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Sample records for murine leukemia efficacy

  1. Antileukemic Efficacy of Continuous vs Discontinuous Dexamethasone in Murine Models of Acute Lymphoblastic Leukemia

    PubMed Central

    Ramsey, Laura B.; Janke, Laura J.; Payton, Monique A.; Cai, Xiangjun; Paugh, Steven W.; Karol, Seth E.; Kamdem, Landry Kamdem; Cheng, Cheng; Williams, Richard T.; Jeha, Sima; Pui, Ching-Hon; Evans, William E.; Relling, Mary V.

    2015-01-01

    Osteonecrosis is one of the most common, serious, toxicities resulting from the treatment of acute lymphoblastic leukemia. In recent years, pediatric acute lymphoblastic leukemia clinical trials have used discontinuous rather than continuous dosing of dexamethasone in an effort to reduce the incidence of osteonecrosis. However, it is not known whether discontinuous dosing would compromise antileukemic efficacy of glucocorticoids. Therefore, we tested the efficacy of discontinuous dexamethasone against continuous dexamethasone in murine models bearing human acute lymphoblastic leukemia xenografts (n = 8 patient samples) or murine BCR-ABL+ acute lymphoblastic leukemia. Plasma dexamethasone concentrations (7.9 to 212 nM) were similar to those achieved in children with acute lymphoblastic leukemia using conventional dosages. The median leukemia-free survival ranged from 16 to 59 days; dexamethasone prolonged survival from a median of 4 to 129 days in all seven dexamethasone-sensitive acute lymphoblastic leukemias. In the majority of cases (7 of 8 xenografts and the murine BCR-ABL model) we demonstrated equal efficacy of the two dexamethasone dosing regimens; whereas for one acute lymphoblastic leukemia sample, the discontinuous regimen yielded inferior antileukemic efficacy (log-rank p = 0.002). Our results support the clinical practice of using discontinuous rather than continuous dexamethasone dosing in patients with acute lymphoblastic leukemia. PMID:26252865

  2. Immunotherapy of murine leukemia. Efficacy of passive serum therapy of Friend leukemia virus-induced disease in immunocompromised mice

    SciTech Connect

    Genovesi, E.V.; Livnat, D.; Collins, J.J.

    1983-02-01

    Previous studies have demonstrated that the passive therapy of Friend murine leukemia virus (F-MuLV)-induced disease with chimpanzee anti-F-MuLV serum is accompanied by the development of host antiviral humoral and cellular immunity, the latter measurable in adoptive transfer protocols and by the ability of serum-protected mice to resist virus rechallenge. The present study was designed to further examine the contribution of various compartments of the host immune system to serum therapy itself, as well as to the acquired antiviral immunity that develops in serum-protected mice, through the use of naturally immunocompromised animals (e.g., nude athymic mice and natural killer (NK)-deficient beige mutant mice) or mice treated with immunoabrogating agents such as sublethal irradiation, cyclophosphamide (Cytoxan (Cy)), cortisone, and /sup 89/Sr. The studies in nude mice indicate that while mature T-cells are not needed for effective serum therapy, they do appear to be necessary for the long-term resistance of serum-protected mice to virus rechallenge and for the generation of the cell population(s) responsible for adoptive transfer of antiviral immunity. Furthermore, this acquired resistance is not due to virus neutralization by serum antibodies since antibody-negative, Cy-treated, serum-protected mice still reject the secondary virus infection. Lastly, while the immunocompromise systems examined did effect various host antiviral immune responses, none of them, including the NK-deficient beige mutation, significantly diminished the efficacy of the passive serum therapy of F-MuLV-induced disease.

  3. Efficacy of JAK/STAT pathway inhibition in murine xenograft models of early T-cell precursor (ETP) acute lymphoblastic leukemia

    PubMed Central

    Maude, Shannon L.; Dolai, Sibasish; Delgado-Martin, Cristina; Vincent, Tiffaney; Robbins, Alissa; Selvanathan, Arthavan; Ryan, Theresa; Hall, Junior; Wood, Andrew C.; Tasian, Sarah K.; Hunger, Stephen P.; Loh, Mignon L.; Mullighan, Charles G.; Wood, Brent L.; Hermiston, Michelle L.; Grupp, Stephan A.; Lock, Richard B.

    2015-01-01

    Early T-cell precursor (ETP) acute lymphoblastic leukemia (ALL) is a recently described subtype of T-ALL characterized by a unique immunophenotype and genomic profile, as well as a high rate of induction failure. Frequent mutations in cytokine receptor and Janus kinase (JAK)/signal transducer and activator of transcription (STAT) signaling pathways led us to hypothesize that ETP-ALL is dependent on JAK/STAT signaling. Here we demonstrate aberrant activation of the JAK/STAT pathway in ETP-ALL blasts relative to non-ETP T-ALL. Moreover, ETP-ALL showed hyperactivation of STAT5 in response to interleukin-7, an effect that was abrogated by the JAK1/2 inhibitor ruxolitinib. In vivo, ruxolitinib displayed activity in 6 of 6 patient-derived murine xenograft models of ETP-ALL, with profound single-agent efficacy in 5 models. Ruxolitinib treatment decreased peripheral blast counts relative to pretreatment levels and compared with control (P < .01) in 5 of 6 ETP-ALL xenografts, with marked reduction in mean splenic blast counts (P < .01) in 6 of 6 samples. Surprisingly, both JAK/STAT pathway activation and ruxolitinib efficacy were independent of the presence of JAK/STAT pathway mutations, raising the possibility that the therapeutic potential of ruxolitinib in ETP-ALL extends beyond those cases with JAK mutations. These findings establish the preclinical in vivo efficacy of ruxolitinib in ETP-ALL, a biologically distinct subtype for which novel therapies are needed. PMID:25645356

  4. ESCRT Requirements for Murine Leukemia Virus Release

    PubMed Central

    Bartusch, Christina; Prange, Reinhild

    2016-01-01

    The Murine Leukemia Virus (MLV) is a gammaretrovirus that hijack host components of the endosomal sorting complex required for transport (ESCRT) for budding. To determine the minimal requirements for ESCRT factors in MLV viral and viral-like particles (VLP) release, an siRNA knockdown screen of ESCRT(-associated) proteins was performed in MLV-producing human cells. We found that MLV VLPs and virions primarily engage the ESCRT-I factor Tsg101 and marginally the ESCRT-associated adaptors Nedd4-1 and Alix to enter the ESCRT pathway. Conversely, the inactivation of ESCRT-II had no impact on VLP and virion egress. By analyzing the effects of individual ESCRT-III knockdowns, VLP and virion release was profoundly inhibited in CHMP2A- and CHMP4B-knockdown cells. In contrast, neither the CHMP2B and CHMP4A isoforms nor CHMP3, CHMP5, and CHMP6 were found to be essential. In case of CHMP1, we unexpectedly observed that the CHMP1A isoform was specifically required for virus budding, but dispensable for VLP release. Hence, MLV utilizes only a subset of ESCRT factors, and viral and viral-like particles differ in ESCRT-III factor requirements. PMID:27096867

  5. ESCRT Requirements for Murine Leukemia Virus Release.

    PubMed

    Bartusch, Christina; Prange, Reinhild

    2016-01-01

    The Murine Leukemia Virus (MLV) is a gammaretrovirus that hijack host components of the endosomal sorting complex required for transport (ESCRT) for budding. To determine the minimal requirements for ESCRT factors in MLV viral and viral-like particles (VLP) release, an siRNA knockdown screen of ESCRT(-associated) proteins was performed in MLV-producing human cells. We found that MLV VLPs and virions primarily engage the ESCRT-I factor Tsg101 and marginally the ESCRT-associated adaptors Nedd4-1 and Alix to enter the ESCRT pathway. Conversely, the inactivation of ESCRT-II had no impact on VLP and virion egress. By analyzing the effects of individual ESCRT-III knockdowns, VLP and virion release was profoundly inhibited in CHMP2A- and CHMP4B-knockdown cells. In contrast, neither the CHMP2B and CHMP4A isoforms nor CHMP3, CHMP5, and CHMP6 were found to be essential. In case of CHMP1, we unexpectedly observed that the CHMP1A isoform was specifically required for virus budding, but dispensable for VLP release. Hence, MLV utilizes only a subset of ESCRT factors, and viral and viral-like particles differ in ESCRT-III factor requirements. PMID:27096867

  6. Oncogene Activation in Myeloid Leukemias by Graffi Murine Leukemia Virus Proviral Integration

    PubMed Central

    Denicourt, Catherine; Edouard, Elsy; Rassart, Eric

    1999-01-01

    The Graffi murine leukemia virus (MuLV) is a nondefective retrovirus that induces granulocytic leukemia in BALB/c and NFS mice. To identify genes involved in Graffi MuLV-induced granulocytic leukemia, tumor cell DNAs were examined for genetic alterations at loci described as common proviral integration sites in MuLV-induced myeloid, lymphoid, and erythroid leukemias. Southern blot analysis revealed rearrangements in c-myc, Fli-1, Pim-1, and Spi-1/PU.1 genes in 20, 10, 3.3, and 3.3% of the tumors tested, respectively. These results demonstrate for the first time the involvement of those genes in granulocytic leukemia. PMID:10196342

  7. Murine models of acute leukemia: important tools in current pediatric leukemia research.

    PubMed

    Jacoby, Elad; Chien, Christopher D; Fry, Terry J

    2014-01-01

    Leukemia remains the most common diagnosis in pediatric oncology and, despite dramatic progress in upfront therapy, is also the most common cause of cancer-related death in children. Much of the initial improvement in outcomes for acute lymphoblastic leukemia (ALL) was due to identification of cytotoxic agents that are active against leukemia followed by the recognition that combination of these cytotoxic agents and prolonged therapy are essential for cure. Recent data demonstrating lack of progress in patients for whom standard chemotherapy fails suggests that the ability to improve outcome for these children will not be dramatically impacted through more intensive or newer cytotoxic agents. Thus, much of the recent research focus has been in the area of improving our understanding of the genetics and the biology of leukemia. Although in vitro studies remain critical, given the complexity of a living system and the increasing recognition of the contribution of leukemia extrinsic factors such as the bone marrow microenvironment, in vivo models have provided important insights. The murine systems that are used can be broadly categorized into syngeneic models in which a murine leukemia can be studied in immunologically intact hosts and xenograft models where human leukemias are studied in highly immunocompromised murine hosts. Both of these systems have limitations such that neither can be used exclusively to study all aspects of leukemia biology and therapeutics for humans. This review will describe the various ALL model systems that have been developed as well as discuss the advantages and disadvantages inherent to these systems that make each particularly suitable for specific types of studies. PMID:24847444

  8. Murine Models of Acute Leukemia: Important Tools in Current Pediatric Leukemia Research

    PubMed Central

    Jacoby, Elad; Chien, Christopher D.; Fry, Terry J.

    2014-01-01

    Leukemia remains the most common diagnosis in pediatric oncology and, despite dramatic progress in upfront therapy, is also the most common cause of cancer-related death in children. Much of the initial improvement in outcomes for acute lymphoblastic leukemia (ALL) was due to identification of cytotoxic agents that are active against leukemia followed by the recognition that combination of these cytotoxic agents and prolonged therapy are essential for cure. Recent data demonstrating lack of progress in patients for whom standard chemotherapy fails suggests that the ability to improve outcome for these children will not be dramatically impacted through more intensive or newer cytotoxic agents. Thus, much of the recent research focus has been in the area of improving our understanding of the genetics and the biology of leukemia. Although in vitro studies remain critical, given the complexity of a living system and the increasing recognition of the contribution of leukemia extrinsic factors such as the bone marrow microenvironment, in vivo models have provided important insights. The murine systems that are used can be broadly categorized into syngeneic models in which a murine leukemia can be studied in immunologically intact hosts and xenograft models where human leukemias are studied in highly immunocompromised murine hosts. Both of these systems have limitations such that neither can be used exclusively to study all aspects of leukemia biology and therapeutics for humans. This review will describe the various ALL model systems that have been developed as well as discuss the advantages and disadvantages inherent to these systems that make each particularly suitable for specific types of studies. PMID:24847444

  9. Efficacy of Posaconazole in Murine Experimental Sporotrichosis

    PubMed Central

    Fernández-Silva, Fabiola; Capilla, Javier; Mayayo, Emilio

    2012-01-01

    We developed a murine model of systemic sporotrichosis by using three strains of each of the two commonest species causing sporotrichosis, i.e., Sporothrix schenckii sensu stricto and Sporothrix brasiliensis, in order to evaluate the efficacy of posaconazole (PSC). The drug was administered at a dose of 2.5 or 5 mg/kg of body weight twice a day by gavage, and one group was treated with amphotericin B (AMB) as a control treatment. Posaconazole, especially at 5 mg/kg, showed good efficacy against all the strains tested, regardless of their MICs, as measured by prolonged survival, tissue burden reduction, and histopathology. PMID:22330929

  10. Replication of the Moloney murine sarcoma-leukemia virus in XC cells.

    PubMed

    Trowbridge, S T; Benyesh-Melnick, M; Biswal, N

    1973-01-01

    The XC rat cell line was found to support the replication of a strain of the Moloney murine sarcoma-leukemia virus. In growth curve experiments cytopathology was paralleled by the production of murine sarcoma virus and leukemia virus progeny having the biologic, antigenic, and biophysical properties of the infecting virus. PMID:4346280

  11. An HSEF for murine myeloid leukemia

    SciTech Connect

    Bond, V.P.; Cronkite, E.P.; Bullis, J.E.; Wuu, C.S.; Marino, S.A.; Zaider, M.

