment of HIV-1 infection, or are currently in clinical development. The role of NNRTIs in prevention of HIV transmission is also addressed. This article forms part of a special issue of Antiviral Research marking the 25th anniversary of antiretroviral drug discovery and development, vol. 85, issue 1, 2010. Copyright 2009 Elsevier B.V. All rights reserved.

Curious discoveries in antiviral drug development: the role of serendipity.

PubMed

De Clercq, Erik

2015-07-01

Antiviral drug development has often followed a curious meandrous route, guided by serendipity rather than rationality. This will be illustrated by ten examples. The polyanionic compounds (i) polyethylene alanine (PEA) and (ii) suramin were designed as an antiviral agent (PEA) or known as an antitrypanosomal agent (suramin), before they emerged as, respectively, a depilatory agent, or reverse transcriptase inhibitor. The 2',3'-dideoxynucleosides (ddNs analogues) (iii) have been (and are still) used in the "Sanger" DNA sequencing technique, although they are now commercialized as nucleoside reverse transcriptase inhibitors (NRTIs) in the treatment of HIV infections. (E)-5-(2-Bromovinyl)-2'-deoxyuridine (iv) was discovered as a selective anti-herpes simplex virus compound and is now primarily used for the treatment of varicella-zoster virus infections. The prototype of the acyclic nucleoside phosphonates (ANPs), (S)-9-(3-hydroxy-2-phosphonylmethoxypropyl)adenine [(S)-HPMPA], (v) was never commercialized, although it gave rise to several marketed products (cidofovir, adefovir, and tenofovir). 1-[2-(Hydroxyethoxy)methyl]-6-(phenylthio)thymine (vi) and TIBO (tetrahydroimidazo[4,5,1-jk][1,4-benzodiazepin-2(1H)]-one and -thione) (vii) paved the way to a number of compounds (i.e., nevirapine, delavirdine, etravirine, and rilpivirine), which are now collectively called non-NRTIs. The bicyclam AMD3100 (viii) was originally described as an anti-HIV agent before it became later marketed as a stem cell mobilizer. The S-adenosylhomocysteine hydrolase inhibitors (ix), while active against a broad range of (-)RNA viruses and poxviruses may be particularly effective against Ebola virus, and for (x) the O-ANP derivatives, the potential application range encompasses virtually all DNA viruses. © 2015 Wiley Periodicals, Inc.

Prevalence and patterns of HIV transmitted drug resistance in Guatemala.

PubMed

Avila-Ríos, Santiago; Mejía-Villatoro, Carlos R; García-Morales, Claudia; Soto-Nava, Maribel; Escobar, Ingrid; Mendizabal, Ricardo; Girón, Amalia; García, Leticia; Reyes-Terán, Gustavo

2011-12-01

To assess human immunodeficiency virus (HIV) diversity and the prevalence of transmitted drug resistance (TDR) in Guatemala. One hundred forty-five antiretroviral treatment-naïve patients referred to the Roosevelt Hospital in Guatemala City were enrolled from October 2010 to March 2011. Plasma HIV pol sequences were obtained and TDR was assessed with the Stanford algorithm and the World Health Organization (WHO) TDR surveillance mutation list. HIV subtype B was highly prevalent in Guatemala (96.6%, 140/145), and a 2.8% (4/145) prevalence of BF1 recombinants and 0.7% (1/145) prevalence of subtype C viruses were found. TDR prevalence for the study period was 8.3% (12/145) with the Stanford database algorithm (score > 15) and the WHO TDR surveillance mutation list. Most TDR cases were associated with non-nucleoside reverse transcriptase inhibitors (NNRTIs) (83.3%, 10/12); a low prevalence of nucleoside reverse transcriptase inhibitors and protease inhibitors was observed in the cohort (< 1% for both families). Low selection of antiretroviral drug resistance mutations was found, except for NNRTI-associated mutations. Major NNRTI mutations such as K101E, K103N, and E138K showed higher frequencies than expected in ART-naïve populations. Higher literacy was associated with a greater risk of TDR (odds ratio 4.14, P = 0.0264). This study represents one of the first efforts to describe HIV diversity and TDR prevalence and trends in Guatemala. TDR prevalence in Guatemala was at the intermediate level. Most TDR cases were associated with NNRTIs. Further and continuous TDR surveillance is necessary to gain more indepth knowledge about TDR spread and trends in Guatemala and to optimize treatment outcomes in the country.

[Recommendations for non-occupational postexposure HIV prophylaxis. Spanish Working Group on Non-Occupational Postexposure HIV Prophylaxis of the Catalonian Center for Epidemiological Studies on AIDS and the AIDS Study Group].

PubMed

Almeda, Jesús; Casabona, Jordi; Allepuz, Alejandro; García-Alcaide, Felipe; del Romero, Jorge; Tural, Cristina; Colm, Joan; Bolao, Ferrán; Campins, Magda; Domínguez, Angela; Force, Lluís; Giménez, Albert; Guerra-Romero, Luis

2002-10-01

Evidence is lacking on the possible efficacy and effectiveness of non-occupational postexposure prophylaxis (PEP). However, because of its biological plausibility, the use of antiretroviral (ARV) drugs to prevent the development of infection in certain cases of accidental or sporadic exposure has begun to be considered as common clinical practice. Previous studies performed in Spain have demonstrated both the demand and the prescription of ARV as PEP and especially the diversity and inconsistency in the criteria used. In this context, in April of 2000 the Centre for Epidemiological Studies on AIDS of Catalonia (CEESCAT) (Department of Health and Social Security of the Autonomous Government of Catalonia), in collaboration with the National AIDS Plan and the AIDS Study Group (GESIDA), promoted the creation of a working group for the drafting of recommendations for PEP against HIV outside the occupational health context. The recommendations have been made bearing in mind the exceptional character of the exposure, the time elapsed since exposure, as well as evaluation of the risk of infection according to the type of exposure and the information available on the source of infection. In addition, the recommendations include the immediate measures necessary, as well as the preventive measures and clinical follow-up required both for HIV and for other infectious agents. All PEP regimens should be started within 72 hours of exposure and appropriate daily doses of two nucleoside reverse transcriptase inhibitors (NRTIs) and a protease inhibitor (PI), or two NRTIs and a non-nucleoside reverse transcriptase inhibitor (NNRTIs), should be administered for four weeks, bearing in mind the pharmacological and clinical situation of the source person. These recommendations should be updated periodically.

Minority Human Immunodeficiency Virus Type 1 Variants in Antiretroviral-Naive Persons with Reverse Transcriptase Codon 215 Revertant Mutations▿ †

PubMed Central

Mitsuya, Yumi; Varghese, Vici; Wang, Chunlin; Liu, Tommy F.; Holmes, Susan P.; Jayakumar, Prerana; Gharizadeh, Baback; Ronaghi, Mostafa; Klein, Daniel; Fessel, W. Jeffrey; Shafer, Robert W.

2008-01-01

T215 revertant mutations such as T215C/D/E/S that evolve from the nucleoside reverse transcriptase (RT) inhibitor mutations T215Y/F have been found in about 3% of human immunodeficiency virus type 1 (HIV-1) isolates from newly diagnosed HIV-1-infected persons. We used a newly developed sequencing method—ultradeep pyrosequencing (UDPS; 454 Life Sciences)—to determine the frequency with which T215Y/F or other RT inhibitor resistance mutations could be detected as minority variants in samples from untreated persons that contain T215 revertants (“revertant” samples) compared with samples from untreated persons that lack such revertants (“control” samples). Among the 22 revertant and 29 control samples, UDPS detected a mean of 3.8 and 4.8 additional RT amino acid mutations, respectively. In 6 of 22 (27%) revertant samples and in 4 of 29 control samples (14%; P = 0.4), UDPS detected one or more RT inhibitor resistance mutations. T215Y or T215F was not detected in any of the revertant or control samples; however, 4 of 22 revertant samples had one or more T215 revertants that were detected by UDPS but not by direct PCR sequencing. The failure to detect viruses with T215Y/F in the 22 revertant samples in this study may result from the overwhelming replacement of transmitted T215Y variants by the more fit T215 revertants or from the primary transmission of a T215 revertant in a subset of persons with T215 revertants. PMID:18715933

Antiretroviral medication prescribing errors are common with hospitalization of HIV-infected patients.

PubMed

Commers, Tessa; Swindells, Susan; Sayles, Harlan; Gross, Alan E; Devetten, Marcel; Sandkovsky, Uriel

2014-01-01

Errors in prescribing antiretroviral therapy (ART) often occur with the hospitalization of HIV-infected patients. The rapid identification and prevention of errors may reduce patient harm and healthcare-associated costs. A retrospective review of hospitalized HIV-infected patients was carried out between 1 January 2009 and 31 December 2011. Errors were documented as omission, underdose, overdose, duplicate therapy, incorrect scheduling and/or incorrect therapy. The time to error correction was recorded. Relative risks (RRs) were computed to evaluate patient characteristics and error rates. A total of 289 medication errors were identified in 146/416 admissions (35%). The most common was drug omission (69%). At an error rate of 31%, nucleoside reverse transcriptase inhibitors were associated with an increased risk of error when compared with protease inhibitors (RR 1.32; 95% CI 1.04-1.69) and co-formulated drugs (RR 1.59; 95% CI 1.19-2.09). Of the errors, 31% were corrected within the first 24 h, but over half (55%) were never remedied. Admissions with an omission error were 7.4 times more likely to have all errors corrected within 24 h than were admissions without an omission. Drug interactions with ART were detected on 51 occasions. For the study population (n = 177), an increased risk of admission error was observed for black (43%) compared with white (28%) individuals (RR 1.53; 95% CI 1.16-2.03) but no significant differences were observed between white patients and other minorities or between men and women. Errors in inpatient ART were common, and the majority were never detected. The most common errors involved omission of medication, and nucleoside reverse transcriptase inhibitors had the highest rate of prescribing error. Interventions to prevent and correct errors are urgently needed.

Consensus Induced Fit Docking (cIFD): methodology, validation, and application to the discovery of novel Crm1 inhibitors

NASA Astrophysics Data System (ADS)

Kalid, Ori; Toledo Warshaviak, Dora; Shechter, Sharon; Sherman, Woody; Shacham, Sharon

2012-11-01

We present the Consensus Induced Fit Docking (cIFD) approach for adapting a protein binding site to accommodate multiple diverse ligands for virtual screening. This novel approach results in a single binding site structure that can bind diverse chemotypes and is thus highly useful for efficient structure-based virtual screening. We first describe the cIFD method and its validation on three targets that were previously shown to be challenging for docking programs (COX-2, estrogen receptor, and HIV reverse transcriptase). We then demonstrate the application of cIFD to the challenging discovery of irreversible Crm1 inhibitors. We report the identification of 33 novel Crm1 inhibitors, which resulted from the testing of 402 purchased compounds selected from a screening set containing 261,680 compounds. This corresponds to a hit rate of 8.2 %. The novel Crm1 inhibitors reveal diverse chemical structures, validating the utility of the cIFD method in a real-world drug discovery project. This approach offers a pragmatic way to implicitly account for protein flexibility without the additional computational costs of ensemble docking or including full protein flexibility during virtual screening.

Naringin Ameliorates HIV-1 Nucleoside Reverse Transcriptase Inhibitors- Induced Mitochondrial Toxicity.

PubMed

Oluwafeyisetan, Adebiyi; Olubunmi, Adebiyi; Peter, Owira

2016-01-01

Mitochondrial reactive oxygen species (ROS) generation and defective oxidative phosphorylation (OXPHOS) have been proposed as possible mechanisms underlying the development of nucleoside reverse transcriptase inhibitors (NRTIs)-induced mitochondrial toxicities. Available options in managing these complications have, so far, produced controversial results, thus necessitating further research into newer agents with promise. Antioxidant and free-radical scavenging effects of naringin, a plant-derived flavonoid, have previously been demonstrated. This study was designed to investigate the effects of naringin on NRTIs-induced mitochondrial toxicity. Wistar rats were randomly divided into Zidovudine (AZT)-only (100 mg/kg body weight BW); AZT+Naringin (100+50 mg/kg BW); AZT+Vitamin E (100+100 mg/kg BW); Stavudine (d4T)- only (50 mg/kg BW); d4T+Naringin (50+50 mg/kg BW); d4T+Vitamin E (50+100 mg/kg BW) and Vehicle (3.0 mL/kg BW)-treated groups, respectively. After 56 days of oral daily dosing, rats were euthanized by halothane overdose, blood collected by cardiac puncture and livers promptly excised for further biochemical and ultrastructural analyses. </p> Results: AZT- or d4T-only caused significant mitochondrial dysfunction and mitochondrial ultrastructural damage compared to controls, while either naringin or vitamin E reversed indices of mitochondrial dysfunction evidenced by significantly reduced mitochondrial malondialdehyde (MDA) and blood lactate concentrations, increased liver manganese superoxide dismutase (MnSOD) activity and upregulate expression of mitochondrial-encoded subunit of electron transport chain (ETC) complex IV protein compared to AZT- or d4T-only treated rats. Furthermore, naringin or vitamin E, respectively, ameliorated mitochondrial damage observed in AZT- or d4T-only treated rats. Naringin ameliorated oxidative stress and NRTI-induced mitochondrial damage and might, therefore, be beneficial in managing toxicities and complications arising from NRTI use.

Efavirenz or nevirapine in three-drug combination therapy with two nucleoside-reverse transcriptase inhibitors for initial treatment of HIV infection in antiretroviral-naïve individuals.

PubMed

Mbuagbaw, Lawrence Ce; Irlam, James H; Spaulding, Alicen; Rutherford, George W; Siegfried, Nandi

2010-12-08

The advent of highly active antiretroviral therapy (HAART) has reduced the morbidity and mortality due to HIV. The World Health Organisation (WHO) antiretroviral treatment (ART) guidelines focus on three classes of antiretroviral drugs, namely: nucleoside/nucleotide reverse transcriptase inhibitors (NRTI), non-nucleoside reverse transcriptase inhibitors (NNRTI) and protease inhibitors (PI). Two of the most common medications given in first-line treatment are the NNRTIs, efavirenz (EFV) and nevirapine (NVP). It is unclear which NNRTI is more efficacious for initial therapy. To determine which NNRTI, EFV or NVP, is more efficacious when given in combination with two NRTIs as part of initial ART for HIV infection in adults and children. We used a comprehensive and exhaustive strategy in an attempt to identify all relevant studies, regardless of language or publication status, in electronic databases and conference proceedings from 1996 to 2009. All randomised controlled trials comparing EFV to NVP in HIV-infected individuals without prior exposure to ART, irrespective of the dosage or NRTI backbone.The primary outcome of interest was virologic response to ART. Other primary outcomes included mortality, clinical progression, severe adverse events, and discontinuation of therapy for any reason. Secondary outcomes were immunologic response to ART, treatment failure, development of ART drug resistance, and prevention of sexual transmission of HIV. Two authors assessed each reference for inclusion and exclusion criteria established a priori. Data were abstracted independently using a standardised abstraction form. Data were analysed on an intention-to-treat basis and reported as per dosage of NVP. We identified seven randomised controlled trials that met our inclusion criteria.The trials were pooled as per dosage of NVP. None of these trials included children.The seven trials enrolled 1,688 participants and found no critical differences between EFV and NVP, except for different toxicity profiles. EFV is more likely to cause central nervous system side-effects, while NVP is more likely to result in raised transaminases and neutropoenia. There was a higher mortality rate in the NVP 400mg once daily arm.The quality of literature to support these conclusions is moderate to high. Drug resistance was slightly less common with EFV than NVP, but the quality of this literature is low since only one of the seven studies reported on this outcome. No studies reported on sexual transmission of HIV. The length of follow-up time, study settings, and NRTI backbone varied greatly. Both drugs have equivalent efficacies in initial treatment of HIV infection when combined with two NRTIs, but different side effects.

Computational Analysis of Molecular Interaction Networks Underlying Change of HIV-1 Resistance to Selected Reverse Transcriptase Inhibitors

PubMed Central

Kierczak, Marcin; Dramiński, Michał; Koronacki, Jacek; Komorowski, Jan

2010-01-01

Motivation Despite more than two decades of research, HIV resistance to drugs remains a serious obstacle in developing efficient AIDS treatments. Several computational methods have been developed to predict resistance level from the sequence of viral proteins such as reverse transcriptase (RT) or protease. These methods, while powerful and accurate, give very little insight into the molecular interactions that underly acquisition of drug resistance/hypersusceptibility. Here, we attempt at filling this gap by using our Monte Carlo feature selection and interdependency discovery method (MCFS-ID) to elucidate molecular interaction networks that characterize viral strains with altered drug resistance levels. Results We analyzed a number of HIV-1 RT sequences annotated with drug resistance level using the MCFS-ID method. This let us expound interdependency networks that characterize change of drug resistance to six selected RT inhibitors: Abacavir, Lamivudine, Stavudine, Zidovudine, Tenofovir and Nevirapine. The networks consider interdependencies at the level of physicochemical properties of mutating amino acids, eg,: polarity. We mapped each network on the 3D structure of RT in attempt to understand the molecular meaning of interacting pairs. The discovered interactions describe several known drug resistance mechanisms and, importantly, some previously unidentified ones. Our approach can be easily applied to a whole range of problems from the domain of protein engineering. Availability A portable Java implementation of our MCFS-ID method is freely available for academic users and can be obtained at: http://www.ipipan.eu/staff/m.draminski/software.htm. PMID:21234299

Different degree of immune recovery using antiretroviral regimens with Vonavir or Zidovudine/Lamivudine and Efavirenz in Patients receiving first line treatment in Iran.

PubMed

Sadeghi, Leila; Moallemi, Samaneh; Tabatabai, Reza Adl; Esmaeilzadeh, Ali; Ahsani-Nasab, Sara; Ahmadi, Niloofar Eghbal; Bayanolhagh, Saeed; Lolaie, Mahin; Narouei, Arsalan; Alinaghi, Ahmad; Mohraz, Minoo

2018-01-07

The initial ART regimens used by most national treatment guidelines in resource-limited settings include 2 nucleoside reverse-transcriptase inhibitors (NRTIs) and 1 non-nucleoside reverse-transcriptase inhibitor (NNRTI). The NRTIshave consistedof Zidovudine (AZT) or Stavudine (d4T) with Lamivudine (3TC); the NNRTI component hasbeen Nevirapine (NVP) or Efavirenz (EFV). There exists few data indicating if Vonavir is more effective in increasing CD4+ T cell counts or other first-line drugs of ART.Immunological outcomes of 134 individuals who just started antiretroviral therapy with Vonavir or Zidovudine/Lamivudine and Efavirenzanalyzed. Immunological response was then assessed during 28 weeks and indicated significant CD4+ T cell increase in both groups. The increase in CD4+ T cell counts was greater in Zidovudine/Lamivudine and Efavirenz treated group. A rapid CD4+ T cell increase occurred after the first 12 weeks of therapy in Vonavir treated group with initial CD4+ T cell counts ≤100 despite the individuals with higher CD4+ T cell counts. As previous studies, we observed the noticeable increase rate of CD4+ Tcells is in the first three months of therapy. A rapid CD4+ Tcell increase occurred shortly after beginning anti retroviraltherapy using either Vonaviror Zidovudine/Lamivudine and Efavirenz. Late increases in CD4+ T cell counts are more pronounced in therapy using Zidovudine/Lamivudine and Efavirenz. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

Computational Analysis of Molecular Interaction Networks Underlying Change of HIV-1 Resistance to Selected Reverse Transcriptase Inhibitors.

PubMed

Kierczak, Marcin; Dramiński, Michał; Koronacki, Jacek; Komorowski, Jan

2010-12-12

Despite more than two decades of research, HIV resistance to drugs remains a serious obstacle in developing efficient AIDS treatments. Several computational methods have been developed to predict resistance level from the sequence of viral proteins such as reverse transcriptase (RT) or protease. These methods, while powerful and accurate, give very little insight into the molecular interactions that underly acquisition of drug resistance/hypersusceptibility. Here, we attempt at filling this gap by using our Monte Carlo feature selection and interdependency discovery method (MCFS-ID) to elucidate molecular interaction networks that characterize viral strains with altered drug resistance levels. We analyzed a number of HIV-1 RT sequences annotated with drug resistance level using the MCFS-ID method. This let us expound interdependency networks that characterize change of drug resistance to six selected RT inhibitors: Abacavir, Lamivudine, Stavudine, Zidovudine, Tenofovir and Nevirapine. The networks consider interdependencies at the level of physicochemical properties of mutating amino acids, eg,: polarity. We mapped each network on the 3D structure of RT in attempt to understand the molecular meaning of interacting pairs. The discovered interactions describe several known drug resistance mechanisms and, importantly, some previously unidentified ones. Our approach can be easily applied to a whole range of problems from the domain of protein engineering. A portable Java implementation of our MCFS-ID method is freely available for academic users and can be obtained at: http://www.ipipan.eu/staff/m.draminski/software.htm.

Antitumor Activity and Mechanism of a Reverse Transcriptase Inhibitor, Dapivirine, in Glioblastoma.

PubMed

Liu, Weiwen; Song, Xian-Lu; Zhao, Shan-Chao; He, Minyi; Wang, Hai; Chen, Ziyang; Xiang, Wei; Yi, Guozhong; Qi, Songtao; Liu, Yawei

2018-01-01

Dapivirine is one of reverse transcriptase inhibitors (RTIs). It is the prototype of diarylpyrimidines (DAPY), formerly known as TMC120 or DAPY R147681 (IUPAC name: 4- [[4-(2, 4, 6-trimethylphenyl) amino]-2-pyrimidinyl] amino]-benzonitrile; CAS no.244767-67-7). The purpose of this study is to investigate the antitumor activity of dapivirine, one of the RTIs, on U87 glioblastoma (GBM) cells in vitro and in vivo . U87 GBM cells were cultured and treated with or without dapivirine. Cell viability was evaluated by CCK-8 (Cell Counting Kit 8, CCK-8) assay; apoptosis was analyzed by flow cytometry; cell migration was evaluated by Boyden Chamber assay; Western blotting was performed to detect proteins related to apoptosis, epithelial-to-mesenchymal transition and autophagy. PathScan intracellular signaling array kit was used to detect important and well-characterized signaling molecules. Tumor xenograft model in nude mice was used to evaluate the antitumorigenic effect in vivo . Dapivirine weakened proliferation of glioma cells and induced the apoptosis of U87 glioblastoma cells. Furthermore, dapivirine regulated autophagy and induced Akt, Bad and SAPK/JNK activations. Moreover, the inhibition of glioma cell growth by dapivirine was also observed in nude mice in vivo . In summary, in our study dapivirine exposure induces stress, resulting in JNK and PI3K/Akt pathway activation through diminished inhibition of the apoptosis and autophagy cascade in U87 GBM cells, which inhibits cell growth in vitro and in vivo .

Development and Characterization of a Vaginal Film Containing Dapivirine, a Non- nucleoside Reverse Transcriptase Inhibitor (NNRTI), for prevention of HIV-1 sexual transmission

PubMed Central

Akil, Ayman; Parniak, Michael A.; Dezzuitti, Charlene S.; Moncla, Bernard J.; Cost, Marilyn R.; Li, Mingguang; Rohan, Lisa Cencia

2012-01-01

Dapivirine, a non-nucleoside reverse transcriptase inhibitor, is a potent and promising anti-HIV molecule. It is currently being investigated for use as a vaginal microbicide in two dosage forms, a semi-solid gel and a silicone elastomer ring. Quick-dissolving films are promising and attractive dosage forms that may provide an alternative platform for the vaginal delivery of microbicide drug candidates. Vaginal films may provide advantages such as discreet use, no product leakage during use, lack of requirement for an applicator for insertion, rapid drug release and minimal packaging and reduced wastage. Within this study the in vitro bioactivity of dapivirine as compared to the NNRTI UC781 was further established and a quick dissolve film was developed for vaginal application of dapivirine for prevention of HIV infection. The developed film was characterized with respect to its physical and chemical attributes including water content, mechanical strength, drug release profile, permeability, compatibility with lactobacilli and bioactivity. The anti-HIV activity of the formulated dapivirine film was confirmed in in vitro and ex vivo models. Importantly the physical and chemical properties of the film as well as its bioactivity were maintained for a period of 18 months. In conclusion, a vaginal film containing dapivirine was developed and characterized. The film was shown to prevent HIV-1 infection in vitro and ex vivo and have acceptable characteristics which make this film a promising candidate for testing as vaginal microbicide. PMID:22708075

Development and Characterization of a Vaginal Film Containing Dapivirine, a Non- nucleoside Reverse Transcriptase Inhibitor (NNRTI), for prevention of HIV-1 sexual transmission.

PubMed

Akil, Ayman; Parniak, Michael A; Dezzuitti, Charlene S; Moncla, Bernard J; Cost, Marilyn R; Li, Mingguang; Rohan, Lisa Cencia

2011-06-01

Dapivirine, a non-nucleoside reverse transcriptase inhibitor, is a potent and promising anti-HIV molecule. It is currently being investigated for use as a vaginal microbicide in two dosage forms, a semi-solid gel and a silicone elastomer ring. Quick-dissolving films are promising and attractive dosage forms that may provide an alternative platform for the vaginal delivery of microbicide drug candidates. Vaginal films may provide advantages such as discreet use, no product leakage during use, lack of requirement for an applicator for insertion, rapid drug release and minimal packaging and reduced wastage. Within this study the in vitro bioactivity of dapivirine as compared to the NNRTI UC781 was further established and a quick dissolve film was developed for vaginal application of dapivirine for prevention of HIV infection. The developed film was characterized with respect to its physical and chemical attributes including water content, mechanical strength, drug release profile, permeability, compatibility with lactobacilli and bioactivity. The anti-HIV activity of the formulated dapivirine film was confirmed in in vitro and ex vivo models. Importantly the physical and chemical properties of the film as well as its bioactivity were maintained for a period of 18 months. In conclusion, a vaginal film containing dapivirine was developed and characterized. The film was shown to prevent HIV-1 infection in vitro and ex vivo and have acceptable characteristics which make this film a promising candidate for testing as vaginal microbicide.

Characterization of Nucleoside Reverse Transcriptase Inhibitor-Associated Mutations in the RNase H Region of HIV-1 Subtype C Infected Individuals.

PubMed

Ngcapu, Sinaye; Theys, Kristof; Libin, Pieter; Marconi, Vincent C; Sunpath, Henry; Ndung'u, Thumbi; Gordon, Michelle L

2017-11-08

The South African national treatment programme includes nucleoside reverse transcriptase inhibitors (NRTIs) in both first and second line highly active antiretroviral therapy regimens. Mutations in the RNase H domain have been associated with resistance to NRTIs but primarily in HIV-1 subtype B studies. Here, we investigated the prevalence and association of RNase H mutations with NRTI resistance in sequences from HIV-1 subtype C infected individuals. RNase H sequences from 112 NRTI treated but virologically failing individuals and 28 antiretroviral therapy (ART)-naive individuals were generated and analysed. In addition, sequences from 359 subtype C ART-naive sequences were downloaded from Los Alamos database to give a total of 387 sequences from ART-naive individuals for the analysis. Fisher's exact test was used to identify mutations and Bayesian network learning was applied to identify novel NRTI resistance mutation pathways in RNase H domain. The mutations A435L, S468A, T470S, L484I, A508S, Q509L, L517I, Q524E and E529D were more prevalent in sequences from treatment-experienced compared to antiretroviral treatment naive individuals, however, only the E529D mutation remained significant after correction for multiple comparison. Our findings suggest a potential interaction between E529D and NRTI-treatment; however, site-directed mutagenesis is needed to understand the impact of this RNase H mutation.

Dynamics of drug resistance-associated mutations in HIV-1 DNA reverse transcriptase sequence during effective ART.

PubMed

Nouchi, A; Nguyen, T; Valantin, M A; Simon, A; Sayon, S; Agher, R; Calvez, V; Katlama, C; Marcelin, A G; Soulie, C

2018-05-29

To investigate the dynamics of HIV-1 variants archived in cells harbouring drug resistance-associated mutations (DRAMs) to lamivudine/emtricitabine, etravirine and rilpivirine in patients under effective ART free from selective pressure on these DRAMs, in order to assess the possibility of recycling molecules with resistance history. We studied 25 patients with at least one DRAM to lamivudine/emtricitabine, etravirine and/or rilpivirine identified on an RNA sequence in their history and with virological control for at least 5 years under a regimen excluding all drugs from the resistant class. Longitudinal ultra-deep sequencing (UDS) and Sanger sequencing of the reverse transcriptase region were performed on cell-associated HIV-1 DNA samples taken over the 5 years of follow-up. Viral variants harbouring the analysed DRAMs were no longer detected by UDS over the 5 years in 72% of patients, with viruses susceptible to the molecules of interest found after 5 years in 80% of patients with UDS and in 88% of patients with Sanger. Residual viraemia with <50 copies/mL was detected in 52% of patients. The median HIV DNA level remained stable (2.4 at baseline versus 2.1 log10 copies/106 cells 5 years later). These results show a clear trend towards clearance of archived DRAMs to reverse transcriptase inhibitors in cell-associated HIV-1 DNA after a long period of virological control, free from therapeutic selective pressure on these DRAMs, reflecting probable residual replication in some reservoirs of the fittest viruses and leading to persistent evolution of the archived HIV-1 DNA resistance profile.

Antioxidants inhibit nuclear export of telomerase reverse transcriptase and delay replicative senescence of endothelial cells.

PubMed

Haendeler, Judith; Hoffmann, Jörg; Diehl, J Florian; Vasa, Mariuca; Spyridopoulos, Ioakim; Zeiher, Andreas M; Dimmeler, Stefanie

2004-04-02

Aging is associated with a rise in intracellular reactive oxygen species (ROS) and a loss of telomerase reverse transcriptase activity. Incubation with H2O2 induced the nuclear export of telomerase reverse transcriptase (TERT) into the cytosol in a Src-family kinase-dependent manner. Therefore, we investigated the hypothesis that age-related increase in reactive oxygen species (ROS) may induce the nuclear export of TERT and contribute to endothelial cell senescence. Continuous cultivation of endothelial cells resulted in an increased endogenous formation of ROS starting after 29 population doublings (PDL). This increase was accompanied by mitochondrial DNA damage and preceded the onset of replicative senescence at PDL 37. Along with the enhanced formation of ROS, we detected an export of nuclear TERT protein from the nucleus into the cytoplasm and an activation of the Src-kinase. Moreover, the induction of premature senescence by low concentrations of H2O2 was completely blocked with the Src-family kinase inhibitor PP2, suggesting a crucial role for Src-family kinases in the induction of endothelial cell aging. Incubation with the antioxidant N-acetylcysteine, from PDL 26, reduced the intracellular ROS formation and prevented mitochondrial DNA damage. Likewise, nuclear export of TERT protein, loss in the overall TERT activity, and the onset of replicative senescence were delayed by incubation with N-acetylcysteine. Low doses of the statin, atorvastatin (0.1 micromol/L), had also effects similar to those of N-acetylcysteine. We conclude that both antioxidants and statins can delay the onset of replicative senescence by counteracting the increased ROS production linked to aging of endothelial cells.

Comparative analysis of drug resistance mutations in the human immunodeficiency virus reverse transcriptase gene in patients who are non-responsive, responsive and naive to antiretroviral therapy.

PubMed

Misbah, Mohammad; Roy, Gaurav; Shahid, Mudassar; Nag, Nalin; Kumar, Suresh; Husain, Mohammad

2016-05-01

Drug resistance mutations in the Pol gene of human immunodeficiency virus 1 (HIV-1) are one of the critical factors associated with antiretroviral therapy (ART) failure in HIV-1 patients. The issue of resistance to reverse transcriptase inhibitors (RTIs) in HIV infection has not been adequately addressed in the Indian subcontinent. We compared HIV-1 reverse transcriptase (RT) gene sequences to identify mutations present in HIV-1 patients who were ART non-responders, ART responders and drug naive. Genotypic drug resistance testing was performed by sequencing a 655-bp region of the RT gene from 102 HIV-1 patients, consisting of 30 ART-non-responding, 35 ART-responding and 37 drug-naive patients. The Stanford HIV Resistance Database (HIVDBv 6.2), IAS-USA mutation list, ANRS_09/2012 algorithm, and Rega v8.02 algorithm were used to interpret the pattern of drug resistance. The majority of the sequences (96 %) belonged to subtype C, and a few of them (3.9 %) to subtype A1. The frequency of drug resistance mutations observed in ART-non-responding, ART-responding and drug-naive patients was 40.1 %, 10.7 % and 20.58 %, respectively. It was observed that in non-responders, multiple mutations were present in the same patient, while in responders, a single mutation was found. Some of the drug-naive patients had more than one mutation. Thymidine analogue mutations (TAMs), however, were found in non-responders and naive patients but not in responders. Although drug resistance mutations were widely distributed among ART non-responders, the presence of resistance mutations in the viruses of drug-naive patients poses a big concern in the absence of a genotyping resistance test.

Immunovirological response to triple nucleotide reverse-transcriptase inhibitors and ritonavir-boosted protease inhibitors in treatment-naive HIV-2-infected patients: The ACHIEV2E Collaboration Study Group.

PubMed

Benard, Antoine; van Sighem, Ard; Taieb, Audrey; Valadas, Emilia; Ruelle, Jean; Soriano, Vicente; Calmy, Alexandra; Balotta, Claudia; Damond, Florence; Brun-Vezinet, Françoise; Chene, Geneviève; Matheron, Sophie

2011-05-01

Triple nucleoside reverse-transcriptase inhibitors (NRTIs) are recommended by the World Health Organization as first-line regimen in treatment-naïve HIV-2-infected patients. However, ritonavir-boosted protease inhibitor (PI/r)-containing regimens are frequently prescribed. In the absence of previous randomized trials, we retrospectively compared these regimens in observational cohorts. HIV-2-infected patients from 7 European cohorts who started triple NRTI or PI/r since January 1998 were included. Piecewise linear models were used to estimate CD4 cell count and plasma HIV-2 RNA level slopes, differentiating an early phase (until end of month 3) and a second phase (months 4-12). On-treatment analyses censored data at major treatment modification and systematically at month 12. Forty-four patients started triple NRTI therapy and 126 started PI/r therapy. Overall, the median CD4 cell count was 191 cells/mm(3) and the median plasma HIV-2 RNA level was ≥2.7 log(10) copies/ml in 61% of the patients at combination antiretroviral therapy (cART) initiation; the median duration of the first cART was 20 months, not differing between groups. PI/r regimens were associated with better CD4 cell count and HIV-2 RNA level outcomes, compared with NRTI regimens. Estimated CD4 cell count slopes were +6 and +12 cells/mm(3)/month during the early phase (P = .22), and -60 cells/mm(3)/year versus +76 cells/mm(3)/year during the second phase (P = .002), for triple NRTI and PI/r, respectively. Estimated mean HIV-2 RNA levels at month 12 in patients with detectable viremia at cART initiation were 4.0 and 2.2 log(10) copies/ml, respectively (P = .005). In this observational study, PI/r-containing regimens showed superior efficacy over triple NRTI regimens as first-line therapy in HIV-2-infected patients.

Trends in Drug Resistance Prevalence, HIV-1 Variants and Clinical Status in HIV-1-infected Pediatric Population in Madrid: 1993 to 2015 Analysis.

PubMed

Rojas Sánchez, Patricia; Domínguez, Sara; Jiménez De Ory, Santiago; Prieto, Luis; Rojo, Pablo; Mellado, Pepa; Navarro, Marisa; Delgado, Rafael; Ramos, José Tomas; Holguín, África

2018-03-01

The expanded use of long-term antiretroviral treatments in infected children may exacerbate the problem of drug resistance mutations selection, which can compromise treatment efficiency. We describe the temporal trends of HIV drug resistance mutations and the HIV-1 variants during 23 years (1993 to March 2016) in the Madrid cohort of HIV-infected children and adolescents. We selected patients with at least one available HIV-1 pol sequence/genotypic resistance profile, establishing different groups according to the sampling year of first resistance data. We determined the prevalence of transmitted drug resistance mutations or acquired drug resistance mutations (DRM), the drug susceptibility among resistant viruses and HIV-1 variants characterized by phylogeny across time. A total of 245 pediatric patients were selected, being mainly female, Spanish native, perinatally infected and carrying HIV-1 subtype B. At first sampling, most pediatric patients were on antiretroviral therapy and heavily pretreated. During 1993 to 2016, transmitted drug resistance mutations was found in 13 (26%) of 50 naive children [non-nucleoside reverse transcriptase inhibitors (NNRTI), 14.6%; nucleoside reverse transcriptase inhibitors (NRTI), 10.4%; protease inhibitors, 8.7%]. DRM appeared in 139 (73.2%) of 190 pretreated patients (NRTI, 64.5%; NNRTI, 36%; protease inhibitors, 35.1%). DRM to NNRTI was higher in last 5 years. Non-B variants infected 14.5% of children and adolescents of the Madrid Cohort, being mainly intersubtype recombinants (76.5%), including complex unique recombinant strains. They caused 3.4% infections before 2000, rising to 85.7% during 2011 to 2016. Periodic surveillance resistance and molecular epidemiology studies in long-term pretreated HIV-infected pediatric populations are required to optimize treatment regimens. Results will permit a better understanding of long-time dynamics of viral resistance and HIV-1 variants in Spain.

Differences in National Antiretroviral Prescribing Patterns between Black and White Patients with HIV/AIDS, 1996–2006

PubMed Central

Oramasionwu, Christine U.; Brown, Carolyn M.; Lawson, Kenneth A.; Ryan, Laurajo; Skinner, Jeff; Frei, Christopher R.

2011-01-01

Objectives The benefit of improved health outcomes for blacks receiving highly active antiretroviral therapy (HAART) lags behind that of whites. This project therefore sought to determine whether the reason for this discrepancy in health outcomes could be attributed to disparities in use of antiretroviral therapy between black and white patients with HIV. Materials and Methods The 1996–2006 National Hospital Ambulatory Medical Care Surveys were used to identify hospital outpatient visits that documented antiretrovirals. Patients younger than 18 years, of nonblack or nonwhite race, and lacking documentation of antiretrovirals were excluded. A multivariable logistic regression model was constructed with race as the independent variable and use of HAART as the dependent variable. Results Approximately 3 million HIV/AIDS patient visits were evaluated. Blacks were less likely than whites to use HAART and protease inhibitors (odds ratio, 95% CI 0.81 [0.81–0.82] and 0.67 [0.67–0.68], respectively). More blacks than whites used non-nucleoside reverse transcriptase inhibitors (odds ratio, 95% CI 1.18 [1.17–1.18]). In 1996, the crude rates of HAART were relatively low for both black and white cohorts (5% vs 6%). The rise in HAART for blacks appeared to lag behind that of whites for several years, until 2002, when the proportion of blacks receiving HAART slightly exceeded the proportion of whites receiving HAART. In later years, the rates of HAART were similar for blacks and whites (81% vs 82% in 2006). Blacks appeared less likely than whites to use protease inhibitors and more likely than whites to use non-nucleoside reverse transcriptase inhibitors from 2000 to 2004. Conclusions Blacks experienced a lag in the use of antiretrovirals at the beginning of the study; this discrepancy dissipated in more recent years. PMID:22089356

Recent trends and patterns in HIV-1 transmitted drug resistance in the United Kingdom.

PubMed

Tostevin, A; White, E; Dunn, D; Croxford, S; Delpech, V; Williams, I; Asboe, D; Pozniak, A; Churchill, D; Geretti, A M; Pillay, D; Sabin, C; Leigh-Brown, A; Smit, E

2017-03-01

Transmission of drug-resistant HIV-1 has decreased in the UK since the early 2000s. This analysis reports recent trends and characteristics of transmitted drug resistance (TDR) in the UK from 2010 to 2013. Resistance tests conducted in antiretroviral treatment (ART)-naïve individuals between 2010 and 2013 were analysed for the presence of transmitted drug resistance mutations (TDRMs), defined as any mutations from a modified 2009 World Health Organization surveillance list, or a modified 2013 International Antiviral Society-USA list for integrase tests. Logistic regression was used to examine associations between demographics and the prevalence of TDRMs. TDRMs were observed in 1223 (7.5%) of 16 425 individuals; prevalence declined from 8.1% in 2010 to 6.6% in 2013 (P = 0.02). The prevalence of TDRMs was higher among men who have sex with men (MSM) compared with heterosexual men and women (8.7% versus 6.4%, respectively) with a trend for decreasing TDRMs among MSM (P = 0.008) driven by a reduction in nucleoside reverse transcriptase inhibitor (NRTI)-related mutations. The most frequently detected TDRMs were K103N (2.2%), T215 revertants (1.6%), M41L (0.9%) and L90M (0.7%). Predicted phenotypic resistance to first-line ART was highest to the nonnucleoside reverse transcriptase inhibitors (NNRTIs) rilpivirine and efavirenz (6.2% and 3.4%, respectively) but minimal to NRTIs, including tenofovir, and protease inhibitors (PIs). No major integrase TDRMs were detected among 101 individuals tested while ART-naïve. We observed a decrease in TDRMs in recent years. However, this was confined to the MSM population and rates remained stable in those with heterosexually acquired HIV infection. Resistance to currently recommended first-line ART, including integrase inhibitors, remained reassuringly low. © 2016 The Authors. HIV Medicine published by John Wiley & Sons Ltd on behalf of British HIV Association.

Prediction of the binding mode and resistance profile for a dual-target pyrrolyl diketo acid scaffold against HIV-1 integrase and reverse-transcriptase-associated ribonuclease H.