    1996-10-01

    In the past decade, a large amount of effort has gone into the development of hit size effectiveness functions (HSEFs), with the ultimate aim of replacing the present absorbed dose-RBE-Q system. However, the absorbed dose determined at the tissue level is incapable of providing information on single hits on (doses to) the single cell. As a result, it is necessary to resort to microdosimetry, which is capable of providing not only the number of hits on cells, but the distribution of hit sizes as well. From this information, an HSEF can be derived. However, to date there have been no sets of data available on animals exposed to radiations of several qualities, and for which microdosimetric data were available. The objective of the present set of experiments was to remedy this situation. Large numbers of mice were exposed to radiations of several different qualities, and were observed throughout their entire lifespan for the appearance of myeloid leukemia. The HSEF developed for this neoplasm is presented and discussed.

  12. Murine leukemia virus envelope gp70 is a shared biomarker for the high-sensitivity quantification of murine tumor burden

    PubMed Central

    Scrimieri, Francesca; Askew, David; Corn, David J; Eid, Saada; Bobanga, Iuliana D; Bjelac, Jaclyn A; Tsao, Matthew L; Allen, Frederick; Othman, Youmna S; Wang, Shih-Chung G; Huang, Alex Y

    2013-01-01

    The preclinical development of anticancer drugs including immunotherapeutics and targeted agents relies on the ability to detect minimal residual tumor burden as a measure of therapeutic efficacy. Real-time quantitative (qPCR) represents an exquisitely sensitive method to perform such an assessment. However, qPCR-based applications are limited by the availability of a genetic defect associated with each tumor model under investigation. Here, we describe an off-the-shelf qPCR-based approach to detect a broad array of commonly used preclinical murine tumor models. In particular, we report that the mRNA coding for the envelope glycoprotein 70 (gp70) encoded by the endogenous murine leukemia virus (MuLV) is universally expressed in 22 murine cancer cell lines of disparate histological origin but is silent in 20 out of 22 normal mouse tissues. Further, we detected the presence of as few as 100 tumor cells in whole lung extracts using qPCR specific for gp70, supporting the notion that this detection approach has a higher sensitivity as compared with traditional tissue histology methods. Although gp70 is expressed in a wide variety of tumor cell lines, it was absent in inflamed tissues, non-transformed cell lines, or pre-cancerous lesions. Having a high-sensitivity biomarker for the detection of a wide range of murine tumor cells that does not require additional genetic manipulations or the knowledge of specific genetic alterations present in a given neoplasm represents a unique experimental tool for investigating metastasis, assessing antitumor therapeutic interventions, and further determining tumor recurrence or minimal residual disease. PMID:24482753

  13. Cross-protective murine graft-versus-leukemia responses to phenotypically distinct myeloid leukemia lines.

    PubMed

    Patterson, A E; Korngold, R

    2000-01-01

    A c-myc retrovirus-transformed myeloid leukemia line, MMB3.19, of C57BL/6 (B6) origin, was developed to investigate graft-versus-leukemia (GVL) activity in murine bone marrow transplantation (BMT) models. It was previously determined that both naive and leukemia-presensitized CD4+-enriched T cells are capable of mediating GVL activity to MMB3.19 challenge in both syngeneic (B6) and allogeneic (C3H.SW-->B6) strain combinations, with the latter coinciding with minimal graft-versus-host disease. In the present study, MMB3.19 and 2 other similarly derived, yet phenotypically diverse, B6 myeloid leukemia lines (MMB1.10 and MMB2.18) were investigated for potential shared tumor antigens in the syngeneic GVL model. Morphologically, all 3 tumor lines are blastic with high cytoplasmic:nuclear ratios, but MMB2.18 displays dendritic processes, whereas MMB1.10 and MMB3.19 have a more rounded appearance. Flow cytometric analysis of the 3 lines revealed constitutive surface molecule expression of Mac-1, Mac-2, F4/80, LFA-1, B7-1, B7-2, H2Kb, H2Db, and macrophage scavenger receptor, consistent with macrophage/monocyte lineages. Furthermore, each of the lines expresses H2I-Ab, but to varying degrees, with MMB2.18 cells having the lowest percentage (31.6%). In vitro 51Cr release assays using MMB3.19-primed T-cell effectors demonstrated equivalent specific lysis of all 3 leukemia-line target cells. In addition, enzyme-linked immunospot analysis of MMB3.19-primed CD4+ T cells revealed significantly increased frequencies of tumor-stimulated interleukin (IL)-2-, IL-4-, and interferon-gamma-secreting cells when restimulated with each of the 3 leukemia lines. Furthermore, when MMB3.19-primed CD4+ T cells were administered in a BMT setting, a protective GVL effect was seen in those mice challenged with MMB1.10, MMB2.18, or MMB3.19. Therefore, in vitro and in vivo experiments indicate that the 3 distinct myeloid leukemia lines share 1 or more common major histocompatibility complex class II

  14. Radiation-induced myeloid leukemia in murine models

    PubMed Central

    2014-01-01

    The use of radiation therapy is a cornerstone of modern cancer treatment. The number of patients that undergo radiation as a part of their therapy regimen is only increasing every year, but this does not come without cost. As this number increases, so too does the incidence of secondary, radiation-induced neoplasias, creating a need for therapeutic agents targeted specifically towards incidence reduction and treatment of these cancers. Development and efficacy testing of these agents requires not only extensive in vitro testing but also a set of reliable animal models to accurately recreate the complex situations of radiation-induced carcinogenesis. As radiation-induced leukemic progression often involves genomic changes such as rearrangements, deletions, and changes in methylation, the laboratory mouse Mus musculus, with its fully sequenced genome, is a powerful tool in cancer research. This fact, combined with the molecular and physiological similarities it shares with man and its small size and high rate of breeding in captivity, makes it the most relevant model to use in radiation-induced leukemia research. In this work, we review relevant M. musculus inbred and F1 hybrid animal models, as well as methods of induction of radiation-induced myeloid leukemia. Associated molecular pathologies are also included. PMID:25062865

  15. A Linkage Map of Endogenous Murine Leukemia Proviruses

    PubMed Central

    Frankel, W. N.; Stoye, J. P.; Taylor, B. A.; Coffin, J. M.

    1990-01-01

    Thirty endogenous proviruses belonging to the modified polytropic (Mpmv) class of murine leukemia virus (MLV) were identified by proviral-cellular DNA junction fragment segregation in several sets of recombinant inbred mice. Twenty-six Mpmv loci were mapped to chromosomal regions by matching proviral strain distribution patterns to those of previously assigned genes. Like other endogenous nonecotropic MLVs, Mpmv loci were present on several chromosomes in all strains examined. We pooled recombinant inbred strain linkage data from 110 MLV loci and selected marker genes in order to construct a chromosomal linkage map. Every mouse chromosome was found to harbor at least one proviral insertion, and several regions contained multiple integrations. However, the overall distribution of the 110 mapped proviruses did not deviate significantly from a random distribution. Because of their polymorphism in inbred strains of mice, and the ability to score as many as 57 proviruses per strain using only three hybridization probes, the nonecotropic MLVs mapped in common strains of mice offer a significant advantage over older methods (e.g., biochemical or individual restriction fragment polymorphisms) as genetic markers. These endogenous insertion elements should also be useful for assessing strain purity, and for studying the relatedness of common and not-so-common inbred strains. PMID:2155154

  16. Human APOBEC3G incorporation into murine leukemia virus particles

    SciTech Connect

    Kremer, Melanie; Schnierle, Barbara S. . E-mail: schba@pei.de

    2005-06-20

    The human APOBEC3G protein exhibits broad antiretroviral activity against a variety of retroviruses. It is packaged into viral particles and executes its antiviral function in the target cell. The packaging of APOBEC3G into different viral particles requires a mechanism that confers this promiscuity. Here, APOBEC3G incorporation into murine leukemia virus (MLV) was studied using retroviral vectors. APOBEC3G uptake did not require either its cytidine deaminase activity or the presence of a retroviral vector genome. Results from immunoprecipitation and co-localization studies of APOBEC3G with a MLV Gag-CFP (cyan fluorescent protein) fusion protein imply an interaction between both proteins. RNase A treatment did not inhibit the co-precipitation of Gag-CFP and APOBEC3G, suggesting that the interaction is RNA independent. Like human immunodeficiency virus (HIV) Gag, the MLV Gag precursor protein appears to interact with APOBEC3G, indicating that Gag contains conserved structures which are used to encapsidate APOBEC3G into different retroviral particles.

  17. Mechanical Properties of Murine Leukemia Virus Particles: Effect of Maturation

    PubMed Central

    Kol, Nitzan; Gladnikoff, Micha; Barlam, David; Shneck, Roni Z.; Rein, Alan; Rousso, Itay

    2006-01-01

    After budding from the host cell, retroviruses undergo a process of internal reorganization called maturation, which is prerequisite to infectivity. Viral maturation is accompanied by dramatic morphological changes, which are poorly understood in physical/mechanistic terms. Here, we study the mechanical properties of live mature and immature murine leukemia virus particles by indentation-type experiments conducted with an atomic force microscope tip. We find that both mature and immature particles have an elastic shell. Strikingly, the virus shell is twofold stiffer in the immature (0.68 N/m) than the mature (0.31 N/m) form. However, finite-element simulation shows that the average Young's modulus of the immature form is more than fourfold lower than that of the mature form. This finding suggests that per length unit, the protein-protein interactions in the mature shell are stronger than those in the immature shell. We also show that the mature virus shell is brittle, since it can be broken by application of large loading forces, by firm attachment to a substrate, or by repeated application of force. Our results are the first analysis of the mechanical properties of an animal virus, and demonstrate a linkage between virus morphology and mechanical properties. PMID:16632508

  18. Resistance to cyclopentenylcytosine in murine leukemia L1210 cells.

    PubMed

    Zhang, H; Cooney, D A; Zhang, M H; Ahluwalia, G; Ford, H; Johns, D G

    1993-12-01

    Cyclopentenyl cytosine (CPEC) exhibits oncological activity in murine and human tumor cells and has now entered Phase I clinical trials. Its mode of action as an antitumor agent appears to be inhibition by its triphosphate (CPEC-TP) of CTP synthase, the enzyme which converts UTP to CTP. In an attempt to elucidate the mechanism of resistance to CPEC, a murine leukemia cell line resistant to CPEC (L1210/CPEC) was developed by N-methyl-N-nitro-N-nitrosoguanidine-induced mutagenesis and subsequent selection by cultivation of the L1210 cells in the presence of 2 microM CPEC. Resistant clones were maintained in CPEC-free medium for 6 generations before biochemical studies were performed. The resistant clone selected for further studies was approximately 13,000-fold less sensitive to growth inhibition by CPEC than the parental cells, and the concentration of CPEC required to deplete CTP in the resistant cells was 50-fold higher than in the sensitive cells. A comparison of the kinetic properties of CTP synthase from sensitive and resistant cells indicated alteration in the properties of the enzyme from the latter; the median inhibitory concentration for CPEC-TP increased from 2 to 14 microM, Km for UTP decreased from 126 to 50 microM, and Vmax increased 12-fold from 0.2 to 2.3 nmol/mg/min. Northern blot analyses of polyadenylated RNA from the resistant and sensitive cells indicated a 3-fold increase in transcripts of the CTP synthase gene in the resistant line. Consistent with these alterations in the properties of the enzyme, the resistant cells exhibited significantly expanded CTP and dCTP pools (4- 5-fold) when compared with the sensitive cells. No change was observed, however, in the properties of uridine-cytidine kinase, the enzyme responsible for the initial phosphorylation of CPEC; despite this, however, cellular uptake of CPEC was greatly decreased, and phosphorylation of CPEC and its incorporation into RNA were 10-fold less than in the parental cells. These latter

  19. Leukemia induction by a new strain of Friend mink cell focus-inducing virus: synergistic effect of Friend ecotropic murine leukemia virus.