PubMed

Yang, Fengyuan; Zheng, Guoxun; Fu, Tingting; Li, Xiaofeng; Tu, Gao; Li, Ying Hong; Yao, Xiaojun; Xue, Weiwei; Zhu, Feng

2018-06-27

The rapid emergence of drug-resistant variants is one of the most common causes of highly active antiretroviral therapeutic (HAART) failure in patients infected with HIV-1. Compared with the existing HAART, the recently developed pyrrolyl diketo acid scaffold targeting both HIV-1 integrase (IN) and reverse transcriptase-associated ribonuclease H (RNase H) is an efficient approach to counteract the failure of anti-HIV treatment due to drug resistance. However, the binding mode and potential resistance profile of these inhibitors with important mechanistic principles remain poorly understood. To address this issue, an integrated computational method was employed to investigate the binding mode of inhibitor JMC6F with HIV-1 IN and RNase H. By using per-residue binding free energy decomposition analysis, the following residues: Asp64, Thr66, Leu68, Asp116, Tyr143, Gln148 and Glu152 in IN, Asp443, Glu478, Trp536, Lys541 and Asp549 in RNase H were identified as key residues for JMC6F binding. And then computational alanine scanning was carried to further verify the key residues. Moreover, the resistance profile of the currently known major mutations in HIV-1 IN and 2 mutations in RNase H against JMC6F was predicted by in silico mutagenesis studies. The results demonstrated that only three mutations in HIV-1 IN (Y143C, Q148R and N155H) and two mutations in HIV-1 RNase H (Y501R and Y501W) resulted in a reduction of JMC6F potency, thus indicating their potential role in providing resistance to JMC6F. These data provided important insights into the binding mode and resistance profile of the inhibitors with a pyrrolyl diketo acid scaffold in HIV-1 IN and RNase H, which would be helpful for the development of more effective dual HIV-1 IN and RNase H inhibitors.

Development of a highly sensitive screen for influenza A in guano and its application in the search for ancient RNA preserved under Antarctic Adelie penguin colonies.

PubMed

Briggs, L C; Ashton, R M; Metcalf, P

2003-01-01

We have developed a reverse transcriptase polymerase chain reaction (RT-PCR)-based assay to detect influenza A in guano samples as part of our program to investigate ancient viral RNA from under Antarctic Adelie penguin (Pygoscelis adeliae) colonies. Of five extraction protocols tested (RNeasy, GTC TRIZOL, GTC Silica, Rnaid, and AGPC), AGPC proved to be the most consistent and sensitive to low concentrations of influenza, but further purification with commercial viral nucleic acid spin filter system was still required to remove remaining PCR inhibitors. RT-PCR was then performed on the eluent and 650 bases of the M1 gene were amplified. The assay was found to be able to detect as little as 100 microl of 0.1 hemagglutination units (HU)/ml influenza.

3-Hydroxypyrimidine-2,4-dione-5-N-benzylcarboxamides potently inhibit HIV-1 integrase and RNase H

PubMed Central

Wu, Bulan; Tang, Jing; Wilson, Daniel J.; Huber, Andrew D.; Casey, Mary C.; Ji, Juan; Kankanala, Jayakanth; Xie, Jiashu; Sarafianos, Stefan G.; Wang, Zhengqiang

2016-01-01

Resistance selection by human immunodeficiency virus (HIV) towards known drug regimens necessitates the discovery of structurally novel antivirals with a distinct resistance profile. Based on our previously reported 3-hydroxypyrimidine-2,4-dione (HPD) core we have designed and synthesized a new integrase strand transfer (INST) inhibitor type featuring a 5-N-benzylcarboxamide moiety. Significantly, the 6-alkylamino variant of this new chemotype consistently conferred low nanomolar inhibitory activity against HIV-1. Extended antiviral testing against a few raltegravir-resistant HIV-1 clones revealed a resistance profile similar to that of the second generation INST inhibitor (INSTIs) dolutegravir. Although biochemical testing and molecular modeling also strongly corroborate the inhibition of INST as the antiviral mechanism of action, selected antiviral analogues also potently inhibited reverse transcriptase (RT) associated RNase H, implying potential dual target inhibition. In vitro ADME assays demonstrated that this novel chemotype possesses largely favorable physicochemical properties suitable for further development. PMID:27283261

Initiating therapy: when to start, what to use.

PubMed

Hirsch, Martin S

2008-05-15

Decisions regarding whether to start combination antiretroviral therapy (cART) during primary infection and when to initiate treatment during chronic infection continue to evolve. Although current data suggest that there may be a benefit to therapy during primary infection, results are inconclusive. Once begun, treatment probably should be continued indefinitely, since its potential advantages disappear over time if treatment is stopped. Recent studies suggest that cART may be useful at higher CD4 cell count thresholds than are currently recommended in several guidelines. Several regimens are acceptable as initial therapy, with tenofovir/emtricitabine/efavirenz favored by many because of potency and ease of administration. Other favored regimens include combinations of 2 nucleoside (or nucleotide) reverse-transcriptase inhibitors and a ritonavir-boosted protease inhibitor. Some new antiretroviral drugs under study, particularly integrase inhibitors, may prove useful in treatment-naive patients.

Soft shell clams Mya arenaria with disseminated neoplasia demonstrate reverse transcriptase activity

USGS Publications Warehouse

House, M.L.; Kim, C.H.; Reno, P.W.

1998-01-01

Disseminated neoplasia (DN), a proliferative cell disorder of the circulatory system of bivalves, was first reported in oysters in 1969. Since that time, the disease has been determined to be transmissible through water-borne exposure, but the etiological agent has not been unequivocally identified. In order to determine if a viral agent, possibly a retrovirus, could be the causative agent of DN, transmission experiments were performed, using both a cell-free filtrate and a sucrose gradient-purified preparation of a cell-free filtrate of DN positive materials. Additionally, a PCR-enhanced reverse transcriptase assay was used to determine if reverse transcriptase was present in tissues or hemolymph from DN positive soft shell clams Mya arenaria. DN was transmitted to healthy clams by injection with whole DN cells, but not with cell-free flitrates prepared from either tissues from DN positive clams, or DN cells. The cell-free preparations from DN-positive tissues and hemolymph having high levels of DN cells in circulation exhibited positive reactions in the PCR-enhanced reverse transcriptase assay. Cell-free preparations of hemolymph from clams having low levels of DN (<0.1% of cells abnormal), hemocytes from normal soft shell clams, and normal soft shell clam tissues did not produce a positive reaction in the PCR enhanced reverse transcriptase assay.

Characterizing Class-Specific Exposure-Viral Load Suppression Response of HIV Antiretrovirals Using A Model-Based Meta-Analysis.

PubMed

Xu, Y; Li, Y F; Zhang, D; Dockendorf, M; Tetteh, E; Rizk, M L; Grobler, J A; Lai, M-T; Gobburu, J; Ankrom, W

2016-08-01

We applied model-based meta-analysis of viral suppression as a function of drug exposure and in vitro potency for short-term monotherapy in human immunodeficiency virus type 1 (HIV-1)-infected treatment-naïve patients to set pharmacokinetic targets for development of nonnucleoside reverse transcriptase inhibitors (NNRTIs) and integrase strand transfer inhibitors (InSTIs). We developed class-specific models relating viral load kinetics from monotherapy studies to potency normalized steady-state trough plasma concentrations. These models were integrated with a literature assessment of doses which demonstrated to have long-term efficacy in combination therapy, in order to set steady-state trough concentration targets of 6.17- and 2.15-fold above potency for NNRTIs and InSTIs, respectively. Both the models developed and the pharmacokinetic targets derived can be used to guide compound selection during preclinical development and to predict the dose-response of new antiretrovirals to inform early clinical trial design. © 2016 The Authors. Clinical and Translational Science published by Wiley Periodicals, Inc. on behalf of American Society for Clinical Pharmacology and Therapeutics.

HIV-1 drug resistance prevalence, drug susceptibility and variant characterization in the Jacobi Medical Center paediatric cohort, Bronx, NY, USA.

PubMed

de Mulder, M; York, V A; Wiznia, A A; Michaud, H A; Nixon, D F; Holguin, A; Rosenberg, M G

2014-03-01

With the advent of combined antiretroviral therapy (cART), perinatally HIV-infected children are surviving into adolescence and beyond. However, drug resistance mutations (DRMs) compromise viral control, affecting the long-term effectiveness of ART. The aims of this study were to detect and identify DRMs in a HIV-1 infected paediatric cohort. Paired plasma and dried blood spots (DBSs) specimens were obtained from HIV-1 perinatally infected patients attending the Jacobi Medical Center, New York, USA. Clinical, virological and immunological data for these patients were analysed. HIV-1 pol sequences were generated from samples to identify DRMs according to the International AIDS Society (IAS) 2011 list. Forty-seven perinatally infected patients were selected, with a median age of 17.7 years, of whom 97.4% were carrying subtype B. They had a mean viral load of 3143 HIV-1 RNA copies/mL and a mean CD4 count of 486 cells/μL at the time of sampling. Nineteen patients (40.4%) had achieved undetectable viraemia (< 50 copies/mL) and 40.5% had a CD4 count of > 500 cells/μL. Most of the patients (97.9%) had received cART, including protease inhibitor (PI)-based regimens in 59.6% of cases. The DRM prevalence was 54.1, 27.6 and 27.0% for nucleoside reverse transcriptase inhibitors (NRTIs), PIs and nonnucleoside reverse transcriptase inhibitors (NNRTIs), respectively. Almost two-thirds (64.9%) of the patients harboured DRMs to at least one drug class and 5.4% were triple resistant. The mean nucleotide similarity between plasma and DBS sequences was 97.9%. Identical DRM profiles were present in 60% of plasma-DBS paired sequences. A total of 30 DRMs were detected in plasma and 26 in DBSs, with 23 present in both. Although more perinatally HIV-1-infected children are reaching adulthood as a result of advances in cART, our study cohort presented a high prevalence of resistant viruses, especially viruses resistant to NRTIs. DBS specimens can be used for DRM detection. © 2013 British HIV Association.

Perinatal acquisition of drug-resistant HIV-1 infection: mechanisms and long-term outcome

PubMed Central

Delaugerre, Constance; Chaix, Marie-Laure; Blanche, Stephane; Warszawski, Josiane; Cornet, Dorine; Dollfus, Catherine; Schneider, Veronique; Burgard, Marianne; Faye, Albert; Mandelbrot, Laurent; Tubiana, Roland; Rouzioux, Christine

2009-01-01

Background Primary-HIV-1-infection in newborns that occurs under antiretroviral prophylaxis that is a high risk of drug-resistance acquisition. We examine the frequency and the mechanisms of resistance acquisition at the time of infection in newborns. Patients and Methods We studied HIV-1-infected infants born between 01 January 1997 and 31 December 2004 and enrolled in the ANRS-EPF cohort. HIV-1-RNA and HIV-1-DNA samples obtained perinatally from the newborn and mother were subjected to population-based and clonal analyses of drug resistance. If positive, serial samples were obtained from the child for resistance testing. Results Ninety-two HIV-1-infected infants were born during the study period. Samples were obtained from 32 mother-child pairs and from another 28 newborns. Drug resistance was detected in 12 newborns (20%): drug resistance to nucleoside reverse transcriptase inhibitors was seen in 10 cases, non-nucleoside reverse transcriptase inhibitors in two cases, and protease inhibitors in one case. For 9 children, the detection of the same resistance mutations in mothers' samples (6 among 10 available) and in newborn lymphocytes (6/8) suggests that the newborn was initially infected by a drug-resistant strain. Resistance variants were either transmitted from mother-to-child or selected during subsequent temporal exposure under suboptimal perinatal prophylaxis. Follow-up studies of the infants showed that the resistance pattern remained stable over time, regardless of antiretroviral therapy, suggesting the early cellular archiving of resistant viruses. The absence of resistance in the mother of the other three children (3/10) and neonatal lymphocytes (2/8) suggests that the newborns were infected by a wild-type strain without long-term persistence of resistance when suboptimal prophylaxis was stopped. Conclusion This study confirms the importance of early resistance genotyping of HIV-1-infected newborns. In most cases (75%), drug resistance was archived in the cellular reservoir and persisted during infancy, with or without antiretroviral treatment. This finding stresses the need for effective antiretroviral treatment of pregnant women. PMID:19765313

Docking and 3-D QSAR studies on indolyl aryl sulfones. Binding mode exploration at the HIV-1 reverse transcriptase non-nucleoside binding site and design of highly active N-(2-hydroxyethyl)carboxamide and N-(2-hydroxyethyl)carbohydrazide derivatives.

PubMed

Ragno, Rino; Artico, Marino; De Martino, Gabriella; La Regina, Giuseppe; Coluccia, Antonio; Di Pasquali, Alessandra; Silvestri, Romano

2005-01-13

Three-dimensional quantitative structure-activity relationship (3-D QSAR) studies and docking simulations were developed on indolyl aryl sulfones (IASs), a class of novel HIV-1 non-nucleoside reverse transcriptase (RT) inhibitors (Silvestri, et al. J. Med. Chem. 2003, 46, 2482-2493) highly active against wild type and some clinically relevant resistant strains (Y181C, the double mutant K103N-Y181C, and the K103R-V179D-P225H strain, highly resistant to efavirenz). Predictive 3-D QSAR models using the combination of GRID and GOLPE programs were obtained using a receptor-based alignment by means of docking IASs into the non-nucleoside binding site (NNBS) of RT. The derived 3-D QSAR models showed conventional correlation (r(2)) and cross-validated (q(2)) coefficients values ranging from 0.79 to 0.93 and from 0.59 to 0.84, respectively. All described models were validated by an external test set compiled from previously reported pyrryl aryl sulfones (Artico, et al. J. Med. Chem. 1996, 39, 522-530). The most predictive 3-D QSAR model was then used to predict the activity of novel untested IASs. The synthesis of six designed derivatives (prediction set) allowed disclosure of new IASs endowed with high anti-HIV-1 activities.

Selective killing of human immunodeficiency virus infected cells by non-nucleoside reverse transcriptase inhibitor-induced activation of HIV protease.

PubMed

Jochmans, Dirk; Anders, Maria; Keuleers, Inge; Smeulders, Liesbeth; Kräusslich, Hans-Georg; Kraus, Günter; Müller, Barbara

2010-10-15

Current antiretroviral therapy against human immunodeficiency virus (HIV-1) reduces viral load and thereby prevents viral spread, but it cannot eradicate proviral genomes from infected cells. Cells in immunological sanctuaries as well as cells producing low levels of virus apparently contribute to a reservoir that maintains HIV persistence in the presence of highly active antiretroviral therapy. Thus, accelerated elimination of virus producing cells may represent a complementary strategy to control HIV infection. Here we sought to exploit HIV protease (PR) related cytotoxicity in order to develop a strategy for drug induced killing of HIV producing cells. PR processes the viral Gag and Gag-Pol polyproteins during virus maturation, but is also implicated in killing of virus producing cells through off-target cleavage of host proteins. It has been observed previously that micromolar concentrations of certain non-nucleoside reverse transcriptase inhibitors (NNRTIs) can stimulate intracellular PR activity, presumably by enhancing Gag-Pol dimerization. Using a newly developed cell-based assay we compared the degree of PR activation displayed by various NNRTIs. We identified inhibitors showing higher potency with respect to PR activation than previously described for NNRTIs, with the most potent compounds resulting in ~2-fold increase of the Gag processing signal at 250 nM. The degree of enhancement of intracellular Gag processing correlated with the compound's ability to enhance RT dimerization in a mammalian two-hybrid assay. Compounds were analyzed for their potential to mediate specific killing of chronically infected MT-4 cells. Levels of cytotoxicity on HIV infected cells determined for the different NNRTIs corresponded to the relative degree of drug induced intracellular PR activation, with CC50 values ranging from ~0.3 μM to above the tested concentration range (10 μM). Specific cytotoxicity was reverted by addition of PR inhibitors. Two of the most active compounds, VRX-480773 and GW-678248, were also tested in primary human cells and mediated cytotoxicity on HIV-1 infected peripheral blood mononuclear cells. These data present proof of concept for targeted drug induced elimination of HIV producing cells. While NNRTIs themselves may not be sufficiently potent for therapeutic application, the results provide a basis for the development of drugs exploiting this mechanism of action.

Selective killing of human immunodeficiency virus infected cells by non-nucleoside reverse transcriptase inhibitor-induced activation of HIV protease

PubMed Central

2010-01-01

Background Current antiretroviral therapy against human immunodeficiency virus (HIV-1) reduces viral load and thereby prevents viral spread, but it cannot eradicate proviral genomes from infected cells. Cells in immunological sanctuaries as well as cells producing low levels of virus apparently contribute to a reservoir that maintains HIV persistence in the presence of highly active antiretroviral therapy. Thus, accelerated elimination of virus producing cells may represent a complementary strategy to control HIV infection. Here we sought to exploit HIV protease (PR) related cytotoxicity in order to develop a strategy for drug induced killing of HIV producing cells. PR processes the viral Gag and Gag-Pol polyproteins during virus maturation, but is also implicated in killing of virus producing cells through off-target cleavage of host proteins. It has been observed previously that micromolar concentrations of certain non-nucleoside reverse transcriptase inhibitors (NNRTIs) can stimulate intracellular PR activity, presumably by enhancing Gag-Pol dimerization. Results Using a newly developed cell-based assay we compared the degree of PR activation displayed by various NNRTIs. We identified inhibitors showing higher potency with respect to PR activation than previously described for NNRTIs, with the most potent compounds resulting in ~2-fold increase of the Gag processing signal at 250 nM. The degree of enhancement of intracellular Gag processing correlated with the compound's ability to enhance RT dimerization in a mammalian two-hybrid assay. Compounds were analyzed for their potential to mediate specific killing of chronically infected MT-4 cells. Levels of cytotoxicity on HIV infected cells determined for the different NNRTIs corresponded to the relative degree of drug induced intracellular PR activation, with CC50 values ranging from ~0.3 μM to above the tested concentration range (10 μM). Specific cytotoxicity was reverted by addition of PR inhibitors. Two of the most active compounds, VRX-480773 and GW-678248, were also tested in primary human cells and mediated cytotoxicity on HIV-1 infected peripheral blood mononuclear cells. Conclusion These data present proof of concept for targeted drug induced elimination of HIV producing cells. While NNRTIs themselves may not be sufficiently potent for therapeutic application, the results provide a basis for the development of drugs exploiting this mechanism of action. PMID:20950436

Diarylpyrimidine-dihydrobenzyloxopyrimidine hybrids: new, wide-spectrum anti-HIV-1 agents active at (sub)-nanomolar level.

PubMed

Rotili, Dante; Tarantino, Domenico; Artico, Marino; Nawrozkij, Maxim B; Gonzalez-Ortega, Emmanuel; Clotet, Bonaventura; Samuele, Alberta; Esté, José A; Maga, Giovanni; Mai, Antonello

2011-04-28

Here, we describe a novel small series of non-nucleoside reverse transcriptase inhibitors (NNRTIs) that combine peculiar structural features of diarylpyrimidines (DAPYs) and dihydro-alkoxy-benzyl-oxopyrimidines (DABOs). These DAPY-DABO hybrids (1-4) showed a characteristic SAR profile and a nanomolar anti-HIV-1 activity at both enzymatic and cellular level. In particular, the two compounds 4d and 2d, with a (sub)nanomolar activity against wild-type and clinically relevant HIV-1 mutant strains, were selected as lead compounds for next optimization studies.

Suicide Inhibitors of Reverse Transcriptase in the Therapy of AIDS and Other Retroviruses

DTIC Science & Technology

1991-07-01

procedures and characterized by IR and NMR spectroscopy . 140 OH py’ridine Ho t O0Tr OHr01 9 10 1 croi 0 0 Aco uO (CH43)2 -CN TOOH pyridja. 0 OxTr D?450...NMR spectroscopy . The observed chemical shifts are comparable to those reported ’in the literature. The spectrum is of the ABX type where the AB part...characterized by NMR spectroscopy . The presence of the characteristic triphosphate group was confirmed by NMR as indicated in the figure below. NMR

Inhibitory Effect of 2,3,5,6-Tetrafluoro-4-[4-(aryl)-1H-1,2,3-triazol-1-yl]benzenesulfonamide Derivatives on HIV Reverse Transcriptase Associated RNase H Activities

PubMed Central

Pala, Nicolino; Esposito, Francesca; Rogolino, Dominga; Carcelli, Mauro; Sanna, Vanna; Palomba, Michele; Naesens, Lieve; Corona, Angela; Grandi, Nicole; Tramontano, Enzo; Sechi, Mario

2016-01-01

The HIV-1 ribonuclease H (RNase H) function of the reverse transcriptase (RT) enzyme catalyzes the selective hydrolysis of the RNA strand of the RNA:DNA heteroduplex replication intermediate, and represents a suitable target for drug development. A particularly attractive approach is constituted by the interference with the RNase H metal-dependent catalytic activity, which resides in the active site located at the C-terminus p66 subunit of RT. Herein, we report results of an in-house screening campaign that allowed us to identify 4-[4-(aryl)-1H-1,2,3-triazol-1-yl]benzenesulfonamides, prepared by the “click chemistry” approach, as novel potential HIV-1 RNase H inhibitors. Three compounds (9d, 10c, and 10d) demonstrated a selective inhibitory activity against the HIV-1 RNase H enzyme at micromolar concentrations. Drug-likeness, predicted by the calculation of a panel of physicochemical and ADME properties, putative binding modes for the active compounds, assessed by computational molecular docking, as well as a mechanistic hypothesis for this novel chemotype are reported. PMID:27556447

Expression of an Mg2+-Dependent HIV-1 RNase H Construct for Drug Screening▿†

PubMed Central

Farias, Richard V.; Vargas, Deborah A.; Castillo, Andres E.; Valenzuela, Beatriz; Coté, Marie L.; Roth, Monica J.; Leon, Oscar

2011-01-01

A single polypeptide of the HIV-1 reverse transcriptase that reconstituted Mg2+-dependent RNase H activity has been made. Using molecular modeling, the construct was designed to encode the p51 subunit joined by a linker to the thumb (T), connection (C), and RNase H (R) domains of p66. This p51-G-TCR construct was purified from the soluble fraction of an Escherichia coli strain, MIC2067(DE3), lacking endogenous RNase HI and HII. The p51-G-TCR RNase H construct displayed Mg2+-dependent activity using a fluorescent nonspecific assay and showed the same cleavage pattern as HIV-1 reverse transcriptase (RT) on substrates that mimic the tRNA removal required for second-strand transfer reactions. The mutant E706Q (E478Q in RT) was purified under similar conditions and was not active. The RNase H of the p51-G-TCR RNase H construct and wild type HIV-1 RT had similar Kms for an RNA-DNA hybrid substrate and showed similar inhibition kinetics to two known inhibitors of the HIV-1 RT RNase H. PMID:21768506

Mechanical allodynia induced by nucleoside reverse transcriptase inhibitor is suppressed by p55TNFSR mediated by herpes simplex virus vector through the SDF1α/CXCR4 system in rats.

PubMed

Huang, Wan; Zheng, Wenwen; Ouyang, Handong; Yi, Hyun; Liu, Shue; Zeng, Weian; Levitt, Roy C; Candiotti, Keith A; Lubarsky, David A; Hao, Shuanglin

2014-03-01

In the human immunodeficiency virus (HIV)-associated sensory neuropathy, neuropathic pain associated with the use of nucleoside reverse transcriptase inhibitors (NRTIs) in patients with HIV/acquired immunodeficiency syndrome is clinically common. While evidence demonstrates that neuropathic pain is influenced by neuroinflammatory events that include the proinflammatory molecules, tumor necrosis factor-α (TNF-α), stromal cell-derived factor 1-α (SDF1-α), and C-X-C chemokine receptor type 4 (CXCR4), the detailed mechanisms by which NRTIs contribute to the development of neuropathic pain are not known. In this study, we investigated the role of these proinflammatory molecules in the dorsal root ganglion (DRG) and the spinal dorsal horn in NRTIs-mediated neuropathic pain state. Neuropathic pain was induced by intraperitoneal administration of 2',3'-dideoxycytidine (ddC, one of the NRTIs). Mechanical threshold was tested using von Frey filament fibers. Nonreplicating herpes simplex virus (HSV) vectors expressing p55 TNF soluble receptor (p55TNFSR) were inoculated into hindpaw of rats. The expression of TNF-α, SDF1-α, and CXCR4 in both the lumbar spinal cord and the L4/5 DRG was examined using Western blots. Intrathecal CXCR4 antagonist was administered. The present study demonstrated that (1) systemic ddC induced upregulation of TNF-α, SDF1-α, and CXCR4 in both the lumbar spinal cord and the L4/5 DRG; (2) p55TNFSR mediated by a nonreplicating HSV vector reversed mechanical allodynia induced by systemic ddC; (3) intrathecal administration of the CXCR4 antagonist AMD3100 increased mechanical threshold; and (4) HSV vector expressing p55TNFSR reversed upregulation of TNF-α, SDF1-α, and CXCR4 induced by ddC in the lumbar spinal dorsal horn and the DRG. Our studies demonstrate that TNF-α through the SDF1/CXCR4 system is involved in the NRTIs-related neuropathic pain state and that blocking the signaling of these proinflammatory molecules is able to reduce NRTIs-related neuropathic pain. These results provide a novel mechanism-based approach (gene therapy) to treating HIV-associated neuropathic pain.

Quantitative Assessment of the Sensitivity of Various Commercial Reverse Transcriptases Based on Armored HIV RNA

PubMed Central

Okello, John B. A.; Rodriguez, Linda; Poinar, Debi; Bos, Kirsten; Okwi, Andrew L.; Bimenya, Gabriel S.; Sewankambo, Nelson K.; Henry, Kenneth R.; Kuch, Melanie; Poinar, Hendrik N.

2010-01-01

Background The in-vitro reverse transcription of RNA to its complementary DNA, catalyzed by the enzyme reverse transcriptase, is the most fundamental step in the quantitative RNA detection in genomic studies. As such, this step should be as analytically sensitive, efficient and reproducible as possible, especially when dealing with degraded or low copy RNA samples. While there are many reverse transcriptases in the market, all claiming to be highly sensitive, there is need for a systematic independent comparison of their applicability in quantification of rare RNA transcripts or low copy RNA, such as those obtained from archival tissues. Methodology/Principal Findings We performed RT-qPCR to assess the sensitivity and reproducibility of 11 commercially available reverse transcriptases in cDNA synthesis from low copy number RNA levels. As target RNA, we used a serially known number of Armored HIV RNA molecules, and observed that 9 enzymes we tested were consistently sensitive to ∼1,000 copies, seven of which were sensitive to ∼100 copies, while only 5 were sensitive to ∼10 RNA template copies across all replicates tested. Despite their demonstrated sensitivity, these five best performing enzymes (Accuscript, HIV-RT, M-MLV, Superscript III and Thermoscript) showed considerable variation in their reproducibility as well as their overall amplification efficiency. Accuscript and Superscript III were the most sensitive and consistent within runs, with Accuscript and Superscript II ranking as the most reproducible enzymes between assays. Conclusions/Significance We therefore recommend the use of Accuscript or Superscript III when dealing with low copy number RNA levels, and suggest purification of the RT reactions prior to downstream applications (eg qPCR) to augment detection. Although the results presented in this study were based on a viral RNA surrogate, and applied to nucleic acid lysates derived from archival formalin-fixed paraffin embedded tissue, their relative performance on RNA obtained from other tissue types may vary, and needs future evaluation. PMID:21085668

Towards novel therapeutics for HIV through fragment-based screening and drug design.

PubMed

Tiefendbrunn, Theresa; Stout, C David

2014-01-01

Fragment-based drug discovery has been applied with varying levels of success to a number of proteins involved in the HIV (Human Immunodeficiency Virus) life cycle. Fragment-based approaches have led to the discovery of novel binding sites within protease, reverse transcriptase, integrase, and gp41. Novel compounds that bind to known pockets within CCR5 have also been identified via fragment screening, and a fragment-based approach to target the TAR-Tat interaction was explored. In the context of HIV-1 reverse transcriptase (RT), fragment-based approaches have yielded fragment hits with mid-μM activity in an in vitro activity assay, as well as fragment hits that are active against drug-resistant variants of RT. Fragment-based drug discovery is a powerful method to elucidate novel binding sites within proteins, and the method has had significant success in the context of HIV proteins.

Coexistence of BRAF V600E and TERT Promoter Mutations in Low-grade Serous Carcinoma of Ovary Recurring as Carcinosarcoma in a Lymph Node: Report of a Case.

PubMed

Tavallaee, Mahkam; Steiner, David F; Zehnder, James L; Folkins, Ann K; Karam, Amer K

2018-04-03

Low-grade serous carcinomas only rarely coexist with or progress to high-grade tumors. We present a case of low-grade serous carcinoma with transformation to carcinosarcoma on recurrence in the lymph node. Identical BRAF V600E and telomerase reverse transcriptase promoter mutations were identified in both the original and recurrent tumor. Given that telomerase reverse transcriptase promotor mutations are thought to play a role in progression of other tumor types, the function of telomerase reverse transcriptase mutations in BRAF mutated low-grade serous carcinoma deserves investigation.

Intravaginal ring delivery of the reverse transcriptase inhibitor TMC 120 as an HIV microbicide.

PubMed

Woolfson, A David; Malcolm, R Karl; Morrow, Ryan J; Toner, Clare F; McCullagh, Stephen D

2006-11-15

TMC 120 (Dapivirine) is a potent non-nucleoside reverse transcriptase inhibitor that is presently being developed as a vaginal HIV microbicide. To date, most vaginal microbicides under clinical investigation have been formulated as single-dose semi-solid gels, designed for application to the vagina before each act of intercourse. However, a clear rationale exists for providing long-term, controlled release of vaginal microbicides in order to afford continuous protection against heterosexually transmitted HIV infection and to improve user compliance. In this study we report on the incorporation of various pharmaceutical excipients into TMC 120 silicone, reservoir-type intravaginal rings (IVRs) in order to modify the controlled release characteristics of the microbicide. The results demonstrate that TMC 120 is released in zero-order fashion from the rings over a 28-day period and that release parameters could be modified by the inclusion of release-modifying excipients in the IVR. The hydrophobic liquid excipient isopropyl myristate had little effect on steady-state daily release rates, but did increase the magnitude and duration of burst release in proportion to excipient loading in the IVR. By comparison, the hydrophobic liquid poly(dimethylsiloxane) had little effect on TMC 120 release parameters. A hydrophilic excipient, lactose, had the surprising effect of decreasing TMC 120 burst release while increasing the apparent steady-state daily release in a concentration-dependent manner. Based on previous cell culture data and vaginal physiology, TMC120 is released from the various ring formulations in amounts potentially capable of maintaining a protective vaginal concentration. It is further predicted that the observed release rates may be maintained for at least a period of 1 year from a single ring device. TMC 120 release profiles and the mechanical properties of rings could be modified by the physicochemical nature of hydrophobic and hydrophilic excipients incorporated into the IVRs.

Neurocognitive Function at the First-Line Failure and on the Second-Line Antiretroviral Therapy in Africa: Analyses From the EARNEST Trial.

PubMed

Kambugu, Andrew; Thompson, Jennifer; Hakim, James; Tumukunde, Dinah; van Oosterhout, Joep J; Mwebaze, Raymond; Hoppe, Anne; Abach, James; Kwobah, Charles; Arenas-Pinto, Alejandro; Walker, Sarah A; Paton, Nicholas I

2016-04-15

To assess neurocognitive function at the first-line antiretroviral therapy failure and change on the second-line therapy. Randomized controlled trial was conducted in 5 sub-Saharan African countries. Patients failing the first-line therapy according to WHO criteria after >12 months on non-nucleoside reverse transcriptase inhibitors-based regimens were randomized to the second-line therapy (open-label) with lopinavir/ritonavir (400 mg/100 mg twice daily) plus either 2-3 clinician-selected nucleoside reverse transcriptase inhibitors, raltegravir, or as monotherapy after 12-week induction with raltegravir. Neurocognitive function was tested at baseline, weeks 48 and 96 using color trails tests 1 and 2, and the Grooved Pegboard test. Test results were converted to an average of the 3 individual test z-scores. A total of 1036 patients (90% of those >18 years enrolled at 13 evaluable sites) had valid baseline tests (58% women, median: 38 years, viral load: 65,000 copies per milliliter, CD4 count: 73 cells per cubic millimeter). Mean (SD) baseline z-score was -2.96 (1.74); lower baseline z-scores were independently associated with older age, lower body weight, higher viral load, lower hemoglobin, less education, fewer weekly working hours, previous central nervous system disease, and taking fluconazole (P < 0.05 in multivariable model). Z-score was increased by mean (SE) of +1.23 (0.04) after 96 weeks on the second-line therapy (P < 0.001; n = 915 evaluable), with no evidence of difference between the treatment arms (P = 0.35). Patients in sub-Saharan Africa failing the first-line therapy had low neurocognitive function test scores, but performance improved on the second-line therapy. Regimens with more central nervous system-penetrating drugs did not enhance neurocognitive recovery indicating this need not be a primary consideration in choosing a second-line regimen.

API consensus guidelines for use of antiretroviral therapy in adults (API-ART guidelines). Endorsed by the AIDS Society of India.

PubMed

Gupta, S B; Pujari, S N; Joshi, S R; Patel, A K

2006-01-01

With rational use of antiretroviral therapy (ART), human immunodeficiency virus (HIV) infection has been transformed into a chronic manageable illness like diabetes and hypertension. These guidelines provide information on state of art, evidence based approach for use of ART in Indian context. When to initiate ART? Antiretroviral therapy is indicated for all symptomatic HIV infected persons regardless of CD4 counts and plasma viral load (PVL) levels. In asymptomatic patients, ART should be offered when the CD4 counts < 200/mm3 and should be considered in patients with CD4 counts between 200-250/mm3. Therapy is not recommended for patients with CD4 count more than 350/ mm3. Involvement of patient in all treatment decisions and assessing readiness is critical before initiating ART. What to start with? A non-nucleoside reverse transcriptase inhibitor (NNRTI) based regimen is recommended for antiretroviral naïve patients. The choice between nevirapine and efavirenz is based on differences in adverse events profiles; cost and availability of convenient fixed dose combinations and need for concomitant use of rifampicin. A backbone of 2-nucleoside reverse transcriptase inhibitors (NRTIs) is combined with the NNRTI. Various combinations and ART strategies not to be used in clinical practice has been enlisted. How to follow up? Recommendations have been made for baseline evaluation and monitoring of patients on ART. These include guidelines on laboratory and clinical evaluation. A plasma viral load at 6 months after initiation of first-line ART is strongly recommended. Yearly estimation of lipid profile has been recommended. How to identify and manage ART failure? The guidelines recognize the issue of identifying ART failure late if only CD4 counts are used for monitoring. In the absence of resistance testing various second-line regimens have been enlisted. A boosted protease inhibitor based regimen is recommended in this situation to be combined with 2-NRTIs. Special situations Recommendations have been made for use of ART in HIV-TB, HIV-HBV, and HIV-HCV co-infected patients. In patients with active TB and a CD4 count < 200/mm3, initiation of ART is recommended as soon as the anti-TB treatment is tolerated. Efavirenz is the only ARV drug, which can be safely used with rifampicin. In pregnancy use of single dose nevirapine for reducing risk of mother to child transmission of HIV is not recommended, because of the risk of development of resistance. For post-exposure prophylaxis taking ART treatment history of the source patient is crucial in designing an effective regimen.

High-throughput screening using pseudotyped lentiviral particles: a strategy for the identification of HIV-1 inhibitors in a cell-based assay.

PubMed

Garcia, Jean-Michel; Gao, Anhui; He, Pei-Lan; Choi, Joyce; Tang, Wei; Bruzzone, Roberto; Schwartz, Olivier; Naya, Hugo; Nan, Fa-Jun; Li, Jia; Altmeyer, Ralf; Zuo, Jian-Ping

2009-03-01

Two decades after its discovery the human immunodeficiency virus (HIV) is still spreading worldwide and killing millions. There are 25 drugs formally approved for HIV currently on the market, but side effects as well as the emergence of HIV strains showing single or multiple resistances to current drug-therapy are causes for concern. Furthermore, these drugs target only 4 steps of the viral cycle, hence the urgent need for new drugs and also new targets. In order to tackle this problem, we have devised a cell-based assay using lentiviral particles to look for post-entry inhibitors of HIV-1. We report here the assay development, validation as well as confirmation of the hits using both wild-type and drug-resistant HIV-1 viruses. The screening was performed on an original library, rich in natural compounds and pure molecules from Traditional Chinese Medicine pharmacopoeia, which had never been screened for anti-HIV activity. The identified hits belong to four chemical sub-families that appear to be all non-nucleoside reverse transcriptase inhibitors (NNRTIs). Secondary tests with live viruses showed that there was good agreement with pseudotyped particles, confirming the validity of this approach for high-throughput drug screens. This assay will be a useful tool that can be easily adapted to screen for inhibitors of viral entry.

Elevated Human telomerase reverse transcriptase gene expression in blood cells associated with chronic and arsenic exposure in Inner Mongolia, China

EPA Science Inventory

BACKGROUND: Arsenic exposure is associated with human cancer. Telomerase containing the catalytic subunit, human telomerase reverse transcriptase (hTERT), can extend telomeres of chromosomes, delay senescence and promoting cell proliferation leading to tumorigenesis. OBJECTIVE:...

Multiple nucleotide preferences determine cleavage-site recognition by the HIV-1 and M-MuLV RNases H.

PubMed

Schultz, Sharon J; Zhang, Miaohua; Champoux, James J

2010-03-19

The RNase H activity of reverse transcriptase is required during retroviral replication and represents a potential target in antiviral drug therapies. Sequence features flanking a cleavage site influence the three types of retroviral RNase H activity: internal, DNA 3'-end-directed, and RNA 5'-end-directed. Using the reverse transcriptases of HIV-1 (human immunodeficiency virus type 1) and Moloney murine leukemia virus (M-MuLV), we evaluated how individual base preferences at a cleavage site direct retroviral RNase H specificity. Strong test cleavage sites (designated as between nucleotide positions -1 and +1) for the HIV-1 and M-MuLV enzymes were introduced into model hybrid substrates designed to assay internal or DNA 3'-end-directed cleavage, and base substitutions were tested at specific nucleotide positions. For internal cleavage, positions +1, -2, -4, -5, -10, and -14 for HIV-1 and positions +1, -2, -6, and -7 for M-MuLV significantly affected RNase H cleavage efficiency, while positions -7 and -12 for HIV-1 and positions -4, -9, and -11 for M-MuLV had more modest effects. DNA 3'-end-directed cleavage was influenced substantially by positions +1, -2, -4, and -5 for HIV-1 and positions +1, -2, -6, and -7 for M-MuLV. Cleavage-site distance from the recessed end did not affect sequence preferences for M-MuLV reverse transcriptase. Based on the identified sequence preferences, a cleavage site recognized by both HIV-1 and M-MuLV enzymes was introduced into a sequence that was otherwise resistant to RNase H. The isolated RNase H domain of M-MuLV reverse transcriptase retained sequence preferences at positions +1 and -2 despite prolific cleavage in the absence of the polymerase domain. The sequence preferences of retroviral RNase H likely reflect structural features in the substrate that favor cleavage and represent a novel specificity determinant to consider in drug design. Copyright (c) 2010 Elsevier Ltd. All rights reserved.

Lopinavir plus nucleoside reverse-transcriptase inhibitors, lopinavir plus raltegravir, or lopinavir monotherapy for second-line treatment of HIV (EARNEST): 144-week follow-up results from a randomised controlled trial.

PubMed

Hakim, James G; Thompson, Jennifer; Kityo, Cissy; Hoppe, Anne; Kambugu, Andrew; van Oosterhout, Joep J; Lugemwa, Abbas; Siika, Abraham; Mwebaze, Raymond; Mweemba, Aggrey; Abongomera, George; Thomason, Margaret J; Easterbrook, Philippa; Mugyenyi, Peter; Walker, A Sarah; Paton, Nicholas I

2018-01-01

Millions of HIV-infected people worldwide receive antiretroviral therapy (ART) in programmes using WHO-recommended standardised regimens. Recent WHO guidelines recommend a boosted protease inhibitor plus raltegravir as an alternative second-line combination. We assessed whether this treatment option offers any advantage over the standard protease inhibitor plus two nucleoside reverse-transcriptase inhibitors (NRTIs) second-line combination after 144 weeks of follow-up in typical programme settings. We analysed the 144-week outcomes at the completion of the EARNEST trial, a randomised controlled trial done in HIV-infected adults or adolescents in 14 sites in five sub-Saharan African countries (Uganda, Zimbabwe, Malawi, Kenya, Zambia). Participants were those who were no longer responding to non-NRTI-based first-line ART, as assessed with WHO criteria, confirmed by viral-load testing. Participants were randomly assigned to receive a ritonavir-boosted protease inhibitor (lopinavir 400 mg with ritonavir 100 mg, twice per day) plus two or three clinician-selected NRTIs (protease inhibitor plus NRTI group), protease inhibitor plus raltegravir (400 mg twice per day; protease inhibitor plus raltegravir group), or protease inhibitor monotherapy (plus raltegravir induction for first 12 weeks, re-intensified to combination therapy after week 96; protease inhibitor monotherapy group). Randomisation was by computer-generated randomisation sequence, with variable block size. The primary outcome was viral load of less than 400 copies per mL at week 144, for which we assessed non-inferiority with a one-sided α of 0·025, and superiority with a two-sided α of 0·025. The EARNEST trial is registered with ISRCTN, number 37737787. Between April 12, 2010, and April 29, 2011, 1837 patients were screened for eligibility, of whom 1277 patients were randomly assigned to an intervention group. In the primary (complete-case) analysis at 144 weeks, 317 (86%) of 367 in the protease inhibitor plus NRTI group had viral loads of less than 400 copies per mL compared with 312 (81%) of 383 in the protease inhibitor plus raltegravir group (p=0·07; lower 95% confidence limit for difference 10·2% vs specified non-inferiority margin 10%). In the protease inhibitor monotherapy group, 292 (78%) of 375 had viral loads of less than 400 copies per mL; p=0·003 versus the protease inhibitor plus NRTI group at 144 weeks. There was no difference between groups in serious adverse events, grade 3 or 4 adverse events (total or ART-related), or events that resulted in treatment modification. Protease inhibitor plus raltegravir offered no advantage over protease inhibitor plus NRTI in virological efficacy or safety. In the primary analysis, protease inhibitor plus raltegravir did not meet non-inferiority criteria. A regimen of protease inhibitor with NRTIs remains the best standardised second-line regimen for use in programmes in resource-limited settings. European and Developing Countries Clinical Trials Partnership (EDCTP), UK Medical Research Council, Instituto de Salud Carlos III, Irish Aid, Swedish International Development Cooperation Agency, Instituto Superiore di Sanita, Merck, ViiV Healthcare, WHO. Copyright © 2018 The Author(s). Published by Elsevier Ltd. This is an Open Access article under the CC BY 4.0 license. Published by Elsevier Ltd.. All rights reserved.