    PubMed Central

    Chesebro, B; Wehrly, K; Nishio, J; Evans, L

    1984-01-01

    A new strain of Friend recombinant mink cell focus-inducing retrovirus, FMCF -1-E, was found to induce leukemias in NFS and IRW mice. Although the isolate was obtained from a stock of FMCF -1 ( Troxler et al., J. Exp. Med. 148:639-653, 1978), FMCF -1-E was distinguishable from FMCF -1 by oligonucleotide fingerprinting and antigenic analysis, using monoclonal antibodies. These analyses suggested that FMCF -1-E is a distinct FMCF isolate rather than a simple variant of FMCF -1. After neonatal inoculation, the latency for leukemia induction was 3 to 8 months. A similar long latency was also seen when Friend murine leukemia virus 57 was inoculated into adult (6-week-old) IRW mice. However, sequential inoculation of FMCF -1-E at birth followed by Friend murine leukemia 57 at 6 weeks of age led to a shortened latency period (2.5 to 4 months). Only neonatal inoculation of Friend murine leukemia virus 57 was able to induce a more rapid appearance of leukemia. The leukemia cell type in the majority of cases, regardless of virus inoculation protocol, was erythroid, but occasional myeloid, lymphoid, and mixed leukemias were also observed. In contrast to NFS and IRW mice, BALB/c mice were resistant to leukemia induction by FMCF -1-E and also showed some transient resistance to leukemia induction by Friend murine leukemia virus 57. Images PMID:6202886

  20. Unstable resistance of G mouse fibroblasts to ecotropic murine leukemia virus infection.

    PubMed Central

    Yoshikura, H; Naito, Y; Moriwaki, K

    1979-01-01

    G mouse cells were resistant to N- and NB-tropic Friend leukemia viruses and to B-tropic WN 1802B. Though the cells were resistant to focus formation by the Moloney isolate of murine sarcoma virus, they were relatively sensitive to helper component murine leukemia virus. To amphotropic murine leukemia virus and to focus formation by amphotropic murine sarcoma virus, G mouse cells were fully permissive. When the cell lines were established starting from the individual embryos, most cell lines were not resistant to the murine leukemia viruses. Only one resistant line was established. Cloning of this cell line indicated that the resistant cells constantly segregated sensitive cells during the culture; i.e., the G mouse cell cultures were probably always mixtures of sensitive and resistant cells. Among the sensitive cell clones, some were devoid of Fv-1 restriction. Such dually permissive cells, and also feral mouse-derived SC-1 cells, retained glucose-6-phosphate dehydrogenase-1 and apparently normal number 4 chromosomes. The loss of Fv-1 restriction in these mouse cells was not brought about by any gross structural changes in the vicinity of Fv-1 on number 4 chromosomes. Images PMID:221667

  1. The target cell of transformation is distinct from the leukemia stem cell in murine CALM/AF10 leukemia models.

    PubMed

    Dutta, S; Krause, A; Vosberg, S; Herold, T; Ksienzyk, B; Quintanilla-Martinez, L; Tizazu, B; Chopra, M; Graf, A; Krebs, S; Blum, H; Greif, P A; Vetter, A; Metzeler, K; Rothenberg-Thurley, M; Schneider, M R; Dahlhoff, M; Spiekermann, K; Zimber-Strobl, U; Wolf, E; Bohlander, S K

    2016-05-01

    The CALM/AF10 fusion gene is found in various hematological malignancies including acute myeloid leukemia (AML), T-cell acute lymphoblastic leukemia and malignant lymphoma. We have previously identified the leukemia stem cell (LSC) in a CALM/AF10-driven murine bone marrow transplant AML model as B220+ lymphoid cells with B-cell characteristics. To identify the target cell for leukemic transformation or 'cell of origin of leukemia' (COL) in non-disturbed steady-state hematopoiesis, we inserted the CALM/AF10 fusion gene preceded by a loxP-flanked transcriptional stop cassette into the Rosa26 locus. Vav-Cre-induced panhematopoietic expression of the CALM/AF10 fusion gene led to acute leukemia with a median latency of 12 months. Mice expressing CALM/AF10 in the B-lymphoid compartment using Mb1-Cre or CD19-Cre inducer lines did not develop leukemia. Leukemias had a predominantly myeloid phenotype but showed coexpression of the B-cell marker B220, and had clonal B-cell receptor rearrangements. Using whole-exome sequencing, we identified an average of two to three additional mutations per leukemia, including activating mutations in known oncogenes such as FLT3 and PTPN11. Our results show that the COL for CALM/AF10 leukemia is a stem or early progenitor cell and not a cell of B-cell lineage with a phenotype similar to that of the LSC in CALM/AF10+ leukemia. PMID:26686248

  2. A Multicenter Blinded Analysis Indicates No Association between Chronic Fatigue Syndrome/Myalgic Encephalomyelitis and either Xenotropic Murine Leukemia Virus-Related Virus or Polytropic Murine Leukemia Virus

    PubMed Central

    Alter, Harvey J.; Mikovits, Judy A.; Switzer, William M.; Ruscetti, Francis W.; Lo, Shyh-Ching; Klimas, Nancy; Komaroff, Anthony L.; Montoya, Jose G.; Bateman, Lucinda; Levine, Susan; Peterson, Daniel; Levin, Bruce; Hanson, Maureen R.; Genfi, Afia; Bhat, Meera; Zheng, HaoQiang; Wang, Richard; Li, Bingjie; Hung, Guo-Chiuan; Lee, Li Ling; Sameroff, Stephen; Heneine, Walid; Coffin, John; Hornig, Mady; Lipkin, W. Ian

    2012-01-01

    ABSTRACT The disabling disorder known as chronic fatigue syndrome or myalgic encephalomyelitis (CFS/ME) has been linked in two independent studies to infection with xenotropic murine leukemia virus-related virus (XMRV) and polytropic murine leukemia virus (pMLV). Although the associations were not confirmed in subsequent studies by other investigators, patients continue to question the consensus of the scientific community in rejecting the validity of the association. Here we report blinded analysis of peripheral blood from a rigorously characterized, geographically diverse population of 147 patients with CFS/ME and 146 healthy subjects by the investigators describing the original association. This analysis reveals no evidence of either XMRV or pMLV infection. PMID:22991430

  3. Sox4 cooperates with PU.1 haploinsufficiency in murine myeloid leukemia

    PubMed Central

    Aue, Georg; Du, Yang; Cleveland, Susan M.; Smith, Stephen B.; Davé, Utpal P.; Liu, Delong; Weniger, Marc A.; Metais, Jean Yves; Jenkins, Nancy A.; Copeland, Neal G.

    2011-01-01

    Cooperation of multiple mutations is thought to be required for cancer development. In previous studies, murine myeloid leukemias induced by transducing wild-type bone marrow progenitors with a SRY sex determining region Y-box 4 (Sox4)–expressing retrovirus frequently carried proviral insertions at Sfpi1, decreasing its mRNA levels, suggesting that reduced Sfpi1 expression cooperates with Sox4 in myeloid leukemia induction. In support of this hypothesis, we show here that mice receiving Sox4 virus-infected Sfpi1ko/+ bone marrow progenitors developed myeloid leukemia with increased penetrance and shortened latency. Interestingly, Sox4 expression further decreased Sfpi1 transcription. Ectopic SOX4 expression reduced endogenous PU.1 mRNA levels in HL60 promyelocytes, and decreased Sfpi1 mRNA levels were also observed in the spleens of leukemic and preleukemic mice receiving Sox4 virus-infected wild-type bone marrow cells. In addition, Sox4 protein bound to a critical upstream regulatory element of Sfpi1 in ChIP assays. Such cooperation probably occurs in de novo human acute myeloid leukemias, as an analysis of 285 acute myeloid leukemia patient samples found a significant negative correlation between SOX4 and PU.1 expression. Our results establish a novel cooperation between Sox4 and reduced Sfpi1 expression in myeloid leukemia development and suggest that SOX4 could be an important new therapeutic target in human acute myeloid leukemia. PMID:21878674

  4. Anti-CD45 radioimmunotherapy using 211At with bone marrow transplantation prolongs survival in a disseminated murine leukemia model

    SciTech Connect

    Orozco, Johnnie J.; Back, Tom; Kenoyer, Aimee L.; Balkin, Ethan R.; Hamlin, Donald K.; Wilbur, D. Scott; Fisher, Darrell R.; Frayo, Shani; Hylarides, Mark; Green, Damian J.; Gopal, Ajay K.; Press, Oliver W.; Pagel, John M.

    2013-05-15

    Anti-CD45 Radioimmunotherapy using an Alpha-Emitting Radionuclide 211At Combined with Bone Marrow Transplantation Prolongs Survival in a Disseminated Murine Leukemia Model ABSTRACT Despite aggressive chemotherapy combined with hematopoietic cell transplant (HCT), many patients with acute myeloid leukemia (AML) relapse. Radioimmunotherapy (RIT) using antibodies (Ab) labeled primarily with beta-emitting radionuclides has been explored to reduce relapse.

  5. Biochemical analysis of murine leukemia viruses isolated from radiation-induced leukemias of strain BALB/c

    SciTech Connect

    Ellis, R.W.; Hopkins, N.; Fleissner, E.

    1980-02-01

    Murine leukemia viruses isolated from radiation-induced BALB/c leukemias were characterized with respect to viral proteins and RNA. Analysis by sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the viral structural proteins revealed that for p12, p15, p30, and gp70, three to four electrophoretic variants of each could be detected. There was no correlation found between any of these mobilities and N- or B-tropism of the viruses. Proteins of all xenotropic viral isolates were identical in their gel electrophoretic profiles. The similar phenotypes of multiple viral clones from individual leukemias and of isolates grown in different cells suggest that the polymorphism of ecotropic viruses was generated in vivo rather than during in vitro virus growth. By two-dimensional fingerprinting of RNase T1-resistant oligonucleotides from 70S viral RNA, the previously reported association of N- and B-tropism with two distinct oligonucleotides was confirmed. The presence of two other oligonucleotides was correlated with positive and negative phenotypes of the virus-coded G/sub IX/ cell surface antigen. The RNAs of two B-tropic isolates with distinctive p15 and p12 phenotypes differed from the RNA of a prototype N-tropic virus by the absence of three oligonucleotides mapping in the 5' portion (gag region) of the prototype RNA. In addition, one small-plaque B-tropic virus displayed extensive changes in the RNA sequences associated with the env region of the prototype.

  6. Isolation of Naturally Occurring Viruses of the Murine Leukemia Virus Group in Tissue Culture

    PubMed Central

    Hartley, Janet W.; Rowe, Wallace P.; Capps, Worth I.; Huebner, Robert J.

    1969-01-01

    A tissue culture cell system for isolation and identification of members of the murine leukemia virus group (the complement fixation for murine leukemia test) was modified to permit the isolation of naturally occurring virus from leukemic and normal mice. The important factors for increasing the sensitivity of the test were the use of National Institutes of Health (NIH) strain Webster Swiss embryo cell cultures and the selection of rat-immune sera having complement-fixing antibodies to tissue culture antigens of both the Gross and FMR subgroups. In all, 163 strains of mouse leukemia virus, from 11 inbred mouse strains, have been isolated. Representative virus isolates were shown to possess the properties of the murine leukemia virus group; i.e., they were chloroform-sensitive, noncytopathic agents which replicated in mouse embryo tissue culture and produced group-reactive, complement-fixing antigen and budding C-type particles visible by electron microscopy. These viruses could serve as helpers in the rescue of Moloney sarcoma virus genome from non-producer hamster sarcoma cells, yielding pseudotypes. All of the 19 field isolates tested were neutralized by Gross passage A antiserum but not by potent antisera to the Moloney, Rauscher, and Friend strains. Virus was recovered regularly from embryos and from the plasma and spleen of adult mice of high leukemic strains. In low leukemic mouse strains, different patterns of virus detection were observed. In C3H/He mice, virus was occasionally present in embryos and was found in 40% of adult spleens. BALB/c mice were virus-negative as fetuses or weanlings, but spleens of more than half of the mice over 6 months of age yielded virus. NIH mice have never yielded virus. In reciprocal matings between AKR and BALB/c mice, virus recovery from embryos was maternally determined. The development of tissue culture isolation procedures made possible for the first time the application of classical infectious disease methods to the

  7. Anti-CD45 radioimmunotherapy using 211At with bone marrow transplantation prolongs survival in a disseminated murine leukemia model

    PubMed Central

    Orozco, Johnnie J.; Bäck, Tom; Kenoyer, Aimee; Balkin, Ethan R.; Hamlin, Donald K.; Wilbur, D. Scott; Fisher, Darrell R.; Frayo, Shani L.; Hylarides, Mark D.; Green, Damian J.; Gopal, Ajay K.; Press, Oliver W.

    2013-01-01

    Despite aggressive chemotherapy combined with hematopoietic stem cell transplantation (HSCT), many patients with acute myeloid leukemia (AML) relapse. Radioimmunotherapy (RIT) using monoclonal antibodies labeled with β-emitting radionuclides has been explored to reduce relapse. β emitters are limited by lower energies and nonspecific cytotoxicity from longer path lengths compared with α emitters such as 211At, which has a higher energy profile and shorter path length. We evaluated the efficacy and toxicity of anti-CD45 RIT using 211At in a disseminated murine AML model. Biodistribution studies in leukemic SJL/J mice showed excellent localization of 211At-anti-murine CD45 mAb (30F11) to marrow and spleen within 24 hours (18% and 79% injected dose per gram of tissue [ID/g], respectively), with lower kidney and lung uptake (8.4% and 14% ID/g, respectively). In syngeneic HSCT studies, 211At-B10-30F11 RIT improved the median survival of leukemic mice in a dose-dependent fashion (123, 101, 61, and 37 days given 24, 20, 12, and 0 µCi, respectively). This approach had minimal toxicity with nadir white blood cell counts >2.7 K/µL 2 weeks after HSCT and recovery by 4 weeks. These data suggest that 211At-anti-CD45 RIT in conjunction with HSCT may be a promising therapeutic option for AML. PMID:23471305

  8. Anti-CD45 radioimmunotherapy using (211)At with bone marrow transplantation prolongs survival in a disseminated murine leukemia model.

    PubMed

    Orozco, Johnnie J; Bäck, Tom; Kenoyer, Aimee; Balkin, Ethan R; Hamlin, Donald K; Wilbur, D Scott; Fisher, Darrell R; Frayo, Shani L; Hylarides, Mark D; Green, Damian J; Gopal, Ajay K; Press, Oliver W; Pagel, John M

    2013-05-01

    Despite aggressive chemotherapy combined with hematopoietic stem cell transplantation (HSCT), many patients with acute myeloid leukemia (AML) relapse. Radioimmunotherapy (RIT) using monoclonal antibodies labeled with β-emitting radionuclides has been explored to reduce relapse. β emitters are limited by lower energies and nonspecific cytotoxicity from longer path lengths compared with α emitters such as (211)At, which has a higher energy profile and shorter path length. We evaluated the efficacy and toxicity of anti-CD45 RIT using (211)At in a disseminated murine AML model. Biodistribution studies in leukemic SJL/J mice showed excellent localization of (211)At-anti-murine CD45 mAb (30F11) to marrow and spleen within 24 hours (18% and 79% injected dose per gram of tissue [ID/g], respectively), with lower kidney and lung uptake (8.4% and 14% ID/g, respectively). In syngeneic HSCT studies, (211)At-B10-30F11 RIT improved the median survival of leukemic mice in a dose-dependent fashion (123, 101, 61, and 37 days given 24, 20, 12, and 0 µCi, respectively). This approach had minimal toxicity with nadir white blood cell counts >2.7 K/µL 2 weeks after HSCT and recovery by 4 weeks. These data suggest that (211)At-anti-CD45 RIT in conjunction with HSCT may be a promising therapeutic option for AML. PMID:23471305

  9. Targeting of a Nuclease to Murine Leukemia Virus Capsids Inhibits Viral Multiplication

    NASA Astrophysics Data System (ADS)

    Natsoulis, Georges; Seshaiah, Partha; Federspiel, Mark J.; Rein, Alan; Hughes, Stephen H.; Boeke, Jef D.

    1995-01-01

    Capsid-targeted viral inactivation is an antiviral strategy in which toxic fusion proteins are targeted to virions, where they inhibit viral multiplication by destroying viral components. These fusion proteins consist of a virion structural protein moiety and an enzymatic moiety such as a nuclease. Such fusion proteins can severely inhibit transposition of yeast retrotransposon Ty1, an element whose transposition mechanistically resembles retroviral multiplication. We demonstrate that expression of a murine retrovirus capsid-staphylococcal nuclease fusion protein inhibits multiplication of the corresponding murine leukemia virus by 30- to 100-fold. Staphylococcal nuclease is apparently inactive intracellularly and hence nontoxic to the host cell, but it is active extracellularly because of its requirement for high concentrations of Ca2+ ions. Virions assembled in and shed from cells expressing the fusion protein contain very small amounts of intact viral RNA, as would be predicted for nuclease-mediated inhibition of viral multiplication.

  10. Genomic complexities of murine leukemia and sarcoma, reticuloendotheliosis, and visna viruses.

    PubMed Central

    Beemon, K L; Faras, A J; Hasse, A T; Duesberg, P H; Maisel, J E

    1976-01-01

    The genetic complexities of several ribodeoxyviruses were measured by quantitative analysis of unique RNase T1-resistant oligonucleotides from 60-70S viral RNAs. Moloney murine leukemia virus was found to have an RNA complexity of 3.5 x 10(6) daltons, whereas Moloney murine sarcoma virus had a significantly smaller genome size of 2.3 x 10(6). Reticuleondotheliosis and visna virus RNAs had complexities of 3.9 x 10(6), respectively. Analysis of RNase A-resistant oligonucleotides of Rous sarcoma virus RNA gave a complexity of 3.6 x 10(6), similar to that previously obtained with RNase T1-resistant oligonucleotides. Since each of these viruses was found to have a unique sequence genomic complexity near the molecular weight of a single 30-40S viral RNA subunit, it was concluded that ribodeoxyvirus genomes are at least largely polyploid. Images PMID:176429

  11. Efficacy of echinocandins against murine infections by Diutina (Candida) rugosa.

    PubMed

    Sanchis, Marta; Sutton, Deanna A; Wiederhold, Nathan P; Guarro, Josep; Capilla, Javier

    2016-09-01

    Echinocandins are recommended as a first-line therapy for invasive candidiasis. Candida rugosa was recently transferred to the new genus Diutina. We have determined the in vitro killing kinetics of two echinocandins, anidulafungin, and caspofungin and their in vivo efficacy, administering doses of 5 or 10 mg/kg, and 1 or 5 mg/kg, respectively against 2 clinical strains of D. rugosa. Both drugs showed a fungicidal concentration-dependent activity and, in a neutropenic murine model of disseminated infection, were able to reduce tissue burden and to prolong survival of mice. These results suggest that both echinocandins could be useful to treat infections by this fungus when isolates show minimal inhibitory concentrations within the range of susceptibility for both drugs. PMID:27342787

  12. Histological and In Vivo Microscopic Analysis of the Bone Marrow Microenvironment in a Murine Model of Chronic Myelogenous Leukemia.

    PubMed

    Weissenberger, Eva S; Krause, Daniela S

    2016-01-01

    Imaging of the leukemic bone marrow microenvironment, also called the leukemic bone marrow niche, is an essential method to determine and to evaluate the progression of chronic myelogenous leukemia (CML) and other leukemias in murine models. In this chapter we introduce the murine model of CML primarily used in our laboratory by describing blood and bone marrow analysis as well as the method of histological sectioning and immunohistochemistry in combination with various stainings that can help to understand the complex interaction between leukemic cells, their normal hematopoietic counterparts, and the bone marrow microenvironment. We conclude with describing how to image the bone marrow niche using in vivo microscopy. PMID:27581139

  13. Typhonium flagelliforme inhibits the proliferation of murine leukemia WEHI-3 cells in vitro and induces apoptosis in vivo.

    PubMed

    Mohan, Syam; Abdul, Ahmad Bustamam; Abdelwahab, Siddig Ibrahim; Al-Zubairi, Adel S; Aspollah Sukari, Mohamed; Abdullah, Rasedee; Taha, Manal Mohamed Elhassan; Beng, Ng Kuan; Isa, Nurbaity Mohd

    2010-11-01

    Typhonium flagelliforme (TF) is a tropical plant, traditionally used by the ethnic population of Malaysia for the cure of various cancers. This plant had shown to induce antiproliferative effect as well as apoptosis in cancer cells. However, there is no available information to address that TF affects murine leukemia cells in vitro and in vivo. Here, we investigated in vitro and in vivo effects of TF on murine leukemia WEHI-3 cells. It was found that the growth of leukemia cells in vitro was inhibited by the various extracts of TF. Among these fractions, the dichloromethane (DCM) tuber extracts of TF showed the lowest IC(50) (24.0 ± 5.2 μg/ml) and had demonstrated apoptogenic effect when observed under fluorescent microscope. We investigated the in vivo effects of DCM tuber extracts of TF on murine leukemia cells, and the results showed that the counts of immature granulocytes and monocytes were significantly decreased in peripheral blood of BALB/c leukemia mice after the oral administration of DCM tuber extracts of TF for 28 days with three doses (200, 400 and 800 mg/kg). These results were confirmed by observing the spleen histopathology and morphology of enlarged spleen and liver in leukemia mice when compared with the control. Furthermore, the cell death mechanism in the spleen tissue of treated mice was found via apoptosis. PMID:20569984

  14. Antileukemic effect of zerumbone-loaded nanostructured lipid carrier in WEHI-3B cell-induced murine leukemia model

    PubMed Central

    Rahman, Heshu Sulaiman; Rasedee, Abdullah; How, Chee Wun; Zeenathul, Nazariah Allaudin; Chartrand, Max Stanley; Yeap, Swee Keong; Abdul, Ahmad Bustamam; Tan, Sheau Wei; Othman, Hemn Hassan; Ajdari, Zahra; Namvar, Farideh; Arulselvan, Palanisamy; Fakurazi, Sharida; Mehrbod, Parvaneh; Daneshvar, Nasibeh; Begum, Hasina

    2015-01-01

    Cancer nanotherapy is progressing rapidly with the introduction of many innovative drug delivery systems to replace conventional therapy. Although the antitumor activity of zerumbone (ZER) has been reported, there has been no information available on the effect of ZER-loaded nanostructured lipid carrier (NLC) (ZER-NLC) on murine leukemia cells. In this study, the in vitro and in vivo effects of ZER-NLC on murine leukemia induced with WEHI-3B cells were investigated. The results from 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide, Hoechst 33342, Annexin V, cell cycle, and caspase activity assays showed that the growth of leukemia cells in vitro was inhibited by ZER-NLC. In addition, outcomes of histopathology, transmission electron microscopy, and Tdt-mediated dUTP nick-end labeling analyses revealed that the number of leukemia cells in the spleen of BALB/c leukemia mice significantly decreased after 4 weeks of oral treatment with various doses of ZER-NLC. Western blotting and reverse-transcription quantitative polymerase chain reaction assays confirmed the antileukemia effects of ZER-NLC. In conclusion, ZER-NLC was shown to induce a mitochondrial-dependent apoptotic pathway in murine leukemia. Loading of ZER in NLC did not compromise the anticancer effect of the compound, suggesting ZER-NLC as a promising and effective delivery system for treatment of cancers. PMID:25767386

  15. Receptor choice determinants in the envelope glycoproteins of amphotropic, xenotropic, and polytropic murine leukemia viruses.

    PubMed Central

    Battini, J L; Heard, J M; Danos, O

    1992-01-01

    The envelope glycoproteins (SU) of mammalian type C retroviruses possess an amino-terminal domain of about 200 residues, which is involved in binding a cell surface receptor. In this domain, highly conserved amino acid sequences are interrupted by two segments of variable length and sequence, VRA and VRB. We have studied the role of these variable regions in receptor recognition and binding by constructing chimeric molecules in which portions of the amino-terminal domains from amphotropic (4070A), xenotropic (NZB), and polytropic (MCF 247) murine leukemia virus SU proteins were permuted. These chimeras, which exchanged either one or two variable regions, were expressed at the surface of replication-defective viral particles by a pseudotyping assay. Wild-type or recombinant env genes were transfected into a cell line producing Moloney murine leukemia virus particles devoid of envelope glycoproteins in which a retrovirus vector genome carrying an Escherichia coli lacZ gene was packaged. The host range and sensitivity to interference of pseudotyped virions were assayed, and we observed which permutations resulted in receptor switch or loss of function. Our results indicate that the determinants of receptor choice are found within the just 120 amino acids of SU proteins. Downstream sequences contribute to the stabilization of the receptor-specific structure. PMID:1310758

  16. Viral genome RNA serves as messenger early in the infectious cycle of murine leukemia virus.

    PubMed Central

    Shurtz, R; Dolev, S; Aboud, M; Salzberg, S

    1979-01-01

    When NIH/3T3 mouse fibroblasts were infected with the Moloney strain of murine leukemia virus, part of the viral genome RNA molecules were detected in polyribosomes of the infected cells early in the infectious cycle. The binding appears to be specific, since we could demonstrate the release of viral RNA from polyribosomes with EDTA. Moreover, when infection occurred in the presence of cycloheximide, most viral RNA molecules were detected in the free cytoplasm. Size analysis on polyribosomal viral RNA molecules indicated that two size class molecules, 38S and 23S, are present in polyribosomes at 3 h after infection. Analysis of the polyriboadenylate [poly(rA)] content of viral RNA extracted from infected polyribosomes demonstrated that such molecules bind with greatest abundance at 3 h after infection, as has been detected with total viral RNA. No molecules lacking poly(rA) stretches could be detected in polyribosomes. Furthermore, when a similar analysis was performed on unbound molecules present in the free cytoplasm, identical results were obtained. We conclude that no selection towards poly(rA)-containing viral molecules is evident on binding to polyribosomes. These findings suggest that the incoming viral genome of the Moloney strain of murine leukemia virus may serve as a messenger for the synthesis of one or more virus-specific proteins early after infection of mouse fibroblasts. PMID:117118

  17. BCL1, a murine model of prolymphocytic leukemia. I. Effect of splenectomy on growth kinetics and organ distribution.

    PubMed Central

    Muirhead, M. J.; Isakson, P. C.; Krolick, K. A.; Uhr, J. W.; Vitetta, E. S.