Podoscyphic acid, a new inhibitor of avian myeloblastosis virus and Moloney murine leukemia virus reverse transcriptase from a Podoscypha species.

PubMed

Erkel, G; Anke, T; Velten, R; Steglich, W

1991-01-01

A novel enzyme inhibitor of RNA-directed DNA-polymerases of avian myeloblastosis and murine leukemia virus was isolated from fermentations of an tasmanian Podoscypha species. Its structure was elucidated by spectroscopic methods and oxidative degradation as (E)-4,5-dioxo-2-hexadecenoic acid (1). The enzyme inhibitor, which was named podoscyphic acid, did not inhibit DNA and RNA synthesis in permeabilized L 1210 cells nor did it affect RNA synthesis in isolated nuclei of L 1210 cells. 1 inhibits protein synthesis in whole L 1210 cells and rabbit reticulocyte lysate and shows very weak antimicrobial and cytotoxic properties. The testing of ethyl (E)-4,5-dioxo-2-hexadecenoate (2) and (E)-4-oxo-2-tetradecenoic acid (11) revealed the importance of the free gamma-oxoacrylic acid unit for the biological activities of 1.

Application of 3D-QSAR, Pharmacophore, and Molecular Docking in the Molecular Design of Diarylpyrimidine Derivatives as HIV-1 Nonnucleoside Reverse Transcriptase Inhibitors.

PubMed

Liu, Genyan; Wang, Wenjie; Wan, Youlan; Ju, Xiulian; Gu, Shuangxi

2018-05-11

Diarylpyrimidines (DAPYs), acting as HIV-1 nonnucleoside reverse transcriptase inhibitors (NNRTIs), have been considered to be one of the most potent drug families in the fight against acquired immunodeficiency syndrome (AIDS). To better understand the structural requirements of HIV-1 NNRTIs, three-dimensional quantitative structure⁻activity relationship (3D-QSAR), pharmacophore, and molecular docking studies were performed on 52 DAPY analogues that were synthesized in our previous studies. The internal and external validation parameters indicated that the generated 3D-QSAR models, including comparative molecular field analysis (CoMFA, q 2 = 0.679, R 2 = 0.983, and r pred 2 = 0.884) and comparative molecular similarity indices analysis (CoMSIA, q 2 = 0.734, R 2 = 0.985, and r pred 2 = 0.891), exhibited good predictive abilities and significant statistical reliability. The docking results demonstrated that the phenyl ring at the C₄-position of the pyrimidine ring was better than the cycloalkanes for the activity, as the phenyl group was able to participate in π⁻π stacking interactions with the aromatic residues of the binding site, whereas the cycloalkanes were not. The pharmacophore model and 3D-QSAR contour maps provided significant insights into the key structural features of DAPYs that were responsible for the activity. On the basis of the obtained information, a series of novel DAPY analogues of HIV-1 NNRTIs with potentially higher predicted activity was designed. This work might provide useful information for guiding the rational design of potential HIV-1 NNRTI DAPYs.

Incomplete IgG response to HIV-1 proteins and low avidity levels in recently converted HIV patients treated with early antiretroviral therapy.

PubMed

Re, Maria Carla; Schiavone, Pasqua; Bon, Isabella; Vitone, Francesca; De Crignis, Elisa; Biagetti, Carlo; Gibellini, Davide

2010-11-01

To evaluate the evolution of antibody avidity and Western blot reactivity in recently infected HIV-1 subjects and to study the impact of highly active antiretroviral therapy (HAART) on avidity maturation of HIV-1-specific immunoglobulin G (IgG) in patients with recent HIV-1 infection. Thirty-six HIV-1 seroconverters were enrolled in this study and followed longitudinally over 24 months to evaluate if the administration of antiretroviral therapy during primary infection affects Western blot reactivity and the evolution of antibody avidity. The patients were divided into two groups; group A consisted of 19 HIV-1-untreated patients who did not receive any drug treatment during our follow-up period; group B consisted of 17 subjects who were treated early with an association of two nucleoside reverse transcriptase inhibitors (NRTI) and one non-nucleoside reverse transcriptase inhibitor (NNRTI) within 3 months after seroconversion. At diagnosis, Western blot analysis and avidity index (mean value) were exactly matched in untreated and treated patients; subsequently, however, a significantly lower reactivity to HIV-1 pol and gag proteins and a lower avidity index (mean values) were observed in HAART-treated patients up until the end of the follow-up period. The impaired production and maturation of the humoral immunological response in antiretroviral-treated patients might be related to a rapid suppression of HIV replication, driven by HAART. These results could have important implications in understanding the complex mechanism of the immune response during HIV infection. Copyright © 2010 International Society for Infectious Diseases. Published by Elsevier Ltd. All rights reserved.

Off-Target Effects of Drugs that Disrupt Human Mitochondrial DNA Maintenance

PubMed Central

Young, Matthew J.

2017-01-01

Nucleoside reverse transcriptase inhibitors (NRTIs) were the first drugs used to treat human immunodeficiency virus (HIV) the cause of acquired immunodeficiency syndrome. Development of severe mitochondrial toxicity has been well documented in patients infected with HIV and administered NRTIs. In vitro biochemical experiments have demonstrated that the replicative mitochondrial DNA (mtDNA) polymerase gamma, Polg, is a sensitive target for inhibition by metabolically active forms of NRTIs, nucleotide reverse transcriptase inhibitors (NtRTIs). Once incorporated into newly synthesized daughter strands NtRTIs block further DNA polymerization reactions. Human cell culture and animal studies have demonstrated that cell lines and mice exposed to NRTIs display mtDNA depletion. Further complicating NRTI off-target effects on mtDNA maintenance, two additional DNA polymerases, Pol beta and PrimPol, were recently reported to localize to mitochondria as well as the nucleus. Similar to Polg, in vitro work has demonstrated both Pol beta and PrimPol incorporate NtRTIs into nascent DNA. Cell culture and biochemical experiments have also demonstrated that antiviral ribonucleoside drugs developed to treat hepatitis C infection act as off-target substrates for POLRMT, the mitochondrial RNA polymerase and primase. Accompanying the above-mentioned topics, this review examines: (1) mtDNA maintenance in human health and disease, (2) reports of DNA polymerases theta and zeta (Rev3) localizing to mitochondria, and (3) additional drugs with off-target effects on mitochondrial function. Lastly, mtDNA damage may induce cell death; therefore, the possibility of utilizing compounds that disrupt mtDNA maintenance to kill cancer cells is discussed. PMID:29214156

A novel nonnucleoside analogue that inhibits human immunodeficiency virus type 1 isolates resistant to current nonnucleoside reverse transcriptase inhibitors.

PubMed

Zhang, Zhijun; Xu, Wen; Koh, Yung-Hyo; Shim, Jae Hoon; Girardet, Jean-Luc; Yeh, Li-Tain; Hamatake, Robert K; Hong, Zhi

2007-02-01

Nonnucleoside reverse transcriptase (RT) inhibitors (NNRTIs) are important components of current combination therapies for human immunodeficiency virus type 1 (HIV-1) infection. However, their low genetic barriers against resistance development, cross-resistance, and serious side effects can compromise the benefits of the two current drugs in this class (efavirenz and nevirapine). In this study, we report a novel and potent NNRTI, VRX-480773, that inhibits viruses from efavirenz-resistant molecular clones and most NNRTI-resistant clinical HIV-1 isolates tested. In vitro mutation selection experiments revealed that longer times were required for viruses to develop resistance to VRX-480773 than to efavirenz. RT mutations selected by VRX-480773 after 3 months of cell culture in the presence of 1 nM VRX-480773 carried the Y181C mutation, resulting in a less-than-twofold increase in resistance to the compound. A virus containing the double mutation V106I-Y181C emerged after 4 months, causing a sixfold increase in resistance. Viruses containing additional mutations of D123G, F227L, and T369I emerged when the cultures were incubated with increasing concentrations of VRX-480773. Most of the resistant viruses selected by VRX-480773 are susceptible to efavirenz. Oral administration of VRX-480773 to dogs resulted in plasma concentrations that were significantly higher than those required for the inhibition of wild-type and mutant viruses. These results warrant further clinical development of VRX-480773 for the treatment of HIV infection in both NNRTI-naive and -experienced patients.

3D-QSAR CoMFA of a series of DABO derivatives as HIV-1 reverse transcriptase non-nucleoside inhibitors.

PubMed

de Brito, Monique Araújo; Rodrigues, Carlos Rangel; Cirino, José Jair Vianna; de Alencastro, Ricardo Bicca; Castro, Helena Carla; Albuquerque, Magaly Girão

2008-08-01

A series of 74 dihydroalkoxybenzyloxopyrimidines (DABOs), a class of highly potent non-nucleoside reverse transcriptase inhibitors (NNRTIs), was retrieved from the literature and studied by comparative molecular field analysis (CoMFA) in order to derive three-dimensional quantitative structure-activity relationship (3D-QSAR) models. The CoMFA study has been performed with a training set of 59 compounds, testing three alignments and four charge schemes (DFT, HF, AM1, and PM3) and using defaults probe atom (Csp (3), +1 charge), cutoffs (30 kcal.mol (-1) for both steric and electrostatic fields), and grid distance (2.0 A). The best model ( N = 59), derived from Alignment 1 and PM3 charges, shows q (2) = 0.691, SE cv = 0.475, optimum number of components = 6, r (2) = 0.930, SEE = 0.226, and F-value = 115.544. The steric and electrostatic contributions for the best model were 43.2% and 56.8%, respectively. The external predictive ability (r (2) pred = 0.918) of the resultant best model was evaluated using a test set of 15 compounds. In order to design more potent DABO analogues as anti-HIV/AIDS agents, attention should be taken in order to select a substituent for the 4-oxopyrimidine ring, since, as revealed by the best CoMFA model, there are a steric restriction at the C2-position, a electron-rich group restriction at the C6-position ( para-substituent of the 6-benzyl group), and a steric allowed region at the C5-position.

Antitumor Activity and Mechanism of a Reverse Transcriptase Inhibitor, Dapivirine, in Glioblastoma

PubMed Central

Liu, Weiwen; Song, Xian-lu; Zhao, Shan-chao; He, Minyi; Wang, Hai; Chen, Ziyang; Xiang, Wei; Yi, Guozhong; Qi, Songtao; Liu, Yawei

2018-01-01

Ethnopharmacological relevance: Dapivirine is one of reverse transcriptase inhibitors (RTIs). It is the prototype of diarylpyrimidines (DAPY), formerly known as TMC120 or DAPY R147681 (IUPAC name: 4- [[4-(2, 4, 6-trimethylphenyl) amino]-2-pyrimidinyl] amino]-benzonitrile; CAS no.244767-67-7). Aim: The purpose of this study is to investigate the antitumor activity of dapivirine, one of the RTIs, on U87 glioblastoma (GBM) cells in vitro and in vivo. Materials and Methods: U87 GBM cells were cultured and treated with or without dapivirine. Cell viability was evaluated by CCK-8 (Cell Counting Kit 8, CCK-8) assay; apoptosis was analyzed by flow cytometry; cell migration was evaluated by Boyden Chamber assay; Western blotting was performed to detect proteins related to apoptosis, epithelial-to-mesenchymal transition and autophagy. PathScan intracellular signaling array kit was used to detect important and well-characterized signaling molecules. Tumor xenograft model in nude mice was used to evaluate the antitumorigenic effect in vivo. Results: Dapivirine weakened proliferation of glioma cells and induced the apoptosis of U87 glioblastoma cells. Furthermore, dapivirine regulated autophagy and induced Akt, Bad and SAPK/JNK activations. Moreover, the inhibition of glioma cell growth by dapivirine was also observed in nude mice in vivo. Conclusion: In summary, in our study dapivirine exposure induces stress, resulting in JNK and PI3K/Akt pathway activation through diminished inhibition of the apoptosis and autophagy cascade in U87 GBM cells, which inhibits cell growth in vitro and in vivo. PMID:29290776

Selective serotonin reuptake inhibitor suppression of HIV infectivity and replication.

PubMed

Benton, Tami; Lynch, Kevin; Dubé, Benoit; Gettes, David R; Tustin, Nancy B; Ping Lai, Jian; Metzger, David S; Blume, Joshua; Douglas, Steven D; Evans, Dwight L

2010-11-01

To test the hypothesis that the selective serotonin reuptake inhibitor (SSRI) citalopram would down-regulate human immunodeficiency virus (HIV) infectivity and that the greatest effects would be seen in people with depression. Depression is a risk factor for morbidity and mortality in HIV/acquired immune deficiency syndrome. Serotonin (5-HT) neurotransmission has been implicated in the pathobiology of depression, and pharmacologic therapies for depression target this system. The 5-HT transporter and 5-HT receptors are widely distributed throughout the central nervous and immune systems. Depression has been associated with suppression of natural killer cells and CD8(+) lymphocytes, key regulators of HIV infection. Ex vivo models for acute and chronic HIV infection were used to study the effects of citalopram on HIV viral infection and replication in 48 depressed and nondepressed women. For both the acute and chronic infection models, HIV reverse transcriptase activity was measured in the citalopram treatment condition and the control condition. The SSRI significantly down-regulated the reverse transcriptase response in both the acute and chronic infection models. Specifically, citalopram significantly decreased the acute HIV infectivity of macrophages. Citalopram also significantly decreased HIV viral replication in the latently infected T-cell line and in the latently infected macrophage cell line. There was no difference in down-regulation by depression status. These studies suggest that an SSRI enhances natural killer/CD8 noncytolytic HIV suppression in HIV/acquired immune deficiency syndrome and decreases HIV viral infectivity of macrophages, ex vivo, suggesting the need for in vivo studies to determine a potential role for agents targeting serotonin in the host defense against HIV.

Standardized comparison of the relative impacts of HIV-1 reverse transcriptase (RT) mutations on nucleoside RT inhibitor susceptibility.

PubMed

Melikian, George L; Rhee, Soo-Yon; Taylor, Jonathan; Fessel, W Jeffrey; Kaufman, David; Towner, William; Troia-Cancio, Paolo V; Zolopa, Andrew; Robbins, Gregory K; Kagan, Ron; Israelski, Dennis; Shafer, Robert W

2012-05-01

Determining the phenotypic impacts of reverse transcriptase (RT) mutations on individual nucleoside RT inhibitors (NRTIs) has remained a statistical challenge because clinical NRTI-resistant HIV-1 isolates usually contain multiple mutations, often in complex patterns, complicating the task of determining the relative contribution of each mutation to HIV drug resistance. Furthermore, the NRTIs have highly variable dynamic susceptibility ranges, making it difficult to determine the relative effect of an RT mutation on susceptibility to different NRTIs. In this study, we analyzed 1,273 genotyped HIV-1 isolates for which phenotypic results were obtained using the PhenoSense assay (Monogram, South San Francisco, CA). We used a parsimonious feature selection algorithm, LASSO, to assess the possible contributions of 177 mutations that occurred in 10 or more isolates in our data set. We then used least-squares regression to quantify the impact of each LASSO-selected mutation on each NRTI. Our study provides a comprehensive view of the most common NRTI resistance mutations. Because our results were standardized, the study provides the first analysis that quantifies the relative phenotypic effects of NRTI resistance mutations on each of the NRTIs. In addition, the study contains new findings on the relative impacts of thymidine analog mutations (TAMs) on susceptibility to abacavir and tenofovir; the impacts of several known but incompletely characterized mutations, including E40F, V75T, Y115F, and K219R; and a tentative role in reduced NRTI susceptibility for K64H, a novel NRTI resistance mutation.

In vitro transport characteristics of EFdA, a novel nucleoside reverse transcriptase inhibitor using Caco-2 and MDCKII cell monolayers.

PubMed

Zhang, Wei; Parniak, Michael A; Sarafianos, Stefan G; Empey, Philip E; Rohan, Lisa C

2014-06-05

4'-Ethynyl-2-fluoro-2'-deoxyadenosine (EFdA) is a novel nucleoside reverse transcriptase inhibitor with a unique mechanism of action and highly potent activity against both wild-type and clinically relevant drug resistant HIV-1 variants. Furthermore, in vivo efficacy and safety evaluations have shown EFdA to be a promising therapeutic candidate for use in the treatment of HIV infection. However, little is known about the pharmacokinetic and biopharmaceutical properties of EFdA. In this study, we evaluated cellular EFdA transport using Caco-2 and Madin-Darby Canine Kidney II (MDCKII) in vitro cell models. Studies using Caco-2 cell monolayers showed that EFdA efflux ratios were >2.0, suggesting that active drug transport mechanisms may play a role in EFdA flux. ABCB1 transporter (PGP1) inhibition was assessed using the acetomethoxy derivate of calcein (calcein-AM) as a fluorescent probe in both wild-type MDCKII and PGP1 overexpressing MDCKII cells. Nonetheless, our data showed that EFdA is not a substrate of PGP1. Additionally, comparative bidirectional flux of EFdA and Lucifer yellow (LY, a well-known paracellular marker) was studied over a range of EFdA concentrations. In MDCKII monolayers, EFdA had an apparent permeability coefficient (Papp) (a-b) of <1×10(-6)cm/s. The Papp values significantly increased in the presence of the paracellular permeability enhancer, indicating that EFdA primarily permeates via the paracellular route. Copyright © 2014 Elsevier B.V. All rights reserved.

Evaluation and Management of Dyslipidemia in Patients with HIV Infection

PubMed Central

Green, Michael L

2002-01-01

OBJECTIVE Persons with HIV infection develop metabolic abnormalities related to their antiretroviral therapy and HIV infection itself. The objective of this study was to summarize the emerging evidence for the incidence, etiology, health risks, and treatment of dyslipidemias in HIV disease. DESIGN Systematic review of original research with quantitative synthesis. MAIN RESULTS Dyslipidemia is common in persons with HIV infection on highly active antiretroviral therapy (HAART), but methodologic differences between studies preclude precise estimates of prevalence and incidence. The typical pattern includes elevated total cholesterol, low-density lipoprotein cholesterol, and triglycerides, which may be markedly elevated. The dyslipidemia may be associated with lipodystrophy, insulin resistance, and, rarely, frank diabetes mellitus. Exposure to protease inhibitors (PIs) is associated with this entire range of metabolic abnormalities. PI-naïve patients on nucleoside reverse transcriptase inhibitors (NRTIs) may develop lipodystrophy, insulin resistance, hypercholesterolemia, and possibly modest elevations in triglycerides but not severe hypertriglyceridemia, which appears to be linked to PIs alone. Most studies have not found an association between CD4 lymphocyte count or HIV viral load and lipid abnormalities. The pathogenesis is incompletely understood and appears to be multifactorial. There are insufficient data to definitively support an increased coronary heart disease risk in patients with HIV-related dyslipidemia. However, some of the same metabolic abnormalities remain firmly established risk factors in other populations. Patients on HAART with severe hypertriglyceridemia may develop pancreatitis or other manifestations of the chylomicronemia syndrome. Some of the metabolic derangements (particularly hypertriglyceridemia) may improve upon replacing a PI with a non-nucleoside reverse transcriptase inhibitor. The limited experience suggests that fibrates, pravastatin, and atorvastatin can safely treat lipid abnormalities in HIV-infected patients. CONCLUSIONS Patients with HIV infection on HAART should be screened for lipid disorders, given their incidence, potential for morbidity, and possible long-term cardiovascular risk. Treatment decisions are complex and must include assessments of cardiac risk, HIV infection status, reversibility of the dyslipidemia, and the effectiveness and toxicities of lipid-lowering medications. The multiple potential drug interactions with antiretroviral or other HIV-related medications should be considered in lipid-lowering drug selection and monitoring. PMID:12390557

6-(1-Benzyl-1H-pyrrol-2-yl)-2,4-dioxo-5-hexenoic acids as dual inhibitors of recombinant HIV-1 integrase and ribonuclease H, synthesized by a parallel synthesis approach.

PubMed

Costi, Roberta; Métifiot, Mathieu; Esposito, Francesca; Cuzzucoli Crucitti, Giuliana; Pescatori, Luca; Messore, Antonella; Scipione, Luigi; Tortorella, Silvano; Zinzula, Luca; Novellino, Ettore; Pommier, Yves; Tramontano, Enzo; Marchand, Christophe; Di Santo, Roberto

2013-11-14

The increasing efficiency of HAART has helped to transform HIV/AIDS into a chronic disease. Still, resistance and drug-drug interactions warrant the development of new anti-HIV agents. We previously discovered hit 6, active against HIV-1 replication and targeting RNase H in vitro. Because of its diketo-acid moiety, we speculated that this chemotype could serve to develop dual inhibitors of both RNase H and integrase. Here, we describe a new series of 1-benzyl-pyrrolyl diketohexenoic derivatives, 7a-y and 8a-y, synthesized following a parallel solution-phase approach. Those 50 analogues have been tested on recombinant enzymes (RNase H and integrase) and in cell-based assays. Approximately half (22) exibited inhibition of HIV replication. Compounds 7b, 7u, and 8g were the most active against the RNase H activity of reverse-transcriptase, with IC50 values of 3, 3, and 2.5 μM, respectively. Compound 8g was also the most potent integrase inhibitor with an IC50 value of 26 nM.

Broadening the use of antiretroviral therapy: the case for feline leukemia virus

PubMed Central

Greggs, Willie M; Clouser, Christine L; Patterson, Steven E; Mansky, Louis M

2011-01-01

Antiretroviral drugs have saved and extended the lives of millions of individuals infected with HIV. The major classes of anti-HIV drugs include reverse transcriptase inhibitors, protease inhibitors, integrase inhibitors, and entry/fusion inhibitors. While antiretroviral drug regimens are not commonly used to treat other types of retroviral infections, there are instances where there is a perceived need for re-evaluation of the benefits of antiretroviral therapy. One case in point is that of feline leukemia virus (FeLV), an infection of companion felines. While vaccines exist to prevent FeLV infection and spread, they have not eliminated FeLV infection. For FeLV-infected felines and their human companions, antiretroviral therapy would be desirable and of practical importance if good options were available. Here, we discuss FeLV biology and current treatment options, and propose that there is a need for antiretroviral treatment options for FeLV infection. The comparative use and analysis of antiretroviral therapy can provide new insights into the mechanism of antiretroviral drug action. PMID:21479142

Generation and Characterization of a Defective HIV-1 Virus as an Immunogen for a Therapeutic Vaccine

PubMed Central

García-Pérez, Javier; García, Felipe; Blanco, Julia; Escribà-García, Laura; Gatell, Jose Maria; Alcamí, Jose; Plana, Montserrat; Sánchez-Palomino, Sonsoles

2012-01-01

Background The generation of new immunogens able to elicit strong specific immune responses remains a major challenge in the attempts to obtain a prophylactic or therapeutic vaccine against HIV/AIDS. We designed and constructed a defective recombinant virus based on the HIV-1 genome generating infective but non-replicative virions able to elicit broad and strong cellular immune responses in HIV-1 seropositive individuals. Results Viral particles were generated through transient transfection in producer cells (293-T) of a full length HIV-1 DNA carrying a deletion of 892 base pairs (bp) in the pol gene encompassing the sequence that codes for the reverse transcriptase (NL4-3/ΔRT clone). The viral particles generated were able to enter target cells, but due to the absence of reverse transcriptase no replication was detected. The immunogenic capacity of these particles was assessed by ELISPOT to determine γ-interferon production in a cohort of 69 chronic asymptomatic HIV-1 seropositive individuals. Surprisingly, defective particles produced from NL4-3/ΔRT triggered stronger cellular responses than wild-type HIV-1 viruses inactivated with Aldrithiol-2 (AT-2) and in a larger proportion of individuals (55% versus 23% seropositive individuals tested). Electron microscopy showed that NL4-3/ΔRT virions display immature morphology. Interestingly, wild-type viruses treated with Amprenavir (APV) to induce defective core maturation also induced stronger responses than the same viral particles generated in the absence of protease inhibitors. Conclusions We propose that immature HIV-1 virions generated from NL4-3/ΔRT viral clones may represent new prototypes of immunogens with a safer profile and stronger capacity to induce cellular immune responses than wild-type inactivated viral particles. PMID:23144996

Substrate mimicry: HIV-1 reverse transcriptase recognizes 6-modified-3′-azido-2′,3′-dideoxyguanosine-5′-triphosphates as adenosine analogs

PubMed Central

Herman, Brian D.; Schinazi, Raymond F.; Zhang, Hong-wang; Nettles, James H.; Stanton, Richard; Detorio, Mervi; Obikhod, Aleksandr; Pradère, Ugo; Coats, Steven J.; Mellors, John W.; Sluis-Cremer, Nicolas

2012-01-01

β-D-3′-Azido-2′,3′-dideoxyguanosine (3′-azido-ddG) is a potent inhibitor of HIV-1 replication with a superior resistance profile to zidovudine. Recently, we identified five novel 6-modified-3′-azido-ddG analogs that exhibit similar or superior anti-HIV-1 activity compared to 3′-azido-ddG in primary cells. To gain insight into their structure–activity–resistance relationships, we synthesized their triphosphate (TP) forms and assessed their ability to inhibit HIV-1 reverse transcriptase (RT). Steady-state and pre-steady-state kinetic experiments show that the 6-modified-3′-azido-ddGTP analogs act as adenosine rather than guanosine mimetics in DNA synthesis reactions. The order of potency of the TP analogs against wild-type RT was: 3′-azido-2,6-diaminopurine >3′-azido-6-chloropurine; 3′-azido-6-N-allylaminopurine > 2-amino-6-N,N-dimethylaminopurine; 2-amino-6-methoxypurine. Molecular modeling studies reveal unique hydrogen-bonding interactions between the nucleotide analogs and the template thymine base in the active site of RT. Surprisingly, the structure–activity relationship of the analogs differed in HIV-1 RT ATP-mediated excision assays of their monophosphate forms, suggesting that it may be possible to rationally design a modified base analog that is efficiently incorporated by RT but serves as a poor substrate for ATP-mediated excision reactions. Overall, these studies identify a promising strategy to design novel nucleoside analogs that exert profound antiviral activity against both WT and drug-resistant HIV-1. PMID:21914723

Reverse transcriptase activity and particles of retroviral density in cultured canine lymphosarcoma supernatants.

PubMed Central

Tomley, F. M.; Armstrong, S. J.; Mahy, B. W.; Owen, L. N.

1983-01-01

Lymphoid tissue from 43 cases of canine lymphosarcoma and from 40 clinically normal dogs have been examined for markers of retrovirus infection. From 69-76% of culture supernatants from lymphosarcomas were shown to contain particles of retroviral density and to possess poly rC-oligo dG templated polymerase (reverse transcriptase) activity compared with 17-24% of culture supernatants from normal canine lymphoid cells. In 6 culture supernatants from cases of lymphosarcoma, high molecular weight 60-70S RNA was detected and shown to be found in association with this particulate reverse transcriptase activity. No such RNA was detected in 6 culture supernatants from normal canine lymphoid cells. PMID:6186265

Development and validation of a rapid reversed-phase HPLC method for the determination of the non-nucleoside reverse transcriptase inhibitor dapivirine from polymeric nanoparticles.

PubMed

das Neves, José; Sarmento, Bruno; Amiji, Mansoor M; Bahia, Maria Fernanda

2010-06-05

The objective of this work was to develop and validate a rapid reversed-phase (RP) high-performance liquid chromatography (HPLC) method for the in vitro pharmaceutical characterization of dapivirine-loaded polymeric nanoparticles. Chromatographic runs were performed on a RP C18 column with a mobile phase comprising acetonitrile-0.5% (w/v) triethanolamine solution in isocratic mode (80:20, v/v) at a flow rate of 1 ml/min. Dapivirine was detected at a wavelength of 290 nm. The method was shown to be specific, linear in the range of 1-50 microg/ml (R(2)=0.9998), precise at the intra-day and inter-day levels as reflected by the relative standard deviation values (less than 0.85%), accurate (recovery rate of 100.17+/-0.35%), and robust to changes in the mobile phase and column brand. The detection and quantitation limits were 0.08 and 0.24 microg/ml, respectively. The method was successfully used to determine the loading capacity and association efficiency of dapivirine in poly(lactic-co-glycolic acid)-based nanoparticles and its in vitro release. Copyright (c) 2010 Elsevier B.V. All rights reserved.

Broad advances in understanding HIV resistance to antiretrovirals: report on the XVII International HIV Drug Resistance Workshop.

PubMed

Mascolini, Mark; Larder, Brendan A; Boucher, Charles A B; Richman, Douglas D; Mellors, John W

2008-01-01

The 2008 International HIV Drug Resistance Workshop explored six topics on viral resistance: new antiretrovirals; clinical implications; epidemiology; new technologies and interpretations; HIV pathogenesis, fitness, and resistance; and mechanisms of resistance. The last of these topics provided a forum for new work on resistance of hepatitis B and C viruses, which were also explored in two poster sessions. Much work focused on resistance to the two most recent antiretroviral classes (integrase inhibitors and CCR5 antagonists), a new set of entry inhibitor candidates and one new class represented by the maturation inhibitor bevirimat. Other research explored two novel non-nucleoside reverse transcriptase inhibitors, etravirine and IDX899. Epidemiological work analysed rates of transmitted resistant virus, multiclass resistance in antiretroviral-experienced patients and a heightened resistance risk in injecting drug users regardless of adherence. New research on resistance technologies involved an enhanced assay for HIV-1 coreceptor determination and improved gene-based tools for predicting coreceptor use. In the pathogenesis arena, a small study of intensification shed light on the likely source of residual viraemia in patients on successful antiretroviral therapy. A large study in Mozambique correlated the timing of infant infection with selection, transmission and persistence of nevirapine resistance mutations. Mechanistic research explored resistance to the integrase inhibitor raltegravir, K65R-mediated resistance to tenofovir and the role of connection domain mutations in resistance to zidovudine.

Past, Current, and Future Developments of Therapeutic Agents for Treatment of Chronic Hepatitis B Virus Infection.

PubMed

Pei, Yameng; Wang, Chunting; Yan, S Frank; Liu, Gang

2017-08-10

For decades, treatment of hepatitis B virus (HBV) infection has been relying on interferon (IFN)-based therapies and nucleoside/nucleotide analogues (NAs) that selectively target the viral polymerase reverse transcriptase (RT) domain and thereby disrupt HBV viral DNA synthesis. We have summarized here the key steps in the HBV viral life cycle, which could potentially be targeted by novel anti-HBV therapeutics. A wide range of next-generation direct antiviral agents (DAAs) with distinct mechanisms of actions are discussed, including entry inhibitors, transcription inhibitors, nucleoside/nucleotide analogues, inhibitors of viral ribonuclease H (RNase H), modulators of viral capsid assembly, inhibitors of HBV surface antigen (HBsAg) secretion, RNA interference (RNAi) gene silencers, antisense oligonucleotides (ASOs), and natural products. Compounds that exert their antiviral activities mainly through host factors and immunomodulation, such as host targeting agents (HTAs), programmed cell death protein 1 (PD-1)/programmed death ligand 1 (PD-L1) inhibitors, and Toll-like receptor (TLR) agonists, are also discussed. In this Perspective, we hope to provide an overview, albeit by no means being comprehensive, for the recent development of novel therapeutic agents for the treatment of chronic HBV infection, which not only are able to sustainably suppress viral DNA but also aim to achieve functional cure warranted by HBsAg loss and ultimately lead to virus eradication and cure of hepatitis B.

Use of propidium monoazide in reverse transcriptase PCR to distinguish between infectious and noninfectious enteric viruses in water samples

EPA Science Inventory

Human enteric viruses can be present in untreated and inadequately treated drinking water. Molecular methods, such as the reverse transcriptase PCR (RT-PCR), can detect viral genomes in a few hours, but they cannot distinguish between infectious and noninfectious viruses. Since o...

Interaction of aurintricarboxylic acid (ATA) with four nucleic acid binding proteins DNase I, RNase A, reverse transcriptase and Taq polymerase

NASA Astrophysics Data System (ADS)

Ghosh, Utpal; Giri, Kalyan; Bhattacharyya, Nitai P.

2009-12-01

In the investigation of interaction of aurintricarboxylic acid (ATA) with four biologically important proteins we observed inhibition of enzymatic activity of DNase I, RNase A, M-MLV reverse transcriptase and Taq polymerase by ATA in vitro assay. As the telomerase reverse transcriptase (TERT) is the main catalytic subunit of telomerase holoenzyme, we also monitored effect of ATA on telomerase activity in vivo and observed dose-dependent inhibition of telomerase activity in Chinese hamster V79 cells treated with ATA. Direct association of ATA with DNase I ( Kd = 9.019 μM)), RNase A ( Kd = 2.33 μM) reverse transcriptase ( Kd = 0.255 μM) and Taq polymerase ( Kd = 81.97 μM) was further shown by tryptophan fluorescence quenching studies. Such association altered the three-dimensional conformation of DNase I, RNase A and Taq polymerase as detected by circular dichroism. We propose ATA inhibits enzymatic activity of the four proteins through interfering with DNA or RNA binding to the respective proteins either competitively or allosterically, i.e. by perturbing three-dimensional structure of enzymes.

The successes and failures of HIV drug discovery.

PubMed

Hashimoto, Chie; Tanaka, Tomohiro; Narumi, Tetsuo; Nomura, Wataru; Tamamura, Hirokazu

2011-10-01

To date, several anti-human immunodeficiency virus (HIV) drugs, including reverse transcriptase inhibitors and protease inhibitors, have been developed and used clinically for the treatment of patients infected with HIV. Recently, novel drugs have been discovered which have different mechanisms of action from those of the above inhibitors, including entry inhibitors and integrase (IN) inhibitors; the clinical use of three of these inhibitors has been approved. Other inhibitors are still in development. This review article summarizes the history of the development of anti-HIV drugs and also focuses on successes in the development of these entry and IN inhibitors, along with looking at exploratory approaches for the development of other inhibitors. Currently used highly active antiretroviral therapy can be subject to a loss of efficacy, due to the emergence of multi-drug resistant (MDR) strains; a change of regimens of the drug combination is required to combat this, along with careful monitoring of the virus and CD4 in the blood, by methods such as cellular tropism testing. In such a situation, entry inhibitors such as CCR5/CXCR4 antagonists, CD4 mimics, fusion inhibitors and IN inhibitors might be optional agents for an expansion of the drug repertoire available to patients at all stages of HIV infection.

Drug-induced gynecomastia.

PubMed

Bowman, John D; Kim, Hyunah; Bustamante, Juan J

2012-12-01

Drugs account for about 20% of gynecomastia cases in men. As a number of factors can alter the estrogen:androgen ratio, several pathophysiologic mechanisms are associated with drugs causing this disorder. Antiandrogens, protease inhibitors, and nucleoside reverse transcriptase inhibitors are the most common drug causes of gynecomastia, whereas first-generation antipsychotics, spironolactone, verapamil, and cimetidine are less common causes. Other drugs have been reported rarely as causes. Treatment may involve switching to an alternative agent or may require surgery or irradiation if the causative agent cannot be discontinued. We reviewed the literature on drug-induced gynecomastia and provided another perspective by reviewing data from the United States Food and Drug Administration's Adverse Event Reporting System. Epidemiologic studies are needed to provide a more accurate description of the frequency of drug-induced gynecomastia. © 2012 Pharmacotherapy Publications, Inc.

Tenofovir-induced kidney injury.

PubMed

Gitman, Michael D; Hirschwerk, David; Baskin, Cindy H; Singhal, Pravin C

2007-03-01

Tenofovir disoproxil fumarate is a nucleotide reverse transcriptase inhibitor with activity against both HIV and the hepatitis B virus. It has had minimal nephrotoxic effects in early clinical trials, but as clinical use has widened, case reports describing tenofovir-induced renal tubular damage, Fanconi's syndrome and diabetes insipidus have been described. The authors review the pharmacokinetics, mechanism of action and clinical uses of tenofovir disoproxil fumarate. The large clinical trials, as well as the case reports of tenofovir-induced kidney injury, are also reviewed. The potential mechanism of renal damage is discussed and recommendations for evaluation and treatment of tenofovir-induced kidney injury are given.

Time trends in drug resistant HIV-1 infections in the United Kingdom up to 2009: multicentre observational study.

PubMed

Dolling, David; Sabin, Caroline; Delpech, Valerie; Smit, Erasmus; Pozniak, Anton; Asboe, David; Brown, Andrew Leigh; Churchill, Duncan; Williams, Ian; Geretti, Anna Maria; Phillips, Andrew; Mackie, Nicola; Murphy, Gary; Castro, Hannah; Pillay, Deenan; Cane, Patricia; Dunn, David; Dolling, David

2012-08-21

To evaluate whether the prevalence of HIV-1 transmitted drug resistance has continued to decline in infections probably acquired within the United Kingdom. Multicentre observational study. All UK public laboratories conducting tests for genotypic HIV resistance as a part of routine care. 14,584 patients infected with HIV-1 subtype B virus, who were first tested for resistance before receiving antiretroviral therapy between January 2002 and December 2009. Prevalence of transmitted drug resistance, defined as one or more resistance mutations from the surveillance list recommended by the World Health Organization. 1654 (11.3%, 95% confidence interval 10.8% to 11.9%) patients had one or more mutations associated with transmitted HIV-1 drug resistance; prevalence was found to decline from 15.5% in 2002 to 9.6% in 2007, followed by a slight increase to 10.9% in 2009 (P=0.21). This later rise was mainly a result of increases in resistance to nucleos(t)ide reverse transcriptase inhibitors (from 5.4% in 2007 to 6.6% in 2009, P=0.24) and protease inhibitors (1.5% to 2.1%, P=0.12). Thymidine analogue mutations, including T215 revertants, remained the most frequent mutations associated with nucleos(t)ide reverse transcriptase inhibitors, despite a considerable fall in stavudine and zidovudine use between 2002 and 2009 (from 29.4% of drug regimens in 2002 to 0.8% in 2009, from 47.9% to 8.8%, respectively). The previously observed decline in the prevalence of transmitted drug resistance in HIV-1 infections probably acquired in the UK seems to have stabilised. The continued high prevalence of thymidine analogue mutations suggests that the source of this resistance may be increasingly from patients who have not undergone antiretroviral therapy and who harbour resistant viruses. Testing of all newly diagnosed HIV-1 positive people should be continued.

2004: which HIV-1 drug resistance mutations are common in clinical practice?

PubMed

Cheung, Peter K; Wynhoven, Brian; Harrigan, P Richard

2004-01-01

The emergence of drug resistance remains a major problem for the treatment of HIV-infected patients. However, the variety of mutational patterns that evolve in clinical practice have made the application of resistance data to clinical decision-making challenging. Despite (or because of) an abundance of drug-resistance data from disparate sources, there is only limited information available describing the patterns of drug resistance which usually appear in the clinic. Here we attempt to address this issue by reviewing HIV drug resistance in the population of patients failing antiretroviral therapy in British Columbia, Canada from June 1996 to December 2003 as an example. Our findings suggest that, although hundreds of mutations have been associated with resistance, relatively few key mutations occur at a high frequency. For example, only the nucleoside reverse transcriptase inhibitor (NRTI) mutations M184V, M41L T215Y, D67N, K70R and L210W, non-nucleoside reverse transcriptase inhibitor (NNRTI) mutations K103N and Y181C, and protease inhibitor (PI) mutation L90M, occur in more than 10% of samples tested for resistance in this population. The introduction of new drugs allows for the selection of new mutations. Trends in the prevalence of resistance-associated mutations have generally followed trends in drug usage, but have not always mirrored them. The phenomenon of cross-resistance can play an important role in the efficacy of new antiretroviral agents, even before they become available. The extent of this cross-resistance depends in part on the prevalence of specific mutations in the population of individuals who have previously received antiretroviral therapy. Hence there is a need to determine which mutations are prevalent in the treated population. The tremendous capacity of HIV to adapt means that common resistance pathways are likely to change over time, and new pathways to resistance are likely to continue to be discovered in the future.

Analysis of drug resistance during HIV RNA viraemia in the MONET trial of darunavir/ritonavir monotherapy.

PubMed

Pulido, Federico; Arribas, José R; Hill, Andrew; Van Delft, Yvon; Moecklinghoff, Christiane

2011-01-01

When patients have HIV RNA suppressed to <50 copies/ml on current treatment, switching to darunavir (DRV)/ritonavir (DRV/r) monotherapy could prevent the development of resistance to other drug classes. In the MONET trial, 256 patients with HIV RNA<50 copies/ml on current highly active antiretroviral therapy (57% with protease inhibitors [PIs] and 43% with non-nucleoside reverse transcriptase inhibitors) and no history of virological failure were randomized to DRV/r 800/100 mg once daily, either as monotherapy (monotherapy arm) or with two nucleoside reverse transcriptase inhibitors (NRTIs; triple therapy arm). All samples with HIV RNA ≥ 50 copies/ml were genotyped, and a virtual phenotype was calculated (VircoType HIV-1 assays; Virco BVBA, Mechelen, Belgium). A total of 63 patients had ≥ 1 HIV RNA result ≥ 50 copies/ml, of whom 38 were successfully genotyped. Most HIV RNA increases were transient and in the range of 50-200 copies/ml. Overall, 36 of the 38 (95%) successfully genotyped patients showed no International AIDS Society-USA major PI mutations, DRV mutations or NRTI mutations. Two patients showed some evidence of PI resistance during transient HIV RNA elevations: one patient in the monotherapy arm had a single DRV mutation (L33F) when HIV RNA was 63 copies/ml (the virus was phenotypically sensitive to DRV [fold change 0.8]) and one PI pretreated patient taking tenofovir disoproxil fumarate/emtricitabine/DRV/r had re-emergence of pre-existing NRTI (M184V) and PI (V82I and L90M) mutations after a short treatment interruption (this virus remained phenotypically sensitive to DRV/r). Both patients showed sustained HIV RNA suppression to week 48 remaining with the same treatment. Emergence of drug resistance after changing a suppressive triple antiretroviral therapy to DRV/r with or without nucleoside analogues is uncommon.