    1981-01-01

    BCL1 is a transplantable murine B-cell leukemia that closely resembles human prolymphocytic leukemia (PLL). Syngeneic mice injected with BCL1 cells develop massively enlarged spleens followed by leukemia. Splenectomy performed either prior to BCL1 transplantation or prior to the leukemic phase of transplanted BCL1 results in a markedly altered clinical syndrome: the onset of leukemia is delayed by about 2 months; the leukemia is low-grade; and the lymph nodes, which are not prominently involved in leukemic animals with intact spleens, are massively infiltrated in the splenectomized transplant recipient. The immunologic phenotype of the BCL1 cell is not altered by splenectomy and thus does not appear to account for the altered tissue distribution of BCL1 in the splenectomized host. However, the results indicate a striking dependence of BCL1 on microenvironmental influences of the host lymphoid tissues. Images Figure 1 Figure 2 PMID:7032308

  18. p27kip1 Maintains a Subset of Leukemia Stem Cells in the Quiescent State in Murine MLL-Leukemia

    PubMed Central

    Zhang, Jun; Seet, Christopher; Sun, Clare; Li, Jing; You, Dewen; Volk, Andrew; Breslin, Peter; Li, Xingyu; Wei, Wei; Qian, Zhijian; Zeleznik-Le, Nancy J.; Zhang, Zhou; Zhang, Jiwang

    2013-01-01

    MLL (mixed-lineage leukemia)-fusion genes induce the development of leukemia through deregulation of normal MLL target genes, such as HOXA9 and MEIS1. Both HOXA9 and MEIS1 are required for MLL-fusion gene-induced leukemogenesis. Co-expression of HOXA9 and MEIS1 induces acute myeloid leukemia (AML) similar to that seen in mice in which MLL-fusion genes are over-expressed. p27kip1 (p27 hereafter), a negative regulator of the cell cycle, has also been defined as an MLL target, the expression of which is up-regulated in MLL leukemic cells (LCs). To investigate whether p27 plays a role in the pathogenesis of MLL-leukemia, we examined the effects of p27 deletion (p27-/-) on MLL-AF9 (MA9)-induced murine AML development. HOXA9/MEIS1 (H/M)-induced, p27 wild-type (p27+/+) and p27-/- AML were studied in parallel as controls. We found that LCs from both MA9-AML and H/M-AML can be separated into three fractions, a CD117-CD11bhi differentiated fraction as well as CD117+CD11bhi and CD117+CD11blo, two less differentiated fractions. The CD117+CD11blo fraction, comprising only 1-3% of total LCs, expresses higher levels of early hematopoietic progenitor markers but lower levels of mature myeloid cell markers compared to other populations of LCs. p27 is expressed and is required for maintaining the quiescent and drug-resistant states of the CD117+CD11blo fraction of MA9-LCs but not of H/M-LCs. p27 deletion significantly compromises the leukemogenic capacity of CD117+CD11blo MA9-LCs by reducing the frequency of leukemic stem cells (LSCs) but does not do so in H/M-LCs. In addition, we found that p27 is highly expressed and required for cell cycle arrest in the CD117-CD11bhi fraction in both types of LCs. Furthermore, we found that c-Myc expression is required for maintaining LCs in an undifferentiated state independently of proliferation. We concluded that p27 represses the proliferation of LCs, which is specifically required for maintaining the quiescent and drug-resistant states of a

  19. Leukemia

    MedlinePlus

    ... version of this page please turn Javascript on. Leukemia What Is Leukemia? Leukemia is a cancer of the blood cells. ... diagnosed with leukemia are over 50 years old. Leukemia Starts in Bone Marrow Click for more information ...

  20. Preclinical activity of the novel B-cell-specific Moloney murine leukemia virus integration site 1 inhibitor PTC-209 in acute myeloid leukemia: Implications for leukemia therapy.

    PubMed

    Nishida, Yuki; Maeda, Aya; Chachad, Dhruv; Ishizawa, Jo; Qiu, Yi Hua; Kornblau, Steven M; Kimura, Shinya; Andreeff, Michael; Kojima, Kensuke

    2015-12-01

    Curing patients with acute myeloid leukemia (AML) remains a therapeutic challenge. The polycomb complex protein B-cell-specific Moloney murine leukemia virus integration site 1 (BMI-1) is required for the self-renewal and maintenance of leukemia stem cells. We investigated the prognostic significance of BMI-1 in AML and the effects of a novel small molecule selective inhibitor of BMI-1, PTC-209. BMI-1 protein expression was determined in 511 newly diagnosed AML patients together with 207 other proteins using reverse-phase protein array technology. Patients with unfavorable cytogenetics according to Southwest Oncology Group criteria had higher levels of BMI-1 compared to those with favorable (P = 0.0006) or intermediate cytogenetics (P = 0.0061), and patients with higher levels of BMI-1 had worse overall survival (55.3 weeks vs. 42.8 weeks, P = 0.046). Treatment with PTC-209 reduced protein level of BMI-1 and its downstream target mono-ubiquitinated histone H2A and triggered several molecular events consistent with the induction of apoptosis, this is, loss of mitochondrial membrane potential, caspase-3 cleavage, BAX activation, and phosphatidylserine externalization. PTC-209 induced apoptosis in patient-derived CD34(+)CD38(low/-) AML cells and, less prominently, in CD34(-) differentiated AML cells. BMI-1 reduction by PTC-209 directly correlated with apoptosis induction in CD34(+) primary AML cells (r = 0.71, P = 0.022). However, basal BMI-1 expression was not a determinant of AML sensitivity. BMI-1 inhibition, which targets a primitive AML cell population, might offer a novel therapeutic strategy for AML. PMID:26450753

  1. Removal of xenotropic murine leukemia virus by nanocellulose based filter paper.

    PubMed

    Asper, M; Hanrieder, T; Quellmalz, A; Mihranyan, A

    2015-11-01

    The removal of xenotrpic murine leukemia virus (xMuLV) by size-exclusion filter paper composed of 100% naturally derived cellulose was validated. The filter paper was produced using cellulose nanofibers derived from Cladophora sp. algae. The filter paper was characterized using atomic force microscopy, scanning electron microscopy, helium pycnometry, and model tracer (100 nm latex beads and 50 nm gold nanoparticles) retention tests. Following the filtration of xMuLV spiked solutions, LRV ≥5.25 log10 TCID50 was observed, as limited by the virus titre in the feed solution and sensitivity of the tissue infectivity test. The results of the validation study suggest that the nanocellulose filter paper is useful for removal of endogenous rodent retroviruses and retrovirus-like particles during the production of recombinant proteins. PMID:26328471

  2. Purification of the Moloney and Rauscher Murine Leukemia Viruses by Use of Zonal Ultracentrifuge Systems

    PubMed Central

    Toplin, I.

    1967-01-01

    The B-IV and B-IX zonal ultracentrifuge rotors were applied to the concentration and purification of the Moloney and Rauscher murine leukemia viruses from large volumes of infected tissue culture fluids and animal materials. Potassium tartrate, potassium citrate and sucrose gradients were used to obtain viral concentrates from the density 1.16 to 1.18 zone. Proteolytic enzyme digestion of tissue culture preparations prior to zonal ultracentrifuge processing was effective in releasing virus from cell debris and producing highly purified, though nonleukemogenic, viral concentrates. Infected Rauscher mouse plasma was processed to give highly purified infectious virus fractions. A single centrifugation of crude Rauscher mouse spleen homogenates resulted in partially purified infectious concentrates with high virus particle counts. Images Fig. 4 PMID:6035050

  3. A flavone derivative from Sesbania sesban leaves and its cytotoxicity against murine leukemia P-388 cells

    NASA Astrophysics Data System (ADS)

    Dianhar, Hanhan; Syah, Yana Maolana; Mujahidin, Didin; Hakim, Euis Holisotan; Juliawaty, Lia Dewi

    2014-03-01

    Sesbania sesban, locally named as Jayanti, is one of Indonesia plants belonging to Fabaceae family. This species is traditionally used by Indonesian people to cure digestive disorders, fever, or headache. Jayanti can grow well in tropical to subtropical region, such as in Asia and Africa. Based on literature, qualitative analysis of the methanol extract of leaves of S. sesban showed that it contained flavonoids, alkaloids, saponins and glycosides. In addition, the activity assay of extracts of different tissues of this species showed antitumor, antimalarial, and antidiabetic activityies (leaves and seed extracts), antioxidants (flower extract), and analgesic (wood extract). Though the extracts of S. sesban parts showed interesting activities, chemical study of those extracts have not been widely reported. Therefore, the objective of this research was to isolate the secondary metabolites from methanol extract of leaves of S. sesban and to determine their cytotoxicity against murine leukemia P-388 cells. One compound has been obtained and identified as 3-hydroxy-4',7-dimethoxyflavone (1), a new isolated compound from nature. This compound was obtained through separation of methanol extract using various chromatographic techniques, such as vacuum liquid chromatography and radial chromatography. The structure elucidation of isolated compound was based on 1D NMR (1H-NMR and 13C-NMR) and 2D NMR (HMBC). The cytotoxicity of methanol extract and compound 1 against murine leukemia P-388 cells examined through MTT assay showed IC50 value of 60.04 μg/mL and 5.40 μg/mL, respectively.

  4. A flavone derivative from Sesbania sesban leaves and its cytotoxicity against murine leukemia P-388 cells

    SciTech Connect

    Dianhar, Hanhan Syah, Yana Maolana Mujahidin, Didin Hakim, Euis Holisotan Juliawaty, Lia Dewi

    2014-03-24

    Sesbania sesban, locally named as Jayanti, is one of Indonesia plants belonging to Fabaceae family. This species is traditionally used by Indonesian people to cure digestive disorders, fever, or headache. Jayanti can grow well in tropical to subtropical region, such as in Asia and Africa. Based on literature, qualitative analysis of the methanol extract of leaves of S. sesban showed that it contained flavonoids, alkaloids, saponins and glycosides. In addition, the activity assay of extracts of different tissues of this species showed antitumor, antimalarial, and antidiabetic activityies (leaves and seed extracts), antioxidants (flower extract), and analgesic (wood extract). Though the extracts of S. sesban parts showed interesting activities, chemical study of those extracts have not been widely reported. Therefore, the objective of this research was to isolate the secondary metabolites from methanol extract of leaves of S. sesban and to determine their cytotoxicity against murine leukemia P-388 cells. One compound has been obtained and identified as 3-hydroxy-4',7-dimethoxyflavone (1), a new isolated compound from nature. This compound was obtained through separation of methanol extract using various chromatographic techniques, such as vacuum liquid chromatography and radial chromatography. The structure elucidation of isolated compound was based on 1D NMR ({sup 1}H-NMR and {sup 13}C-NMR) and 2D NMR (HMBC). The cytotoxicity of methanol extract and compound 1 against murine leukemia P-388 cells examined through MTT assay showed IC{sub 50} value of 60.04 μg/mL and 5.40 μg/mL, respectively.

  5. Characterization of mouse cellular deoxyribonucleic acid homologous to Abelson murine leukemia virus-specific sequences.

    PubMed Central

    Dale, B; Ozanne, B

    1981-01-01

    The genome of Abelson murine leukemia virus (A-MuLV) consists of sequences derived from both BALB/c mouse deoxyribonucleic acid and the genome of Moloney murine leukemia virus. Using deoxyribonucleic acid linear intermediates as a source of retroviral deoxyribonucleic acid, we isolated a recombinant plasmid which contained 1.9 kilobases of the 3.5-kilobase mouse-derived sequences found in A-MuLV (A-MuLV-specific sequences). We used this clone, designated pSA-17, as a probe restriction enzyme and Southern blot analyses to examine the arrangement of homologous sequences in BALB/c deoxyribonucleic acid (endogenous Abelson sequences). The endogenous Abelson sequences within the mouse genome were interrupted by noncoding regions, suggesting that a rearrangement of the cell sequences was required to produce the sequence found in the virus. Endogenous Abelson sequences were arranged similarly in mice that were susceptible to A-MuLV tumors and in mice that were resistant to A-MuLV tumors. An examination of three BALB/c plasmacytomas and a BALB/c early B-cell tumor likewise revealed no alteration in the arrangement of the endogenous Abelson sequences. Homology to pSA-17 was also observed in deoxyribonucleic acids prepared from rat, hamster, chicken, and human cells. An isolate of A-MuLV which encoded a 160,000-dalton transforming protein (P160) contained 700 more base pairs of mouse sequences than the standard A-MuLV isolate, which encoded a 120,000-dalton transforming protein (P120). Images PMID:9279386

  6. Murine Leukemias with Retroviral Insertions at Lmo2 Are Predictive of the Leukemias Induced in SCID-X1 Patients Following Retroviral Gene Therapy

    PubMed Central

    Davé, Utpal P.; Akagi, Keiko; Tripathi, Rati; Cleveland, Susan M.; Thompson, Mary A.; Yi, Ming; Stephens, Robert; Downing, James R.; Jenkins, Nancy A.; Copeland, Neal G.

    2009-01-01

    Five X-linked severe combined immunodeficiency patients (SCID-X1) successfully treated with autologous bone marrow stem cells infected ex vivo with an IL2RG-containing retrovirus subsequently developed T-cell leukemia and four contained insertional mutations at LMO2. Genetic evidence also suggests a role for IL2RG in tumor formation, although this remains controversial. Here, we show that the genes and signaling pathways deregulated in murine leukemias with retroviral insertions at Lmo2 are similar to those deregulated in human leukemias with high LMO2 expression and are highly predictive of the leukemias induced in SCID-X1 patients. We also provide additional evidence supporting the notion that IL2RG and LMO2 cooperate in leukemia induction but are not sufficient and require additional cooperating mutations. The highly concordant nature of the genetic events giving rise to mouse and human leukemias with mutations at Lmo2 are an encouraging sign to those wanting to use mice to model human cancer and may help in designing safer methods for retroviral gene therapy. PMID:19461887

  7. Expression of murine leukemia viruses in the highly lymphomatous BXH-2 recombinant inbred mouse strain.

    PubMed Central

    Bedigian, H G; Taylor, B A; Meier, H

    1981-01-01

    Among 12 recombinant inbred strains of mice derived from crossing two strains, C57BL/6J and C3H/HeJ, which have a low incidence of neoplastic disease, one strain (BXH-2) has been found to have a high incidence of lymphoma, of non-T-cell origin, at an early age. The BXH-2 strain carries the Fv-1b allele and spontaneously expresses a B-tropic murine leukemia virus beginning at as early as 10 days of gestation and continuing throughout their life. No significant differences in ecotropic virus titers were observed at any age tested (16 to 17 days of gestation through 7 months), whereas xenotropic virus was first detected in lymphoid tissues of 2-month-old mice and virus titers increased with age. Dual tropic virus(es), which induced cytopathic changes on mink lung cells, was isolated from BXH-2 lymphomatous tissues. Unlike AKR mink lung focus-forming virus (N-tropic recombinant), BXH-2 dual tropic virus is B tropic and induces cytopathic changes in mouse fibroblast cultures as well. The BXH-2 mouse provides a model system for studying the role of replication-competent viruses in spontaneously occurring leukemias of non-T-cell lineage and neurological disease. Images PMID:6268848

  8. Effects of murine leukemia virus env gene proteins on macrophage-mediated cytotoxicity in vitro

    NASA Technical Reports Server (NTRS)

    Chapes, S. K.; Takemoto, L. J.; Spooner, B. S. (Principal Investigator)

    1991-01-01

    F5b Tumor cells were incubated with concentrated culture supernatants taken from cells resistant (F5m) or sensitive (F5b) to contact-dependent macrophage cytotoxicity. Macrophage cell line B6MP102 and murine peritoneal macrophages killed targets incubated with supernatants taken from sensitive cells but poorly killed cells incubated in supernatants isolated from resistant cells. Membranes from cells resistant to macrophage killing, F5m, were fused into F5b cells. The fused F5b cells were killed significantly less than F5b cells fused with F5b cell membranes or untreated F5b cells. The decreased killing of F5b cells corresponded to increased concentrations of gp70(a) molecules on F5b cells. Affinity purified gp70(a) was added to cytotoxicity assays but failed to inhibit macrophage cytotoxicity. P15E molecules were detectable on both F5b and F5m cells. In addition, a synthetic peptide found to exhibit the inhibitory properties of p15E was added to cytotoxicity assays. P15E synthetic peptide also did not inhibit macrophage cytotoxicity. Therefore, env gene proteins of murine leukemia virus do not appear responsible for inducing tumor cell resistance to activated macrophage contact-dependent cytotoxicity.