Increase of Transmitted Drug Resistance among HIV-Infected Sub-Saharan Africans Residing in Spain in Contrast to the Native Population

PubMed Central

Yebra, Gonzalo; de Mulder, Miguel; Pérez-Elías, María Jesús; Pérez-Molina, José Antonio; Galán, Juan Carlos; Llenas-García, Jara; Moreno, Santiago; Holguín, África

2011-01-01

Background The prevalence of transmitted HIV drug resistance (TDR) is stabilizing or decreasing in developed countries. However, this trend is not specifically evaluated among immigrants from regions without well-implemented antiretroviral strategies. Methods TDR trends during 1996–2010 were analyzed among naïve HIV-infected patients in Spain, considering their origin and other factors. TDR mutations were defined according to the World Health Organization list. Results Pol sequence was available for 732 HIV-infected patients: 292 native Spanish, 226 sub-Saharan Africans (SSA), 114 Central-South Americans (CSA) and 100 from other regions. Global TDR prevalence was 9.7% (10.6% for Spanish, 8.4% for SSA and 7.9% for CSA). The highest prevalences were found for protease inhibitors (PI) in Spanish (3.1%), for non-nucleoside reverse transcriptase inhibitors (NNRTI) in SSA (6.5%) and for nucleoside reverse transcriptase inhibitors (NRTI) in both Spanish and SSA (6.5%). The global TDR rate decreased from 11.3% in 2004–2006 to 8.4% in 2007–2010. Characteristics related to a decreasing TDR trend in 2007-10 were Spanish and CSA origin, NRTI- and NNRTI-resistance, HIV-1 subtype B, male sex and infection through injection drug use. TDR remained stable for PI-resistance, in patients infected through sexual intercourse and in those carrying non-B variants. However, TDR increased among SSA and females. K103N was the predominant mutation in all groups and periods. Conclusion TDR prevalence tended to decrease among HIV-infected native Spanish and Central-South Americans, but it increased up to 13% in sub-Saharan immigrants in 2007–2010. These results highlight the importance of a specific TDR surveillance among immigrants to prevent future therapeutic failures, especially when administering NNRTIs. PMID:22046345

Transmitted HIV Drug Resistance Is High and Longstanding in Metropolitan Washington, DC

PubMed Central

Kassaye, Seble G.; Grossman, Zehava; Balamane, Maya; Johnston-White, Betsy; Liu, Chenglong; Kumar, Princy; Young, Mary; Sneller, Michael C.; Sereti, Irini; Dewar, Robin; Rehm, Catherine; Meyer, William; Shafer, Robert; Katzenstein, David; Maldarelli, Frank

2016-01-01

Background. Washington, DC, has 2.5% human immunodeficiency virus (HIV) prevalence, 3.9% among African Americans. Antiretrovirals (ARTs) are the cornerstone for treatment and prevention. Monitoring changes in transmitted drug resistance (TDR) is critical for effective HIV care. Methods. HIV genotype data for individuals enrolled in research studies in metropolitan Washington, D.C., were used to identify TDR using the World Health Organization mutation list [Bennett DE, Camacho RJ, Otelea D, et al. Drug resistance mutations for surveillance of transmitted HIV-1 drug-resistance: 2009 update. PloS One 2009; 4:e4724]. HIV phylogenies were reconstructed using maximum likelihood and Bayesian methods. HIV transmission clusters were supported by 1000 bootstrap values >0.70 and posterior probability >0.95 of having a common ancestor. Results. Among 710 individuals enrolled in 1994–2013, the median age was 38.6 years, 46.2% were female, and 53.3% were African-American. TDR was 22.5% among 566 treatment-naive individuals; 15.8% had nucleoside/nucleotide reverse transcriptase inhibitor (NRTI) resistance, 9.8% had nonnucleoside reverse-transcriptase inhibitor (NNRTI) resistance, and 4.2% had protease inhibitor (PI) resistance. Single class TDR was 10.0%, 5.1%, and 1.6% to NRTIs, NNRTIs, and PIs. Dual TDR to PI and NRTI was seen in 1.6%, NRTI and NNRTI in 3.4%, and triple class TDR in 0.9%. TDR frequency decreased from 1994–2006 (27.1%) to 2007–2013 (19.4%; P = .02). Only 6/79 (7.6%) individuals within transmission clusters had evidence of TDR. Discussions. We identified high prevalence of TDR among HIV-infected individuals in metropolitan Washington, DC, regardless of gender. Active surveillance for TDR is needed to guide ART usage and analyses of risk group contributions to HIV transmission and resistance. PMID:27307507

An assessment of the relationship between the World Health Organization HIV drug resistance early warning indicators and HIV drug resistance acquisition.

PubMed

St-Jean, M; Harrigan, P R; Sereda, P; Montaner, Jsg; Lima, V D

2017-05-01

The World Health Organization (WHO)'s HIV drug resistance (HIVDR) early warning indicators (EWIs) measure antiretroviral therapy (ART)-site factors associated with HIVDR prevention, without HIVDR laboratory testing. We assessed the relationship between EWIs and HIVDR acquisition using data from British Columbia, Canada. Eligible patients were ART-naïve, were ≥ 19 years old, had initiated ART between 1 January 2000 and 31 December 2012, had ≥ 15 months of follow-up, and were without transmitted HIVDR. Patients were followed for acquired HIVDR until 31 March 2014, the last contact date, or death. We built logistic regression models to assess the associations and predictive ability of individual indicators and of the EWI Score (the number of indicators for which a patient did not meet the criteria) on HIVDR acquisition (to any class of HIVDR, lamivudine (3TC)/emtricitabine (FTC), nonnucleoside reverse transcriptase inhibitors (NNRTIs), nucleoside reverse transcriptase inhibitors (NRTIs) or protease inhibitors (PIs)]). All explored EWIs were associated with at least one class of HIVDR, with the exception of 'ART prescribing practices'. We observed a dose-response relationship between acquiring HIVDR to any antiretroviral class and an increasing EWI score in our predictive logistic regression model. The area under the curve was 0.848 (excellent discrimination). The adjusted odds ratios for acquiring any class of HIVDR for an EWI score of 1, 2 and ≥ 3 versus 0 were 2.30 [95% confidence Interval (CI) 1.21-4.38], 3.35 (95% CI: 1.86-6.03) and 7.26 (95% CI: 4.18-12.61), respectively. Several EWIs were associated with and predictive of HIVDR, supporting the WHO EWIs as a component of the HIVDR prevention method in settings where HIVDR testing is not routinely or widely available. © 2016 British HIV Association.

Structural and Preclinical Studies of Computationally Designed Non-Nucleoside Reverse Transcriptase Inhibitors for Treating HIV infection

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kudalkar, Shalley N.; Beloor, Jagadish; Chan, Albert H.

The clinical benefits of HIV-1 non-nucleoside reverse transcriptase (RT) inhibitors (NNRTIs) are hindered by their unsatisfactory pharmacokinetic (PK) properties along with the rapid development of drug-resistant variants. However, the clinical efficacy of these inhibitors can be improved by developing compounds with enhanced pharmacological profiles and heightened antiviral activity. We used computational and structure-guided design to develop two next-generation NNRTI drug candidates, compounds I and II, which are members of a class of catechol diethers. We evaluated the preclinical potential of these compounds in BALB/c mice because of their high solubility (510 µg/ml for compound I and 82.9 µg/ml for compoundmore » II), low cytotoxicity, and enhanced antiviral activity against wild-type (WT) HIV-1 RT and resistant variants. Additionally, crystal structures of compounds I and II with WT RT suggested an optimal binding to the NNRTI binding pocket favoring the high anti-viral potency. A single intraperitoneal dose of compounds I and II exhibited a prolonged serum residence time of 48 hours and concentration maximum (Cmax) of 4000- to 15,000-fold higher than their therapeutic/effective concentrations. These Cmax values were 4- to 15-fold lower than their cytotoxic concentrations observed in MT-2 cells. Compound II showed an enhanced area under the curve (0–last) and decreased plasma clearance over compound I and efavirenz, the standard of care NNRTI. Hence, the overall (PK) profile of compound II was excellent compared with that of compound I and efavirenz. Furthermore, both compounds were very well tolerated in BALB/c mice without any detectable acute toxicity. Taken together, these data suggest that compounds I and II possess improved anti-HIV-1 potency, remarkable in vivo safety, and prolonged in vivo circulation time, suggesting strong potential for further development as new NNRTIs for the potential treatment of HIV infection.« less

Human Immunodeficiency Virus Type 1 (HIV-1) Integration: a Potential Target for Microbicides To Prevent Cell-Free or Cell-Associated HIV-1 Infection ▿

PubMed Central

Terrazas-Aranda, Katty; Van Herrewege, Yven; Hazuda, Daria; Lewi, Paul; Costi, Roberta; Di Santo, Roberto; Cara, Andrea; Vanham, Guido

2008-01-01

Conceptually, blocking human immunodeficiency virus type 1 (HIV-1) integration is the last possibility for preventing irreversible cellular infection. Using cocultures of monocyte-derived dendritic cells and CD4+ T cells, which represent primary targets in sexual transmission, we demonstrated that blocking integration with integrase strand transfer inhibitors (InSTIs), particularly L-870812, could consistently block cell-free and cell-associated HIV-1 infection. In a pretreatment setting in which the compound was present before and during infection and was afterwards gradually diluted during the culture period, the naphthyridine carboxamide L-870812 blocked infection with the cell-free and cell-associated HIV-1 Ba-L strain at concentrations of, respectively, 1,000 and 10,000 nM. The potency of L-870812 was similar to that of the nucleotide reverse transcriptase inhibitor R-9-(2-phosphonylmethoxypropyl) adenine (PMPA) but one or two orders of magnitude lower than those of the nonnucleoside reverse transcriptase inhibitors UC781 and TMC120. In contrast, the diketo acid RDS derivative InSTIs showed clear-cut but weaker antiviral activity than L-870812. Moreover, L-870812 completely blocked subtype C and CRFO2_AG primary isolates, which are prevalent in the African heterosexual epidemic. Furthermore, the addition of micromolar concentrations of L-870812 even 24 h after infection could still block both cell-free and cell-associated Ba-L, opening the prospect of postexposure prophylaxis. Finally, an evaluation of the combined activity of L-870812 with either T20, zidovudine, PMPA, UC781, or TMC120 against replication-deficient HIV-1 Ba-L (env) pseudovirus suggested synergistic activity for all combinations. Importantly, compounds selected for the study by using the coculture model were devoid of acute or delayed cytotoxic effects at HIV-blocking concentrations. Therefore, these findings provide evidence supporting consideration of HIV-1 integration as a target for microbicide development. PMID:18474579

Novel Bifunctional Quinolonyl Diketo Acid Derivatives as HIV-1 Integrase Inhibitors: Design, Synthesis, Biological Activities and Mechanism of Action

PubMed Central

Di Santo, Roberto; Costi, Roberta; Roux, Alessandra; Artico, Marino; Lavecchia, Antonio; Marinelli, Luciana; Novellino, Ettore; Palmisano, Lucia; Andreotti, Mauro; Amici, Roberta; Galluzzo, Clementina Maria; Nencioni, Lucia; Palamara, Anna Teresa; Pommier, Yves; Marchand, Christophe

2008-01-01

The virally encoded integrase protein is an essential enzyme in the life cycle of the HIV-1 virus and represents an attractive and validated target in the development of therapeutics against HIV infection. Drugs that selectively inhibit this enzyme, when used in combination with inhibitors of reverse transcriptase and protease, are believed to be highly effective in suppressing the viral replication. Among the HIV-1 integrase inhibitors, the β-diketo acids (DKAs) represent a major lead for anti-HIV-1drug development. In this study, novel bifunctional quinolonyl diketo acid derivatives were designed, synthesized and tested for their inhibitory ability against HIV-1 integrase. The compounds are potent inhibitors of integrase activity. Particularly, derivative 8 is a potent IN inhibitor for both steps of the reaction (3′-processing and strand transfer) and exhibits both high antiviral activity against HIV-1 infected cells and low cytotoxicity. Molecular modeling studies provide a plausible mechanism of action, which is consistent with ligand SARs and enzyme photo-crosslinking experiments. PMID:16539381

Development and evaluation of a culture-independent method for source determination of fecal wastes in surface and storm waters using reverse transcriptase-PCR detection of FRNA coliphage genogroup gene sequences.

EPA Science Inventory

A complete method, incorporating recently improved reverse transcriptase-PCR primer/probe assays and including controls for determining interferences to phage recoveries from water sample concentrates and for detecting interferences to their analysis, was developed for the direct...

Novel Structure of Ty3 Reverse Transcriptase | Center for Cancer Research

Cancer.gov

Retrotransposons are mobile genetic elements that self amplify via a single-stranded RNA intermediate, which is converted to double-stranded DNA by an encoded reverse transcriptase (RT) with both DNA polymerase (pol) and ribonuclease H (RNase) activities. Categorized by whether they contain flanking long terminal repeat (LTR) sequences, retrotransposons play a critical role in

Development and evaluation of a culture-independent method for source determination of fecal wastes in surface and storm waters using reverse transcriptase-PCR detection of FRNA coliphage genogroup gene sequences

EPA Science Inventory

A complete method, incorporating recently improved reverse transcriptase-PCR primer/probe assays and including controls for determining interferences to phage recoveries from water sample concentrates and for detecting interferences to their analysis, was developed for the direct...

Executive summary of the GESIDA/National AIDS Plan Consensus Document on Antiretroviral Therapy in Adults Infected by the Human Immunodeficiency Virus (Updated January 2017).

PubMed

2017-05-31

Antiretroviral therapy (ART) is recommended for all patients infected by HIV-1. The objective of ART is to achieve an undetectable plasma viral load (PVL). Initial ART should be based on a combination of 3 drugs, including 2 nucleoside reverse transcriptase inhibitors (tenofovir in either of its two formulations plus emtricitabine or abacavir plus lamivudine) and another drug from a different family. Four of the recommended regimens, all of which have an integrase inhibitor as the third drug (dolutegravir, elvitegravir boosted with cobicistat or raltegravir), are considered preferential, whereas a further 3 regimens (based on elvitegravir/cobicistat, rilpivirine, or darunavir boosted with cobicistat or ritonavir) are considered alternatives. We present the reasons and criteria for switching ART in patients with an undetectable PVL and in those who present virological failure, in which case salvage ART should include 3 (or at least 2) drugs that are fully active against HIV. We also update the criteria for ART in specific situations (acute infection, HIV-2 infection, pregnancy) and comorbidities (tuberculosis or other opportunistic infections, kidney disease, liver disease and cancer). Copyright © 2017. Publicado por Elsevier España, S.L.U.

Substrate-induced stable enzyme-inhibitor complex formation allows tight binding of novel 2-aminopyrimidin-4(3H)-ones to drug-resistant HIV-1 reverse transcriptase mutants.

PubMed

Samuele, Alberta; Facchini, Marcella; Rotili, Dante; Mai, Antonello; Artico, Marino; Armand-Ugón, Mercedes; Esté, José A; Maga, Giovanni

2008-09-01

We recently reported the synthesis and biological evaluation of a novel series of 5-alkyl-2-(N,N-disubstituted)amino-6-(2,6-difluorophenylalkyl)-3,4-dihydropyrimidin-4(3H)-ones (F(2)-N,N-DABOs). These compounds are highly active against both wild-type HIV-1 and the K103N, Y181C, and Y188L mutant strains. Herein we present novel 6-(2-chloro-6-fluorophenylalkyl)-N,N-DABO (2-Cl-6-F-N,N-DABO) derivatives and investigate the molecular basis for their high-affinity binding to HIV-1 reverse transcriptase (RT). Our results show that the new compounds display higher association rates than the difluoro derivatives toward wild-type HIV-1 RT or drug-resistant RT mutant forms. We also show that they preferentially associate to either the free enzyme or the enzyme-nucleic acid binary complex, and that this binding is stabilized upon formation of the ternary complex between HIV-1 RT and both the nucleic acid and nucleotide substrates. Interestingly, one compound showed dissociation rates from the ternary complex with RT mutants K103N and Y181I 10-20-fold slower than from the corresponding complex with wild-type RT.

Hypersusceptibility to substrate analogs conferred by mutations in human immunodeficiency virus type 1 reverse transcriptase.

PubMed

Smith, Robert A; Anderson, Donovan J; Preston, Bradley D

2006-07-01

Human immunodeficiency virus type 1 (HIV-1) reverse transcriptase (RT) contains four structural motifs (A, B, C, and D) that are conserved in polymerases from diverse organisms. Motif B interacts with the incoming nucleotide, the template strand, and key active-site residues from other motifs, suggesting that motif B is an important determinant of substrate specificity. To examine the functional role of this region, we performed "random scanning mutagenesis" of 11 motif B residues and screened replication-competent mutants for altered substrate analog sensitivity in culture. Single amino acid replacements throughout the targeted region conferred resistance to lamivudine and/or hypersusceptibility to zidovudine (AZT). Substitutions at residue Q151 increased the sensitivity of HIV-1 to multiple nucleoside analogs, and a subset of these Q151 variants was also hypersusceptible to the pyrophosphate analog phosphonoformic acid (PFA). Other AZT-hypersusceptible mutants were resistant to PFA and are therefore phenotypically similar to PFA-resistant variants selected in vitro and in infected patients. Collectively, these data show that specific amino acid replacements in motif B confer broad-spectrum hypersusceptibility to substrate analog inhibitors. Our results suggest that motif B influences RT-deoxynucleoside triphosphate interactions at multiple steps in the catalytic cycle of polymerization.

Inhibition of Human Immunodeficiency Virus Type 1 Infection by the Candidate Microbicide Dapivirine, a Nonnucleoside Reverse Transcriptase Inhibitor▿

PubMed Central

Fletcher, P.; Harman, S.; Azijn, H.; Armanasco, N.; Manlow, P.; Perumal, D.; de Bethune, M.-P.; Nuttall, J.; Romano, J.; Shattock, R.

2009-01-01

Heterosexual transmission of human immunodeficiency virus (HIV) remains the major route of infection worldwide; thus, there is an urgent need for additional prevention strategies, particularly strategies that could be controlled by women, such as topical microbicides. Potential microbicide candidates must be both safe and effective. Using cellular and tissue explant models, we have evaluated the activity of the nonnucleoside reverse transcriptase inhibitor (NNRTI) dapivirine as a vaginal microbicide. In tissue compatibility studies, dapivirine was well tolerated by epithelial cells, T cells, macrophages, and cervical tissue explants. Dapivirine demonstrated potent dose-dependent inhibitory effects against a broad panel of HIV type 1 isolates from different clades. Furthermore, dapivirine demonstrated potent activity against a wide range of NNRTI-resistant isolates. In human cervical explant cultures, dapivirine was able not only to inhibit direct infection of mucosal tissue but also to prevent the dissemination of the virus by migratory cells. Activity was retained in the presence of semen or a cervical mucus simulant. Furthermore, dapivirine demonstrated prolonged inhibitory effects: it was able to prevent both localized and disseminated infection for as long as 6 days posttreatment. The prolonged protection observed following pretreatment of genital tissue and the lack of observable toxicity suggest that dapivirine has considerable promise as a potential microbicide candidate. PMID:19029331

Impact of the Central Polypurine Tract on the Kinetics of Human Immunodeficiency Virus Type 1 Vector Transduction

PubMed Central

Van Maele, Bénédicte; De Rijck, Jan; De Clercq, Erik; Debyser, Zeger

2003-01-01

Lentiviral vectors derived from human immunodeficiency virus type 1 (HIV-1) show great promise as gene carriers for future gene therapy. Insertion of a fragment containing the central polypurine tract (cPPT) in HIV-1 vector constructs is known to enhance transduction efficiency drastically, reportedly by facilitating the nuclear import of HIV-1 cDNA through a central DNA flap. We have studied the impact of the cPPT on the kinetics of HIV-1 vector transduction by real-time PCR. The kinetics of total HIV-1 DNA, two-long-terminal-repeat (2-LTR) circles, and, by an Alu-PCR, integrated proviral DNA were monitored. About 6 to 12 h after transduction, the total HIV-1 DNA reached a maximum level, followed by a steep decrease. The 2-LTR circles peaked after 24 to 48 h and were diluted upon cell division. Integration of HIV-1 DNA was first detected at 12 h postinfection. When HIV-1 vectors that contained the cPPT were used, DNA synthesis was similar but a threefold higher amount of 2-LTR circles was detected, confirming the impact on nuclear import. Moreover, a 10-fold increase in the amount of integrated DNA was observed in the presence of the cPPT. Only in the absence of the cPPT was a saturation in 2-LTR circle formation seen at a high multiplicity of infection, suggesting a role for the cPPT in overcoming a barrier to the nuclear import of HIV-1 DNA. A major effect of the central DNA flap on the juxtaposition of both LTRs is unlikely, since transduction with HIV-1 vectors containing ectopic cPPT fragments resulted in increased amounts of 2-LTR circles as well as integrated DNA. Inhibitors of transduction by cPPT-containing HIV vectors were also studied by real-time PCR. The reverse transcriptase inhibitor azidothymidine (AZT) and the nonnucleoside reverse transcriptase inhibitor α-APA clearly inhibited viral DNA synthesis, whereas integrase inhibitors such as the diketo acid L-708,906 and the pyranodipyrimidine V-165 specifically inhibited integration. PMID:12663775

Emergence of uncommon HIV-1 non-B subtypes and circulating recombinant forms and trends in transmission of antiretroviral drug resistance in patients with primary infection during the 2013-2015 period in Marseille, Southeastern France.

PubMed

Tamalet, Catherine; Tissot-Dupont, Hervé; Motte, Anne; Tourrès, Christian; Dhiver, Catherine; Ravaux, Isabelle; Poizot-Martin, Isabelle; Dieng, Thérèse; Tomei, Christelle; Bregigeon, Sylvie; Zaegel-Faucher, Olivia; Laroche, Hélène; Aherfi, Sarah; Mokhtari, Saadia; Chaudet, Hervé; Ménard, Amelie; Brouqui, Philippe; Stein, Andreas; Colson, Philippe

2018-05-24

Primary HIV-1 infections (PHI) with non-B subtypes are increasing in developed countries while transmission of HIV-1 harboring antiretroviral resistance-associated mutations (RAMs) remains a concern. This study assessed non-B HIV-1 subtypes and RAMs prevalence among patients with PHI in university hospitals of Marseille, Southeastern France, in 2005-2015 (11 years). HIV-1 sequences were obtained by in-house protocols from 115 patients with PHI, including 38 for the 2013-2015 period. On the basis of the phylogenetic analysis of the reverse transcriptase region, non-B subtypes were identified in 31% of these patients. They included 3 different subtypes (3A, 1C, 4F), 23 circulating recombinant forms (CRFs) (CRF02_AG, best BLAST hits being CRF 36_cpx and CRF30 in 7 and 1 cases, respectively), and 5 unclassified sequences (U). Non-B subtypes proportion increased significantly, particularly in 2011-2013 vs in 2005-2010 (P = .03). CRF02_AG viruses largely predominated in 2005-2013 whereas atypical strains more difficult to classify and undetermined recombinants emerged recently (2014-2015). The prevalence of protease, nucleos(t)ide reverse transcriptase, and first-generation nonnucleoside reverse transcriptase inhibitors-associated RAMs were 1.7% (World Health Organization [WHO] list, 2009/2.6% International AIDS Society [IAS] list, 2017), 5.2%/4.3%, and 5.2%/5.2%, respectively. Etravirine/rilpivirine-associated RAM (IAS) prevalence was 4.3%. Men who have sex with men (MSM) were more frequently infected with drug-resistant viruses than other patients (26% vs 7%; P = .011). The recent increase of these rare HIV-1 strains and the spread of drug-resistant HIV-1 among MSM in Southeastern France might be considered when implementing prevention strategies and starting therapies. © 2018 Wiley Periodicals, Inc.

A Combination Microbicide Gel Protects Macaques Against Vaginal Simian Human Immunodeficiency Virus-Reverse Transcriptase Infection, But Only Partially Reduces Herpes Simplex Virus-2 Infection After a Single High-Dose Cochallenge

PubMed Central

Hsu, Mayla; Aravantinou, Meropi; Menon, Radhika; Seidor, Samantha; Goldman, Daniel; Kenney, Jessica; Derby, Nina; Gettie, Agegnehu; Blanchard, James; Piatak, Michael; Lifson, Jeffrey D.; Fernández-Romero, Jose A.; Zydowsky, Thomas M.

2014-01-01

Abstract Herpes simplex virus-2 (HSV-2) infection increases HIV susceptibility. We previously established a rhesus macaque model of vaginal HSV-2 preexposure followed by cochallenge with HSV-2 and simian/human immunodeficiency virus-reverse transcriptase (SHIV-RT). Using this model, we showed that a gel containing the nonnucleoside reverse transcriptase inhibitor (NNRTI) MIV-150 in carrageenan (CG) reduced SHIV-RT infection. To evaluate the efficacy of new generation microbicides against both viruses, we first established dual infection after single vaginal cochallenge with SHIV-RT and HSV-2 in HSV-2-naive macaques. All animals (6/6) became HSV-2 infected, with 4/6 coinfected with SHIV-RT. In a control group cochallenged with SHIV-RT and UV-inactivated HSV-2, 2/4 became SHIV-RT infected, and none had detectable HSV-2. Low-level HSV-2-specific antibody and T cell responses were detected in some HSV-2-infected animals. To test a CG gel containing MIV-150 and zinc acetate (MZC), which provided naive animals full protection from SHIV-RT for at least 8 h, MZC (vs. CG) was applied daily for 14 days followed by cochallenge 8 h later. MZC prevented SHIV-RT infection (0/9 infected, p=0.04 vs. 3/6 in CG controls), but only reduced HSV-2 infection by 20% (6/9 infected vs. 5/6 in CG, p=0.6). In HSV-2-infected animals, none of the gel-treated animals seroconverted, and only the CG controls had measurable HSV-2-specific T cell responses. This study shows the promise of MZC to prevent immunodeficiency virus infection (even in the presence of HSV-2) and reduce HSV-2 infection after exposure to a high-dose inoculum. Additionally, it demonstrates the potential of a macaque coinfection model to evaluate broad-spectrum microbicides. PMID:24117013

A combination microbicide gel protects macaques against vaginal simian human immunodeficiency virus-reverse transcriptase infection, but only partially reduces herpes simplex virus-2 infection after a single high-dose cochallenge.

PubMed

Hsu, Mayla; Aravantinou, Meropi; Menon, Radhika; Seidor, Samantha; Goldman, Daniel; Kenney, Jessica; Derby, Nina; Gettie, Agegnehu; Blanchard, James; Piatak, Michael; Lifson, Jeffrey D; Fernández-Romero, Jose A; Zydowsky, Thomas M; Robbiani, Melissa

2014-02-01

Herpes simplex virus-2 (HSV-2) infection increases HIV susceptibility. We previously established a rhesus macaque model of vaginal HSV-2 preexposure followed by cochallenge with HSV-2 and simian/human immunodeficiency virus-reverse transcriptase (SHIV-RT). Using this model, we showed that a gel containing the nonnucleoside reverse transcriptase inhibitor (NNRTI) MIV-150 in carrageenan (CG) reduced SHIV-RT infection. To evaluate the efficacy of new generation microbicides against both viruses, we first established dual infection after single vaginal cochallenge with SHIV-RT and HSV-2 in HSV-2-naive macaques. All animals (6/6) became HSV-2 infected, with 4/6 coinfected with SHIV-RT. In a control group cochallenged with SHIV-RT and UV-inactivated HSV-2, 2/4 became SHIV-RT infected, and none had detectable HSV-2. Low-level HSV-2-specific antibody and T cell responses were detected in some HSV-2-infected animals. To test a CG gel containing MIV-150 and zinc acetate (MZC), which provided naive animals full protection from SHIV-RT for at least 8 h, MZC (vs. CG) was applied daily for 14 days followed by cochallenge 8 h later. MZC prevented SHIV-RT infection (0/9 infected, p=0.04 vs. 3/6 in CG controls), but only reduced HSV-2 infection by 20% (6/9 infected vs. 5/6 in CG, p=0.6). In HSV-2-infected animals, none of the gel-treated animals seroconverted, and only the CG controls had measurable HSV-2-specific T cell responses. This study shows the promise of MZC to prevent immunodeficiency virus infection (even in the presence of HSV-2) and reduce HSV-2 infection after exposure to a high-dose inoculum. Additionally, it demonstrates the potential of a macaque coinfection model to evaluate broad-spectrum microbicides.

Mechanisms Used for Genomic Proliferation by Thermophilic Group II Introns

PubMed Central

Mohr, Georg; Ghanem, Eman; Lambowitz, Alan M.

2010-01-01

Mobile group II introns, which are found in bacterial and organellar genomes, are site-specific retroelments hypothesized to be evolutionary ancestors of spliceosomal introns and retrotransposons in higher organisms. Most bacteria, however, contain no more than one or a few group II introns, making it unclear how introns could have proliferated to higher copy numbers in eukaryotic genomes. An exception is the thermophilic cyanobacterium Thermosynechococcus elongatus, which contains 28 closely related copies of a group II intron, constituting ∼1.3% of the genome. Here, by using a combination of bioinformatics and mobility assays at different temperatures, we identified mechanisms that contribute to the proliferation of T. elongatus group II introns. These mechanisms include divergence of DNA target specificity to avoid target site saturation; adaptation of some intron-encoded reverse transcriptases to splice and mobilize multiple degenerate introns that do not encode reverse transcriptases, leading to a common splicing apparatus; and preferential insertion within other mobile introns or insertion elements, which provide new unoccupied sites in expanding non-essential DNA regions. Additionally, unlike mesophilic group II introns, the thermophilic T. elongatus introns rely on elevated temperatures to help promote DNA strand separation, enabling access to a larger number of DNA target sites by base pairing of the intron RNA, with minimal constraint from the reverse transcriptase. Our results provide insight into group II intron proliferation mechanisms and show that higher temperatures, which are thought to have prevailed on Earth during the emergence of eukaryotes, favor intron proliferation by increasing the accessibility of DNA target sites. We also identify actively mobile thermophilic introns, which may be useful for structural studies, gene targeting in thermophiles, and as a source of thermostable reverse transcriptases. PMID:20543989

The allosteric HIV-1 integrase inhibitor BI-D affects virion maturation but does not influence packaging of a functional RNA genome.

PubMed

van Bel, Nikki; van der Velden, Yme; Bonnard, Damien; Le Rouzic, Erwann; Das, Atze T; Benarous, Richard; Berkhout, Ben

2014-01-01

The viral integrase (IN) is an essential protein for HIV-1 replication. IN inserts the viral dsDNA into the host chromosome, thereby aided by the cellular co-factor LEDGF/p75. Recently a new class of integrase inhibitors was described: allosteric IN inhibitors (ALLINIs). Although designed to interfere with the IN-LEDGF/p75 interaction to block HIV DNA integration during the early phase of HIV-1 replication, the major impact was surprisingly found on the process of virus maturation during the late phase, causing a reverse transcription defect upon infection of target cells. Virus particles produced in the presence of an ALLINI are misformed with the ribonucleoprotein located outside the virus core. Virus assembly and maturation are highly orchestrated and regulated processes in which several viral proteins and RNA molecules closely interact. It is therefore of interest to study whether ALLINIs have unpredicted pleiotropic effects on these RNA-related processes. We confirm that the ALLINI BI-D inhibits virus replication and that the produced virus is non-infectious. Furthermore, we show that the wild-type level of HIV-1 genomic RNA is packaged in virions and these genomes are in a dimeric state. The tRNAlys3 primer for reverse transcription was properly placed on this genomic RNA and could be extended ex vivo. In addition, the packaged reverse transcriptase enzyme was fully active when extracted from virions. As the RNA and enzyme components for reverse transcription are properly present in virions produced in the presence of BI-D, the inhibition of reverse transcription is likely to reflect the mislocalization of the components in the aberrant virus particle.

The group II intron maturase: a reverse transcriptase and splicing factor go hand in hand.

PubMed

Zhao, Chen; Pyle, Anna Marie

2017-12-01

The splicing of group II introns in vivo requires the assistance of a multifunctional intron encoded protein (IEP, or maturase). Each IEP is also a reverse-transcriptase enzyme that enables group II introns to behave as mobile genetic elements. During splicing or retro-transposition, each group II intron forms a tight, specific complex with its own encoded IEP, resulting in a highly reactive holoenzyme. This review focuses on the structural basis for IEP function, as revealed by recent crystal structures of an IEP reverse transcriptase domain and cryo-EM structures of an IEP-intron complex. These structures explain how the same IEP scaffold is utilized for intron recognition, splicing and reverse transcription, while providing a physical basis for understanding the evolutionary transformation of the IEP into the eukaryotic splicing factor Prp8. Copyright © 2017 Elsevier Ltd. All rights reserved.

Telomerase reverse transcriptase (TERT) expression and role of vincristine sulfate in mouse model of malignancy related peritoneal ascites: an experimental metastatic condition.

PubMed

Chaklader, M; Das, P; Pereira, J A; Chatterjee, S; Basak, P; Law, A; Banerjee, T; Chauhan, S; Law, S

2011-06-01

To evaluate the efficacy of intraperitoneal vincristine administration into ascitic sarcoma-180 bearing mice as a model of human malignant ascites regarding various peritoneal/retroperitoneal sarcomatosis, and to evaluate the flowcytometric telomerase reverse transcriptase expression for the diagnostic and prognostic purposes. Present study included disease induction by intraperitoneal homologous ascitic sarcoma-180 transplantation followed by in vivo intraperitoneal drug administration to study mitotic index, flowcytometric cell cycle and telomerase reverse transcriptase expression pattern, erythrosin-B dye exclusion study for malignant cell viability assessment. Besides, in vitro malignant ascite culture in presence and absence of vincristine sulfate and survival study were also taken into consideration. Intraperitoneal vincristine administration (concentration 0.5 mg/kg body weight) significantly diminished the mitotic index in diseased subjects in comparison to untreated control subjects. Treated group of animals showed increased life span and median survival time. Cell viability assessment during the course of drug administration also revealed gradual depression on cell viability over time. Flowcytometric cell cycle analysis showed a good prognostic feature of chemotherapeutic administration schedule by representing high G2/M phase blocked cells along with reduced telomerase reverse transcriptase positive cells in treated animals. We conclude that long term administration of vincristine sulfate in small doses could be a good pharmacological intervention in case of malignant peritoneal ascites due to sarcomatosis as it indirectly reduced the level of telomerase reverse transcriptase expression in malignant cells by directly regulating cell cycle and simultaneously increased the life expectancy of the diseased subjects.

Adverse Drug Reaction Reporting in Africa and a Comparison of Individual Case Safety Report Characteristics Between Africa and the Rest of the World: Analyses of Spontaneous Reports in VigiBase®.

PubMed

Ampadu, Haggar H; Hoekman, Jarno; de Bruin, Marieke L; Pal, Shanthi N; Olsson, Sten; Sartori, Daniele; Leufkens, Hubert G M; Dodoo, Alexander N O

2016-04-01

Following the start of the World Health Organization (WHO) Programme for International Drug Monitoring (PIDM) by 10 member countries in 1968, it took another 24 years for the first two African countries to join in 1992, by which time the number of member countries in the PIDM had grown to 33. Whilst pharmacovigilance (PV), including the submission of individual case safety reports (ICSR) to VigiBase(®), the WHO global ICSR database, is growing in Africa, no data have been published on the growth of ICSR reporting from Africa and how the features of ICSRs from Africa compare with the rest of the world (RoW). The objective of this paper was to provide an overview of the growth of national PV centres in Africa, the reporting of ICSRs by African countries, and the features of ICSRs from Africa, and to compare ICSRs from Africa with the RoW. The search and analysis interface of VigiBase(®)--VigiLyze(®)--was used to characterise ICSRs submitted by African countries and the RoW. The distribution of ICSRs by African countries was listed and characterised by anatomic therapeutic chemical (ATC) code, Medical Dictionary for Regulatory Activities (MedDRA(®)) system organ class (SOC) classification, and patient age and sex. The case-defining features of ICSRs between Africa and the RoW were also compared. The number of African countries in the PIDM increased from 2 in 1992 to 35 at the end of September 2015, and African PIDM members have cumulatively submitted 103,499 ICSRs (0.88 % of global ICSRs) to VigiBase(®). The main class of products in African ICSRs are nucleoside and nucleotide reverse transcriptase inhibitors (14.04 %), non-nucleoside reverse transcriptase inhibitors (9.09 %), antivirals for the treatment of HIV infections (5.50 %), combinations of sulfonamides and trimethoprim (2.98 %) and angiotensin-converting enzyme (ACE) inhibitors (2.42 %). The main product classes implicated in ICSRs from the RoW are tumour necrosis factor-α (TNFα) inhibitors (5.29 %), topical nonsteroidal anti-inflammatory preparations (2.26 %), selective immunosuppressants (2.08 %), selective serotonin reuptake inhibitors (2.04 %) and HMG CoA reductase inhibitors (1.85 %). The main SOCs reported from Africa versus the RoW include skin and subcutaneous tissue disorders (31.14 % vs. 19.58 %), general disorders and administration site conditions (20.91 % vs. 30.49 %) and nervous system disorders (17.48 % vs. 19.13 %). The 18-44 years age group dominated ICSRs from Africa, while the 45-64 years age group dominated the RoW. Identical proportions of females (57 % Africa and the RoW) and males (37 % Africa and the RoW) were represented. As at the end of September 2015, 35 of 54 African countries were Full Member countries of the PIDM. Although the number of ICSRs from Africa has increased substantially, ICSRs from Africa still make up <1 % of the global total in VigiBase(®). The features of ICSRs from Africa differ to those from the RoW in relation to the classes of products as well as age group of patients affected. The gender of patients represented in these ICSRs are identical.

Applicability of integrated cell culture reverse transcriptase quantitative PCR (ICC-RTqPCR) for the simultaneous detection of the four human enteric enterovirus species in disinfection studies

EPA Science Inventory

A newly developed integrated cell culture reverse transcriptase quantitative PCR (ICC-RTqPCR) method and its applicability in UV disinfection studies is described. This method utilizes a singular cell culture system coupled with four RTqPCR assays to detect infectious serotypes t...

Discovery of genetic variants of the kinases that activate tenofovir among individuals in the United States, Thailand, and South Africa: HPTN067.

PubMed

Figueroa, Dominique B; Tillotson, Joseph; Li, Maoji; Piwowar-Manning, Estelle; Hendrix, Craig W; Holtz, Timothy H; Bokoch, Kevin; Bekker, Linda-Gail; van Griensven, Frits; Mannheimer, Sharon; Hughes, James P; Grant, Robert M; Bumpus, Namandjé N

2018-01-01

Tenofovir (TFV), a nucleotide reverse transcriptase inhibitor, requires two phosphorylation steps to form a competitive inhibitor of HIV reverse transcriptase. Adenylate kinase 2 (AK2) has been previously demonstrated to phosphorylate tenofovir to tenofovir-monophosphate, while creatine kinase, muscle (CKM), pyruvate kinase, muscle (PKM) and pyruvate kinase, liver and red blood cell (PKLR) each have been found to phosphorylate tenofovir-monophosphate to the pharmacologically active tenofovir-diphosphate. In the present study, genomic DNA isolated from dried blood spots collected from 505 participants from Bangkok, Thailand; Cape Town, South Africa; and New York City, USA were examined for variants in AK2, CKM, PKM, and PKLR using next-generation sequencing. The bioinformatics tools SIFT and PolyPhen predicted that 19 of the 505 individuals (3.7% frequency) carried variants in at least one kinase that would result in a decrease or loss of enzymatic activity. To functionally test these predictions, AK2 and AK2 variants were expressed in and purified from E. coli, followed by investigation of their activities towards tenofovir. Interestingly, we found that purified AK2 had the ability to phosphorylate tenofovir-monophosphate to tenofovir-diphosphate in addition to phosphorylating tenofovir to tenofovir-monophosphate. Further, four of the six AK2 variants predicted to result in a loss or decrease of enzyme function exhibited a ≥30% decrease in activity towards tenofovir in our in vitro assays. Of note, an AK2 K28R variant resulted in a 72% and 81% decrease in the formation of tenofovir-monophosphate and tenofovir-diphosphate, respectively. These data suggest that there are naturally occurring genetic variants that could potentially impact TFV activation.

Detection of EML4-ALK fusion gene in Chinese non-small cell lung cancer by using a sensitive quantitative real-time reverse transcriptase PCR technique.

PubMed

Fu, Sha; Wang, Fang; Shao, Qiong; Zhang, Xu; Duan, Li-Ping; Zhang, Xiao; Zhang, Li; Shao, Jian-Yong

2015-04-01

Anaplastic lymphoma kinase (ALK) rearrangement is present in approximately 5% of lung adenocarcinoma. Clinical trials on ALK inhibitor phase I to III have shown an interesting disease control rate and acceptable tolerability in ALK rearrangement patients. In clinical application, the precise diagnostic strategy for identifying ALK rearrangements remains to be determined. In this study, ALK rearrangement was screened by using quantitative real-time reverse transcriptase polymerase chain reaction (qRT-PCR), direct sequencing, 2 fluorescence in situ hybridization (FISH) assays, and immunohistochemistry in 173 lung adenocarcinomas. We identified 18 cases (10.4%) with EML4-ALK fusion-positive by qRT-PCR, and all were positive for EML4-ALK fusion gene validated by direct sequencing. The result was consistent with that of other methods. Furthermore, of the 18 EML4-ALK fusion-positive cases, 16 (9.2%) were positive by using EML4-ALK fusion probe FISH, and 15 (8.7%) were positive by using ALK break-apart probe FISH and immunohistochemistry staining. Of the 18 ALK fusion-positive lung adenocarcinomas, 8 cases (44.4%) were histologically diagnosed as subtypes of cribriform adenocarcinoma, 7 cases (38.9%) as cribriform adenocarcinoma mixed with papillary and/or mucinous pattern, 2 cases (11.1%) as papillary adenocarcinoma, and 1 case (5.6%) as mucinous adenocarcinoma. In the present study, the ALK rearrangement frequency detected by qRT-PCR in Chinese NSCLC patients was higher than that in the western populations. QRT-PCR is a rapid, sensitive technology that could be used as a screening tool for identifying EML4-ALK fusion-positive NSCLC patients who would be sensitive for receiving ALK inhibitor therapy.