  9. Pretargeted Radioimmunotherapy Using Anti-CD45 Monoclonal Antibodies to Deliver Radiation to Murine Hematolymphoid Tissues and Human Myeloid Leukemia

    SciTech Connect

    Pagel, John M.; Matthews, Dana C.; Kenoyer, Aimee L.; Hamlin, Donald K.; Wilbur, D. Scott; Fisher, Darrell R.; Gopal, Ajay K.; Lin, Yukang; Saganic, Laura; Appelbaum, Frederick R.; Press, Oliver W.

    2009-01-01

    The efficacy of radioimmunotherapy (RIT) for treatment of patients with hematological malignancies frequently fails because of disease recurrence. We therefore conducted pretargeted RIT studies to augment the efficacy in mice of therapy using a pretargeted anti-human (h)CD45 antibody (Ab)-streptavidin (SA) conjugate followed by delivery of a biotinylated clearing agent and radiolabeled-DOTA-biotin. Tumor-to-blood ratios at 24 hours were 20:1 using pretargeted anti-hCD45 RIT and <1:1 with conventional RIT. In vivo imaging studies confirmed that the pretargeted RIT approach provided high-contrast tumor images with minimal blood-pool activity, whereas directly-labeled anti-hCD45 Ab produced distinct tumor images but the blood pool retained a large amount of labeled antibody for a prolonged time. Therapy experiments demonstrated that 90Y-DOTA-biotin significantly prolonged survival of mice treated pretargeted with anti-hCD45 Ab-SA compared to mice treated with conventional RIT using 90Y-labeled anti-hCD45 Ab at the maximally tolerated dose (400 µCi). Since human CD45 antigens are confined to xenograft tumor cells in this model, and all murine tissues are devoid of hCD45 and will not bind anti-hCD45 Ab, we also compared one-step and pretargeted RIT using an anti-murine (m)CD45 Ab (A20 ) in a model where the target antigen is present on normal hematopoietic tissues. After 24 hours, 27.3 ± 2.8% of the injected dose of radionuclide was delivered per gram (% ID/g) of lymph node using 131I-A20-Ab compared with 40.0 ± 5.4% ID/g for pretargeted 111In-DOTA-biotin (p value). These data suggest that multi-step pretargeted methods for delivering RIT are superior to conventional RIT when targeting CD45 for the treatment of leukemia and may allow for the intensification of therapy, while minimizing toxicities.

  10. Anti-CD45 Radioimmunotherapy with 90Y but Not 177Lu Is Effective Treatment in a Syngeneic Murine Leukemia Model

    PubMed Central

    Orozco, Johnnie J.; Balkin, Ethan R.; Gooley, Ted A.; Kenoyer, Aimee; Hamlin, Donald K.; Wilbur, D. Scott; Fisher, Darrell R.; Hylarides, Mark D.; Shadman, Mazyar; Green, Damian J.; Gopal, Ajay K.; Press, Oliver W.; Pagel, John M.

    2014-01-01

    Radioimmunotherapy (RIT) for treatment of hematologic malignancies has primarily employed monoclonal antibodies (Ab) labeled with 131I or 90Y which have limitations, and alternative radionuclides are needed to facilitate wider adoption of RIT. We therefore compared the relative therapeutic efficacy and toxicity of anti-CD45 RIT employing 90Y and 177Lu in a syngeneic, disseminated murine myeloid leukemia (B6SJLF1/J) model. Biodistribution studies showed that both 90Y- and 177Lu-anti-murine CD45 Ab conjugates (DOTA-30F11) targeted hematologic tissues, as at 24 hours 48.8±21.2 and 156±14.6% injected dose per gram of tissue (% ID/g) of 90Y-DOTA-30F11 and 54.2±9.5 and 199±11.7% ID/g of 177Lu-DOTA-30F11 accumulated in bone marrow (BM) and spleen, respectively. However, 90Y-DOTA-30F11 RIT demonstrated a dose-dependent survival benefit: 60% of mice treated with 300 µCi 90Y-DOTA-30F11 lived over 180 days after therapy, and mice treated with 100 µCi 90Y-DOTA-30F11 had a median survival 66 days. 90Y-anti-CD45 RIT was associated with transient, mild myelotoxicity without hepatic or renal toxicity. Conversely, 177Lu- anti-CD45 RIT yielded no long-term survivors. Thus, 90Y was more effective than 177Lu for anti-CD45 RIT of AML in this murine leukemia model. PMID:25460570

  11. Structural and biochemical characterization of the inhibitor complexes of xenotropic murine leukemia virus-related virus protease

    SciTech Connect

    Li, Mi; Gustchina, Alla; Matúz, Krisztina; Tözsér, Jozsef; Namwong, Sirilak; Goldfarb, Nathan E.; Dunn, Ben M.; Wlodawer, Alexander

    2012-10-23

    Interactions between the protease (PR) encoded by the xenotropic murine leukemia virus-related virus and a number of potential inhibitors have been investigated by biochemical and structural techniques. It was observed that several inhibitors used clinically against HIV PR exhibit nanomolar or even subnanomolar values of K{sub i}, depending on the exact experimental conditions. Both TL-3, a universal inhibitor of retroviral PRs, and some inhibitors originally shown to inhibit plasmepsins were also quite potent, whereas inhibition by pepstatin A was considerably weaker. Crystal structures of the complexes of xenotropic murine leukemia virus-related virus PR with TL-3, amprenavir and pepstatin A were solved at high resolution and compared with the structures of complexes of these inhibitors with other retropepsins. Whereas TL-3 and amprenavir bound in a predictable manner, spanning the substrate-binding site of the enzyme, two molecules of pepstatin A bound simultaneously in an unprecedented manner, leaving the catalytic water molecule in place.

  12. Free and integrated recombinant murine leukemia virus DNAs appear in preleukemic thymuses of AKR/J mice.

    PubMed Central

    Herr, W; Gilbert, W

    1984-01-01

    We studied the appearance and structure of murine leukemia viral genomes in preleukemic AKR/J mice by Southern hybridization. Up to an average of one to two copies per thymocyte of unintegrated murine leukemia virus DNA appears in the thymuses of preleukemic mice beginning at 4 to 5 months of age and disappears in leukemic thymuses. The free viral genomes are absent in the spleens, livers, and brains of preleukemic mice. Using a series of ecotropic and nonecotropic murine leukemia virus hybridization probes, we showed that the unintegrated viral genomes are structurally analogous to those of recombinant mink cell focus-forming viruses that appear as proviruses in leukemic AKR thymocytes, suggesting that these free viral DNAs are the direct precursors to the leukemia-specific proviruses. The mosaic of ecotropic and nonecotropic sequences within these unintegrated viral DNAs varies from one preleukemic thymus to another but often appears structurally homogeneous within individual thymuses, indicating that often each thymus was being infected by a unique mink cell focus-forming virus. Analysis of high-molecular-weight DNA shows that recombinant proviruses reside in the chromosomal DNA of thymocytes within the preleukemic thymus, with the number rising to an average of several copies per thymocyte, but we do not detect any preferred integration sites. These results suggest that, in general, before the development of thymic leukemias in AKR mice there is a massive infection by a unique mink cell focus-forming virus which then integrates into many different sites of individual thymocytes, one of which grows out to become a tumor. Images PMID:6321787

  13. NMR study of xenotropic murine leukemia virus-related virus protease in a complex with amprenavir

    SciTech Connect

    Furukawa, Ayako; Okamura, Hideyasu; Morishita, Ryo; Matsunaga, Satoko; Kobayashi, Naohiro; Ikegami, Takahisa; Kodaki, Tsutomu; Takaori-Kondo, Akifumi; Ryo, Akihide; Nagata, Takashi; Katahira, Masato

    2012-08-24

    Highlights: Black-Right-Pointing-Pointer Protease (PR) of XMR virus (XMRV) was successfully synthesized with cell-free system. Black-Right-Pointing-Pointer Interface of XMRV PR with an inhibitor, amprenavir (APV), was identified with NMR. Black-Right-Pointing-Pointer Structural heterogeneity is induced for two PR protomers in the APV:PR = 1:2 complex. Black-Right-Pointing-Pointer Structural heterogeneity is transmitted even to distant regions from the interface. Black-Right-Pointing-Pointer Long-range transmission of structural change may be utilized for drug discovery. -- Abstract: Xenotropic murine leukemia virus-related virus (XMRV) is a virus created through recombination of two murine leukemia proviruses under artificial conditions during the passage of human prostate cancer cells in athymic nude mice. The homodimeric protease (PR) of XMRV plays a critical role in the production of functional viral proteins and is a prerequisite for viral replication. We synthesized XMRV PR using the wheat germ cell-free expression system and carried out structural analysis of XMRV PR in a complex with an inhibitor, amprenavir (APV), by means of NMR. Five different combinatorially {sup 15}N-labeled samples were prepared and backbone resonance assignments were made by applying Otting's method, with which the amino acid types of the [{sup 1}H, {sup 15}N] HSQC resonances were automatically identified using the five samples (Wu et al., 2006) . A titration experiment involving APV revealed that one APV molecule binds to one XMRV PR dimer. For many residues, two distinct resonances were observed, which is thought to be due to the structural heterogeneity between the two protomers in the APV:XMRV PR = 1:2 complex. PR residues at the interface with APV have been identified on the basis of chemical shift perturbation and identification of the intermolecular NOEs by means of filtered NOE experiments. Interestingly, chemical shift heterogeneity between the two protomers of XMRV PR has

  14. Effect of Doxorubicin/Pluronic SP1049C on Tumorigenicity, Aggressiveness, DNA Methylation and Stem Cell Markers in Murine Leukemia

    PubMed Central

    Li, Shu; Kabanov, Alexander V.

    2013-01-01

    Purpose Pluronic block copolymers are potent sensitizers of multidrug resistant cancers. SP1049C, a Pluronic-based micellar formulation of doxorubicin (Dox) has completed Phase II clinical trial and demonstrated safety and efficacy in patients with advanced adenocarcinoma of the esophagus and gastroesophageal junction. This study elucidates the ability of SP1049C to deplete cancer stem cells (CSC) and decrease tumorigenicity of cancer cells in vivo. Experimental Design P388 murine leukemia ascitic tumor was grown in BDF1 mice. The animals were treated with: (a) saline, (b) Pluronics alone, (c) Dox or (d) SP1049C. The ascitic cancer cells were isolated at different passages and examined for 1) in vitro colony formation potential, 2) in vivo tumorigenicity and aggressiveness, 3) development of drug resistance and Wnt signaling activation 4) global DNA methylation profiles, and 5) expression of CSC markers. Results SP1049C treatment reduced tumor aggressiveness, in vivo tumor formation frequency and in vitro clonogenic potential of the ascitic cells compared to drug, saline and polymer controls. SP1049C also prevented overexpression of BCRP and activation of Wnt-β-catenin signaling observed with Dox alone. Moreover, SP1049C significantly altered the DNA methylation profiles of the cells. Finally, SP1049C decreased CD133+ P388 cells populations, which displayed CSC-like properties and were more tumorigenic compared to CD133− cells. Conclusions SP1049C therapy effectively suppresses the tumorigenicity and aggressiveness of P388 cells in a mouse model. This may be due to enhanced activity of SP1049C against CSC and/or altered epigenetic regulation restricting appearance of malignant cancer cell phenotype. PMID:23977261

  15. The BET family of proteins targets Moloney Murine Leukemia Virus integration near transcription start sites

    PubMed Central

    De Rijck, Jan; de Kogel, Christine; Demeulemeester, Jonas; Vets, Sofie; Ashkar, Sara El; Malani, Nirav; Bushman, Frederic D; Landuyt, Bart; Husson, Steven J.; Busschots, Katrien; Gijsbers, Rik; Debyser, Zeger

    2014-01-01

    Summary A hallmark of retroviral replication is integration of the viral genome in the host cell DNA. This characteristic makes retrovirus-based vectors attractive delivery vehicles for gene therapy. However, adverse events in gene therapeutic trials, caused by activation of proto-oncogenes due to Murine Leukemia Virus (MLV)-derived vector integration, hamper their application. Here we show that bromodomain and extraterminal (BET) proteins (BRD2, BRD3 and BRD4) and MLV integrase specifically interact and co-localize within the nucleus of the cell. Inhibition of the BET proteins chromatin interaction via specific bromodomain inhibitors blocks MLV virus replication at the integration step. MLV integration site distribution parallels the chromatin binding profile of BET proteins, and expression of an artificial fusion protein of the BET integrase binding domain with the chromatin interaction domain of the lentiviral targeting factor LEDGF/p75, retargets MLV integration away from TSS and into the body of actively transcribed genes, conform to the Human Immunodeficiency Virus (HIV) integration pattern. Together these data validate BET proteins as MLV integration targeting factors. PMID:24183673

  16. Bimodal high-affinity association of Brd4 with murine leukemia virus integrase and mononucleosomes.

    PubMed

    Larue, Ross C; Plumb, Matthew R; Crowe, Brandon L; Shkriabai, Nikoloz; Sharma, Amit; DiFiore, Julia; Malani, Nirav; Aiyer, Sriram S; Roth, Monica J; Bushman, Frederic D; Foster, Mark P; Kvaratskhelia, Mamuka

    2014-04-01

    The importance of understanding the molecular mechanisms of murine leukemia virus (MLV) integration into host chromatin is highlighted by the development of MLV-based vectors for human gene-therapy. We have recently identified BET proteins (Brd2, 3 and 4) as the main cellular binding partners of MLV integrase (IN) and demonstrated their significance for effective MLV integration at transcription start sites. Here we show that recombinant Brd4, a representative of the three BET proteins, establishes complementary high-affinity interactions with MLV IN and mononucleosomes (MNs). Brd4(1-720) but not its N- or C-terminal fragments effectively stimulate MLV IN strand transfer activities in vitro. Mass spectrometry- and NMR-based approaches have enabled us to map key interacting interfaces between the C-terminal domain of BRD4 and the C-terminal tail of MLV IN. Additionally, the N-terminal fragment of Brd4 binds to both DNA and acetylated histone peptides, allowing it to bind tightly to MNs. Comparative analyses of the distributions of various histone marks along chromatin revealed significant positive correlations between H3- and H4-acetylated histones, BET protein-binding sites and MLV-integration sites. Our findings reveal a bimodal mechanism for BET protein-mediated MLV integration into select chromatin locations. PMID:24520112

  17. Xenotropic Murine Leukemia Virus-Related Virus (XMRV) and the Safety of the Blood Supply.

    PubMed

    Johnson, Andrew D; Cohn, Claudia S

    2016-10-01

    In 2006, a new virus, xenotropic murine leukemia virus-related virus (XMRV), was discovered in a cohort of U.S. men with prostate cancer. Soon after this initial finding, XMRV was also detected in samples from patients with chronic fatigue syndrome (CFS). The blood community, which is highly sensitive to the threat of emerging infectious diseases since the HIV/AIDS crisis, recommended indefinite deferral of all blood donors with a history of CFS. As XMRV research progressed, conflicting results emerged regarding the importance of this virus in the pathophysiology of prostate cancer and/or CFS. Molecular biologists traced the development of XMRV to a recombination event in a laboratory mouse that likely occurred circa 1993. The virus was propagated via cell lines derived from a tumor present in this mouse and spread through contamination of laboratory samples. Well-controlled experiments showed that detection of XMRV was due to contaminated samples and was not a marker of or a causal factor in prostate cancer or CFS. This paper traces the development of XMRV in the prostate and CFS scientific communities and explores the effect it had on the blood community. PMID:27358491

  18. Expression of mink cell focus-forming murine leukemia virus-related transcripts in AKR mice

    SciTech Connect

    Khan, A.S.; Laigret, F.; Rodi, C.P.

    1987-03-01

    The authors used a synthetic 16-base-pair mink cell focus-forming (MCF) env-specific oligomer as radiolabeled probe to study MCF murine leukemia virus (MuLV)-related transcripts in brain, kidney, liver, spleen, and thymus tissues of AKR mice ranging from 5 weeks to 6 months (mo) of age. Tissue-specific expression of poly(A)/sup +/ RNAs was seen. In addition, all the tissues tested contained 3.0-kb messages. The transcription of these MCF-related mRNAs was independent of the presence of ecotropic and xenotropic MuLVs. In general, expression of the MCF env-related transcripts appeared to peak at 2 mo of age; these messages were barely detectable in brain, kidney, liver, and spleen tissues after 2 mo and in thymus tissue after 4 mo of age. All of the subgenomic MCF env-related mRNAs appeared to contain the 190-base-pair cellular DNA insert, characteristic of the long terminal repeats associated with endogenous MCF env-related proviruses. No genomic-size (8.4-kb) transcripts corresponding to endogenous MCF-related proviruses were detected. An 8.4-kb MCF env-related mRNA was first seen at 3 mo of age, exclusively in thymus tissue. This species most likely represents the first appearance of a recombinant MCF-related MuLV genome. The transcripts which were detected in thymus tissue might be involved in the generation of leukemogenic MCF viruses.

  19. Functional dissection of the Moloney murine leukemia virus envelope protein gp70.

    PubMed

    Bae, Y; Kingsman, S M; Kingsman, A J

    1997-03-01

    The envelope protein of Moloney murine leukemia virus (Mo-MLV) is a complex glycoprotein that mediates receptor binding and entry via fusion with cell membranes. By using a series of substitution mutations and truncations in the Mo-MLV external envelope surface protein gp70, we have identified regions important for these processes. Firstly, truncations of gp70 revealed that the minimal continuous receptor-binding region is amino acids 9 to 230, in broad agreement with other studies. Secondly, within this region there are two key basic amino acids, Arg-83 and Arg-95, that are essential for receptor binding and may interact with a negatively charged residue(s) or with the pi electrons of the aromatic ring on a hydrophobic residue(s) in the basic amino acid transporter protein that is the Mo-MLV ecotropic receptor. Finally, we showed that outside the minimal receptor-binding region at amino acids 2 to 8, there is a region that is essential for postbinding fusion events. PMID:9032341

  20. Insights into the nuclear export of murine leukemia virus intron-containing RNA

    PubMed Central

    Pessel-Vivares, Lucie; Houzet, Laurent; Lainé, Sébastien; Mougel, Marylène

    2015-01-01

    The retroviral genome consists of an intron-containing transcript that has essential cytoplasmic functions in the infected cell. This viral transcript can escape splicing, circumvent the nuclear checkpoint mechanisms and be transported to the cytoplasm by hijacking the host machinery. Once in the cytoplasm, viral unspliced RNA acts as mRNA to be translated and as genomic RNA to be packaged into nascent viruses. The murine leukemia virus (MLV) is among the first retroviruses discovered and is classified as simple Retroviridae due to its minimal encoding capacity. The oncogenic and transduction abilities of MLV are extensively studied, whereas surprisingly the crucial step of its nuclear export has remained unsolved until 2014. Recent work has revealed the recruitment by MLV of the cellular NXF1/Tap-dependent pathway for export. Unconventionally, MLV uses of Tap to export both spliced and unspliced viral RNAs. Unlike other retroviruses, MLV does not harbor a unique RNA signal for export. Indeed, multiple sequences throughout the MLV genome appear to promote export of the unspliced MLV RNA. We review here the current understanding of the export mechanism and highlight the determinants that influence MLV export. As the molecular mechanism of MLV export is elucidated, we will gain insight into the contribution of the export pathway to the cytoplasmic fate of the viral RNA. PMID:26158194

  1. Solution Properties of Murine Leukemia Virus Gag Protein: Differences from HIV-1 Gag▿

    PubMed Central

    Datta, Siddhartha A. K.; Zuo, Xiaobing; Clark, Patrick K.; Campbell, Stephen J.; Wang, Yun-Xing; Rein, Alan

    2011-01-01

    Immature retrovirus particles are assembled from the multidomain Gag protein. In these particles, the Gag proteins are arranged radially as elongated rods. We have previously characterized the properties of HIV-1 Gag in solution. In the absence of nucleic acid, HIV-1 Gag displays moderately weak interprotein interactions, existing in monomer-dimer equilibrium. Neutron scattering and hydrodynamic studies suggest that the protein is compact, and biochemical studies indicate that the two ends can approach close in three-dimensional space, implying the need for a significant conformational change during assembly. We now describe the properties of the Gag protein of Moloney murine leukemia virus (MLV), a gammaretrovirus. We found that this protein is very different from HIV-1 Gag: it has much weaker protein-protein interaction and is predominantly monomeric in solution. This has allowed us to study the protein by small-angle X-ray scattering and to build a low-resolution molecular envelope for the protein. We found that MLV Gag is extended in solution, with an axial ratio of ∼7, comparable to its dimensions in immature particles. Mutational analysis suggests that runs of prolines in its matrix and p12 domains and the highly charged stretch at the C terminus of its capsid domain all contribute to this extended conformation. These differences between MLV Gag and HIV-1 Gag and their implications for retroviral assembly are discussed. PMID:21917964

  2. Murine leukemia virus in organs of senescence-prone and -resistant mouse strains.

    PubMed

    Carp, R I; Meeker, H C; Chung, R; Kozak, C A; Hosokawa, M; Fujisawa, H

    2002-03-31

    A series of inbred strains of mice have been developed that are either prone (SAMP) or resistant (SAMR) to accelerated senescence. All of these strains originated from an inadvertent cross or crosses between the AKR/J mouse strain and an unknown strain(s). The characteristics of the nine senescence-prone lines differ, with all strains showing generalized aspects of accelerated aging but with each line having a specific aging-related change that is emphasized, e.g. learning and memory deficits, osteoporosis and senile amyloidosis. The senescence-resistant strains have normal patterns of aging and do not show the specific aging-related changes seen in SAMP strains. The fact that AKR mice have high levels of endogenous, ecotropic murine leukemia virus (MuLV) prompted an examination of the expression levels of MuLV in SAM strains. Analysis of brain, spleen and thymus samples revealed that seven of nine SAMP strains had high levels of MuLV and contained the Emv11 provirus (previously termed Akv1) that encodes the predominant MuLV found in AKR mice. In contrast, none of the SAMR strains had Emv11 or significant amounts of virus. The current findings represent an initial step in determining the role of MuLV in the accelerated senescence seen in SAMP strains. PMID:11850021

  3. Fv-1 restriction and its effects on murine leukemia virus integration in vivo and in vitro.

    PubMed Central

    Pryciak, P M; Varmus, H E

    1992-01-01

    We have investigated the mechanisms by which alleles at the mouse Fv-1 locus restrict replication of murine leukemia viruses. Inhibition of productive infection is closely paralleled by reduced accumulation of integrated proviral DNA as well as by reduced levels of linear viral DNA in a cytoplasmic fraction. Nevertheless, viral DNA is present at nearly normal levels in a nuclear fraction, and total amounts of viral DNA are only mildly affected in restrictive infections, suggesting a block in integration to account for reduced levels of proviral DNA. However, integrase (IN)-dependent trimming of 3' ends of viral DNA occurs normally in vivo during restrictive infections, demonstrating that not all IN-mediated events are prevented in vivo. Furthermore, viral integration complexes present in nuclear extracts of infected restrictive cells are fully competent to integrate their DNA into a heterologous target in vitro. Thus, the Fv-1-dependent activity that restricts integration in vivo may be lost in vitro; alternatively, Fv-1 restriction may prevent a step required for integration in vivo that is bypassed in vitro. Images PMID:1326652

  4. Solution Properties of Murine Leukemia Virus Gag Protein: Differences from HIV-1 Gag

    SciTech Connect

    Datta, Siddhartha A.K.; Zuo, Xiaobing; Clark, Patrick K.; Campbell, Stephen J.; Wang, Yun-Xing; Rein, Alan

    2012-05-09

    Immature retrovirus particles are assembled from the multidomain Gag protein. In these particles, the Gag proteins are arranged radially as elongated rods. We have previously characterized the properties of HIV-1 Gag in solution. In the absence of nucleic acid, HIV-1 Gag displays moderately weak interprotein interactions, existing in monomer-dimer equilibrium. Neutron scattering and hydrodynamic studies suggest that the protein is compact, and biochemical studies indicate that the two ends can approach close in three-dimensional space, implying the need for a significant conformational change during assembly. We now describe the properties of the Gag protein of Moloney murine leukemia virus (MLV), a gammaretrovirus. We found that this protein is very different from HIV-1 Gag: it has much weaker protein-protein interaction and is predominantly monomeric in solution. This has allowed us to study the protein by small-angle X-ray scattering and to build a low-resolution molecular envelope for the protein. We found that MLV Gag is extended in solution, with an axial ratio of {approx}7, comparable to its dimensions in immature particles. Mutational analysis suggests that runs of prolines in its matrix and p12 domains and the highly charged stretch at the C terminus of its capsid domain all contribute to this extended conformation. These differences between MLV Gag and HIV-1 Gag and their implications for retroviral assembly are discussed.