Adenosine 3′,5′-cyclic monophosphate (cAMP)-dependent phosphoregulation of mitochondrial complex I is inhibited by nucleoside reverse transcriptase inhibitors

PubMed Central

Lund, Kaleb C.; Wallace, Kendall B.

2008-01-01

Nucleoside analog reverse transcriptase inhibitors (NRTI) are known to directly inhibit mitochondrial complex I activity as well as various mitochondrial kinases. Recent observations that complex I activity and superoxide production are modulated through cAMP-dependent phosphorylation suggests a mechanism through which NRTIs may affect mitochondrial respiration via kinase-dependent protein phosphorylation. In the current study we examine the potential for NRTIs to inhibit the cAMP-dependent phosphorylation of complex I and the associated NADH:CoQ oxidoreductase activities and rates of superoxide production using HepG2 cells. Phosphoprotein staining of immunocaptured complex I revealed that 3′-azido-3′-deoxythymidine (AZT; 10 and 50 μM), AZT monophosphate (150 μM), and 2′,3′-dideoxycytidine (ddC; 1μM) prevented the phosphorylation of the NDUFB11 subunit of complex I. This was associated with a decrease in complex I activity with AZT and AZT monophosphate only. In the presence of succinate, superoxide production was increased with 2′,3′-dideoxyinosine (ddI; 10 μM) and ddC (1 μM). In the presence of succinate + cAMP AZT showed an inverse dose-dependent effect on superoxide production. None of the NRTIs examined inhibit PKA activity suggesting that the observed effects are due to a direct interaction with complex I. These data demonstrate a direct effect of NRTIs on cAMP-dependent regulation of mitochondrial bioenergetics independent of DNA polymerase-γ activity; in the case of AZT these observations may provide a mechanism for the observed long-term toxicity with this drug. PMID:17904600

Transcription of G-protein coupled receptors in corporeal smooth muscle is regulated by the endogenous neutral endopeptidase inhibitor sialorphin.

PubMed

Tong, Yuehong; Tiplitsky, Scott I; Tar, Moses; Melman, Arnold; Davies, Kelvin P

2008-08-01

Several reports suggest that the rat Vcsa1 gene is down-regulated in models of erectile dysfunction. The Vcsa protein product sialorphin is an endogenous neutral endopeptidase inhibitor and its down-regulation could result in prolonged activation of G-protein activated signaling pathways by their peptide agonists. We investigated whether Vcsa1 down-regulation could result in an adaptive change in GPCR (G-protein coupled receptor) expression. Gene expression in cultured rat corporeal smooth muscle cells following treatment with siRNA directed against Vcsa1 or the neutral endopeptidase gene was analyzed using microarray and quantitative reverse transcriptase-polymerase chain reaction. In rats Vcsa1 is one of the most down-regulated genes following bilateral transection of the cavernous nerves. In that animal model we also investigated whether Vcsa1 down-regulation was accompanied by similar changes in gene expression in corporeal smooth muscle cells in which Vcsa1 was knocked down in vitro. Microarray analysis and quantitative reverse transcriptase-polymerase chain reaction demonstrated that corporeal smooth muscle cells treated in vitro with siRNA against Vcsa1 resulted in GPCR up-regulation as a functional group. In contrast, treatment of corporeal smooth muscle cells that lowered neutral endopeptidase activity resulted in decreased GPCR expression. These results suggest that the peptide product of Vcsa1, sialorphin, can effect GPCR expression by acting on neutral endopeptidase. In animals with bilaterally transected cavernous nerves the decreased Vcsa1 expression is accompanied by increased GPCR expression in cavernous tissue. These experiments suggest that the mechanism by which Vcsa1 modulates erectile function is partly mediated through changes in GPCR expression.

HIV-1 drug resistance in recently HIV-infected pregnant mother's naïve to antiretroviral therapy in Dodoma urban, Tanzania.

PubMed

Vairo, Francesco; Nicastri, Emanuele; Liuzzi, Giuseppina; Chaula, Zainab; Nguhuni, Boniface; Bevilacqua, Nazario; Forbici, Federica; Amendola, Alessandra; Fabeni, Lavinia; De Nardo, Pasquale; Perno, Carlo Federico; Cannas, Angela; Sakhoo, Calistus; Capobianchi, Maria Rosaria; Ippolito, Giuseppe

2013-09-21

HIV resistance affects virological response to therapy and efficacy of prophylaxis in mother-to-child-transmission. The study aims to assess the prevalence of HIV primary resistance in pregnant women naïve to antiretrovirals. Cross sectional baseline analysis of a cohort of HIV + pregnant women (HPW) enrolled in the study entitled Antiretroviral Management of Antenatal and Natal HIV Infection (AMANI, peace in Kiswahili language). The AMANI study began in May 2010 in Dodoma, Tanzania. In this observational cohort, antiretroviral treatment was provided to all women from the 28th week of gestation until the end of the breastfeeding period. Baseline CD4 cell count, viral load and HIV drug-resistance genotype were collected. Drug-resistance analysis was performed on 97 naïve infected-mothers. The prevalence of all primary drug resistance and primary non-nucleoside reverse-transcriptase inhibitors resistance was 11.9% and 7.5%, respectively. K103S was found in two women with no M184V detection. HIV-1 subtype A was the most commonly identified, with a high prevalence of subtype A1, followed by C, D, C/D recombinant, A/C recombinant and A/D recombinant. HIV drug- resistance mutations were detected in A1 and C subtypes. Our study reports an 11.9% prevalence rate of primary drug resistance in naïve HIV-infected pregnant women from a remote area of Tanzania. Considering that the non-nucleoside reverse-transcriptase inhibitors are part of the first-line antiretroviral regimen in Tanzania and all of Africa, resistance surveys should be prioritized in settings where antiretroviral therapy programs are scaled up.

HIV Drug Resistance Testing in a Resource Limited Setting with High Viral Diversity: The First Twenty Eight Months Experience.

PubMed

Ngo-Malabo, Elodie Teclaire; Ngoupo, Paul Alain; Sadeuh-Mba, Serge Alain; Akongnwi, Emmanuel; Banaï, Robert; Ngono, Laure; Bilong-Bilong, Charles Felix; Kfutwah, Anfumbom; Njouom, Richard

2017-01-01

First line antiretroviral therapy in a resource-limited setting consists of nucleotide and non-nucleotide reverse transcriptase inhibitors. Protease inhibitors are the hub of second line therapy. The decision to change antiretroviral therapy for a patient is frequently presumptive because of the lack of genotypic resistance tests in routine follow-up. We describe here the resistance profiles observed in patients with varying terms of antiretroviral therapy in Cameroon after implementation of HIV genotypic resistance testing in routine practice. HIV genotypic resistance testing was carried out on consecutive samples received between August 2013 and November 2015. Protease (Prot) and reverse transcriptase (Rt) genes of the HIV genome were amplified, sequenced and analyzed for drug resistance mutations following the algorithm set up by the French National Agency for research on HIV/AIDS and viral hepatitis. Specimens from a total of 167 patients infected with non-B HIV subtypes were received during the study period. Overall 61.7% patients had viral loads of more than 3log copies/ml, suggesting treatment failure. Among the 72 patients on first line, 56 (77.8%) were resistant to Lamivudine, 57 (79.1%) to Efavirenz and 58 (80.6%) to Nevirapine. Overall, more patients (75.0%) on first line antiretroviral therapy harbored multi-drug resistance compared to their counterparts on second line (25.8%). This study revealed that a group of patients with antiretroviral therapy failure harbored multi-drug resistance mutations related to the majority of drugs in the first line regimen. Therefore, HIV resistance testing could be a useful tool to improve HIV care in resource limited settings like Cameroon where treatment options are limited. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

HIV Salvage Therapy Does Not Require Nucleoside Reverse Transcriptase Inhibitors: A Randomized, Controlled Trial.

PubMed

Tashima, Karen T; Smeaton, Laura M; Fichtenbaum, Carl J; Andrade, Adriana; Eron, Joseph J; Gandhi, Rajesh T; Johnson, Victoria A; Klingman, Karin L; Ritz, Justin; Hodder, Sally; Santana, Jorge L; Wilkin, Timothy; Haubrich, Richard H

2015-12-15

Nucleoside reverse transcriptase inhibitors (NRTIs) are often included in antiretroviral regimens in treatment-experienced patients in the absence of data from randomized trials. To compare treatment success between participants who omit versus those who add NRTIs to an optimized antiretroviral regimen of 3 or more agents. Multicenter, randomized, controlled trial. (ClinicalTrials.gov: NCT00537394). Outpatient HIV clinics. Treatment-experienced patients with HIV infection and viral resistance. Open-label optimized regimens (not including NRTIs) were selected on the basis of treatment history and susceptibility testing. Participants were randomly assigned to omit or add NRTIs. The primary efficacy outcome was regimen failure through 48 weeks using a noninferiority margin of 15%. The primary safety outcome was time to initial episode of a severe sign, symptom, or laboratory abnormality before discontinuation of NRTI assignment. 360 participants were randomly assigned, and 93% completed a 48-week visit. The cumulative probability of regimen failure was 29.8% in the omit-NRTIs group versus 25.9% in the add-NRTIs group (difference, 3.2 percentage points [95% CI, -6.1 to 12.5 percentage points]). No significant between-group differences were found in the primary safety end points or the proportion of participants with HIV RNA level less than 50 copies/mL. No deaths occurred in the omit-NRTIs group compared with 7 deaths in the add-NRTIs group. Unblinded study design, and the study may not be applicable to resource-poor settings. Treatment-experienced patients with HIV infection starting a new optimized regimen can safely omit NRTIs without compromising virologic efficacy. Omitting NRTIs will reduce pill burden, cost, and toxicity in this patient population. National Institute of Allergy and Infectious Diseases, Boehringer Ingelheim, Janssen, Merck, ViiV Healthcare, Roche, and Monogram Biosciences (LabCorp).

Higher rates of triple-class virological failure in perinatally HIV-infected teenagers compared with heterosexually infected young adults in Europe.

PubMed

Judd, A; Lodwick, R; Noguera-Julian, A; Gibb, D M; Butler, K; Costagliola, D; Sabin, C; van Sighem, A; Ledergerber, B; Torti, C; Mocroft, A; Podzamczer, D; Dorrucci, M; De Wit, S; Obel, N; Dabis, F; Cozzi-Lepri, A; García, F; Brockmeyer, N H; Warszawski, J; Gonzalez-Tome, M I; Mussini, C; Touloumi, G; Zangerle, R; Ghosn, J; Castagna, A; Fätkenheuer, G; Stephan, C; Meyer, L; Campbell, M A; Chene, G; Phillips, A

2017-03-01

The aim of the study was to determine the time to, and risk factors for, triple-class virological failure (TCVF) across age groups for children and adolescents with perinatally acquired HIV infection and older adolescents and adults with heterosexually acquired HIV infection. We analysed individual patient data from cohorts in the Collaboration of Observational HIV Epidemiological Research Europe (COHERE). A total of 5972 participants starting antiretroviral therapy (ART) from 1998, aged < 20 years at the start of ART for those with perinatal infection and 15-29 years for those with heterosexual infection, with ART containing at least two nucleoside reverse transcriptase inhibitors (NRTIs) and a nonnucleoside reverse transcriptase inhibitor (NNRTI) or a boosted protease inhibitor (bPI), were followed from ART initiation until the most recent viral load (VL) measurement. Virological failure of a drug was defined as VL > 500 HIV-1 RNA copies/mL despite ≥ 4 months of use. TCVF was defined as cumulative failure of two NRTIs, an NNRTI and a bPI. The median number of weeks between diagnosis and the start of ART was higher in participants with perinatal HIV infection compared with participants with heterosexually acquired HIV infection overall [17 (interquartile range (IQR) 4-111) vs. 8 (IQR 2-38) weeks, respectively], and highest in perinatally infected participants aged 10-14 years [49 (IQR 9-267) weeks]. The cumulative proportion with TCVF 5 years after starting ART was 9.6% [95% confidence interval (CI) 7.0-12.3%] in participants with perinatally acquired infection and 4.7% (95% CI 3.9-5.5%) in participants with heterosexually acquired infection, and highest in perinatally infected participants aged 10-14 years when starting ART (27.7%; 95% CI 13.2-42.1%). Across all participants, significant predictors of TCVF were those with perinatal HIV aged 10-14 years, African origin, pre-ART AIDS, NNRTI-based initial regimens, higher pre-ART viral load and lower pre-ART CD4. The results suggest a beneficial effect of starting ART before adolescence, and starting young people on boosted PIs, to maximize treatment response during this transitional stage of development. © 2016 The Authors. HIV Medicine published by John Wiley & Sons Ltd on behalf of British HIV Association.

[GeSIDA/National AIDS Plan: Consensus document on antiretroviral therapy in adults infected by the human immunodeficiency virus (Updated January 2014)].

PubMed

2014-01-01

This consensus document is an update of combined antiretroviral therapy (cART) guidelines for HIV-1 infected adult patients. To formulate these recommendations a panel composed of members of the Grupo de Estudio de Sida and the Plan Nacional sobre el Sida reviewed the efficacy and safety advances in clinical trials, cohort and pharmacokinetic studies published in medical journals (PubMed and Embase) or presented in medical scientific meetings. Recommendations strength and the evidence in which they are supported are based on modified criteria of the Infectious Diseases Society of America. In this update, antiretroviral therapy (ART) is recommended for all patients infected by type 1 human immunodeficiency virus (HIV-1). The strength and grade of the recommendation varies with the clinical circumstances: CDC stage B or C disease (A-I), asymptomatic patients (depending on the CD4+ T-lymphocyte count: <350cells/μL, A-I; 350-500 cells/μL, A-II, and >500 cells/μL, B-III), comorbid conditions (HIV nephropathy, chronic hepatitis caused by HBV or HCV, age >55years, high cardiovascular risk, neurocognitive disorders, and cancer, A-II), and prevention of transmission of HIV (mother-to-child or heterosexual, A-I; men who have sex with men, A-III). The objective of ART is to achieve an undetectable plasma viral load. Initial ART should always comprise a combination of 3 drugs, including 2 nucleoside reverse transcriptase inhibitors and a third drug from a different family (non-nucleoside reverse transcriptase inhibitor, protease inhibitor, or integrase inhibitor). Some of the possible initial regimens have been considered alternatives. This update presents the causes and criteria for switching ART in patients with undetectable plasma viral load and in cases of virological failure where rescue ART should comprise 2 or 3 drugs that are fully active against the virus. An update is also provided for the specific criteria for ART in special situations (acute infection, HIV-2 infection, and pregnancy) and with comorbid conditions (tuberculosis or other opportunistic infections, kidney disease, liver disease, and cancer). These new guidelines updates previous recommendations related to cART (when to begin and what drugs should be used), how to monitor and what to do in case of viral failure or drug adverse reactions. cART specific criteria in comorbid patients and special situations are equally updated. Copyright © 2014 Elsevier España, S.L.U. y Sociedad Española de Enfermedades Infecciosas y Microbiología Clínica. All rights reserved.

Sensitive Deep-Sequencing-Based HIV-1 Genotyping Assay To Simultaneously Determine Susceptibility to Protease, Reverse Transcriptase, Integrase, and Maturation Inhibitors, as Well as HIV-1 Coreceptor Tropism

PubMed Central

Gibson, Richard M.; Meyer, Ashley M.; Winner, Dane; Archer, John; Feyertag, Felix; Ruiz-Mateos, Ezequiel; Leal, Manuel; Robertson, David L.; Schmotzer, Christine L.

2014-01-01

With 29 individual antiretroviral drugs available from six classes that are approved for the treatment of HIV-1 infection, a combination of different phenotypic and genotypic tests is currently needed to monitor HIV-infected individuals. In this study, we developed a novel HIV-1 genotypic assay based on deep sequencing (DeepGen HIV) to simultaneously assess HIV-1 susceptibilities to all drugs targeting the three viral enzymes and to predict HIV-1 coreceptor tropism. Patient-derived gag-p2/NCp7/p1/p6/pol-PR/RT/IN- and env-C2V3 PCR products were sequenced using the Ion Torrent Personal Genome Machine. Reads spanning the 3′ end of the Gag, protease (PR), reverse transcriptase (RT), integrase (IN), and V3 regions were extracted, truncated, translated, and assembled for genotype and HIV-1 coreceptor tropism determination. DeepGen HIV consistently detected both minority drug-resistant viruses and non-R5 HIV-1 variants from clinical specimens with viral loads of ≥1,000 copies/ml and from B and non-B subtypes. Additional mutations associated with resistance to PR, RT, and IN inhibitors, previously undetected by standard (Sanger) population sequencing, were reliably identified at frequencies as low as 1%. DeepGen HIV results correlated with phenotypic (original Trofile, 92%; enhanced-sensitivity Trofile assay [ESTA], 80%; TROCAI, 81%; and VeriTrop, 80%) and genotypic (population sequencing/Geno2Pheno with a 10% false-positive rate [FPR], 84%) HIV-1 tropism test results. DeepGen HIV (83%) and Trofile (85%) showed similar concordances with the clinical response following an 8-day course of maraviroc monotherapy (MCT). In summary, this novel all-inclusive HIV-1 genotypic and coreceptor tropism assay, based on deep sequencing of the PR, RT, IN, and V3 regions, permits simultaneous multiplex detection of low-level drug-resistant and/or non-R5 viruses in up to 96 clinical samples. This comprehensive test, the first of its class, will be instrumental in the development of new antiretroviral drugs and, more importantly, will aid in the treatment and management of HIV-infected individuals. PMID:24468782

Maturation inhibitors: a new therapeutic class targets the virus structure.

PubMed

Salzwedel, Karl; Martin, David E; Sakalian, Michael

2007-01-01

The current standard of care for HIV/AIDS in the developed world is HAART therapy, usually a combination of two reverse transcriptase inhibitors and a protease inhibitor. Despite the success of this regimen, there is a continuing need for new drug options to overcome problems with tolerability and the emergence of viral resistance. In this review we discuss the discovery of a potential new class of antiretroviral therapeutics, known as maturation inhibitors, and the development of the first-in-class compound, bevirimat. Bevirimat is distinguished from the currently available antiretrovirals by its unique target and mode of action. While the specific interactions responsible for activity have yet to be fully characterized, it is clear that the target for bevirimat is the Gag polyprotein precursor, the main structural protein responsible for assembly and budding of virion particles. As basic research continues on the precise mechanism of action of bevirimat, clinical development is progressing, with demonstration of both safety and efficacy in early-stage trials. These encouraging results, coupled with the discovery and development of future generations of maturation inhibitors, suggest that maturation inhibitors may be added to the growing set of tools available to control HIV/AIDS.

Brief Report: Long-Term (96-Week) Efficacy and Safety After Switching From Tenofovir Disoproxil Fumarate to Tenofovir Alafenamide in HIV-Infected, Virologically Suppressed Adults

PubMed Central

Raffi, François; Orkin, Chloe; Clarke, Amanda; Slama, Laurence; Gallant, Joel; Daar, Eric; Henry, Keith; Santana-Bagur, Jorge; Stein, David K.; Bellos, Nicholaos; Scarsella, Anthony; Yan, Mingjin; Abram, Michael E.; Cheng, Andrew

2017-01-01

Abstract: In a double-blind, phase 3 trial, 663 HIV-infected, virologically suppressed adults were randomized to switch to tenofovir alafenamide (TAF; n = 333) vs. remain on tenofovir disoproxil fumarate (TDF; n = 330), each coformulated with emtricitabine (FTC), while continuing their third agent (boosted protease inhibitor or unboosted third agent). At week 96, 88.6% on FTC/TAF and 89.1% on FTC/TDF had HIV-1 RNA <50 copies per milliliter [adjusted difference −0.5% (95% confidence interval: −5.3 to 4.4%)]. Proteinuria, albuminuria, proximal renal tubular function, and bone mineral density improved after switching to TAF- from TDF-containing regimens. These longer-term data support FTC/TAF as a safe, well-tolerated, and durable nucleotide reverse transcriptase inhibitor backbone. PMID:28272164

Phylogenetic analysis of HIV-1 reverse transcriptase sequences from 382 patients recruited in JJ Hospital of Mumbai, India, between 2002 and 2008.

PubMed

Deshpande, Alaka; Jauvin, Valerie; Pinson, Patricia; Jeannot, Anne Cecile; Fleury, Herve J

2009-06-01

Analysis of reverse transcriptase (RT) sequences of 382 HIV-1 isolates from untreated and treated patients recruited in JJ Hospital (Mumbai, India) between 2002 and 2008 shows that subtype C is largely predominant (98%) and that non-C sequences cluster with A1, B, CRF01_AE, and CRF06_cpx.

Targets for inhibition of HIV replication: entry, enzyme action, release and maturation.

PubMed

Sierra-Aragón, Saleta; Walter, Hauke

2012-01-01

Inhibition of HIV replication initially targeted viral enzymes, which are exclusively expressed by the virus and not present in the human cell. The development of reverse transcriptase (RT) inhibitors started with the discovery of antiretroviral activity of the nucleoside analog zidovudine in March 1987. Currently, six major classes of antiretroviral drugs are used for the treatment of HIV-infected patients: the RT inhibitors, nucleoside inhibitors and nonnucleoside inhibitors, the protease inhibitors, the integrase inhibitor raltegravir, the fusion inhibitor enfuvirtide (T-20), and the chemokine receptor 5 antagonist maraviroc. A seventh class, the maturation inhibitors, has not yet been approved as their effectiveness is impaired by HIV-1 polymorphisms naturally occurring in 30-40% of HIV-1 therapy-naive isolates. The use of antiretroviral combination therapy has proven to be effective in delaying progression to AIDS and to reconstitute the immune system of HIV-infected individuals. During the last 5 years, the introduction of the newest antiretrovirals has increased treatment efficacy tremendously. However, the development and accumulation of resistance to all antiretroviral drug classes are still a major problem. Additional targets will have to be defined to achieve the ultimate goal: the eradication of the virus from the infected human body. Copyright © 2012 S. Karger AG, Basel.

Application of Reverse Transcriptase-PCR-DGGE as a rapid method for routine determination of Vibrio spp. in foods.

PubMed

Chahorm, Kanchana; Prakitchaiwattana, Cheunjit

2018-01-02

The aim of this research was to evaluate the feasibility of PCR-DGGE and Reverse Transcriptase-PCR-DGGE techniques for rapid detection of Vibrio species in foods. Primers GC567F and 680R were initially evaluated for amplifying DNA and cDNA of ten references Vibrio species by PCR method. The GC-clamp PCR amplicons were separated according to their sequences by the DGGE using 10% (w/v) polyacrylamide gel containing 45-70% urea and formamide denaturants. Two pair of Vibrio species, which could not be differentiated on the gel, was Vibrio fluvialis - Vibrio furnissii and Vibrio parahaemolyticus - Vibrio harveyi. To determine the detection limit, in the community of 10 reference strains containing the same viable population, distinct DNA bands of 3 species; Vibrio cholerae, Vibrio mimicus and Vibrio alginolyticus were consistently observed by PCR-DGGE technique. In fact, 5 species; Vibrio cholerae, Vibrio mimicus, Vibrio alginolyticus, Vibrio parahaemolyticus and Vibrio fluvialis consistently observed by Reverse Transcriptase-PCR-DGGE. In the community containing different viable population increasing from 10 2 to 10 5 CFU/mL, PCR-DGGE analysis only detected the two most prevalent species, while RT-PCR-DGGE detected the five most prevalent species. Therefore, Reverse Transcriptase-PCR-DGGE was also selected for detection of various Vibrio cell conditions, including viable cell (VC), injured cells from frozen cultures (IVC) and injured cells from frozen cultures with pre-enrichment (PIVC). It was found that cDNA band of all cell conditions gave the same migratory patterns, except that multiple cDNA bands of Plesiomonas shigelloides under IVC and PIVC conditions were found. When Reverse Transcriptase-PCR-DGGE was used for detecting Vibrio parahaemolyticus in the pathogen-spiked food samples, Vibrio parahaemolyticus could be detected in the spiked samples containing at least 10 2 CFU/g of this pathogen. The results obtained also corresponded to standard method (USFDA, 2004). In comparison with the detection of the Vibrio profiles in fourteen food samples using standard method, Reverse Transcriptase-PCR-DGGE resulted in 100%, 75% and 50% similarity in 3, 1 and 6 food samples, respectively. Copyright © 2017 Elsevier B.V. All rights reserved.

Identification of a methylated oligoribonucleotide as a potent inhibitor of HIV-1 reverse transcription complex.

PubMed

Grigorov, Boyan; Bocquin, Anne; Gabus, Caroline; Avilov, Sergey; Mély, Yves; Agopian, Audrey; Divita, Gilles; Gottikh, Marina; Witvrouw, Myriam; Darlix, Jean-Luc

2011-07-01

Upon HIV-1 infection of a target cell, the viral reverse transcriptase (RT) copies the genomic RNA to synthesize the viral DNA. The genomic RNA is within the incoming HIV-1 core where it is coated by molecules of nucleocapsid (NC) protein that chaperones the reverse transcription process. Indeed, the RT chaperoning properties of NC extend from the initiation of cDNA synthesis to completion of the viral DNA. New and effective drugs against HIV-1 continue to be required, which prompted us to search for compounds aimed at inhibiting NC protein. Here, we report that the NC chaperoning activity is extensively inhibited in vitro by small methylated oligoribonucleotides (mODN). These mODNs were delivered intracellularly using a cell-penetrating-peptide and found to impede HIV-1 replication in primary human cells at nanomolar concentrations. Extensive analysis showed that viral cDNA synthesis was severely impaired by mODNs. Partially resistant viruses with mutations in NC and RT emerged after months of passaging in cell culture. A HIV-1 molecular clone (NL4.3) bearing these mutations was found to replicate at high concentrations of mODN, albeit with a reduced fitness. Small, methylated ODNs such as mODN-11 appear to be a new type of highly potent inhibitor of HIV-1.

Identification of a methylated oligoribonucleotide as a potent inhibitor of HIV-1 reverse transcription complex

PubMed Central

Grigorov, Boyan; Bocquin, Anne; Gabus, Caroline; Avilov, Sergey; Mély, Yves; Agopian, Audrey; Divita, Gilles; Gottikh, Marina; Witvrouw, Myriam; Darlix, Jean-Luc

2011-01-01

Upon HIV-1 infection of a target cell, the viral reverse transcriptase (RT) copies the genomic RNA to synthesize the viral DNA. The genomic RNA is within the incoming HIV-1 core where it is coated by molecules of nucleocapsid (NC) protein that chaperones the reverse transcription process. Indeed, the RT chaperoning properties of NC extend from the initiation of cDNA synthesis to completion of the viral DNA. New and effective drugs against HIV-1 continue to be required, which prompted us to search for compounds aimed at inhibiting NC protein. Here, we report that the NC chaperoning activity is extensively inhibited in vitro by small methylated oligoribonucleotides (mODN). These mODNs were delivered intracellularly using a cell-penetrating-peptide and found to impede HIV-1 replication in primary human cells at nanomolar concentrations. Extensive analysis showed that viral cDNA synthesis was severely impaired by mODNs. Partially resistant viruses with mutations in NC and RT emerged after months of passaging in cell culture. A HIV-1 molecular clone (NL4.3) bearing these mutations was found to replicate at high concentrations of mODN, albeit with a reduced fitness. Small, methylated ODNs such as mODN-11 appear to be a new type of highly potent inhibitor of HIV-1. PMID:21447560

High-throughput sequencing of human plasma RNA by using thermostable group II intron reverse transcriptases

PubMed Central

Qin, Yidan; Yao, Jun; Wu, Douglas C.; Nottingham, Ryan M.; Mohr, Sabine; Hunicke-Smith, Scott; Lambowitz, Alan M.

2016-01-01

Next-generation RNA-sequencing (RNA-seq) has revolutionized transcriptome profiling, gene expression analysis, and RNA-based diagnostics. Here, we developed a new RNA-seq method that exploits thermostable group II intron reverse transcriptases (TGIRTs) and used it to profile human plasma RNAs. TGIRTs have higher thermostability, processivity, and fidelity than conventional reverse transcriptases, plus a novel template-switching activity that can efficiently attach RNA-seq adapters to target RNA sequences without RNA ligation. The new TGIRT-seq method enabled construction of RNA-seq libraries from <1 ng of plasma RNA in <5 h. TGIRT-seq of RNA in 1-mL plasma samples from a healthy individual revealed RNA fragments mapping to a diverse population of protein-coding gene and long ncRNAs, which are enriched in intron and antisense sequences, as well as nearly all known classes of small ncRNAs, some of which have never before been seen in plasma. Surprisingly, many of the small ncRNA species were present as full-length transcripts, suggesting that they are protected from plasma RNases in ribonucleoprotein (RNP) complexes and/or exosomes. This TGIRT-seq method is readily adaptable for profiling of whole-cell, exosomal, and miRNAs, and for related procedures, such as HITS-CLIP and ribosome profiling. PMID:26554030

Initial therapy with protease inhibitor-sparing regimens: evaluation of nevirapine and delavirdine.

PubMed

Conway, B

2000-06-01

We have compared the results (on-treatment analyses) of 2 randomized clinical trials of protease inhibitor-sparing regimens in drug-naive patients. In the INCAS (Italy, Netherlands, Canada, Australia) study, the mean decrease in plasma viral load over 52 weeks was 2.2 log(10) copies/mL in 40 patients who were receiving zidovudine/didanosine/nevirapine (18 [45%] had maximal suppression), with a mean increase in CD4 T cell counts of 139 cells/microL. In protocol 0021 Part II, the mean decrease in plasma viral load over 52 weeks was 2.1 log(10) copies/mL in 34 patients who were receiving zidovudine/lamivudine/delavirdine (20 [59%] had maximal suppression), with a mean increase in CD4 T cell counts of 88 cells/microL. The virologic and immunologic efficacy of the 2 triple-drug regimens are similar. Until results of long-term studies are available to establish whether a preferred approach to initial therapy exists, nonnucleoside reverse transcriptase inhibitors may be a valuable alternative to protease inhibitors in the initial therapy of antiretroviral-naive, moderately immunosuppressed patients.

Amarogentin Induces Apoptosis of Liver Cancer Cells via Upregulation of p53 and Downregulation of Human Telomerase Reverse Transcriptase in Mice.

PubMed

Huang, Chun; Li, Runqin; Zhang, Yinglin; Gong, Jianping

2017-10-01

Amarogentin has been reported to have a preventive effect on liver cancer via inducing cancer cell apoptosis. We attempted to elucidate the roles of p53-associated apoptosis pathways in the chemopreventive mechanism of amarogentin. The findings of this study will facilitate the development of a novel supplementary strategy for the treatment of liver cancer. The purity of amarogentin was assessed by high-performance liquid chromatography. The inhibitory ratios of the liver cell lines were determined using a Cell Counting Kit-8 following treatment with a gradient concentration of amarogentin. Cell apoptosis was detected by flow cytometry using annexin V-fluorescein isothiocyanate/propidium iodide kits. The gene and protein expression of p53-associated molecules, such as Akt, human telomerase reverse transcriptase, RelA, and p38, was detected by real-time quantitative polymerase chain reaction, Western blotting, and immunohistochemical staining in liver cancer cells and mouse tumor tissues after treatment with amarogentin. The inhibitory effect of amarogentin on cell proliferation was more obvious in liver cancer cells, and amarogentin was more likely to induce the apoptosis of liver cancer cells than that of normal liver cells. The gene and protein expression levels of Akt, RelA, and human telomerase reverse transcriptase were markedly higher in the control group than in the preventive group and treatment groups. Only the expression of human telomerase reverse transcriptase was downregulated, accompanied by the upregulation of p53. The results of our study suggest that amarogentin promotes apoptosis of liver cancer cells by the upregulation of p53 and downregulation of human telomerase reverse transcriptase and prevents the malignant transformation of these cells.

Amarogentin Induces Apoptosis of Liver Cancer Cells via Upregulation of p53 and Downregulation of Human Telomerase Reverse Transcriptase in Mice

PubMed Central

Li, Runqin; Zhang, Yinglin

2016-01-01

Background and Objective: Amarogentin has been reported to have a preventive effect on liver cancer via inducing cancer cell apoptosis. We attempted to elucidate the roles of p53-associated apoptosis pathways in the chemopreventive mechanism of amarogentin. The findings of this study will facilitate the development of a novel supplementary strategy for the treatment of liver cancer. Materials and Methods: The purity of amarogentin was assessed by high-performance liquid chromatography. The inhibitory ratios of the liver cell lines were determined using a Cell Counting Kit-8 following treatment with a gradient concentration of amarogentin. Cell apoptosis was detected by flow cytometry using annexin V-fluorescein isothiocyanate/propidium iodide kits. The gene and protein expression of p53-associated molecules, such as Akt, human telomerase reverse transcriptase, RelA, and p38, was detected by real-time quantitative polymerase chain reaction, Western blotting, and immunohistochemical staining in liver cancer cells and mouse tumor tissues after treatment with amarogentin. Results: The inhibitory effect of amarogentin on cell proliferation was more obvious in liver cancer cells, and amarogentin was more likely to induce the apoptosis of liver cancer cells than that of normal liver cells. The gene and protein expression levels of Akt, RelA, and human telomerase reverse transcriptase were markedly higher in the control group than in the preventive group and treatment groups. Only the expression of human telomerase reverse transcriptase was downregulated, accompanied by the upregulation of p53. Conclusion: The results of our study suggest that amarogentin promotes apoptosis of liver cancer cells by the upregulation of p53 and downregulation of human telomerase reverse transcriptase and prevents the malignant transformation of these cells. PMID:27402632

Inhibitory effect of aqueous dandelion extract on HIV-1 replication and reverse transcriptase activity

PubMed Central

2011-01-01

Background Acquired immunodeficiency syndrome (AIDS), which is caused by the human immunodeficiency virus (HIV), is an immunosuppressive disease that results in life-threatening opportunistic infections. The general problems in current therapy include the constant emergence of drug-resistant HIV strains, adverse side effects and the unavailability of treatments in developing countries. Natural products from herbs with the abilities to inhibit HIV-1 life cycle at different stages, have served as excellent sources of new anti-HIV-1 drugs. In this study, we aimed to investigate the anti-HIV-1 activity of aqueous dandelion extract. Methods The pseudotyped HIV-1 virus has been utilized to explore the anti-HIV-1 activity of dandelion, the level of HIV-1 replication was assessed by the percentage of GFP-positive cells. The inhibitory effect of the dandelion extract on reverse transcriptase activity was assessed by the reverse transcriptase assay kit. Results Compared to control values obtained from cells infected without treatment, the level of HIV-1 replication and reverse transcriptase activity were decreased in a dose-dependent manner. The data suggest that dandelion extract has a potent inhibitory activity against HIV-1 replication and reverse transcriptase activity. The identification of HIV-1 antiviral compounds from Taraxacum officinale should be pursued. Conclusions The dandelion extract showed strong activity against HIV-1 RT and inhibited both the HIV-1 vector and the hybrid-MoMuLV/MoMuSV retrovirus replication. These findings provide additional support for the potential therapeutic efficacy of Taraxacum officinale. Extracts from this plant may be regarded as another starting point for the development of an antiretroviral therapy with fewer side effects. PMID:22078030

World Health Organization Generic Protocol to Assess Drug-Resistant HIV Among Children <18 Months of Age and Newly Diagnosed With HIV in Resource-Limited Countries

PubMed Central

Penazzato, Martina; Jordan, Michael R.; Persaud, Deborah; Mofenson, Lynne M.; Bennett, Diane E.

2012-01-01

Increased use of nonnucleoside reverse transcriptase inhibitors (NNRTIs) in pregnant and breastfeeding women will result in fewer children infected with human immunodeficiency virus (HIV). However, among children infected despite prevention of mother-to-child transmission (PMTCT), a substantial proportion will acquire NNRTI-resistant HIV, potentially compromising response to NNRTI-based antiretroviral therapy (ART). In countries scaling up PMTCT and pediatric ART programs, it is crucial to assess the proportion of young children with drug-resistant HIV to improve health outcomes and support national and global decision making on optimal selection of pediatric first-line ART. This article summarizes a new World Health Organization surveillance protocol to assess resistance using remnant dried blood spot specimens from a representative sample of children aged <18 months being tested for early infant diagnosis. PMID:22544184

High Prevalence of HIV Drug Resistance Among Newly Diagnosed Infants Aged <18 Months: Results From a Nationwide Surveillance in Nigeria.

PubMed

Inzaule, Seth C; Osi, Samuels J; Akinbiyi, Gbenga; Emeka, Asadu; Khamofu, Hadiza; Mpazanje, Rex; Ilesanmi, Oluwafunke; Ndembi, Nicaise; Odafe, Solomon; Sigaloff, Kim C E; Rinke de Wit, Tobias F; Akanmu, Sulaimon

2018-01-01

WHO recommends protease-inhibitor-based first-line regimen in infants because of risk of drug resistance from failed prophylaxis used in prevention of mother-to-child transmission (PMTCT). However, cost and logistics impede implementation in sub-Saharan Africa, and >75% of children still receive nonnucleoside reverse transcriptase inhibitor-based regimen (NNRTI) used in PMTCT. We assessed the national pretreatment drug resistance prevalence of HIV-infected children aged <18 months in Nigeria, using WHO-recommended HIV drug resistance surveillance protocol. We used remnant dried blood spots collected between June 2014 and July 2015 from 15 early infant diagnosis facilities spread across all the 6 geopolitical regions of Nigeria. Sampling was through a probability proportional-to-size approach. HIV drug resistance was determined by population-based sequencing. Overall, in 48% of infants (205 of 430) drug resistance mutations (DRM) were detected, conferring resistance to predominantly NNRTIs (45%). NRTI and multiclass NRTI/NNRTI resistance were present at 22% and 20%, respectively, while resistance to protease inhibitors was at 2%. Among 204 infants with exposure to drugs for PMTCT, 57% had DRMs, conferring NNRTI resistance in 54% and multiclass NRTI/NNRTI resistance in 29%. DRMs were also detected in 34% of 132 PMTCT unexposed infants. A high frequency of PDR, mainly NNRTI-associated, was observed in a nationwide surveillance among newly diagnosed HIV-infected children in Nigeria. PDR prevalence was equally high in PMTCT-unexposed infants. Our results support the use of protease inhibitor-based first-line regimens in HIV-infected young children regardless of PMTCT history and underscore the need to accelerate implementation of the newly disseminated guideline in Nigeria.

Reverse Transcriptase-Containing Particles Induced in Rous Sarcoma Virus-Transformed Rat Cells by Arginine Deprivation

PubMed Central

Kotler, Moshe; Weinberg, Eynat; Haspel, Osnat; Becker, Yechiel

1972-01-01

Incubation of rat cells transformed by Rous sarcoma virus (RSV) in an arginine-deficient medium resulted in accumulation of particles in the culture medium. Such particles did not appear when the transformed rat cells were incubated in a complete medium nor in the medium of primary rat cells which were incubated either in arginine-deficient or complete media. The particles which were released from the arginine-deprived transformed rat cells resemble C-type particles in their properties. These particles band in sucrose gradients at a density of 1.16 g/ml and contain 35S ribonucleic acid (RNA) molecules and a reverse transcriptase activity. Analysis of the cytoplasm of transformed and primary rat cells, deprived and undeprived of arginine, revealed the presence of reverse transcriptase-containing particles which banded in sucrose gradients at a density of 1.14 g/ml. These particles differed from the particles released into the medium by the arginine-deprived RSV-transformed rat cells. The deoxyribonucleic acid (DNA) molecules synthesized in vitro by the reverse transcriptase present in the particles isolated from the medium of arginine-deprived cells hybridized to RSV RNA, whereas the DNA synthesized by the cell-bound enzyme had no homology to RSV RNA. PMID:4116137

Human Immunodeficiency Virus Type 1 cDNA Integration: New Aromatic Hydroxylated Inhibitors and Studies of the Inhibition Mechanism

PubMed Central

Farnet, C. M.; Wang, B.; Hansen, M.; Lipford, J. R.; Zalkow, L.; Robinson, W. E.; Siegel, J.; Bushman, F.

1998-01-01

Integration of the human immunodeficiency virus type 1 (HIV-1) cDNA is a required step for viral replication. Integrase, the virus-encoded enzyme important for integration, has not yet been exploited as a target for clinically useful inhibitors. Here we report on the identification of new polyhydroxylated aromatic inhibitors of integrase including ellagic acid, purpurogallin, 4,8,12-trioxatricornan, and hypericin, the last of which is known to inhibit viral replication. These compounds and others were characterized in assays with subviral preintegration complexes (PICs) isolated from HIV-1-infected cells. Hypericin was found to inhibit PIC assays, while the other compounds tested were inactive. Counterscreening of these and other integrase inhibitors against additional DNA-modifying enzymes revealed that none of the polyhydroxylated aromatic compounds are active against enzymes that do not require metals (methylases, a pox virus topoisomerase). However, all were cross-reactive with metal-requiring enzymes (restriction enzymes, a reverse transcriptase), implicating metal atoms in the inhibitory mechanism. In mechanistic studies, we localized binding of some inhibitors to the catalytic domain of integrase by assaying competition of binding by labeled nucleotides. These findings help elucidate the mechanism of action of the polyhydroxylated aromatic inhibitors and provide practical guidance for further inhibitor development. PMID:9736543

Development of Reverse Transcription Thermostable Helicase-Dependent DNA Amplification for the Detection of Tomato Spotted Wilt Virus.

PubMed

Wu, Xinghai; Chen, Chanfa; Xiao, Xizhi; Deng, Ming Jun

2016-11-01

A protocol for the reverse transcription-helicase-dependent amplification (RT-HDA) of isothermal DNA was developed for the detection of tomato spotted wilt virus (TSWV). Specific primers, which were based on the highly conserved region of the N gene sequence in TSWV, were used for the amplification of virus's RNA. The LOD of RT-HDA, reverse transcriptase-loop-mediated isothermal amplification (RT-LAMP), and reverse transcriptase-polymerase chain reaction (RT-PCR) assays were conducted using 10-fold serial dilution of RNA eluates. TSWV sensitivity in RT-HDA and RT-LAMP was 4 pg RNA compared with 40 pg RNA in RT-PCR. The specificity of RT-HDA for TSWV was high, showing no cross-reactivity with other tomato and Tospovirus viruses including cucumber mosaic virus (CMV), tomato black ring virus (TBRV), tomato mosaic virus (ToMV), or impatiens necrotic spot virus (INSV). The RT-HDA method is effective for the detection of TSWV in plant samples and is a potential tool for early and rapid detection of TSWV.

Critical Contribution of Tyr15 in the HIV-1 Integrase (IN) in Facilitating IN Assembly and Nonenzymatic Function through the IN Precursor Form with Reverse Transcriptase.

PubMed

Takahata, Tatsuro; Takeda, Eri; Tobiume, Minoru; Tokunaga, Kenzo; Yokoyama, Masaru; Huang, Yu-Lun; Hasegawa, Atsuhiko; Shioda, Tatsuo; Sato, Hironori; Kannagi, Mari; Masuda, Takao

2017-01-01

Nonenzymatic roles for HIV-1 integrase (IN) at steps prior to the enzymatic integration step have been reported. To obtain structural and functional insights into the nonenzymatic roles of IN, we performed genetic analyses of HIV-1 IN, focusing on a highly conserved Tyr15 in the N-terminal domain (NTD), which has previously been shown to regulate an equilibrium state between two NTD dimer conformations. Replacement of Tyr15 with alanine, histidine, or tryptophan prevented HIV-1 infection and caused severe impairment of reverse transcription without apparent defects in reverse transcriptase (RT) or in capsid disassembly kinetics after entry into cells. Cross-link analyses of recombinant IN proteins demonstrated that lethal mutations of Tyr15 severely impaired IN structure for assembly. Notably, replacement of Tyr15 with phenylalanine was tolerated for all IN functions, demonstrating that a benzene ring of the aromatic side chain is a key moiety for IN assembly and functions. Additional mutagenic analyses based on previously proposed tetramer models for IN assembly suggested a key role of Tyr15 in facilitating the hydrophobic interaction among IN subunits, together with other proximal residues within the subunit interface. A rescue experiment with a mutated HIV-1 with RT and IN deleted (ΔRT ΔIN) and IN and RT supplied in trans revealed that the nonenzymatic IN function might be exerted through the IN precursor conjugated with RT (RT-IN). Importantly, the lethal mutations of Tyr15 significantly reduced the RT-IN function and assembly. Taken together, Tyr15 seems to play a key role in facilitating the proper assembly of IN and RT on viral RNA through the RT-IN precursor form. Inhibitors of the IN enzymatic strand transfer function (INSTI) have been applied in combination antiretroviral therapies to treat HIV-1-infected patients. Recently, allosteric IN inhibitors (ALLINIs) that interact with HIV-1 IN residues, the locations of which are distinct from the catalytic sites targeted by INSTI, have been discovered. Importantly, ALLINIs affect the nonenzymatic role(s) of HIV-1 IN, providing a rationale for the development of next-generation IN inhibitors with a mechanism that is distinct from that of INSTI. Here, we demonstrate that Tyr15 in the HIV-1 IN NTD plays a critical role during IN assembly by facilitating the hydrophobic interaction of the NTD with the other domains of IN. Importantly, we found that the functional assembly of IN through its fusion form with RT is critical for IN to exert its nonenzymatic function. Our results provide a novel mechanistic insight into the nonenzymatic function of HIV-1 IN and its prevention. Copyright © 2016 American Society for Microbiology.

Follow-up on long-term antiretroviral therapy for cats infected with feline immunodeficiency virus.

PubMed

Medeiros, Sheila de Oliveira; Abreu, Celina Monteiro; Delvecchio, Rodrigo; Ribeiro, Anísia Praxedes; Vasconcelos, Zilton; Brindeiro, Rodrigo de Moraes; Tanuri, Amilcar

2016-04-01

Feline immunodeficiency virus (FIV) is a lentivirus that induces AIDS-like disease in cats. Some of the antiretroviral drugs available to treat patients with HIV type 1 are used to treat FIV-infected cats; however, antiretroviral therapy (ART) is not used in cats as a long-term treatment. In this study, the effects of long-term ART were evaluated in domestic cats treated initially with the nucleoside transcriptase reverse inhibitor (NTRI) zidovudine (AZT) over a period ranging from 5-6 years, followed by a regimen of the NTRI lamivudine (3TC) plus AZT over 3 years. Viral load, sequencing of pol (reverse transcriptase [RT]) region and CD4:CD8 lymphocyte ratio were evaluated during and after treatment. Untreated cats were evaluated as a control group. CD4:CD8 ratios were lower, and uncharacterized resistance mutations were found in the RT region in the group of treated cats. A slight increase in viral load was observed in some cats after discontinuing treatment. The data strongly suggest that treated cats were resistant to therapy, and uncharacterized resistance mutations in the RT gene of FIV were selected for by AZT. Few studies have been conducted to evaluate the effect of long-term antiretroviral therapy in cats. To date, resistance mutations have not been described in vivo. © ISFM and AAFP 2015.

Targeted transgenic overexpression of mitochondrial thymidine kinase (TK2) alters mitochondrial DNA (mtDNA) and mitochondrial polypeptide abundance: transgenic TK2, mtDNA, and antiretrovirals.

PubMed

Hosseini, Seyed H; Kohler, James J; Haase, Chad P; Tioleco, Nina; Stuart, Tami; Keebaugh, Erin; Ludaway, Tomika; Russ, Rodney; Green, Elgin; Long, Robert; Wang, Liya; Eriksson, Staffan; Lewis, William

2007-03-01

Mitochondrial toxicity limits nucleoside reverse transcriptase inhibitors (NRTIs) for acquired immune deficiency syndrome. NRTI triphosphates, the active moieties, inhibit human immunodeficiency virus reverse transcriptase and eukaryotic mitochondrial DNA polymerase pol-gamma. NRTI phosphorylation seems to correlate with mitochondrial toxicity, but experimental evidence is lacking. Transgenic mice (TGs) with cardiac overexpression of thymidine kinase isoforms (mitochondrial TK2 and cytoplasmic TK1) were used to study NRTI mitochondrial toxicity. Echocardiography and nuclear magnetic resonance imaging defined cardiac performance and structure. TK gene copy and enzyme activity, mitochondrial (mt) DNA and polypeptide abundance, succinate dehydrogenase and cytochrome oxidase histochemistry, and electron microscopy correlated with transgenesis, mitochondrial structure, and biogenesis. Antiretroviral combinations simulated therapy. Untreated hTK1 or TK2 TGs exhibited normal left ventricle mass. In TK2 TGs, cardiac TK2 gene copy doubled, activity increased 300-fold, and mtDNA abundance doubled. Abundance of the 17-kd subunit of complex I, succinate dehydrogenase histochemical activity, and cristae density increased. NRTIs increased left ventricle mass 20% in TK2 TGs. TK activity increased 3 logs in hTK1 TGs, but no cardiac phenotype resulted. NRTIs abrogated functional effects of transgenically increased TK2 activity but had no effect on TK2 mtDNA abundance. Thus, NRTI mitochondrial phosphorylation by TK2 is integral to clinical NRTI mitochondrial toxicity.

Targeted Transgenic Overexpression of Mitochondrial Thymidine Kinase (TK2) Alters Mitochondrial DNA (mtDNA) and Mitochondrial Polypeptide Abundance

PubMed Central

Hosseini, Seyed H.; Kohler, James J.; Haase, Chad P.; Tioleco, Nina; Stuart, Tami; Keebaugh, Erin; Ludaway, Tomika; Russ, Rodney; Green, Elgin; Long, Robert; Wang, Liya; Eriksson, Staffan; Lewis, William

2007-01-01

Mitochondrial toxicity limits nucleoside reverse transcriptase inhibitors (NRTIs) for acquired immune deficiency syndrome. NRTI triphosphates, the active moieties, inhibit human immunodeficiency virus reverse transcriptase and eukaryotic mitochondrial DNA polymerase pol-γ. NRTI phosphorylation seems to correlate with mitochondrial toxicity, but experimental evidence is lacking. Transgenic mice (TGs) with cardiac overexpression of thymidine kinase isoforms (mitochondrial TK2 and cytoplasmic TK1) were used to study NRTI mitochondrial toxicity. Echocardiography and nuclear magnetic resonance imaging defined cardiac performance and structure. TK gene copy and enzyme activity, mitochondrial (mt) DNA and polypeptide abundance, succinate dehydrogenase and cytochrome oxidase histochemistry, and electron microscopy correlated with transgenesis, mitochondrial structure, and biogenesis. Antiretroviral combinations simulated therapy. Untreated hTK1 or TK2 TGs exhibited normal left ventricle mass. In TK2 TGs, cardiac TK2 gene copy doubled, activity increased 300-fold, and mtDNA abundance doubled. Abundance of the 17-kd subunit of complex I, succinate dehydrogenase histochemical activity, and cristae density increased. NRTIs increased left ventricle mass 20% in TK2 TGs. TK activity increased 3 logs in hTK1 TGs, but no cardiac phenotype resulted. NRTIs abrogated functional effects of transgenically increased TK2 activity but had no effect on TK2 mtDNA abundance. Thus, NRTI mitochondrial phosphorylation by TK2 is integral to clinical NRTI mitochondrial toxicity. PMID:17322372

Risks Associated With Lentiviral Vector Exposures and Prevention Strategies

PubMed Central

Schlimgen, Ryan; Howard, John; Wooley, Dawn; Thompson, Maureen; Baden, Lindsey R.; Yang, Otto O.; Christiani, David C.; Mostoslavsky, Gustavo; Diamond, David V.; Duane, Elizabeth Gilman; Byers, Karen; Winters, Thomas; Gelfand, Jeffrey A.; Fujimoto, Gary; Hudson, T. Warner; Vyas, Jatin M.

2016-01-01

Lentiviral vectors (LVVs) are powerful genetic tools that are being used with greater frequency in biomedical laboratories and clinical trials. Adverse events reported from initial clinical studies provide a basis for risk assessment of occupational exposures, yet many questions remain about the potential harm that LVVs may cause. We review those risks and provide a framework for principal investigators, Institutional Biosafety Committees, and occupational health professionals to assess and communicate the risks of exposure to staff. We also provide recommendations to federal research and regulatory agencies for tracking LVV exposures to evaluate long-term outcomes. U.S. Food and Drug Administration approved antiviral drugs for HIV have theoretical benefits in LVV exposures, although evidence to support their use is currently limited. If treatment is appropriate, we recommend a 7-day treatment with an integrase inhibitor with or without a reverse transcriptase inhibitor within 72 hours of exposure. PMID:27930472

Antiretroviral Therapy for HIV-2 Infection: Recommendations for Management in Low-Resource Settings

PubMed Central

Peterson, Kevin; Jallow, Sabelle; Rowland-Jones, Sarah L.; de Silva, Thushan I.

2011-01-01

HIV-2 contributes approximately a third to the prevalence of HIV in West Africa and is present in significant amounts in several low-income countries outside of West Africa with historical ties to Portugal. It complicates HIV diagnosis, requiring more expensive and technically demanding testing algorithms. Natural polymorphisms and patterns in the development of resistance to antiretrovirals are reviewed, along with their implications for antiretroviral therapy. Nonnucleoside reverse transcriptase inhibitors, crucial in standard first-line regimens for HIV-1 in many low-income settings, have no effect on HIV-2. Nucleoside analogues alone are not sufficiently potent enough to achieve durable virologic control. Some protease inhibitors, in particular those without ritonavir boosting, are not sufficiently effective against HIV-2. Following review of the available evidence and taking the structure and challenges of antiretroviral care in West Africa into consideration, the authors make recommendations and highlight the needs of special populations. PMID:21490779

Indolylarylsulfones as HIV-1 non-nucleoside reverse transcriptase inhibitors: new cyclic substituents at indole-2-carboxamide.

PubMed

La Regina, Giuseppe; Coluccia, Antonio; Brancale, Andrea; Piscitelli, Francesco; Gatti, Valerio; Maga, Giovanni; Samuele, Alberta; Pannecouque, Christophe; Schols, Dominique; Balzarini, Jan; Novellino, Ettore; Silvestri, Romano

2011-03-24

New indolylarylsulfone derivatives bearing cyclic substituents at indole-2-carboxamide linked through a methylene/ethylene spacer were potent inhibitors of the WT HIV-1 replication in CEM and PBMC cells with inhibitory concentrations in the low nanomolar range. Against the mutant L100I and K103N RT HIV-1 strains in MT-4 cells, compounds 20, 24-26, 36, and 40 showed antiviral potency superior to that of NVP and EFV. Against these mutant strains, derivatives 20, 24-26, and 40 were equipotent to ETV. Molecular docking experiments on this novel series of IAS analogues have also suggested that the H-bond interaction between the nitrogen atom in the carboxamide chain of IAS and Glu138:B is important in the binding of these compounds. These results are in accordance with the experimental data obtained on the WT and on the mutant HIV-1 strains tested.

Dissimilarities in the Metabolism of Antiretroviral Drugs used in HIV Pre-exposure Prophylaxis in Colon and Vagina Tissues

PubMed Central

To, Elaine E.; Hendrix, Craig W.; Bumpus, Namandjé N.

2013-01-01

Attempts to prevent HIV infection through pre-exposure prophylaxis (PrEP) include topical application of anti-HIV drugs to the mucosal sites of infection; however, a potential role for local drug metabolizing enzymes in modulating the exposure of the mucosal tissues to these drugs has yet to be explored. Here we present the first report that enzymes belonging to the cytochrome P450 (CYP) and UDP-glucuronosyltransferase (UGT) families of drug metabolizing enzymes are expressed and active in vaginal and colorectal tissue using biopsies collected from healthy volunteers. In doing so, we discovered that dapivirine and maraviroc, a non-nucleoside reverse transcriptase inhibitor and an entry inhibitor currently in development as microbicides for HIV PrEP, are differentially metabolized in colorectal tissue and vaginal tissue. Taken together, these data should help to guide the optimization of small molecules being developed for HIV PrEP. PMID:23965226

Antiretroviral Drug Use in a Cohort of HIV-Uninfected Women in the United States: HIV Prevention Trials Network 064.

PubMed

Chen, Iris; Clarke, William; Ou, San-San; Marzinke, Mark A; Breaud, Autumn; Emel, Lynda M; Wang, Jing; Hughes, James P; Richardson, Paul; Haley, Danielle F; Lucas, Jonathan; Rompalo, Anne; Justman, Jessica E; Hodder, Sally L; Eshleman, Susan H

2015-01-01

Antiretroviral (ARV) drug use was analyzed in HIV-uninfected women in an observational cohort study conducted in 10 urban and periurban communities in the United States with high rates of poverty and HIV infection. Plasma samples collected in 2009-2010 were tested for the presence of 16 ARV drugs. ARV drugs were detected in samples from 39 (2%) of 1,806 participants: 27/181 (15%) in Baltimore, MD and 12/179 (7%) in Bronx, NY. The ARV drugs detected included different combinations of non-nucleoside reverse transcriptase inhibitors and protease inhibitors (1-4 drugs/sample). These data were analyzed in the context of self-reported data on ARV drug use. None of the 39 women who had ARV drugs detected reported ARV drug use at any study visit. Further research is needed to evaluate ARV drug use by HIV-uninfected individuals.

Antiretroviral Drug Use in a Cohort of HIV-Uninfected Women in the United States: HIV Prevention Trials Network 064

PubMed Central

Chen, Iris; Clarke, William; Ou, San-San; Marzinke, Mark A.; Breaud, Autumn; Emel, Lynda M.; Wang, Jing; Hughes, James P.; Richardson, Paul; Haley, Danielle F.; Lucas, Jonathan; Rompalo, Anne; Justman, Jessica E.; Hodder, Sally L.; Eshleman, Susan H.

2015-01-01

Antiretroviral (ARV) drug use was analyzed in HIV-uninfected women in an observational cohort study conducted in 10 urban and periurban communities in the United States with high rates of poverty and HIV infection. Plasma samples collected in 2009–2010 were tested for the presence of 16 ARV drugs. ARV drugs were detected in samples from 39 (2%) of 1,806 participants: 27/181 (15%) in Baltimore, MD and 12/179 (7%) in Bronx, NY. The ARV drugs detected included different combinations of non-nucleoside reverse transcriptase inhibitors and protease inhibitors (1–4 drugs/sample). These data were analyzed in the context of self-reported data on ARV drug use. None of the 39 women who had ARV drugs detected reported ARV drug use at any study visit. Further research is needed to evaluate ARV drug use by HIV-uninfected individuals. PMID:26445283

Drug Interactions and Antiretroviral Drug Monitoring

PubMed Central

Foy, Matthew; Sperati, C. John; Lucas, Gregory M.

2014-01-01

Due to the improved longevity afforded by combination antiretroviral therapy (cART), HIV-infected individuals are developing several non-AIDS related comorbid conditions. Consequently, medical management of the HIV-infected population is increasingly complex, with a growing list of potential drug-drug interactions (DDIs). This article reviews some of the most relevant and emerging potential interactions between antiretroviral medications and other agents. The most common DDIs are those involving protease inhibitors or non-nucleoside reverse transcriptase inhibitors which alter the cytochrome P450 enzyme system and/or drug transporters such as p-glycoprotein. Of note are the new agents for the treatment of chronic hepatitis C virus infection. These new classes of drugs and others drugs which are increasingly used in this patient population represent a significant challenge with regard to achieving the goals of effective HIV suppression and minimization of drug-related toxicities. Awareness of DDIs and a multidisciplinary approach are imperative in reaching these goals. PMID:24950731

Molecular Docking and Molecular Dynamics to Identify a Novel Human Immunodeficiency Virus Inhibitor from Alkaloids of Toddalia asiatica.

PubMed

Priya, R; Sumitha, Rajendrarao; Doss, C George Priya; Rajasekaran, C; Babu, S; Seenivasan, R; Siva, R

2015-10-01

Acquired immunodeficiency syndrome caused by human immunodeficiency virus (HIV) is an immunosuppressive disease. Over the past decades, it has plagued human health due to the grave consequences in its harness. For this reason, anti-HIV agents are imperative, and the search for the same from natural resources would assure the safety. In this investigation we have performed molecular docking, molecular property prediction, drug-likeness score, and molecular dynamics (MD) simulation to develop a novel anti-HIV drug. We have screened 12 alkaloids from a medicinal plant Toddalia asiatica for its probabilistic binding with the active site of the HIV-1-reverse transcriptase (HIV-1-RT) domain (the major contributor to the onset of the disease). The docking results were evaluated based on free energies of binding (ΔG), and the results suggested toddanol, toddanone, and toddalenone to be potent inhibitors of HIV-1-RT. In addition, the alkaloids were subjected to molecular property prediction analysis. Toddanol and toddanone with more rotatable bonds were found to have a drug-likeness score of 0.23 and 0.11, respectively. These scores were comparable with the standard anti-HIV drug zidovudine with a model score 0.28. Finally, two characteristic protein-ligand complexes were exposed to MD simulation to determine the stability of the predicted conformations. The toddanol-RT complex showed higher stability and stronger H-bonds than toddanone-RT complex. Based on these observations, we firmly believe that the alkaloid toddanol could aid in efficient HIV-1 drug discovery. In the present study, the molecular docking and MD simulations are performed to explore the possible binding mode of HIV 1 RT with 12 alkaloids of T. asiatica. Molecular docking by AutoDock4 revealed three alkaloids toddanol, toddanone, and toddalenone with highest binding affinity towards HIV 1 RT. The drug likeness model score revealed a positive score for toddanol and toddanone which is comparable to the drug likeness score of the standard anti HIV drug zidovudine. Results from simulation analysis revealed that toddanol RT complex is more stable than toddanone RT complex inferring toddanol as a potential anti HIV drug molecule. Abbreviations used: HIV: Human immunodeficiency virus, HIV 1 RT: HIV 1 reverse transcriptase, RNase H: Ribonuclease H, MD: Molecular dynamics, PDB: Protein databank, RMSD: Root mean square deviation, RMSF: Root mean square fluctuation.

Novel Structure of Ty3 Reverse Transcriptase | Center for Cancer Research

Cancer.gov

Retrotransposons are mobile genetic elements that self amplify via a single-stranded RNA intermediate, which is converted to double-stranded DNA by an encoded reverse transcriptase (RT) with both DNA polymerase (pol) and ribonuclease H (RNase) activities. Categorized by whether they contain flanking long terminal repeat (LTR) sequences, retrotransposons play a critical role in the architecture of eukaryotic genomes and are the evolutionary origin of retroviruses, including human immunodeficiency virus (HIV).

The Role of eIF4E Activity in Breast Cancer

DTIC Science & Technology

2010-08-01

ORF, open reading frame; qPCR, quantitative PCR; RACE, rapid amplification of cDNA ends; RT, reverse transcriptase ; uORF, upstream ORF; UTR...were also performed using template lacking RT ( reverse transcriptase ): products were either undetectable or greatly reduced (>30000-fold less product...have previously shown that a 5’UTR expressed from the human AXIN2 gene contains a sixty nucleotide sequence that is predicted to form a stable stem

Treatment with integrase inhibitor suggests a new interpretation of HIV RNA decay curves that reveals a subset of cells with slow integration

DOE PAGES

Cardozo, Erwing Fabian; Andrade, Adriana; Mellors, John W.; ...

2017-07-05

The kinetics of HIV-1 decay under treatment depends on the class of antiretrovirals used. Mathematical models are useful to interpret the different profiles, providing quantitative information about the kinetics of virus replication and the cell populations contributing to viral decay. We modeled proviral integration in short- and long-lived infected cells to compare viral kinetics under treatment with and without the integrase inhibitor raltegravir (RAL). We fitted the model to data obtained from participants treated with RAL-containing regimes or with a four-drug regimen of protease and reverse transcriptase inhibitors. Our model explains the existence and quantifies the three phases of HIV-1more » RNA decay in RAL-based regimens vs. the two phases observed in therapies without RAL. Our findings indicate that HIV-1 infection is mostly sustained by short-lived infected cells with fast integration and a short viral production period, and by long-lived infected cells with slow integration but an equally short viral production period. We propose that these cells represent activated and resting infected CD4+ T-cells, respectively, and estimate that infection of resting cells represent ~4% of productive reverse transcription events in chronic infection. RAL reveals the kinetics of proviral integration, showing that in short-lived cells the pre-integration population has a half-life of ~7 hours, whereas in long-lived cells this half-life is ~6 weeks. We also show that the efficacy of RAL can be estimated by the difference in viral load at the start of the second phase in protocols with and without RAL. Altogether, we provide a mechanistic model of viral infection that parsimoniously explains the kinetics of viral load decline under multiple classes of antiretrovirals.« less

Treatment with integrase inhibitor suggests a new interpretation of HIV RNA decay curves that reveals a subset of cells with slow integration

DOE Office of Scientific and Technical Information (OSTI.GOV)

Cardozo, Erwing Fabian; Andrade, Adriana; Mellors, John W.

The kinetics of HIV-1 decay under treatment depends on the class of antiretrovirals used. Mathematical models are useful to interpret the different profiles, providing quantitative information about the kinetics of virus replication and the cell populations contributing to viral decay. We modeled proviral integration in short- and long-lived infected cells to compare viral kinetics under treatment with and without the integrase inhibitor raltegravir (RAL). We fitted the model to data obtained from participants treated with RAL-containing regimes or with a four-drug regimen of protease and reverse transcriptase inhibitors. Our model explains the existence and quantifies the three phases of HIV-1more » RNA decay in RAL-based regimens vs. the two phases observed in therapies without RAL. Our findings indicate that HIV-1 infection is mostly sustained by short-lived infected cells with fast integration and a short viral production period, and by long-lived infected cells with slow integration but an equally short viral production period. We propose that these cells represent activated and resting infected CD4+ T-cells, respectively, and estimate that infection of resting cells represent ~4% of productive reverse transcription events in chronic infection. RAL reveals the kinetics of proviral integration, showing that in short-lived cells the pre-integration population has a half-life of ~7 hours, whereas in long-lived cells this half-life is ~6 weeks. We also show that the efficacy of RAL can be estimated by the difference in viral load at the start of the second phase in protocols with and without RAL. Altogether, we provide a mechanistic model of viral infection that parsimoniously explains the kinetics of viral load decline under multiple classes of antiretrovirals.« less

Novel antiretroviral combinations in treatment-experienced patients with HIV infection: rationale and results.

PubMed

Taiwo, Babafemi; Murphy, Robert L; Katlama, Christine

2010-09-10

Novel antiretroviral drugs offer different degrees of improvement in activity against drug-resistant HIV, short- and long-term tolerability, and dosing convenience compared with earlier drugs. Those drugs approved more recently and commonly used in treatment-experienced patients include the entry inhibitor enfuvirtide, protease inhibitors (PIs) [darunavir and tipranavir], a C-C chemokine receptor (CCR) type 5 antagonist (maraviroc), an integrase inhibitor (raltegravir) and etravirine, a non-nucleoside reverse transcriptase inhibitor (NNRTI). Novel agents in earlier stages of development include a CCR5 monoclonal antibody (PRO 140) administered subcutaneously once weekly, once-daily integrase inhibitors (elvitegravir and S/GSK1349572), and several nucleoside (nucleotide) reverse transcriptase inhibitors and NNRTIs. Bevirimat, a maturation inhibitor, has compromised activity in the presence of relatively common Gag polymorphisms. Viral suppression is necessary to control the evolution of drug resistance, reduce chronic immune activation that probably underlies the excess morbidity and mortality in HIV-infected patients, and reduce viral transmission, including transmitted drug resistance. In general, the proportion of viraemic patients who achieve suppression increases with the number of active pharmacokinetically compatible antiretroviral drugs in the regimen. In the ANRS139-TRIO trial, 86% of highly treatment-experienced patients treated with darunavir-ritonavir, etravirine and raltegravir had HIV RNA <50 copies/mL at 48 weeks. In patients who had received at least 12 weeks of a stable regimen and had no darunavir resistance-associated mutations, once-daily darunavir boosted with ritonavir 100 mg was virologically noninferior with better lipid effects than with the twice-daily dosing, which requires a 200 mg total daily dose of ritonavir. Raltegravir plus a boosted PI is being investigated for second-line therapy in patients not responding to NNRTI-based first-line treatment in resource-limited settings (RLS). However, concerns about this potential strategy include the low barrier against resistance of raltegravir, limited penetration of some PIs into the CNS and the unknown impact of integrase polymorphisms seen more commonly in non-B subtype HIV-1. In patients who have already achieved viral suppression, novel agents may be used to simplify the dosing schedule, lower costs (such as by switching to boosted PI monotherapy), reduce adverse events or preserve antiretroviral drug options, especially since the absence of an HIV eradication strategy implies the need for life-long combination antiretroviral therapy. Switching enfuvirtide to raltegravir eliminated painful injection-site reactions without compromising virological suppression. Two studies found different virological outcomes when patients were switched from lopinavir/ritonavir to raltegravir, but there was an improvement in the lipid profile. Simplifying to darunavir-ritonavir monotherapy after suppression of plasma HIV RNA to <50 copies/mL has been found to be safe with no emergence of resistance in cases of viral rebound, but longer-term data are needed. The initial suggestion that maraviroc may possess unique CD4+ T-cell boosting effects was not confirmed in several clinical trials. Improved understanding of HIV pathogenesis has opened new frontiers for research such as identifying the sources, consequences and optimal management of residual viraemia in those with plasma HIV RNA <50 copies/mL. Globally, however, one of the most urgent priorities is providing the increasing number of treatment-experienced virologically failing patients in RLS with access to optimal treatment, including those treatments based on novel antiretroviral agents.

Evaluation of Cytokine Synthesis in Human Whole Blood by Enzyme Linked Immunoassay (ELISA), Reverse Transcriptase Polymerase Chain Reaction (RT-PCR), and Flow Cytometry

DTIC Science & Technology

2007-05-08

deoxynucleotide triphosphates, from Sigma. Sequences for glyceraldehyde-3-phosphate dehydrogenase ( G3PDH ), IL-8,and TNF-a were amplified with primer...This was accomplished by normalizing all samples to the mRNA for the moderately expressed housekeeping function glyceraldehyde-3 -phosphate...without and with isolation of cells before reverse transcription and PCR. G3PDH mRNA target amplifies at 983 base pairs. The 630 base pair band is the

Comprehensive phylogenetic analysis of bacterial reverse transcriptases.

PubMed

Toro, Nicolás; Nisa-Martínez, Rafael

2014-01-01

Much less is known about reverse transcriptases (RTs) in prokaryotes than in eukaryotes, with most prokaryotic enzymes still uncharacterized. Two surveys involving BLAST searches for RT genes in prokaryotic genomes revealed the presence of large numbers of diverse, uncharacterized RTs and RT-like sequences. Here, using consistent annotation across all sequenced bacterial species from GenBank and other sources via RAST, available from the PATRIC (Pathogenic Resource Integration Center) platform, we have compiled the data for currently annotated reverse transcriptases from completely sequenced bacterial genomes. RT sequences are broadly distributed across bacterial phyla, but green sulfur bacteria and cyanobacteria have the highest levels of RT sequence diversity (≤85% identity) per genome. By contrast, phylum Actinobacteria, for which a large number of genomes have been sequenced, was found to have a low RT sequence diversity. Phylogenetic analyses revealed that bacterial RTs could be classified into 17 main groups: group II introns, retrons/retron-like RTs, diversity-generating retroelements (DGRs), Abi-like RTs, CRISPR-Cas-associated RTs, group II-like RTs (G2L), and 11 other groups of RTs of unknown function. Proteobacteria had the highest potential functional diversity, as they possessed most of the RT groups. Group II introns and DGRs were the most widely distributed RTs in bacterial phyla. Our results provide insights into bacterial RT phylogeny and the basis for an update of annotation systems based on sequence/domain homology.

Comprehensive Phylogenetic Analysis of Bacterial Reverse Transcriptases

PubMed Central

Toro, Nicolás; Nisa-Martínez, Rafael

2014-01-01

Much less is known about reverse transcriptases (RTs) in prokaryotes than in eukaryotes, with most prokaryotic enzymes still uncharacterized. Two surveys involving BLAST searches for RT genes in prokaryotic genomes revealed the presence of large numbers of diverse, uncharacterized RTs and RT-like sequences. Here, using consistent annotation across all sequenced bacterial species from GenBank and other sources via RAST, available from the PATRIC (Pathogenic Resource Integration Center) platform, we have compiled the data for currently annotated reverse transcriptases from completely sequenced bacterial genomes. RT sequences are broadly distributed across bacterial phyla, but green sulfur bacteria and cyanobacteria have the highest levels of RT sequence diversity (≤85% identity) per genome. By contrast, phylum Actinobacteria, for which a large number of genomes have been sequenced, was found to have a low RT sequence diversity. Phylogenetic analyses revealed that bacterial RTs could be classified into 17 main groups: group II introns, retrons/retron-like RTs, diversity-generating retroelements (DGRs), Abi-like RTs, CRISPR-Cas-associated RTs, group II-like RTs (G2L), and 11 other groups of RTs of unknown function. Proteobacteria had the highest potential functional diversity, as they possessed most of the RT groups. Group II introns and DGRs were the most widely distributed RTs in bacterial phyla. Our results provide insights into bacterial RT phylogeny and the basis for an update of annotation systems based on sequence/domain homology. PMID:25423096

Endothelial Function in HIV-Infected Antiretroviral Naïve Subjects Before and After Starting Potent Antiretroviral Therapy: AIDS Clinical Trials Group Study 5152s

PubMed Central

Torriani, Francesca J.; Komarow, Lauren; Parker, Robert A.; Cotter, Bruno R.; Currier, Judith S.; Dubé, Michael P.; Fichtenbaum, Carl J.; Gerschenson, Mariana; Mitchell, Carol K.C.; Murphy, Robert L.; Squires, Kathleen; Stein, James H.

2008-01-01

Objectives This study evaluated the effects of three class-sparing antiretroviral therapy (ART) regimens on endothelial function in HIV-infected subjects participating in a randomized trial. Background Endothelial dysfunction has been observed in patients receiving ART for human immunodeficiency virus (HIV) infection. Methods This was a prospective, multicenter study of treatment-naïve subjects who were randomly assigned to receive a protease inhibitor-sparing regimen of nucleoside reverse transcriptase inhibitors (NRTIs) + efavirenz, a non-nucleoside reverse transcriptase inhibitor-sparing regimen of NRTIs + lopinavir/ritonavir, or a NRTI-sparing regimen of efavirenz + lopinavir/ritonavir. NRTIs were lamivudine + stavudine, zidovudine, or tenofovir. Brachial artery flow-mediated dilation (FMD) was determined by B-mode ultrasound before starting on ART, then after 4 and 24 weeks. Results There were 82 subjects (median age 35 years, 91% men, 54% white). Baseline CD4 cell counts and plasma HIV RNA values were 245 cells/mm3 and 4.8 log10 copies/ml, respectively. At baseline, FMD was 3.68% (interquartile range 1.98 – 5.51%). After 4 and 24 weeks of ART, plasma HIV RNA decreased by 2.1 and 3.0 log10 copies/mL, respectively. FMD increased by 0.74% (−0.62 – +2.74, p=0.003) and 1.48% (−0.20 – +4.30%, p< 0.001), respectively, with similar changes in each arm (pKW>0.600). The decrease in plasma HIV RNA at 24 weeks was associated with greater FMD (rs=− 0.30, p=0.017). Conclusions Among treatment-naïve individuals with HIV, three different ART regimens rapidly improved endothelial function. Benefits were similar for all ART regimens, appeared quickly, and persisted at 24 weeks. Condensed Abstract Among 82 treatment-naïve HIV-infected subjects participating in a prospective, multicenter study of three class-sparing antiretroviral therapy regimens, flow-mediated dilation of the brachial artery improved after 4 (+0.74%, p=0.003) and 24 weeks (+1.48%, p< 0.001), with similar changes in each arm (pKW>0.600). PMID:18687253

Abacavir–Lamivudine versus Tenofovir–Emtricitabine for Initial HIV-1 Therapy

PubMed Central

Sax, Paul E.; Tierney, Camlin; Collier, Ann C.; Fischl, Margaret A.; Mollan, Katie; Peeples, Lynne; Godfrey, Catherine; Jahed, Nasreen C.; Myers, Laurie; Katzenstein, David; Farajallah, Awny; Rooney, James F.; Ha, Belinda; Woodward, William C.; Koletar, Susan L.; Johnson, Victoria A.; Geiseler, P. Jan; Daar, Eric S.

2009-01-01

BACKGROUND The use of fixed-dose combination nucleoside reverse-transcriptase inhibitors (NRTIs) with a nonnucleoside reverse-transcriptase inhibitor or a ritonavir-boosted protease inhibitor is recommended as initial therapy in patients with human immunodeficiency virus type 1 (HIV-1) infection, but which NRTI combination has greater efficacy and safety is not known. METHODS In a randomized, blinded equivalence study involving 1858 eligible patients, we compared four once-daily anti retroviral regimens as initial therapy for HIV-1 infection: abacavir–lamivudine or tenofovir disoproxil fumarate (DF)–emtricitabine plus efavirenz or ritonavir-boosted atazanavir. The primary efficacy end point was the time from randomization to virologic failure (defined as a confirmed HIV-1 RNA level ≥1000 copies per milliliter at or after 16 weeks and before 24 weeks, or ≥200 copies per milliliter at or after 24 weeks). RESULTS A scheduled interim review by an independent data and safety monitoring board showed significant differences in virologic efficacy, according to the NRTI combination, among patients with screening HIV-1 RNA levels of 100,000 copies per milliliter or more. At a median follow-up of 60 weeks, among the 797 patients with screening HIV-1 RNA levels of 100,000 copies per milliliter or more, the time to virologic failure was significantly shorter in the abacavir–lamivudine group than in the tenofovir DF–emtricitabine group (hazard ratio, 2.33; 95% confidence interval, 1.46 to 3.72; P<0.001), with 57 virologic failures (14%) in the abacavir–lamivudine group versus 26 (7%) in the tenofovir DF–emtricitabine group. The time to the first adverse event was also shorter in the abacavir–lamivudine group (P<0.001). There was no significant difference between the study groups in the change from the baseline CD4 cell count at week 48. CONCLUSIONS In patients with screening HIV-1 RNA levels of 100,000 copies per milliliter or more, the times to virologic failure and the first adverse event were both significantly shorter in patients randomly assigned to abacavir–lamivudine than in those assigned to tenofovir DF–emtricitabine. (ClinicalTrials.gov number, NCT00118898.) PMID:19952143

Trends in Transmission of Drug Resistance and Prevalence of Non-B Subtypes in Patients with Acute or Recent HIV-1 Infection in Barcelona in the Last 16 Years (1997-2012).

PubMed

Ambrosioni, Juan; Sued, Omar; Nicolas, David; Parera, Marta; López-Diéguez, María; Romero, Anabel; Agüero, Fernando; Marcos, María Ángeles; Manzardo, Christian; Zamora, Laura; Gómez-Carrillo, Manuel; Gatell, José María; Pumarola, Tomás; Miró, José María

2015-01-01

To evaluate the prevalence of transmitted drug resistance (TDR) and non-B subtypes in patients with acute/recent HIV-1 infection in Barcelona during the period 1997-2012. Patients from the "Hospital Clínic Primary HIV-1 Infection Cohort" with a genotyping test performed within 180 days of infection were included. The 2009 WHO List of Mutations for Surveillance of Transmitted HIV-1 Drug Resistance was used for estimating the prevalence of TDR and phylogenetic analysis for subtype determination. 189 patients with acute/recent HIV-1 infection were analyzed in 4 time periods (1997-2000, n=28; 2001-4, n=42; 2005-8, n=55 and 2009-12, n=64). The proportion of patients with acute/recent HIV-1 infection with respect to the total of newly HIV-diagnosed patients in our center increased over the time and was 2.18%, 3.82%, 4.15% and 4.55% for the 4 periods, respectively (p=0.005). The global prevalence of TDR was 9%, or 17.9%, 9.5%, 3.6% and 9.4% by study period (p=0.2). The increase in the last period was driven by protease-inhibitor and nucleoside-reverse-transcriptase-inhibitor resistance mutations while non-nucleoside-reverse-transcriptase inhibitor TDR and TDR of more than one family decreased. The overall prevalence of non-B subtypes was 11.1%, or 0%, 4.8%, 9.1% and 20.3 by study period (p=0.01). B/F recombinants, B/G recombinants and subtype F emerged in the last period. We also noticed an increase in the number of immigrant patients (p=0.052). The proportion of men-who-have-sex-with-men (MSM) among patients with acute/recent HIV-1 infection increased over the time (p=0.04). The overall prevalence of TDR in patients with acute/recent HIV-1 infection in Barcelona was 9%, and it has stayed relatively stable in recent years. Non-B subtypes and immigrants proportions progressively increased.

The Role of elF4E Activity in Breast Cancer

DTIC Science & Technology

2011-08-01

protein; ORF, open reading frame; qPCR, quantitative PCR; RACE, rapid amplification of cDNA ends; RT, reverse transcriptase ; uORF, upstream ORF; UTR...Reactions were also performed using template lacking RT ( reverse transcriptase ): products were either undetectable or greatly reduced (>30000-fold less...that a 5’UTR expressed from the human AXIN2 gene contains a sixty nucleotide sequence that is predicted to form a stable stem-loop structure6. This

A Pragmatic Approach to HIV-1 Drug Resistance Determination in Resource-Limited Settings by Use of a Novel Genotyping Assay Targeting the Reverse Transcriptase-Encoding Region Only

PubMed Central

Bronze, Michelle; Wallis, Carole L.; Stuyver, Lieven; Steegen, Kim; Balinda, Sheila; Kityo, Cissy; Stevens, Wendy; Rinke de Wit, Tobias F.; Schuurman, Rob

2013-01-01

In resource-limited settings (RLS), reverse transcriptase (RT) inhibitors form the backbone of first-line treatment regimens. We have developed a simplified HIV-1 drug resistance genotyping assay targeting the region of RT harboring all major RT inhibitor resistance mutation positions, thus providing all relevant susceptibility data for first-line failures, coupled with minimal cost and labor. The assay comprises a one-step RT-PCR amplification reaction, followed by sequencing using one forward and one reverse primer, generating double-stranded coverage of RT amino acids (aa) 41 to 238. The assay was optimized for all major HIV-1 group M subtypes in plasma and dried blood spot (DBS) samples using a panel of reference viruses for HIV-1 subtypes A to D, F to H, and circulating recombinant form 01_AE (CRF01_AE) and applied to 212 clinical plasma samples and 25 DBS samples from HIV-1-infected individuals from Africa and Europe. The assay was subsequently transferred to Uganda and applied locally on clinical plasma samples. All major HIV-1 subtypes could be detected with an analytical sensitivity of 5.00E+3 RNA copies/ml for plasma and DBS. Application of the assay on 212 clinical samples from African subjects comprising subtypes A to D, F to H (rare), CRF01_AE, and CRF02_AG at a viral load (VL) range of 6.71E+2 to 1.00E+7 (median, 1.48E+5) RNA copies/ml was 94.8% (n = 201) successful. Application on clinical samples in Uganda demonstrated a comparable success rate. Genotyping of clinical DBS samples, all subtype C with a VL range of 1.02E+3 to 4.49E+5 (median, 1.42E+4) RNA copies/ml, was 84.0% successful. The described assay greatly reduces hands-on time and the costs required for genotyping and is ideal for use in RLS, as demonstrated in a reference laboratory in Uganda and its successful application on DBS samples. PMID:23536405

Cardiac effects of 3-iodothyronamine: a new aminergic system modulating cardiac function.

PubMed

Chiellini, Grazia; Frascarelli, Sabina; Ghelardoni, Sandra; Carnicelli, Vittoria; Tobias, Sandra C; DeBarber, Andrea; Brogioni, Simona; Ronca-Testoni, Simonetta; Cerbai, Elisabetta; Grandy, David K; Scanlan, Thomas S; Zucchi, Riccardo

2007-05-01

3-Iodothyronamine T1AM is a novel endogenous thyroid hormone derivative that activates the G protein-coupled receptor known as trace anime-associated receptor 1 (TAAR1). In the isolated working rat heart and in rat cardiomyocytes, T1AM produced a reversible, dose-dependent negative inotropic effect (e.g., 27+/-5, 51+/-3, and 65+/-2% decrease in cardiac output at 19, 25, and 38 microM concentration, respectively). An independent negative chronotropic effect was also observed. The hemodynamic effects of T1AM were remarkably increased in the presence of the tyrosine kinase inhibitor genistein, whereas they were attenuated in the presence of the tyrosine phosphatase inhibitor vanadate. No effect was produced by inhibitors of protein kinase A, protein kinase C, calcium-calmodulin kinase II, phosphatidylinositol-3-kinase, or MAP kinases. Tissue cAMP levels were unchanged. In rat ventricular tissue, Western blot experiments with antiphosphotyrosine antibodies showed reduced phosphorylation of microsomal and cytosolic proteins after perfusion with synthetic T1AM; reverse transcriptase-polymerase chain reaction experiments revealed the presence of transcripts for at least 5 TAAR subtypes; specific and saturable binding of [125I]T1AM was observed, with a dissociation constant in the low micromolar range (5 microM); and endogenous T1AM was detectable by tandem mass spectrometry. In conclusion, our findings provide evidence for the existence of a novel aminergic system modulating cardiac function.

A modified single-tube one-step product-enhanced reverse transcriptase (mSTOS-PERT) assay with heparin as DNA polymerase inhibitor for specific detection of RTase activity.

PubMed

Fan, Xiao-Yong; Lü, Guo-Zhen; Wu, Li-Na; Chen, Jing-Hua; Xu, Wen-Qing; Zhao, Chun-Nü; Guo, Sheng-Qi

2006-12-01

Current regulations and recommendations proposed for the production of vaccines in continuous cell lines of any origin demand that these be free of exogenous viruses, particularly retroviruses. Recently, the ultra-sensitive product-enhanced reverse transcriptase (PERT) assay can be used to detect minute of reverse transcriptase (RTase) in single retroviral particle and is 10(6) times more sensitive than the conventional RTase assays. However, coincidental with this increase in sensitivity is an increase in false-positive reactions derived from contaminating cellular DNA polymerases, which are known to have RTase-like activities. To develop a modified single-tube one-step PERT (mSTOS-PERT) assay with improvements on decreasing significantly the level of false-positive reactions, and to evaluate the mSTOS-PERT assay for sensitivity and specificity. Ampliwaxtrade mark was used to compartmentalize the reverse transcription (RT) and PCR step in the same micro-tube with more efficiency and reproducibility, while maintaining the high sensitivity. The DNA amplification products were separated by 2% agarose gel electrophoresis, and then analyzed by non-isotopic Southern blot hybridization. A wide variety of cell lines used in biologicals production were detected to validate the improved mSTOS-PERT assay. The detection limit for the mSTOS-PERT assay was at least 10(-9)U, when using AMV-RTase as a positive control. Furthermore, heparin involvement in the RT step can eliminate completely the false-positive PERT signals which are exhibited by cellular polymerases such as DNA-dependent DNA polymerase alpha, gamma released by cell death. Most mammalian cells (MRC-5, Vero, WISH, 2BS, RK-13, MDCK, etc.) are PERT-negative in cell supernatants. Some PERT-positive signals in cell lysates were found to be introduced by the cellular DNA polymerases and could be inhibited specifically by heparin. Chick cells derived from either chick embryo fibroblasts (CEF) or allantoic fluid from SPF embryonated eggs, murine hybridoma cell SP2/0, etc., contained authentic RTase activities, which could not be inactivated by heparin. The improved mSTOS-PERT assay described here may distinguish the genuine RTase activity from cellular polymerases with high sensitivity and specificity, and is rapid and easy to perform to screen for the possible contamination of minute retroviruses in the cell substrates used in vaccine production.

Cerebrospinal fluid abacavir concentrations in HIV-positive patients following once-daily administration.

PubMed

Calcagno, A; Pinnetti, C; De Nicolò, A; Scarvaglieri, E; Gisslen, M; Tempestilli, M; D'Avolio, A; Fedele, V; Di Perri, G; Antinori, A; Bonora, S

2018-06-01

Abacavir is a widely used nucleotide reverse transcriptase inhibitor, for which cerebrospinal fluid (CSF) exposure has been previously assessed in twice-daily recipients. We studied abacavir CSF concentrations in 61 and nine HIV-positive patients taking abacavir once daily and twice daily, respectively. Patients on once-daily abacavir had higher plasma and CSF concentrations (96 vs. 22 ng ml -1 , P = 0.038 and 123 vs. 49 ng ml -1 , P = 0.038) but similar CSF-to-plasma ratios (0.8 vs. 0.5, P = 0.500). CSF abacavir concentrations were adequate in patients receiving once-daily treatment. © 2018 The British Pharmacological Society.

ONIOM DFT/PM3 calculations on the interaction between dapivirine and HIV-1 reverse transcriptase, a theoretical study.

PubMed

Liang, Y H; Chen, F E

2007-08-01

Theoretical investigations of the interaction between dapivirine and the HIV-1 RT binding site have been performed by the ONIOM2 (B3LYP/6-31G (d,p): PM3) and B3LYP/6-31G (d,p) methods. The results derived from this study indicate that this inhibitor dapivirine forms two hydrogen bonds with Lys101 and exhibits strong π-π stacking or H…π interaction with Tyr181 and Tyr188. These interactions play a vital role in stabilizing the NNIBP/dapivirine complex. Additionally, the predicted binding energy of the BBF optimized structure for this complex system is -18.20 kcal/mol.

HIVprotI: an integrated web based platform for prediction and design of HIV proteins inhibitors.

PubMed

Qureshi, Abid; Rajput, Akanksha; Kaur, Gazaldeep; Kumar, Manoj

2018-03-09

A number of anti-retroviral drugs are being used for treating Human Immunodeficiency Virus (HIV) infection. Due to emergence of drug resistant strains, there is a constant quest to discover more effective anti-HIV compounds. In this endeavor, computational tools have proven useful in accelerating drug discovery. Although methods were published to design a class of compounds against a specific HIV protein, but an integrated web server for the same is lacking. Therefore, we have developed support vector machine based regression models using experimentally validated data from ChEMBL repository. Quantitative structure activity relationship based features were selected for predicting inhibition activity of a compound against HIV proteins namely protease (PR), reverse transcriptase (RT) and integrase (IN). The models presented a maximum Pearson correlation coefficient of 0.78, 0.76, 0.74 and 0.76, 0.68, 0.72 during tenfold cross-validation on IC 50 and percent inhibition datasets of PR, RT, IN respectively. These models performed equally well on the independent datasets. Chemical space mapping, applicability domain analyses and other statistical tests further support robustness of the predictive models. Currently, we have identified a number of chemical descriptors that are imperative in predicting the compound inhibition potential. HIVprotI platform ( http://bioinfo.imtech.res.in/manojk/hivproti ) would be useful in virtual screening of inhibitors as well as designing of new molecules against the important HIV proteins for therapeutics development.

Novel Codon Insert in HIV Type 1 Clade B Reverse Transcriptase Associated with Low-Level Viremia During Antiretroviral Therapy

PubMed Central

Gianella, Sara; Vazquez, Homero; Ignacio, Caroline; Zweig, Adam C.; Richman, Douglas D.; Smith, Davey M.

2014-01-01

Abstract We investigated the pol genotype in two phylogenetically and epidemiologically linked partners, who were both experiencing persistent low-level viremia during antiretroviral therapy. In one partner we identified a new residue insertion between codon 248 and 249 of the HIV-1 RNA reverse transcriptase (RT) coding region (HXB2 numbering). We then investigated the potential impact of identified mutations in RT and antiretroviral binding affinity using a novel computational approach. PMID:24020934

Synthesis of the highly selective p38 MAPK inhibitor UR-13756 for possible therapeutic use in Werner syndrome.

PubMed

Bagley, Mark C; Davis, Terence; Rokicki, Michal J; Widdowson, Caroline S; Kipling, David

2010-02-01

UR-13756 is a potent and selective p38 mitogen-activated protein kinase (MAPK) inhibitor, reported to have good bioavailability and pharmacokinetic properties and, thus, is of potential use in the treatment of accelerated aging in Werner syndrome. Irradiation of 2-chloroacrylonitrile and methylhydrazine in ethanol at 100 °C gives 1-methyl-3-aminopyrazole, which reacts with 4-fluorobenzaldehyde and a ketone, obtained by Claisen condensation of 4-picoline, in a Hantzsch-type 3-component hereocyclocondensation, to give the pyrazolopyridine UR-13756. UR-13756 shows p38 MAPK inhibitory activity in human telomerase reverse transcriptase-immortalized HCA2 dermal fibroblasts, with an IC(50) of 80 nm, as shown by ELISA, is 100% efficacious for up to 24 h at 1.0 μm and displays excellent kinase selectivity over the related stress-activated c-Jun kinases. In addition, UR-13756 is an effective p38 inhibitor at 1.0 μm in Werner syndrome cells, as shown by immunoblot. The convergent synthesis of UR-13756 is realized using microwave dielectric heating and provides a highly selective inhibitor that shows excellent selectivity for p38 MAPK over c-Jun N-terminal kinase.

Antiretroviral drug resistance and phylogenetic diversity of HIV-1 in Chile.

PubMed

Ríos, Maritza; Delgado, Elena; Pérez-Alvarez, Lucía; Fernández, Jorge; Gálvez, Paula; de Parga, Elena Vázquez; Yung, Verónica; Thomson, Michael M; Nájera, Rafael

2007-06-01

This study reports the analysis of human immunodeficiency virus type 1 (HIV-1) protease (PR) and reverse transcriptase (RT) coding sequences from 136 HIV-1-infected subjects from Chile, 66 (49%) of them under antiretroviral (ARV) treatment. The prevalence of mutations conferring high or intermediate resistance levels to ARVs was 77% among treated patients and 2.5% among drug-naïve subjects. The distribution of resistance prevalence in treated patients by drug class was 61% to nucleoside RT inhibitors, 84% to nonnucleoside RT inhibitors, and 46% to PR inhibitors. Phylogenetic analysis revealed that 115 (85%) subjects were infected with subtype B viruses, 1 with a subtype F1 virus, and 20 (15%) carried BF intersubtype recombinants. Most BF recombinants grouped into two clusters, one related to CRF12_BF, while the other could represent a new circulating recombinant form (CRF). In conclusion, this is the first report analysing the prevalence of ARV resistance which includes patients under HAART from Chile. Additionally, phylogenetic analysis of the PR-RT coding sequences reveals the presence of BF intersubtype recombinants. (c) 2007 Wiley-Liss, Inc.

High HIV-1 Diversity and Prevalence of Transmitted Drug Resistance Among Antiretroviral-Naive HIV-Infected Pregnant Women from Rio de Janeiro, Brazil.

PubMed

Delatorre, Edson; Silva-de-Jesus, Carlos; Couto-Fernandez, José Carlos; Pilotto, Jose H; Morgado, Mariza G

2017-01-01

Antiretroviral (ARV) resistance mutations in human immunodeficiency virus type 1 (HIV-1) infection may reduce the efficacy of prophylactic therapy to prevent mother-to-child transmission (PMTCT) and future treatment options. This study evaluated the diversity and the prevalence of transmitted drug resistance (TDR) in protease (PR) and reverse transcriptase (RT) regions of HIV-1 pol gene among 87 ARV-naive HIV-1-infected pregnant women from Rio de Janeiro, Brazil, between 2012 and 2015. The viral diversity comprised HIV-1 subtypes B (67.8%), F1 (17.2%), and C (4.6%); the circulating recombinant forms 12_BF (2.3%), 28/29_BF, 39_BF, 02_AG (1.1% each) and unique recombinants forms (4.5%). The overall prevalence of any TDR was 17.2%, of which 5.7% for nucleoside RT inhibitors, 5.7% for non-nucleoside RT inhibitors, and 8% for PR inhibitors. The TDR prevalence found in this population may affect the virological outcome of the standard PMTCT ARV-regimens, reinforcing the importance of continuous monitoring.

Resistance Analyses of Integrase Strand Transfer Inhibitors within Phase 3 Clinical Trials of Treatment-Naive Patients

PubMed Central

White, Kirsten L.; Raffi, Francois; Miller, Michael D.

2014-01-01

The integrase (IN) strand transfer inhibitors (INSTIs), raltegravir (RAL), elvitegravir (EVG) and dolutegravir (DTG), comprise the newest drug class approved for the treatment of HIV-1 infection, which joins the existing classes of reverse transcriptase, protease and binding/entry inhibitors. The efficacy of first-line regimens has attained remarkably high levels, reaching undetectable viral loads in 90% of patients by Week 48; however, there remain patients who require a change in regimen due to adverse events, virologic failure with emergent resistance or other issues of patient management. Large, randomized clinical trials conducted in antiretroviral treatment-naive individuals are required for drug approval in this population in the US, EU and other countries, with the primary endpoint for virologic success at Week 48. However, there are differences in the definition of virologic failure and the evaluation of drug resistance among the trials. This review focuses on the methodology and tabulation of resistance to INSTIs in phase 3 clinical trials of first-line regimens and discusses case studies of resistance. PMID:25054884

A full-coordinate model of the polymerase domain of HIV-1 reverse transcriptase and its interaction with a nucleic acid substrate

NASA Technical Reports Server (NTRS)

Setlik, R. F.; Meyer, D. J.; Shibata, M.; Roskwitalski, R.; Ornstein, R. L.; Rein, R.

1994-01-01

We present a full-coordinate model of residues 1-319 of the polymerase domain of HIV-I reverse transcriptase. This model was constructed from the x-ray crystallographic structure of Jacobo-Molina et al. (Jacobo-Molina et al., P.N.A.S. USA 90, 6320-6324 (1993)) which is currently available to the degree of C-coordinates. The backbone and side-chain atoms were constructed using the MAXSPROUT suite of programs (L. Holm and C. Sander, J. Mol. Biol. 218, 183-194 (1991)) and refined through molecular modeling. A seven base pair A-form dsDNA was positioned in the nucleic acid binding cleft to represent the template-primer complex. The orientation of the template-primer complex in the nucleic acid binding cleft was guided by the positions of phosphorus atoms in the crystal structure.

Valproic Acid Induces Telomerase Reverse Transcriptase Expression during Cortical Development.

PubMed

Kim, Ki Chan; Choi, Chang Soon; Gonzales, Edson Luck T; Mabunga, Darine Froy N; Lee, Sung Hoon; Jeon, Se Jin; Hwangbo, Ram; Hong, Minha; Ryu, Jong Hoon; Han, Seol-Heui; Bahn, Geon Ho; Shin, Chan Young

2017-10-01

The valproic acid (VPA)-induced animal model is one of the most widely utilized environmental risk factor models of autism. Autism spectrum disorder (ASD) remains an insurmountable challenge among neurodevelopmental disorders due to its heterogeneity, unresolved pathological pathways and lack of treatment. We previously reported that VPA-exposed rats and cultured rat primary neurons have increased Pax6 expression during post-midterm embryonic development which led to the sequential upregulation of glutamatergic neuronal markers. In this study, we provide experimental evidence that telomerase reverse transcriptase (TERT), a protein component of ribonucleoproteins complex of telomerase, is involved in the abnormal components caused by VPA in addition to Pax6 and its downstream signals. In embryonic rat brains and cultured rat primary neural progenitor cells (NPCs), VPA induced the increased expression of TERT as revealed by Western blot, RT-PCR, and immunostainings. The HDAC inhibitor property of VPA is responsible for the TERT upregulation. Chromatin immunoprecipitation revealed that VPA increased the histone acetylation but blocked the HDAC1 binding to both Pax6 and Tert genes. Interestingly, the VPA-induced TERT overexpression resulted to sequential upregulations of glutamatergic markers such as Ngn2 and NeuroD1, and inter-synaptic markers such as PSD-95, α-CaMKII, vGluT1 and synaptophysin. Transfection of Tert siRNA reversed the effects of VPA in cultured NPCs confirming the direct involvement of TERT in the expression of those markers. This study suggests the involvement of TERT in the VPA-induced autistic phenotypes and has important implications for the role of TERT as a modulator of balanced neuronal development and transmission in the brain.

[Mutations of resistance of HIV-1 in previously untreated patients at penitentiary centers of the Autonomous Community of Valencia, Spain. REPRICOVA study].

PubMed

García-Guerrero, Julio; Herrero, Agustín; Vera, Enrique; Almenara, José M; Araújo, Rosa; Saurí, Vicente V; Castellano, Juan C; Fernández-Clemente, Luis; Bedia, Miguel; Llorente, María I; González-Morán, Francisco

2002-03-02

Our purpose was to determine the prevalence of mutations of resistance to nucleoside inhibitors of reverse transcriptase (NIRT) and protease inhibitors (PI) in the HIV-1 genotype of naïve infected subjects in the prisons of the Autonomous Community of Valencia, Spain. Multicentric, descriptive, cross-sectional study of prevalence including a systematic stratified and randomised sampling by centres. Demographic, clinical, virological and immunological data were collected. The HIV gene of protease and transcriptase was studied in peripheral blood plasma samples by means of double PCR amplification and subsequent automatic sequence. Reference: wild strain HXB2. Plasma was obtained from 133 individuals (119 men and 14 women). 117 samples were selected and the rest did not have enough copies for transcription. With regard to NIRT, 7 samples (5.2% of total) showed some mutation of resistance: M41L, D67N, L210W and K219Q, all them secondary to and associated with resistance to zidovudine, abacavir as well as group B multinucleoside-resistance. With regard to PI, only one sample showed a primary mutation, M46I, which was associated with resistance to indinavir. Moreover, a further 41 samples were found to express some secondary mutation. In our series, there was a low number of primary mutations of resistance. These results allow us to exclude the systematic use of resistance tests before an initiation antiretroviral therapy.

Zidovudine ameliorates pathology in the mouse model of Duchenne muscular dystrophy via P2RX7 purinoceptor antagonism.

PubMed

Al-Khalidi, Rasha; Panicucci, Chiara; Cox, Paul; Chira, Natalia; Róg, Justyna; Young, Christopher N J; McGeehan, Rhiannon E; Ambati, Kameshwari; Ambati, Jayakrishna; Zabłocki, Krzysztof; Gazzerro, Elisabetta; Arkle, Stephen; Bruno, Claudio; Górecki, Dariusz C

2018-04-11

Duchenne muscular dystrophy (DMD) is the most common inherited muscle disorder that causes severe disability and death of young men. This disease is characterized by progressive muscle degeneration aggravated by sterile inflammation and is also associated with cognitive impairment and low bone density. Given that no current treatment can improve the long-term outcome, approaches with a strong translational potential are urgently needed. Duchenne muscular dystrophy (DMD) alters P2RX7 signaling in both muscle and inflammatory cells and inhibition of this receptor resulted in a significant attenuation of muscle and non-muscle symptoms in DMD mdx mouse model. As P2RX7 is an attractive target in a range of human diseases, specific antagonists have been developed. Yet, these will require lengthy safety testing in the pediatric population of Duchenne muscular dystrophy (DMD) patients. In contrast, Nucleoside Reverse Transcriptase Inhibitors (NRTIs) can act as P2RX7 antagonists and are drugs with an established safety record, including in children. We demonstrate here that AZT (Zidovudine) inhibits P2RX7 functions acting via the same allosteric site as other antagonists. Moreover, short-term AZT treatment at the peak of disease in DMD mdx mice attenuated the phenotype without any detectable side effects. Recovery was evident in the key parameters such as reduced sarcolemma permeability confirmed by lower serum creatine kinase levels and IgG influx into myofibres, decreased inflammatory cell numbers and inflammation markers in leg and heart muscles of treated mice. Moreover, this short-term therapy had some positive impact on muscle strength in vivo and no detrimental effect on mitochondria, which is the main side-effect of Nucleoside Reverse Transcriptase Inhibitors (NRTIs). Given these results, we postulate that AZT could be quickly re-purposed for the treatment of this highly debilitating and lethal disease. This approach is not constrained by causative DMD mutations and may be effective in alleviating both muscle and non-muscle abnormalities.

Basic science and pathogenesis of ageing with HIV: potential mechanisms and biomarkers.

PubMed

Lagathu, Claire; Cossarizza, Andrea; Béréziat, Véronique; Nasi, Milena; Capeau, Jacqueline; Pinti, Marcello

2017-06-01

: The increased prevalence of age-related comorbidities and mortality is worrisome in ageing HIV-infected patients. Here, we aim to analyse the different ageing mechanisms with regard to HIV infection. Ageing results from the time-dependent accumulation of random cellular damage. Epigenetic modifications and mitochondrial DNA haplogroups modulate ageing. In antiretroviral treatment-controlled patients, epigenetic clock appears to be advanced, and some haplogroups are associated with HIV infection severity. Telomere shortening is enhanced in HIV-infected patients because of HIV and some nucleoside analogue reverse transcriptase inhibitors. Mitochondria-related oxidative stress and mitochondrial DNA mutations are increased during ageing and also by some nucleoside analogue reverse transcriptase inhibitors. Overall, increased inflammation or 'inflammageing' is a major driver of ageing and could result from cell senescence with secreted proinflammatory mediators, altered gut microbiota, and coinfections. In HIV-infected patients, the level of inflammation and innate immunity activation is enhanced and related to most comorbidities and to mortality. This status could result, in addition to age, from the virus itself or viral protein released from reservoirs, from HIV-enhanced gut permeability and dysbiosis, from antiretroviral treatment, from frequent cytomegalovirus and hepatitis C virus coinfections, and also from personal and environmental factors, as central fat accumulation or smoking. Adaptive immune activation and immunosenescence are associated with comorbidities and mortality in the general population but are less predictive in HIV-infected patients. Biomarkers to evaluate ageing in HIV-infected patients are required. Numerous systemic or cellular inflammatory, immune activation, oxidative stress, or senescence markers can be tested in serum or peripheral blood mononuclear cells. The novel European Study to Establish Biomarkers of Human Ageing MARK-AGE algorithm, evaluating the biological age, is currently assessed in HIV-infected patients and reveals an advanced biological age. Some enhanced inflammatory or innate immune activation markers are interesting but still not validated for the patient's follow-up. To be able to assess patients' biological age is an important objective to improve their healthspan.

Efficacy and safety of rilpivirine in treatment-naive, HIV-1-infected patients with hepatitis B virus/hepatitis C virus coinfection enrolled in the Phase III randomized, double-blind ECHO and THRIVE trials.

PubMed

Nelson, Mark; Amaya, Gerardo; Clumeck, Nathan; Arns da Cunha, Clovis; Jayaweera, Dushyantha; Junod, Patrice; Li, Taisheng; Tebas, Pablo; Stevens, Marita; Buelens, Annemie; Vanveggel, Simon; Boven, Katia

2012-08-01

The efficacy and hepatic safety of the non-nucleoside reverse transcriptase inhibitors rilpivirine (TMC278) and efavirenz were compared in treatment-naive, HIV-infected adults with concurrent hepatitis B virus (HBV) and/or hepatitis C virus (HCV) infection in the pooled week 48 analysis of the Phase III, double-blind, randomized ECHO (NCT00540449) and THRIVE (NCT00543725) trials. Patients received 25 mg of rilpivirine once daily or 600 mg of efavirenz once daily, plus two nucleoside/nucleotide reverse transcriptase inhibitors. At screening, patients had alanine aminotransferase/aspartate aminotransferase levels ≤5× the upper limit of normal. HBV and HCV status was determined at baseline by HBV surface antigen, HCV antibody and HCV RNA testing. HBV/HCV coinfection status was known for 670 patients in the rilpivirine group and 665 in the efavirenz group. At baseline, 49 rilpivirine and 63 efavirenz patients [112/1335 (8.4%)] were coinfected with either HBV [55/1357 (4.1%)] or HCV [57/1333 (4.3%)]. The safety analysis included all available data, including beyond week 48. Eight patients seroconverted during the study (rilpivirine: five; efavirenz: three). A higher proportion of patients achieved viral load <50 copies/mL (intent to treat, time to loss of virological response) in the subgroup without HBV/HCV coinfection (rilpivirine: 85.0%; efavirenz: 82.6%) than in the coinfected subgroup (rilpivirine: 73.5%; efavirenz: 79.4%) (rilpivirine, P = 0.04 and efavirenz, P = 0.49, Fisher's exact test). The incidence of hepatic adverse events (AEs) was low in both groups in the overall population (rilpivirine: 5.5% versus efavirenz: 6.6%) and was higher in HBV/HCV-coinfected patients than in those not coinfected (26.7% versus 4.1%, respectively). Hepatic AEs were more common and response rates lower in HBV/HCV-coinfected patients treated with rilpivirine or efavirenz than in those who were not coinfected.

HIV-1 transmission networks across Cyprus (2010-2012).

PubMed

Kostrikis, Leondios G; Hezka, Johana; Stylianou, Dora C; Kostaki, Evangelia; Andreou, Maria; Kousiappa, Ioanna; Paraskevis, Dimitrios; Demetriades, Ioannis

2018-01-01

A molecular epidemiology study of HIV-1 infection was conducted in one hundred diagnosed and untreated HIV-1-infected patients in Cyprus between 2010 and 2012, representing 65.4% of all the reported HIV-1 infections in Cyprus in this three-year period, using a previously defined enrolment strategy. Eighty-two patients were newly diagnosed (genotypic drug resistance testing within six months from diagnosis), and eighteen patients were HIV-1 diagnosed for a longer period or the diagnosis date was unknown. Phylogenetic trees of the pol sequences obtained in this study with reference sequences indicated that subtypes B and A1 were the most common subtypes present and accounted for 41.0 and 19.0% respectively, followed by subtype C (7.0%), F1 (8.0%), CRF02_AG (4.0%), A2 (2.0%), other circulating recombinant forms (CRFs) (7.0%) and unknown recombinant forms (URFs) (12%). Most of the newly-diagnosed study subjects were Cypriots (63%), males (78%) with median age 39 (Interquartile Range, IQR 33-48) reporting having sex with other men (MSM) (51%). A high rate of clustered transmission of subtype B drug-sensitive strains to reverse transcriptase and protease inhibitors was observed among MSM, twenty-eight out of forty-one MSM study subjects (68.0%) infected were implicated in five transmission clusters, two of which are sub-subtype A1 and three of which are subtype B strains. The two largest MSM subtype B clusters included nine and eight Cypriot men, respectively, living in all major cities in Cyprus. There were only three newly diagnosed patients with transmitted drug resistant HIV-1 strains, one study subject from the United Kingdom infected with subtype B strain and one from Romania with sub-subtype A2 strain, both with PI drug resistance mutation M46L and one from Greece with sub-subtype A1 with non-nucleoside reverse transcriptase inhibitors (NNRTI) drug resistance mutation K103N.

Pharmacokinetics and Pharmacodynamics of Atazanavir in HIV-1-Infected Children Treated With Atazanavir Powder and Ritonavir: Combined Analysis of the PRINCE-1 and -2 Studies.

PubMed

Sevinsky, Heather; Zaru, Luna; Wang, Reena; Xu, Xiaohui; Pikora, Cheryl; Correll, Todd A; Eley, Timothy

2018-06-01

Two clinical studies (PRINCE-1 and -2) in HIV-1-infected children assessed the safety, efficacy and pharmacokinetics of dual nucleos(t)ide reverse transcriptase inhibitor background therapy plus once-daily atazanavir (ATV) powder formulation boosted with ritonavir (ATV + RTV). Here, we present a combined analysis of ATV pharmacokinetics and pharmacodynamics across these studies. Intensive 24-hour pharmacokinetic profiles at steady state compared ATV exposures (area under the concentration-time curve in one dosing interval) in 5 ATV + RTV baseline weight-band dosing categories, with historic data in adults receiving ATV + RTV 300/100 mg capsules. Repeated ATV Ctrough measurements over 48 weeks explored relationships between ATV composite Ctrough quartiles (CCQs) with virologic efficacy and key safety parameters. Of 146 children included in this combined analysis, 49.3% were male, 56.8% were Black/African American and 62.3% were antiretroviral experienced. Proportions with HIV-1 RNA <50 copies/mL at week 48 were 13/32, 24/32, 19/32 and 13/28 in the lowest through highest ATV CCQs, respectively. Mean changes from baseline in total bilirubin at week 48 were +0.3, +0.5, +0.6 and +1.0 mg/dL in the lowest through highest ATV CCQs, respectively. Corresponding proportions with adverse events of hyperbilirubinemia by week 48 were 1/36, 4/36, 5/36 and 13/35, respectively. Changes from baseline in total amylase or electrocardiogram parameters and adverse events of diarrhea did not vary by ATV CCQs. Weight-band dosing of ATV + RTV plus optimized dual nucleos(t)ide reverse transcriptase inhibitors in young HIV-1-infected children achieved similar ATV exposure to that in adults; no unexpected safety findings occurred, and with the exception of lower virologic suppression in the lowest ATV CCQ, there was no apparent trend in virologic suppression across ATV CCQs.

Limited clinical benefit of minority K103N and Y181C-variant detection in addition to routine genotypic resistance testing in antiretroviral therapy-naive patients.

PubMed

Metzner, Karin J; Scherrer, Alexandra U; von Wyl, Viktor; Böni, Jürg; Yerly, Sabine; Klimkait, Thomas; Aubert, Vincent; Furrer, Hansjakob; Hirsch, Hans H; Vernazza, Pietro L; Cavassini, Matthias; Calmy, Alexandra; Bernasconi, Enos; Weber, Rainer; Günthard, Huldrych F

2014-09-24

The presence of minority nonnucleoside reverse transcriptase inhibitor (NNRTI)-resistant HIV-1 variants prior to antiretroviral therapy (ART) has been linked to virologic failure in treatment-naive patients. We performed a large retrospective study to determine the number of treatment failures that could have been prevented by implementing minority drug-resistant HIV-1 variant analyses in ART-naïve patients in whom no NNRTI resistance mutations were detected by routine resistance testing. Of 1608 patients in the Swiss HIV Cohort Study, who have initiated first-line ART with two nucleoside reverse transcriptase inhibitors (NRTIs) and one NNRTI before July 2008, 519 patients were eligible by means of HIV-1 subtype, viral load and sample availability. Key NNRTI drug resistance mutations K103N and Y181C were measured by allele-specific PCR in 208 of 519 randomly chosen patients. Minority K103N and Y181C drug resistance mutations were detected in five out of 190 (2.6%) and 10 out of 201 (5%) patients, respectively. Focusing on 183 patients for whom virologic success or failure could be examined, virologic failure occurred in seven out of 183 (3.8%) patients; minority K103N and/or Y181C variants were present prior to ART initiation in only two of those patients. The NNRTI-containing, first-line ART was effective in 10 patients with preexisting minority NNRTI-resistant HIV-1 variant. As revealed in settings of case-control studies, minority NNRTI-resistant HIV-1 variants can have an impact on ART. However, the implementation of minority NNRTI-resistant HIV-1 variant analysis in addition to genotypic resistance testing (GRT) cannot be recommended in routine clinical settings. Additional associated risk factors need to be discovered.

Zidovudine, abacavir and lamivudine increase the radiosensitivity of human esophageal squamous cancer cell lines.

PubMed

Chen, Xuan; Wang, Cong; Guan, Shanghui; Liu, Yuan; Han, Lihui; Cheng, Yufeng

2016-07-01

Telomerase is a type of reverse transcriptase that is overexpressed in almost all human tumor cells, but not in normal tissues, which provides an opportunity for radiosensitization targeting telomerase. Zidovudine, abacavir and lamivudine are reverse transcriptase inhibitors that have been applied in clinical practice for several years. We sought to explore the radiosensitization effect of these three drugs on human esophageal cancer cell lines. Eca109 and Eca9706 cells were treated with zidovudine, abacavir and lamivudine for 48 h before irradiation was administered. Samples were collected 1 h after irradiation. Clonal efficiency assay was used to evaluate the effect of the combination of these drugs with radiation doses of 2, 4, 6 and 8 Gy. DNA damage was measured by comet assay. Telomerase activity (TA) and relative telomere length (TL) were detected and evaluated by real-time PCR. Apoptosis rates were assessed by flow cytometric analysis. The results showed that all the drugs tested sensitized the esophageal squamous cell carcinoma (ESCC) cell lines to radiation through an increase in radiation-induced DNA damage and cell apoptosis, deregulation of TA and decreasing the shortened TL caused by radiation. Each of the drugs investigated (zidovudine, abacavir and lamivudine) could be used for sensitizing human esophageal cancer cell lines to radiation. Consequently, the present study supports the potential of these three drugs as therapeutic agents for the radiosensitization of esophageal squamous cell cancer.

In-vitro Cell Culture and Real-time Reverse Transcriptase PCR-based Assays to Detect Infective Toxoplas gondii Oocysts

EPA Science Inventory

Toxoplasma gondii is an obligate intracellular, apicomplexan parasite that infects humans. It is ubiquitous in nature and seroprevalence in the United States and in Europe ranges from 25->70%. Although typically associated with causing foodborne outbreaks, recent studies in Canad...

Current lead natural products for the chemotherapy of human immunodeficiency virus (HIV) infection.

PubMed

De Clercq, E

2000-09-01

A large variety of natural products have been described as anti-HIV agents, and for a portion thereof the target of interaction has been identified. Cyanovirin-N, a 11-kDa protein from Cyanobacterium (blue-green alga) irreversibly inactivates HIV and also aborts cell-to-cell fusion and transmission of HIV, due to its high-affinity interaction with gp120. Various sulfated polysaccharides extracted from seaweeds (i.e., Nothogenia fastigiata, Aghardhiella tenera) inhibit the virus adsorption process. Ingenol derivatives may inhibit virus adsorption at least in part through down-regulation of CD4 molecules on the host cells. Inhibition of virus adsorption by flavanoids such as (-)epicatechin and its 3-O-gallate has been attributed to an irreversible interaction with gp120 (although these compounds are also known as reverse transcriptase inhibitors). For the triterpene glycyrrhizin (extracted from the licorice root Glycyrrhiza radix) the mode of anti-HIV action may at least in part be attributed to interference with virus-cell binding. The mannose-specific plant lectins from Galanthus, Hippeastrum, Narcissus, Epipac tis helleborine, and Listera ovata, and the N-acetylgl ucosamine-specific lectin from Urtica dioica would primarily be targeted at the virus-cell fusion process. Various other natural products seem to qualify as HIV-cell fusion inhibitors: the siamycins [siamycin I (BMY-29304), siamycin II (RP 71955, BMY 29303), and NP-06 (FR901724)] which are tricyclic 21-amino-acid peptides isolated from Streptomyces spp that differ from one another only at position 4 or 17 (valine or isoleucine in each case); the betulinic acid derivative RPR 103611, and the peptides tachyplesin and polyphemusin which are highly abundant in hemocyte debris of the horseshoe crabs Tachypleus tridentatus and Limulus polyphemus, i.e., the 18-amino-acid peptide T22 from which T134 has been derived. Both T22 and T134 have been shown to block T-tropic X4 HIV-1 strains through a specific antagonism with the HIV corecept or CXCR4. A number of natural products have been reported to interact with the reverse transcriptase, i.e., baicalin, avarol, avarone, psychotrine, phloroglucinol derivatives, and, in particular, calanolides (from the tropical rainforest tree, Calophyllum lanigerum) and inophyllums (from the Malaysian tree, Calophyllum inophyllum). The natural marine substance illimaquinone would be targeted at the RNase H function of the reverse transcriptase. Curcumin (diferuloylmethane, from turmeric, the roots/rhizomes of Curcuma spp), dicaffeoylquinic and dicaffeoylt artaric acids, L-chicoric acid, and a number of fungal metabolites (equisetin, phomasetin, oteromycin, and integric acid) have all been proposed as HIV-1 integrase inhibitors. Yet, we have recently shown that L-c hicoric acid owes its anti-HIV activity to a specific interaction with the viral envelope gp120 rather than integrase. A number of compounds would be able to inhibit HIV-1 gene expression at the transcription level: the flavonoid chrysin (through inhibition of casein kinase II, the antibacter ial peptides melittin (from bee venom) and cecropin, and EM2487, a novel substance produced by Streptomyces. (ABSTRACT TRUNCATED)

Identification of a Pyridoxine-Derived Small-Molecule Inhibitor Targeting Dengue Virus RNA-Dependent RNA Polymerase.

PubMed

Xu, Hong-Tao; Colby-Germinario, Susan P; Hassounah, Said; Quashie, Peter K; Han, Yingshan; Oliveira, Maureen; Stranix, Brent R; Wainberg, Mark A

2016-01-01

The viral RNA-dependent RNA polymerase (RdRp) activity of the dengue virus (DENV) NS5 protein is an attractive target for drug design. Here, we report the identification of a novel class of inhibitor (i.e., an active-site metal ion chelator) that acts against DENV RdRp activity. DENV RdRp utilizes a two-metal-ion mechanism of catalysis; therefore, we constructed a small library of compounds, through mechanism-based drug design, aimed at chelating divalent metal ions in the catalytic site of DENV RdRp. We now describe a pyridoxine-derived small-molecule inhibitor that targets DENV RdRp and show that 5-benzenesulfonylmethyl-3-hydroxy-4-hydroxymethyl-pyridine-2-carboxylic acid hydroxyamide (termed DMB220) inhibited the RdRp activity of DENV serotypes 1 to 4 at low micromolar 50% inhibitory concentrations (IC50s of 5 to 6.7 μM) in an enzymatic assay. The antiviral activity of DMB220 against DENV infection was also verified in a cell-based assay and showed a 50% effective concentration (EC50) of <3 μM. Enzyme assays proved that DMB220 was competitive with nucleotide incorporation. DMB220 did not inhibit the enzymatic activity of recombinant HIV-1 reverse transcriptase and showed only weak inhibition of HIV-1 integrase strand transfer activity, indicating high specificity for DENV RdRp. S600T substitution in the DENV RdRp, which was previously shown to confer resistance to nucleoside analogue inhibitors (NI), conferred 3-fold hypersusceptibility to DMB220, and enzymatic analyses showed that this hypersusceptibility may arise from the decreased binding/incorporation efficiency of the natural NTP substrate without significantly impacting inhibitor binding. Thus, metal ion chelation at the active site of DENV RdRp represents a viable anti-DENV strategy, and DMB220 is the first of a new class of DENV inhibitor. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Lack of detection of a putative retrovirus associated with haemic neoplasia in the soft shell clam Mya arenaria.

PubMed

AboElkhair, M; Iwamoto, T; Clark, K F; McKenna, P; Siah, A; Greenwood, S J; Berthe, F C J; Casey, J W; Cepica, A

2012-01-01

Haemic neoplasia (HN) is a leukemia-like disease that affects at least 20 species of marine bivalves including soft shell clam, Mya arenaria. Since the disease was discovered in 1969, the etiology remains unknown. A retroviral etiology has been suggested based on the detection of reverse transcriptase activity and electron microscopic observation of retroviral-like particles using negative staining. To date, however no virus isolate and no retroviral sequence from HN has been obtained. Moreover, transmission of the disease by cell-free filtrate from affected clams has not been reproduced. In the current study, we reinvestigated the association of HN with a putative retrovirus. Sucrose gradient centrifugation followed by assessment of reverse transcriptase activity, electrophoretic analysis of protein and RNA, and electron microscopic examinations of fractions corresponding to retroviral density were employed. Detection of retroviral pol sequences using degenerate RT-PCR approaches was also attempted. Our results showed visible bands at the expected density of retrovirus in HN-positive and HN-negative clam tissues and both with reverse transcriptase activity. Electron microscopy, RNA analysis, protein analysis, and PCR systems targeting the pol gene of retroviruses did not however provide clear evidence supporting presence of a retrovirus. We point out that the retrovirus etiology of HN of Mya arenaria proposed some 25 years ago should be reconsidered in the absence of a virus isolate or virus sequences. Copyright © 2011 Elsevier Inc. All rights reserved.

HIV drug resistance surveillance for prioritizing treatment in resource-limited settings

PubMed Central

Walensky, Rochelle P.; Weinstein, Milton C.; Yazdanpanah, Yazdan; Losina, Elena; Mercincavage, Lauren M.; Touré, Siaka; Divi, Nomita; Anglaret, Xavier; Goldie, Sue J.; Freedberg, Kenneth A.

2008-01-01

Background Sentinel testing programs for HIV drug resistance in resource-limited settings can inform policy on antiretroviral therapy (ART) and drug sequencing. Objective To examine the value of resistance surveillance in influencing recommendations toward effective and cost-effective sequencing of ART regimens. Methods A state-transition model of HIV infection was adapted to simulate clinical care in Côte d’Ivoire and evaluate the incremental cost-effectiveness of (1) no ART; (2) ART beginning with a non-nucleoside reverse transcriptase inhibitor (NNRTI)-based regimen followed by a boosted protease inhibitor (PI)-based regimen; and (3) ART beginning with a boosted PI-based regimen followed by an NNRTI-based regimen. Results At a 5% prevalence of NNRTI resistance, a strategy that started with a PI-based regimen had a smaller health benefit and higher cost-effectiveness ratio than a strategy that started with an NNRTI-based regimen (cost-effectiveness ratio $910/year of life saved). Results consistently favored initiation with an NNRTI-based regimen, regardless of the population prevalence of NNRTI resistance (up to 76%) and the efficacy of an NNRTI-based regimen in the setting of resistance. The most influential parameters on the cost-effectiveness of sequencing strategies were boosted PI-based regimen costs and the efficacy of this regimen when used as second-line therapy. Conclusions Drug costs and treatment efficacies, but not NNRTI resistance levels, were most influential in determining optimal HIV drug sequencing in Côte d’Ivoire. Results of surveillance for NNRTI resistance should not be used as a major guide to treatment policy in resource-limited settings. PMID:17457091

Peripheral neuropathy in HIV: an analysis of evidence-based approaches.

PubMed

Nicholas, Patrice K; Corless, Inge B; Evans, Linda A

2014-01-01

Peripheral neuropathy is a common and vexing symptom for people living with HIV infection (PLWH). Neuropathy occurs in several different syndromes and is identified in the literature as distal sensory polyneuropathy or distal sensory peripheral neuropathy. More recently, the HIV literature has focused on the syndrome as painful HIV-associated sensory neuropathy, addressing the symptom rather than the underlying pathophysiology. Assessment of neuropathy in PLWH is critical and must be incorporated into nursing practice for each visit. Neuropathy has been attributed to the direct effects of HIV, exposure to antiretroviral medications (particularly the nucleoside reverse transcriptase inhibitors), advanced immune suppression, and comorbid tuberculosis infection and exposure to antituberculosis medications. Evidence supports the importance of addressing neuropathy in PLWH with pharmacologic treatment regimens and complementary/alternative approaches. This paper examines the pathophysiology, evidence, and approaches to managing peripheral neuropathy. A case study has been included to illustrate a patient's experience with neuropathy symptoms. Copyright © 2014 Association of Nurses in AIDS Care. Published by Elsevier Inc. All rights reserved.

HuR interacts with human immunodeficiency virus type 1 reverse transcriptase, and modulates reverse transcription in infected cells

PubMed Central

Lemay, Julie; Maidou-Peindara, Priscilla; Bader, Thomas; Ennifar, Eric; Rain, Jean-Christophe; Benarous, Richard; Liu, Lang Xia

2008-01-01

Reverse transcription of the genetic material of human immunodeficiency virus type 1 (HIV-1) is a critical step in the replication cycle of this virus. This process, catalyzed by reverse transcriptase (RT), is well characterized at the biochemical level. However, in infected cells, reverse transcription occurs in a multiprotein complex – the reverse transcription complex (RTC) – consisting of viral genomic RNA associated with viral proteins (including RT) and, presumably, as yet uncharacterized cellular proteins. Very little is known about the cellular proteins interacting with the RTC, and with reverse transcriptase in particular. We report here that HIV-1 reverse transcription is affected by the levels of a nucleocytoplasmic shuttling protein – the RNA-binding protein HuR. A direct protein-protein interaction between RT and HuR was observed in a yeast two-hybrid screen and confirmed in vitro by homogenous time-resolved fluorescence (HTRF). We mapped the domain interacting with HuR to the RNAse H domain of RT, and the binding domain for RT to the C-terminus of HuR, partially overlapping the third RRM RNA-binding domain of HuR. HuR silencing with specific siRNAs greatly impaired early and late steps of reverse transcription, significantly inhibiting HIV-1 infection. Moreover, by mutagenesis and immunoprecipitation studies, we could not detect the binding of HuR to the viral RNA. These results suggest that HuR may be involved in and may modulate the reverse transcription reaction of HIV-1, by an as yet unknown mechanism involving a protein-protein interaction with HIV-1 RT. PMID:18544151

Selection of HIV resistance associated with antiretroviral therapy initiated due to pregnancy and suspended postpartum.

PubMed

Ellis, Giovanina M; Huang, Sharon; Hitti, Jane; Frenkel, Lisa M

2011-11-01

Compare the risk of HIV drug resistance in women stopping suppressive nelfinavir (NFV)-based or Nevirapine (NVP)-based antiretroviral therapy (ART) after pregnancy. Specimens collected after stopping ART were tested for drug resistance by an oligonucleotide ligation assay and consensus sequencing. When postpartum drug resistance was detected, specimens obtained at study entry and during ART were evaluated. Sixteen of 38 women with ART-induced suppression of viral replication suspended ART postpartum. Resistance mutations were detected in 75% who stopped NFV-ART and in 50% who stopped NVP-ART. M184V, associated with Lamivudine resistance, was more frequent among those randomized to NFV-ART compared with NVP-ART (6 of 8 versus 1 of 8; P = 0.04), and nonnucleoside reverse transcriptase inhibitor resistance was detected in 4 of 8 stopping NVP-ART. HIV drug resistance was frequently observed among women who stopped suppressive NVP-ART or NFV-ART postpartum. This suggests that NFV-ART may have suboptimal potency, that staggering discontinuation of NVP-ART may be warranted, and/or ART adherence may be lax in women who choose to stop ART postpartum.

Reverse Transcriptase Activity in Mature Spermatozoa of Mouse

PubMed Central

Giordano, Roberto; Magnano, Anna Rosa; Zaccagnini, Germana; Pittoggi, Carmine; Moscufo, Nicola; Lorenzini, Rodolfo; Spadafora, Corrado

2000-01-01

We show here that a reverse transcriptase (RT) activity is present in murine epididymal spermatozoa. Sperm cells incubated with human poliovirus RNA can take up exogenous RNA molecules and internalize them in nuclei. Direct PCR amplification of DNA extracted from RNA-incubated spermatozoa indicate that poliovirus RNA is reverse-transcribed in cDNA fragments. PCR analysis of two-cell embryos shows that poliovirus RNA-challenged spermatozoa transfer retrotranscribed cDNA molecules into eggs during in vitro fertilization. Finally, RT molecules can be visualized on sperm nuclear scaffolds by immunogold electron microscopy. These results, therefore, reveal a novel metabolic function in spermatozoa, which may play a role during early embryonic development. PMID:10725323

Gamma-irradiated bacterial preparation having anti-tumor activity

DOEpatents

Vass, Arpad A.; Tyndall, Richard L.; Terzaghi-Howe, Peggy

1999-01-01

A bacterial preparation from Pseudomonas species isolated #15 ATCC 55638 that has been exposed to gamma radiation exhibits cytotoxicity that is specific for neoplastic carcinoma cells. A method for obtaining a bacterial preparation having antitumor activity consists of suspending a bacterial isolate in media and exposing the suspension to gamma radiation. A bacterial preparation of an aged culture of an amoeba-associated bacteria exhibits anti-reverse transcriptase activity. A method for obtaining a bacterial preparation having anti-reverse transcriptase activity from an amoeba-associated bacterial isolate grown to stationary phase is disclosed.

Global asymptotic stability for HIV-1 dynamics with two distributed delays.

PubMed

Wang, Jinliang; Huang, Gang; Takeuchi, Yasuhiro

2012-09-01

Based on the drugs treatment to control HIV-1 infection and viral replication, we express the intracellular eclipse phase of virions in host cell as distributed delays because of pharmacological actions. In present paper, we investigate a class of HIV-1 infection dynamical model with two distributed delays. One of them describes the period between the time that HIV virion enters (infects) target cell and the time that the infected cell starts to produce new viral particles. The other describes the time for the virion maturation process. They are both allowed to tend to be infinite because of drugs resistent strains. By the Lyapunov direct method of and utilizing the technology of constructing Lyapunov functionals, we identify the basic reproduction number R(0) as a threshold quantity for the stability of equilibria. More precisely, if R(0) ≤ 1, the infection-free equilibrium is globally asymptotically stable; on the contrary, if R(0) > 1, then an infected equilibrium appears which is globally asymptotically stable. The dynamical results indicate that time delays have effect on the global stability of two equilibria through threshold value R(0), which is a decreasing function of delays. The biological meanings imply that any drugs that can prolong the time of viral reproduction through slowing down the reverse transcription of HIV in host and virus maturation process may also help control the HIV-1 infection and virus loads. Another way to increase the efficacy of the protease inhibitor and the reverse transcriptase inhibitor (i.e. increasing n(p) and n(rt)) is also desirable treatment strategies.

Arylazolylthioacetanilide. Part 11: design, synthesis and biological evaluation of 1,2,4-triazole thioacetanilide derivatives as novel non-nucleoside HIV-1 reverse transcriptase inhibitors.

PubMed

Li, Zhenyu; Cao, Yuan; Zhan, Peng; Pannecouque, Christophe; Balzarini, Jan; Clercq, Erik De; Shen, Yuemao; Liu, Xinyong

2013-11-01

A series of novel 1,2,4-triazole thioacetanilide derivatives has been designed, synthesized and evaluated for their anti-HIV activities in MT-4 cells. Half of these compounds showed moderate to potent activities against wild-type HIV-1 with an EC50 ranging from 38.0 μM to 4.08 µM. Among them, 2-(4-(2-fluorobenzyl)-5-isopropyl-4H-1,2,4-triazol- 3-ylthio)-N-(2-nitrophenyl)acetamide 7d was identified as the most promising compound (EC50 = 4.26 µM, SI = 49). However, no compound was active against HIV-2. The preliminary structure-activity relationships among the newly synthesized congeners are discussed.

Surveillance of transmitted HIV drug resistance using matched plasma and dried blood spot specimens from voluntary counseling and testing sites in Ho Chi Minh City, Vietnam, 2007-2008.

PubMed

Duc, Nguyen Bui; Hien, Bui Thu; Wagar, Nick; Tram, Tran Hong; Giang, Le Truong; Yang, Chunfu; Wolfe, Mitchell I; Hien, Nguyen Tran; Tuan, Nguyen Anh

2012-05-01

During 2007-2008, surveillance of transmitted human immunodeficiency virus (HIV) drug resistance (TDR) was performed following World Health Organization guidance among clients with newly diagnosed HIV infection attending voluntary counseling and testing (VCT) sites in Ho Chi Minh City (HCMC), Vietnam. Moderate (5%-15%) TDR to nonnucleoside reverse-transcriptase inhibitors (NNRTIs) was observed among VCT clients aged 18-21 years. Follow-up surveillance of TDR in HCMC and other geographic regions of Vietnam is warranted. Data generated will guide the national HIV drug resistance surveillance strategy and support selection of current and future first-line antiretroviral therapy and HIV prevention programs.

Effectiveness of Non-nucleoside Reverse-Transcriptase Inhibitor-Based Antiretroviral Therapy in Women Previously Exposed to a Single Intrapartum Dose of Nevirapine: A Multi-country, Prospective Cohort Study

PubMed Central

Stringer, Jeffrey S. A.; McConnell, Michelle S.; Kiarie, James; Bolu, Omotayo; Anekthananon, Thanomsak; Jariyasethpong, Tavatchai; Potter, Dara; Mutsotso, Winnie; Borkowf, Craig B.; Mbori-Ngacha, Dorothy; Muiruri, Peter; Ong'ech, John Odero; Zulu, Isaac; Njobvu, Lungowe; Jetsawang, Bongkoch; Pathak, Sonal; Bulterys, Marc; Shaffer, Nathan; Weidle, Paul J.

2010-01-01

Background Intrapartum and neonatal single-dose nevirapine (NVP) reduces the risk of mother-to-child HIV transmission but also induces viral resistance to non-nucleoside reverse transcriptase inhibitor (NNRTI) drugs. This drug resistance largely fades over time. We hypothesized that women with a prior single-dose NVP exposure would have no more than a 10% higher cumulative prevalence of failure of their NNRTI-containing antiretroviral therapy (ART) over the first 48 wk of therapy than would women without a prior exposure. Methods and Findings We enrolled 355 NVP-exposed and 523 NVP-unexposed women at two sites in Zambia, one site in Kenya, and two sites in Thailand into a prospective, non-inferiority cohort study and followed them for 48 wk on ART. Those who died, discontinued NNRTI-containing ART, or had a plasma viral load ≥400 copies/ml at either the 24 wk or 48 wk study visits and confirmed on repeat testing were characterized as having failed therapy. Overall, 114 of 355 NVP-exposed women (32.1%) and 132 of 523 NVP-unexposed women (25.2%) met criteria for treatment failure. The difference in failure rates between the exposure groups was 6.9% (95% confidence interval [CI] 0.8%–13.0%). The failure rates of women stratified by our predefined exposure interval categories were as follows: 47 of 116 women in whom less than 6 mo elapsed between exposure and starting ART failed therapy (40%; p<0.001 compared to unexposed women); 25 of 67 women in whom 7–12 mo elapsed between exposure and starting ART failed therapy (37%; p = 0.04 compared to unexposed women); and 42 of 172 women in whom more than 12 mo elapsed between exposure and starting ART failed therapy (24%; p = 0.82 compared to unexposed women). Locally weighted regression analysis also indicated a clear inverse relationship between virologic failure and the exposure interval. Conclusions Prior exposure to single-dose NVP was associated with an increased risk of treatment failure; however, this risk seems largely confined to women with a more recent exposure. Women requiring ART within 12 mo of NVP exposure should not be prescribed an NNRTI-containing regimen as first-line therapy. Please see later in the article for the Editors' Summary PMID:20169113

Prevalence in the USA of rilpivirine resistance-associated mutations in clinical samples and effects on phenotypic susceptibility to rilpivirine and etravirine.

PubMed

Picchio, Gaston R; Rimsky, Laurence T; Van Eygen, Veerle; Haddad, Mojgan; Napolitano, Laura A; Vingerhoets, Johan

2014-01-01

The prevalence of rilpivirine resistance-associated mutations (RAMs) in the USA, and their effect on phenotypic susceptibility to rilpivirine and etravirine, was evaluated in clinical samples from HIV-1-infected patients. In total, 15,991 samples submitted to Monogram Biosciences (South San Francisco, CA, USA) for routine resistance testing between January 2010 and June 2011 were assessed for the presence of known rilpivirine RAMs K101E/P, E138A/G/K/Q/R, V179L, Y181C/I/V, Y188L, H221Y, F227C and M230I/L; non-nucleoside reverse transcriptase inhibitor (NNRTI) RAMs K103N, L100I and L100I+K103N; and the nucleoside reverse transcriptase inhibitor (NRTI) RAMs M184I/V and their combinations with rilpivirine RAMs. Phenotypic susceptibility (PhenoSenseGT(®) assay; Monogram Biosciences) was evaluated, with reduced susceptibility defined as fold change (FC) in 50% inhibitory concentration (IC50)>2.0 for rilpivirine and FC>2.9 for etravirine. Of the 15,991 samples, 17% harboured ≥1 rilpivirine RAMs. The prevalence of most rilpivirine RAMs and combinations of NNRTI RAMs of interest was low (≤3%), except for Y181C (7%). Rilpivirine RAMs were often associated with reduced rilpivirine phenotypic susceptibility. Median FC values >2.0 were observed for clinical isolates with rilpivirine RAMs K101P, E138Q/R, Y181C/I/V, Y188L or M230L, and for the combination of E138K with M184I/V, and K101E with M184I. Most rilpivirine FC values >2.0 were associated with etravirine FC values >2.9 for individual rilpivirine RAMs and those combined with M184I/V. There was no relationship between the presence of K103N and rilpivirine FC. However, the L100I+K103N combination (without rilpivirine RAMs), at <2% prevalence, was associated with a rilpivirine FC>2.0. Based on 15,991 US clinical samples from HIV-1-infected patients, the frequency of most known rilpivirine RAMs apart from Y181C was low.

HIV-associated lipodystrophy syndrome: A review of clinical aspects

PubMed Central

Baril, Jean-Guy; Junod, Patrice; LeBlanc, Roger; Dion, Harold; Therrien, Rachel; Laplante, François; Falutz, Julian; Côté, Pierre; Hébert, Marie-Nicole; Lalonde, Richard; Lapointe, Normand; Lévesque, Dominic; Pinault, Lyse; Rouleau, Danielle; Tremblay, Cécile; Trottier, Benoît; Trottier, Sylvie; Tsoukas, Chris; Weiss, Karl

2005-01-01

Approximately two years after the introduction of highly active antiretroviral therapy for the treatment of HIV infection, body shape changes and metabolic abnormalities were increasingly observed. Initially, these were ascribed to protease inhibitors, but it is now clear that nucleoside reverse transcriptase inhibitors also contribute to lipodystrophy syndrome. The syndrome groups together clinical conditions describing changes in body fat distribution that include lipoatrophy, lipoaccumulation or both. However, there does not appear to be a direct link between lipoatrophy and lipoaccumulation that would support a single mechanism for the redistribution of body fat. Currently, there is no clear definition of lipodystrophy, which explains the difficulty in determining its prevalence and etiology. There are no current guidelines for the treatment of fat distribution abnormalities that occur in the absence of other metabolic complications. The present article reviews the current state of knowledge of the definition, symptoms, risk factors, pathogenesis, diagnosis and treatment of the morphological changes associated with lipodystrophy syndrome. PMID:18159551

Fosamprenavir calcium plus ritonavir for HIV infection.

PubMed

Torres, Harrys A; Arduino, Roberto C

2007-06-01

Fosamprenavir is a protease inhibitor (PI) approved for the treatment of HIV-1 infection. Fosamprenavir is a prodrug of amprenavir developed to reduce the pill burden yet maintain the unique resistance pattern and efficacy associated with amprenavir. In a head-to-head, noninferiority trial in antiretroviral treatment-naive HIV-infected patients, the antiviral efficacy and tolerability of ritonavir-boosted fosamprenavir was not inferior to ritonavir-boosted lopinavir, when the PIs were combined with two other nucleoside reverse transcriptase inhibitors. There are fewer studies published about fosamprenavir use in antiretroviral treatment-experienced HIV-infected patients. The high genetic barrier to the development of resistance to fosamprenavir and the low level of cross-resistance between ritonavir-boosted fosamprenavir and other PI regimens are notable. As with amprenavir, gastrointestinal disturbance and rash are the most frequent short-term treatment-limiting events with fosamprenavir. Treatment with ritonavir-boosted fosamprenavir can produce a durable response. To date, fosamprenavir is one of the recommended preferred PI components for the treatment of antiretroviral-naive HIV-infected patients.

Xenobiotics that affect oxidative phosphorylation alter differentiation of human adipose-derived stem cells at concentrations that are found in human blood

PubMed Central

Llobet, Laura; Toivonen, Janne M.; Montoya, Julio; Ruiz-Pesini, Eduardo; López-Gallardo, Ester

2015-01-01

ABSTRACT Adipogenesis is accompanied by differentiation of adipose tissue-derived stem cells to adipocytes. As part of this differentiation, biogenesis of the oxidative phosphorylation system occurs. Many chemical compounds used in medicine, agriculture or other human activities affect oxidative phosphorylation function. Therefore, these xenobiotics could alter adipogenesis. We have analyzed the effects on adipocyte differentiation of some xenobiotics that act on the oxidative phosphorylation system. The tested concentrations have been previously reported in human blood. Our results show that pharmaceutical drugs that decrease mitochondrial DNA replication, such as nucleoside reverse transcriptase inhibitors, or inhibitors of mitochondrial protein synthesis, such as ribosomal antibiotics, diminish adipocyte differentiation and leptin secretion. By contrast, the environmental chemical pollutant tributyltin chloride, which inhibits the ATP synthase of the oxidative phosphorylation system, can promote adipocyte differentiation and leptin secretion, leading to obesity and metabolic syndrome as postulated by the obesogen hypothesis. PMID:26398948

Dissimilarities in the metabolism of antiretroviral drugs used in HIV pre-exposure prophylaxis in colon and vagina tissues.

PubMed

To, Elaine E; Hendrix, Craig W; Bumpus, Namandjé N

2013-10-01

Attempts to prevent HIV infection through pre-exposure prophylaxis (PrEP) include topical application of anti-HIV drugs to the mucosal sites of infection; however, a potential role for local drug metabolizing enzymes in modulating the exposure of the mucosal tissues to these drugs has yet to be explored. Here we present the first report that enzymes belonging to the cytochrome P450 (CYP) and UDP-glucuronosyltransferase (UGT) families of drug metabolizing enzymes are expressed and active in vaginal and colorectal tissue using biopsies collected from healthy volunteers. In doing so, we discovered that dapivirine and maraviroc, a non-nucleoside reverse transcriptase inhibitor and an entry inhibitor currently in development as microbicides for HIV PrEP, are differentially metabolized in colorectal tissue and vaginal tissue. Taken together, these data should help to guide the optimization of small molecules being developed for HIV PrEP. Copyright © 2013 Elsevier Inc. All rights reserved.

Altered Oligodendrocyte Maturation and Myelin Maintenance: The Role of Antiretrovirals in HIV-Associated Neurocognitive Disorders.

PubMed

Jensen, Brigid K; Monnerie, Hubert; Mannell, Maggie V; Gannon, Patrick J; Espinoza, Cagla Akay; Erickson, Michelle A; Bruce-Keller, Annadora J; Gelman, Benjamin B; Briand, Lisa A; Pierce, R Christopher; Jordan-Sciutto, Kelly L; Grinspan, Judith B

2015-11-01

Despite effective viral suppression through combined antiretroviral therapy (cART), approximately half of HIV-positive individuals have HIV-associated neurocognitive disorders (HAND). Studies of antiretroviral-treated patients have revealed persistent white matter abnormalities including diffuse myelin pallor, diminished white matter tracts, and decreased myelin protein mRNAs. Loss of myelin can contribute to neurocognitive dysfunction because the myelin membrane generated by oligodendrocytes is essential for rapid signal transduction and axonal maintenance. We hypothesized that myelin changes in HAND are partly due to effects of antiretroviral drugs on oligodendrocyte survival and/or maturation. We showed that primary mouse oligodendrocyte precursor cell cultures treated with therapeutic concentrations of HIV protease inhibitors ritonavir or lopinavir displayed dose-dependent decreases in oligodendrocyte maturation; however, this effect was rapidly reversed after drug removal. Conversely, nucleoside reverse transcriptase inhibitor zidovudine had no effect. Furthermore, in vivo ritonavir administration to adult mice reduced frontal cortex myelin protein levels. Finally, prefrontal cortex tissue from HIV-positive individuals with HAND on cART showed a significant decrease in myelin basic protein compared with untreated HIV-positive individuals with HAND or HIV-negative controls. These findings demonstrate that antiretrovirals can impact myelin integrity and have implications for myelination in juvenile HIV patients and myelin maintenance in adults on lifelong therapy.

Impact of lopinavir/ritonavir use on antiretroviral resistance in recent clinical practice.

PubMed

Lambert-Niclot, Sidonie; Masquelier, Bernard; Cohen Codar, Isabelle; Soulie, Cathia; Delaugerre, Constance; Morand-Joubert, Laurence; Charpentier, Charlotte; Ferre, Virginie; Plantier, Jean-Christophe; Montes, Brigitte; Carret, Sophie; Perrot, Valerie; Peytavin, Gilles; Costagliola, Dominique; Calvez, Vincent; Marcelin, Anne-Geneviève

2012-10-01

This observational study was requested by French health authorities to determine the impact of lopinavir/ritonavir (Kaletra(®)) on antiretroviral resistance in clinical practice. Virological failures of lopinavir/ritonavir and their effects on the resistance to protease inhibitors and reverse transcriptase inhibitors were evaluated in protease inhibitor-experienced patients. Virological failure was defined as an HIV-1 plasma viral load >50 copies/mL after at least 3 months of lopinavir/ritonavir-containing antiretroviral therapy. For all patients, a resistance genotypic test was available at failure and before lopinavir/ritonavir treatment. Data from 72 patients with inclusion criteria were studied. The mean viral load at baseline was 4 log(10) copies/mL (1.6-6.5). Mutations in the protease gene significantly selected between baseline and failure were L10V, K20R, L33F, M36I, I47V, I54V, A71V and I85V (P < 0.05). Patients who had more than seven protease inhibitor mutations at baseline showed a significantly increased risk of occurrence of protease inhibitor mutations. The proportion of viruses susceptible to atazanavir, fosamprenavir and darunavir decreased significantly between baseline and failure (P < 0.05). Among patients with a virus susceptible to atazanavir at day 0, 26% (n = 14) exhibited a virus resistant or possibly resistant at the time of failure. This proportion was 32% (n = 16) for fosamprenavir and 16% (n = 7) for darunavir. A darunavir-based regimen appears to be a sequential option in the case of lopinavir/ritonavir failure. To compare and determine the best treatment sequencing, similar studies should be performed for darunavir/ritonavir and atazanavir/ritonavir.

Antiretroviral Drug Susceptibility Among HIV-Infected Adults Failing Antiretroviral Therapy in Rakai, Uganda

PubMed Central

Laeyendecker, Oliver; Nakigozi, Gertrude; Gallant, Joel E.; Huang, Wei; Hudelson, Sarah E.; Quinn, Thomas C.; Newell, Kevin; Serwadda, David; Gray, Ronald H.; Wawer, Maria J.; Eshleman, Susan H.

2012-01-01

Abstract We analyzed antiretroviral drug susceptibility in HIV-infected adults failing first- and second-line antiretroviral treatment (ART) in Rakai, Uganda. Samples obtained from participants at baseline (pretreatment) and at the time of failure on first-line ART and second-line ART were analyzed using genotypic and phenotypic assays for antiretroviral drug resistance. Test results were obtained from 73 samples from 38 individuals (31 baseline samples, 36 first-line failure samples, and six second-line failure samples). Four (13%) of the 31 baseline samples had mutations associated with resistance to nucleoside or nonnucleoside reverse transcriptase inhibitors (NRTIs and NNRTIs, respectively). Among the 36 first-line failure samples, 31 (86%) had NNRTI resistance mutations and 29 (81%) had lamivudine resistance mutations; only eight (22%) had other NRTI resistance mutations. None of the six individuals failing a second-line protease inhibitor (PI)-based regimen had PI resistance mutations. Six (16%) of the participants had discordant genotypic and phenotypic test results. Genotypic resistance to drugs included in first-line ART regimens was detected prior to treatment and among participants failing first-line ART. PI resistance was not detected in individuals failing second-line ART. Surveillance for transmitted and acquired drug resistance remains a priority for scale-up of ART. PMID:22443282

Molecular Docking and Molecular Dynamics to Identify a Novel Human Immunodeficiency Virus Inhibitor from Alkaloids of Toddalia asiatica

PubMed Central

Priya, R.; Sumitha, Rajendrarao; Doss, C. George Priya; Rajasekaran, C.; Babu, S.; Seenivasan, R.; Siva, R.

2015-01-01

Background: Acquired immunodeficiency syndrome caused by human immunodeficiency virus (HIV) is an immunosuppressive disease. Over the past decades, it has plagued human health due to the grave consequences in its harness. Objective: For this reason, anti-HIV agents are imperative, and the search for the same from natural resources would assure the safety. Materials and Methods: In this investigation we have performed molecular docking, molecular property prediction, drug-likeness score, and molecular dynamics (MD) simulation to develop a novel anti-HIV drug. We have screened 12 alkaloids from a medicinal plant Toddalia asiatica for its probabilistic binding with the active site of the HIV-1-reverse transcriptase (HIV-1-RT) domain (the major contributor to the onset of the disease). Results: The docking results were evaluated based on free energies of binding (ΔG), and the results suggested toddanol, toddanone, and toddalenone to be potent inhibitors of HIV-1-RT. In addition, the alkaloids were subjected to molecular property prediction analysis. Toddanol and toddanone with more rotatable bonds were found to have a drug-likeness score of 0.23 and 0.11, respectively. These scores were comparable with the standard anti-HIV drug zidovudine with a model score 0.28. Finally, two characteristic protein-ligand complexes were exposed to MD simulation to determine the stability of the predicted conformations. Conclusion: The toddanol-RT complex showed higher stability and stronger H-bonds than toddanone-RT complex. Based on these observations, we firmly believe that the alkaloid toddanol could aid in efficient HIV-1 drug discovery. SUMMARY In the present study, the molecular docking and MD simulations are performed to explore the possible binding mode of HIV 1 RT with 12 alkaloids of T. asiatica. Molecular docking by AutoDock4 revealed three alkaloids toddanol, toddanone, and toddalenone with highest binding affinity towards HIV 1 RT. The drug likeness model score revealed a positive score for toddanol and toddanone which is comparable to the drug likeness score of the standard anti HIV drug zidovudine. Results from simulation analysis revealed that toddanol RT complex is more stable than toddanone RT complex inferring toddanol as a potential anti HIV drug molecule. Abbreviations used: HIV: Human immunodeficiency virus, HIV 1 RT: HIV 1 reverse transcriptase, RNase H: Ribonuclease H, MD: Molecular dynamics, PDB: Protein databank, RMSD: Root mean square deviation, RMSF: Root mean square fluctuation. PMID:26929575

Studies on the inhibition of Moloney murine leukemia virus reverse transcriptase by N-tritylamino acids and N-tritylamino acid-nucleotide compounds.

PubMed

Hawtrey, Arthur; Pieterse, Anton; van Zyl, Johann; Van der Bijl, Pieter; Van der Merwe, Marichen; Nel, William; Ariatti, Mario

2008-09-01

N-Acylated derivatives of 8-(6-aminohexyl) amino-adenosine-5 '-phosphate were prepared and studied with regard to their effect on DNA synthesis by the Moloney leukemia virus reverse transcriptase. N-palmitoyl and N-nicotinyl derivatives and bis-8-(6-aminohexyl) amino-5'-AMP inhibited the enzyme partially using poly (rA).oligo d(pT)(16-18) as template-primer with [(3)H]dTTP. In order to increase hydrophobicity in the acyl component tethered to the 8-(6-aminohexyl) amino group on the adenine nucleotide, N-trityl-L-phenylalanine and the N-trityl derivatives of the o, m, and p-fluoro-DL-phenylalanine were initially examined for inhibition of the enzyme using the above template-primer system. The compounds all inhibited the reverse transcriptase with IC(50) values of approximately 60-80 microM. However, when N-trityl-m-fluoro-DL-phenylalanine was coupled to the nucleotide 8-(6-aminohexyl) amino-adenosine-5'-phosphate, the inhibitory activity of this compound increased significantly (IC(50) = 5 microM).

Modified concentration method for the detection of enteric viruses on fruits and vegetables by reverse transcriptase-polymerase chain reaction or cell culture.

PubMed

Dubois, Eric; Agier, Cécilia; Traoré, Ousmane; Hennechart, Catherine; Merle, Ghislaine; Crucière, Catherine; Laveran, Henri

2002-12-01

Fruits and vegetables may act as a vehicle of human enteric virus if they are irrigated with sewage-contaminated water or prepared by infected food handlers. An elution-concentration method was modified to efficiently detect, by reverse transcriptase-polymerase chain reaction (RT-PCR) or by cell culture, contamination by poliovirus, hepatitis A virus (HAV), and Norwalk-like virus (NLV) of various fresh and frozen berries and fresh vegetables. The protocol included washing the fruit or vegetable surface with 100 mM Tris-HCl, 50 mM glycine, and 3% beef extract, pH 9.5 buffer, which favors viral elution from acid-releasing berries, supplemented with 50 mM MgCl2 to reduce the decrease in viral infectivity during the process. The viral concentration method was based on polyethylene glycol precipitation. Copurified RT-PCR inhibitors and cytotoxic compounds were removed from viral concentrates by chloroform-butanol extraction. Viruses from 100 g of vegetal products could be recovered in volumes of 3 to 5 ml. Viral RNAs were isolated by a spin column method before molecular detection or concentrates were filtered (0.22-microm porosity) and inoculated on cell culture for infectious virus detection. About 15% of infectious poliovirus and 20% of infectious HAV were recovered from frozen raspberry surfaces. The percentage of viral RNA recovery was estimated by RT-PCR to be about 13% for NLV, 17% for HAV, and 45 to 100% for poliovirus. By this method, poliovirus and HAV RNA were detected on products inoculated with a titer of about 5 x 10(1) 50% tissue culture infectious dose per 100 g. NLV RNA was detected at an initial inoculum of 1.2 x 10(3) RT-PCR amplifiable units. This method would be useful for the viral analysis of fruits or vegetables during an epidemiological investigation of foodborne diseases.

High prevalence of bevirimat resistance mutations in protease inhibitor-resistant HIV isolates.

PubMed

Verheyen, Jens; Verhofstede, Chris; Knops, Elena; Vandekerckhove, Linos; Fun, Axel; Brunen, Diede; Dauwe, Kenny; Wensing, Annemarie M J; Pfister, Herbert; Kaiser, Rolf; Nijhuis, Monique

2010-03-13

Bevirimat is the first drug of a new class of antivirals that hamper the maturation of HIV. The objective of this study was to evaluate the sequence variability of the gag region targeted by bevirimat in HIV subtype-B isolates. Of 484 HIV subtype-B isolates, the gag region comprising amino acids 357-382 was sequenced. Of the patients included, 270 were treatment naive and 214 were treatment experienced. In the latter group, 48 HIV isolates harboured mutations associated with reverse transcriptase inhibitor resistance only, and 166 HIV isolates carried mutations associated with protease inhibitor resistance. In the treatment-naive patient population, approximately 30% harboured an HIV isolate with at least one mutation associated with a reduced susceptibility to bevirimat (H358Y, L363M, Q369H, V370A/M/del and T371del). In HIV isolates with protease inhibitor resistance, the prevalence of bevirimat resistance mutations increased to 45%. Accumulation of mutations at four positions in the bevirimat target region, S368C, Q369H, V370A and S373P, was significantly observed. Mutations associated with bevirimat resistance were detected more frequently in HIV isolates with three or more protease inhibitor resistance mutations than in those with less than three protease inhibitor mutations. Reduced bevirimat activity can be expected in one-third of treatment-naive HIV subtype-B isolates and significantly more in protease inhibitor-resistant HIV. These data indicate that screening for bevirimat resistance mutations before administration of the drug is essential.

Comparison of reverse transcriptase PCR, reverse transcriptase loop-mediated isothermal amplification, and culture-based assays for Salmonella detection from pork processing environments.

PubMed

Techathuvanan, Chayapa; Draughon, Frances Ann; D'Souza, Doris Helen

2011-02-01

Novel rapid Salmonella detection assays without the need for sophisticated equipment or labor remain in high demand. Real-time reverse transcriptase PCR (RT-PCR) assays, though rapid and sensitive, require expensive thermocyclers, while a novel RT loop-mediated isothermal amplification (RT-LAMP) method requires only a simple water bath. Our objective was to compare the detection sensitivity of Salmonella Typhimurium from the pork processing environment by RT-LAMP, RT-PCR, and culture-based assays. Carcass and surface swabs and carcass rinses were obtained from a local processing plant. Autoclaved carcass rinses (500 ml) were spiked with Salmonella Typhimurium and filtered. Filters were placed in stomacher bags containing tetrathionate broth (TTB) and analyzed with or without 10-h enrichment at 37 °C. Natural swabs were stomached with buffered peptone water, and natural carcass rinses were filtered, preenriched, and further enriched in TTB. Serially-diluted enriched samples were enumerated by spread plating on xylose lysine Tergitol 4 agar. RNA was extracted from 5 ml of enriched TTB with TRIzol. RT-LAMP assay using previously described invA primers was conducted at 62 °C for 90 min in a water bath with visual detection and by gel electrophoresis. SYBR Green I-based-real-time RT-PCR was carried out with invA primers followed by melt temperature analysis. The results of RT-LAMP detection for spiked carcass rinses were comparable to those of RT-PCR and cultural plating, with detection limits of 1 log CFU/ml, although they were obtained significantly faster, within 24 h including preenrichment and enrichment. RT-LAMP showed 4 of 12 rinse samples positive, while RT-PCR showed 1 of 12 rinse samples positive. For swabs, 6 of 27 samples positive by RT-LAMP and 5 of 27 by RT-PCR were obtained. This 1-day RT-LAMP assay shows promise for routine Salmonella screening by the pork industry. Copyright ©, International Association for Food Protection

The CD4/CD8 Ratio is Inversely Associated with Carotid Intima-Media Thickness Progression in Human Immunodeficiency Virus-Infected Patients on Antiretroviral Treatment.

PubMed

Bernal Morell, Enrique; Serrano Cabeza, José; Muñoz, Ángeles; Marín, Irene; Masiá, Mar; Gutiérrez, Félix; Cano, Alfredo

2016-07-01

Inversion of the CD4/CD8 ratio (<1) has been identified as a surrogate marker of immunosenescence and an independent predictor of AIDS events in HIV-infected patients and mortality in the general population. We aimed to assess the association between the CD4/CD8 ratio and carotid intima-media thickness (cIMT) progression in treated HIV-infected patients as a marker of coronary heart disease. A longitudinal study was conducted during 3 years in 96 virally suppressed HIV-infected patients receiving antiretroviral treatment (ART). We analyzed the associations between the CD4/CD8 ratio, cardiovascular risk factors, and antiretroviral treatment (ART) and progression of subclinical atherosclerosis assessed using cIMT at baseline and after 3 years. Finally, 96 patients completed the study. Seventy six (79.1%) patients were male, aged 44 ± 10 years; 39 (40.6%) were on treatment with protease inhibitors; 49 (51.04%) with nonnucleoside reverse transcriptase inhibitors, 6 (6.25%) with integrase inhibitors, 3 (3.12%) with maraviroc, and 2 (2.08%) just with nucleoside reverse transcriptase inhibitors. The mean of ART exposition was 6.9 ± 5.9 years. Twenty six (27%) patients had family history of ischemic heart disease, 51 (53.12%) were smokers, 12 (12.5%) were hypertensive, 4 (4.16%) had type 2 diabetes, 23 (23.9%) had dyslipidemia, and 31 (32.3%) were infected with hepatitis C virus. Baseline cIMT was significantly associated with age (rho = 0.497; p < .001), basal glucemia (rho = 0.323; p = .001), triglycerides (rho = 0.232; p = .023), Framingham risk score (rho = 0.324; p = .001), CD4/CD8 ratio (rho = -0.176; p = .05), and dyslipidemia (0.72 ± 0.16 mm vs. 0.63 ± 0.11 mm; p = .029). In multivariable analysis where cardiovascular risk factor and ART were included, cIMT progression was inversely associated with CD4/CD8 ratio [odds ratio (OR) = 0.283; confidence interval (95% CI) 0.099-0.809; p = .019]. In conclusion, the inversion of CD4/CD8 ratio in treated HIV-infected patients is independently associated with cIMT progression, a marker of coronary heart disease. Therefore, it might be clinically useful as predictor of cardiovascular events.

Antiretroviral Drugs for Treatment and Prevention of HIV Infection in Adults: 2016 Recommendations of the International Antiviral Society-USA Panel.

PubMed

Günthard, Huldrych F; Saag, Michael S; Benson, Constance A; del Rio, Carlos; Eron, Joseph J; Gallant, Joel E; Hoy, Jennifer F; Mugavero, Michael J; Sax, Paul E; Thompson, Melanie A; Gandhi, Rajesh T; Landovitz, Raphael J; Smith, Davey M; Jacobsen, Donna M; Volberding, Paul A

2016-07-12

New data and therapeutic options warrant updated recommendations for the use of antiretroviral drugs (ARVs) to treat or to prevent HIV infection in adults. To provide updated recommendations for the use of antiretroviral therapy in adults (aged ≥18 years) with established HIV infection, including when to start treatment, initial regimens, and changing regimens, along with recommendations for using ARVs for preventing HIV among those at risk, including preexposure and postexposure prophylaxis. A panel of experts in HIV research and patient care convened by the International Antiviral Society-USA reviewed data published in peer-reviewed journals, presented by regulatory agencies, or presented as conference abstracts at peer-reviewed scientific conferences since the 2014 report, for new data or evidence that would change previous recommendations or their ratings. Comprehensive literature searches were conducted in the PubMed and EMBASE databases through April 2016. Recommendations were by consensus, and each recommendation was rated by strength and quality of the evidence. Newer data support the widely accepted recommendation that antiretroviral therapy should be started in all individuals with HIV infection with detectable viremia regardless of CD4 cell count. Recommended optimal initial regimens for most patients are 2 nucleoside reverse transcriptase inhibitors (NRTIs) plus an integrase strand transfer inhibitor (InSTI). Other effective regimens include nonnucleoside reverse transcriptase inhibitors or boosted protease inhibitors with 2 NRTIs. Recommendations for special populations and in the settings of opportunistic infections and concomitant conditions are provided. Reasons for switching therapy include convenience, tolerability, simplification, anticipation of potential new drug interactions, pregnancy or plans for pregnancy, elimination of food restrictions, virologic failure, or drug toxicities. Laboratory assessments are recommended before treatment, and monitoring during treatment is recommended to assess response, adverse effects, and adherence. Approaches are recommended to improve linkage to and retention in care are provided. Daily tenofovir disoproxil fumarate/emtricitabine is recommended for use as preexposure prophylaxis to prevent HIV infection in persons at high risk. When indicated, postexposure prophylaxis should be started as soon as possible after exposure. Antiretroviral agents remain the cornerstone of HIV treatment and prevention. All HIV-infected individuals with detectable plasma virus should receive treatment with recommended initial regimens consisting of an InSTI plus 2 NRTIs. Preexposure prophylaxis should be considered as part of an HIV prevention strategy for at-risk individuals. When used effectively, currently available ARVs can sustain HIV suppression and can prevent new HIV infection. With these treatment regimens, survival rates among HIV-infected adults who are retained in care can approach those of uninfected adults.

Combination antiretroviral use and preterm birth.

PubMed

Watts, D Heather; Williams, Paige L; Kacanek, Deborah; Griner, Raymond; Rich, Kenneth; Hazra, Rohan; Mofenson, Lynne M; Mendez, Hermann A

2013-02-15

Use of antiretroviral drugs (ARVs) during pregnancy has been associated with higher risk of preterm birth. The Pediatric HIV/AIDS Cohort Study network's Surveillance Monitoring for ART Toxicities study is a US-based cohort of human immunodeficiency virus (HIV)-exposed uninfected children. We evaluated maternal ARV use during pregnancy and the risk of any type of preterm birth (ie, birth before 37 completed weeks of gestation), the risk of spontaneous preterm birth (ie, preterm birth that occurred after preterm labor or membrane rupture, without other complications), and the risk of small for gestational age (SGA; ie, a birth weight of <10th percentile for gestational age). Multivariable logistic regression models were used to evaluate the association of ARVs and timing of exposure, while adjusting for maternal characteristics. Among 1869 singleton births, 18.6% were preterm, 10.2% were spontaneous preterm, and 7.3% were SGA. A total of 89% used 3-drug combination ARV regimens during pregnancy. In adjusted models, the odds of preterm birth and spontaneous preterm birth were significantly greater among mothers who used protease inhibitors during the first trimester (adjusted odds ratios, 1.55 and 1.59, respectively) but not among mothers who used nonnucleoside reverse-transcriptase inhibitor or triple-nucleoside regimens during the first trimester. Combination ARV exposure starting later in pregnancy was not associated with increased risk. No associations were observed between SGA and exposure to combination ARV regimens. Protease inhibitor use early in pregnancy may be associated with increased risk for prematurity.