  5. Structural basis of suppression of host translation termination by Moloney Murine Leukemia Virus

    NASA Astrophysics Data System (ADS)

    Tang, Xuhua; Zhu, Yiping; Baker, Stacey L.; Bowler, Matthew W.; Chen, Benjamin Jieming; Chen, Chen; Hogg, J. Robert; Goff, Stephen P.; Song, Haiwei

    2016-06-01

    Retroviral reverse transcriptase (RT) of Moloney murine leukemia virus (MoMLV) is expressed in the form of a large Gag-Pol precursor protein by suppression of translational termination in which the maximal efficiency of stop codon read-through depends on the interaction between MoMLV RT and peptidyl release factor 1 (eRF1). Here, we report the crystal structure of MoMLV RT in complex with eRF1. The MoMLV RT interacts with the C-terminal domain of eRF1 via its RNase H domain to sterically occlude the binding of peptidyl release factor 3 (eRF3) to eRF1. Promotion of read-through by MoMLV RNase H prevents nonsense-mediated mRNA decay (NMD) of mRNAs. Comparison of our structure with that of HIV RT explains why HIV RT cannot interact with eRF1. Our results provide a mechanistic view of how MoMLV manipulates the host translation termination machinery for the synthesis of its own proteins.

  6. Structure of glycosylated and unglycosylated gag polyproteins of Rauscher murine leukemia virus: carbohydrate attachment sites.

    PubMed Central

    Schultz, A M; Lockhart, S M; Rabin, E M; Oroszlan, S

    1981-01-01

    The structural relationships among the gag polyproteins Pr65gag, Pr75gag, and gPr80gag of Rauscher murine leukemia virus were studied by endoglycosidase H digestion and formic acid cleavage. Fragments were identified by precipitation with specific antisera to constituent virion structural proteins followed by one-dimensional mapping. Endoglycosidase H reduced the size of gPr80gag to that of Pr75gag. By comparing fragments of gPr80gag and the apoprotein Pr75gag, the former was shown to contain two mannose-rich oligosaccharide units. By comparing fragments of Pr65gag and Pr75gag, the latter was shown to differ from Pr65gag at the amino terminus by the presence of a leader peptide approximately 7,000 daltons in size. The internal and carboxyl-terminal peptides of the two unglycosylated polyproteins were not detectably different. The location of the two N-linked carbohydrate chains in gPr80gag has been specified. One occurs in the carboxyl-terminal half of the polyprotein at asparagine177 of the p30 sequence and the other is found in a 23,000-dalton fragment located in the amino-terminal region of gPr80gag and containing the additional amino acid sequences not found in Pr65gag plus a substantial portion of p15. Images PMID:7241663

  7. In Vitro Assembly of Virus-Like Particles of a Gammaretrovirus, the Murine Leukemia Virus XMRV

    PubMed Central

    Hadravová, Romana; de Marco, Alex; Ulbrich, Pavel; Štokrová, Jitka; Doležal, Michal; Pichová, Iva; Ruml, Tomáš

    2012-01-01

    Immature retroviral particles are assembled by self-association of the structural polyprotein precursor Gag. During maturation the Gag polyprotein is proteolytically cleaved, yielding mature structural proteins, matrix (MA), capsid (CA), and nucleocapsid (NC), that reassemble into a mature viral particle. Proteolytic cleavage causes the N terminus of CA to fold back to form a β-hairpin, anchored by an internal salt bridge between the N-terminal proline and the inner aspartate. Using an in vitro assembly system of capsid-nucleocapsid protein (CANC), we studied the formation of virus-like particles (VLP) of a gammaretrovirus, the xenotropic murine leukemia virus (MLV)-related virus (XMRV). We show here that, unlike other retroviruses, XMRV CA and CANC do not assemble tubular particles characteristic of mature assembly. The prevention of β-hairpin formation by the deletion of either the N-terminal proline or 10 initial amino acids enabled the assembly of ΔProCANC or Δ10CANC into immature-like spherical particles. Detailed three-dimensional (3D) structural analysis of these particles revealed that below a disordered N-terminal CA layer, the C terminus of CA assembles a typical immature lattice, which is linked by rod-like densities with the RNP. PMID:22090120

  8. Structural basis of suppression of host translation termination by Moloney Murine Leukemia Virus.

    PubMed

    Tang, Xuhua; Zhu, Yiping; Baker, Stacey L; Bowler, Matthew W; Chen, Benjamin Jieming; Chen, Chen; Hogg, J Robert; Goff, Stephen P; Song, Haiwei

    2016-01-01

    Retroviral reverse transcriptase (RT) of Moloney murine leukemia virus (MoMLV) is expressed in the form of a large Gag-Pol precursor protein by suppression of translational termination in which the maximal efficiency of stop codon read-through depends on the interaction between MoMLV RT and peptidyl release factor 1 (eRF1). Here, we report the crystal structure of MoMLV RT in complex with eRF1. The MoMLV RT interacts with the C-terminal domain of eRF1 via its RNase H domain to sterically occlude the binding of peptidyl release factor 3 (eRF3) to eRF1. Promotion of read-through by MoMLV RNase H prevents nonsense-mediated mRNA decay (NMD) of mRNAs. Comparison of our structure with that of HIV RT explains why HIV RT cannot interact with eRF1. Our results provide a mechanistic view of how MoMLV manipulates the host translation termination machinery for the synthesis of its own proteins. PMID:27329342

  9. Assembly and composition of intracellular particles formed by Moloney murine leukemia virus.

    PubMed Central

    Hansen, M; Jelinek, L; Jones, R S; Stegeman-Olsen, J; Barklis, E

    1993-01-01

    Assembly of type C retroviruses such as Moloney murine leukemia virus (M-MuLV) ordinarily occurs at the plasma membranes of infected cells and absolutely requires the particle core precursor protein, Pr65gag. Previously we have shown that Pr65gag is membrane associated and that at least a portion of intracellular Pr65gag protein appears to be routed to the plasma membrane by a vesicular transport pathway. Here we show that intracellular particle formation can occur in M-MuLV-infected cells. M-MuLV immature particles were observed by electron microscopy budding into and within rough endoplasmic reticulum, Golgi, and vacuolar compartments. Biochemical fractionation studies indicated that intracellular Pr65gag was present in nonionic detergent-resistant complexes of greater than 150S. Additionally, viral RNA and polymerase functions appeared to be associated with intracellular particles, as were Gag-beta-galactosidase fusion proteins which have the capacity to be incorporated into virions. Immature intracellular particles in postnuclear lysates could be proteolytically processed in vitro to mature forms, while extracellular immature M-MuLV particles remained immature as long as 10 h during incubations. The occurrence of M-MuLV-derived intracellular particles demonstrates that Pr65gag can associate with intracellular membranes and indicates that if a plasma membrane Pr65gag receptor exists, it also can be found in other membrane compartments. These results support the hypothesis that intracellular particles may serve as a virus reservoir during in vivo infections. Images PMID:8350394

  10. Structural basis of suppression of host translation termination by Moloney Murine Leukemia Virus

    PubMed Central

    Tang, Xuhua; Zhu, Yiping; Baker, Stacey L.; Bowler, Matthew W.; Chen, Benjamin Jieming; Chen, Chen; Hogg, J. Robert; Goff, Stephen P.; Song, Haiwei

    2016-01-01

    Retroviral reverse transcriptase (RT) of Moloney murine leukemia virus (MoMLV) is expressed in the form of a large Gag–Pol precursor protein by suppression of translational termination in which the maximal efficiency of stop codon read-through depends on the interaction between MoMLV RT and peptidyl release factor 1 (eRF1). Here, we report the crystal structure of MoMLV RT in complex with eRF1. The MoMLV RT interacts with the C-terminal domain of eRF1 via its RNase H domain to sterically occlude the binding of peptidyl release factor 3 (eRF3) to eRF1. Promotion of read-through by MoMLV RNase H prevents nonsense-mediated mRNA decay (NMD) of mRNAs. Comparison of our structure with that of HIV RT explains why HIV RT cannot interact with eRF1. Our results provide a mechanistic view of how MoMLV manipulates the host translation termination machinery for the synthesis of its own proteins. PMID:27329342

  11. Effect of internal genomic sequences of the Moloney murine leukemia virus on replication

    SciTech Connect

    Fomin, I.K.; Lobanova, A.B.; Voitenok, N.N.

    1995-11-01

    Construction and use of retrovirus vectors derived from the Moloney murine leukemia virus (MoMuLV) are described. These vectors, designated minimal vectors, contain the left and right long terminal repeats (LTRs), a binding site for proline tRNA, a polypurine tract (PPT), and a dominant marker for selective introduction of vectors into a packaging cell line, but lack the internal sequences of the virus genome. The experiments showed that the minimal vectors can be replicated and that their titer was approximately 1500-fold lower than that of wild-type vectors. The minimal vectors were shown to contain all the cis-acting sequences necessary for correct reverse transcription. One infectious virion, like wild-type viruses, produced only one provirus. Unlike the avian reticuloendotheliosis virus (REV), {Psi}{sup +} and {Psi}{sup {minus}} genomes of MoMuLV did not compete for virion proteins in the {Psi}2 packaging cell line. When an insert was introduced into a central part of the LTR U5 region, the titer of the minimal vector remained the same, while the titer of the wild-type vector decreased approximately 40-fold. 28 refs., 2 figs., 2 tabs.

  12. Xenotropic Murine Leukemia Virus-Related Virus in Chronic Fatigue Syndrome and Prostate Cancer

    PubMed Central

    2010-01-01

    Xenotropic murine leukemia virus-related virus (XMRV) is a γ retrovirus that has been associated with chronic fatigue syndrome (CFS) and prostate cancer. The search for viral causes of these syndromes was reignited by the finding that RNase L activity was low in hereditary prostate cancer and some CFS patients. The six strains of XMRV that have been sequenced have greater than 99% identity, indicating a new human infection rather than laboratory contamination. DNA, RNA, and proteins from XMRV have been detected in 50% to 67% of CFS patients and in about 3.7% of healthy controls. XMRV infections could be transmitted to permissive cell lines from CFS plasma, suggesting the potential for communicable and blood-borne spread of the virus and potentially CFS. This troubling concept is currently under intense evaluation. The most important steps now are to independently confirm the initial findings; develop reliable assays of biomarkers; and to move on to investigations of XMRV pathophysiology and treatment in CFS, prostate cancer, and potentially other virus-related syndromes, if they exist. PMID:20425007

  13. Sequence-specific binding of DNA by the Moloney murine leukemia virus integrase protein.

    PubMed Central

    Krogstad, P A; Champoux, J J

    1990-01-01

    Genetic studies have indicated that integration of retroviral DNA into the host genome depends on the presence of the inverted repeats at the free termini of the long terminal repeats on the unintegrated DNA and on the product of the 3' end of the pol gene (the integrase [IN] protein). While the precise function of the Moloney murine leukemia virus IN protein is uncertain, others have shown that it is a DNA-binding protein and functions in the processing of the inverted repeats prior to integration. By using site-directed mutagenesis, we cloned and expressed the IN protein in Escherichia coli. Crude extracts of total cellular protein were fractionated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, transferred to nitrocellulose filters, denatured in guanidine, renatured, and incubated with oligonucleotide probes. Single- and double-stranded oligonucleotides corresponding to the termini of unintegrated linear viral DNA were specifically bound by the IN protein in this assay. These data suggest that the role of the Moloney IN protein in the early steps of integration involves sequence-specific recognition of the DNA sequences found at the ends of the long terminal repeats. Images PMID:2186176

  14. Construction and characterization of the recombinant Moloney murine leukemia viruses bearing the mouse Fv-4 env gene.

    PubMed Central

    Masuda, M; Yoshikura, H

    1990-01-01

    A nucleotide sequence of the mouse Fv-4 env gene was completed. Structural comparison revealed a close relationship of Fv-4 to the ecotropic Cas-Br-E murine leukemia virus isolated from a wild mouse in southern California. Various portions of the env gene of Moloney murine leukemia virus were replaced by the corresponding Fv-4 env sequence to construct recombinant murine leukemia virus clones. Infectivity of these recombinants was checked by the S+L- cell focus induction assay and the XC cell syncytium formation assay. Recombinants bearing the following Fv-4 env sequence retained ecotropic infectivity; the AccI-BamHI and BamHI-BalI regions coding for the N- and C-terminal halves of Fv-4 gp70SU, respectively; and the BalI-NcoI region encoding the cleavage site between gp70SU and p15(E)TM of the Fv-4 env. However, when the Fv-4 sequence was substituted for the p15(E)TM-coding NcoI-EcoRV region or the AccI-EcoRV region covering almost the entire env gene, infectivity was undetectable in our assays. The recombinant clone containing the Fv-4 AccI-EcoRV region, i.e., almost the entire Fv-4 env sequence, was introduced with pSV2neo into NIH 3T3 cells, and a G418r cell line named NIH(Fv4)-2 was isolated. The NIH(Fv4)-2 cell released viral particles that contained reverse transcriptase, Fv-4 env molecules as well as the other viral proteins, and viral genomic RNA. However, proviral DNA synthesis was not detected upon inoculation of this virus in NIH 3T3 cells. The loss of infectivity of the recombinant virus bearing the Fv-4 AccI-EcoRV region appeared to be caused by failure in an early step of replication. Images PMID:2304138

  15. Splicing of Friend Murine Leukemia Virus env-mRNA Enhances Its Ability to Form Polysomes

    PubMed Central

    Machinaga, Akihito; Ishihara, Syuhei; Shirai, Akiko; Takase-Yoden, Sayaka

    2016-01-01

    Friend murine leukemia virus (MLV) belongs to the gamma retroviruses of the Retroviridae family. The positive-sense RNA of its genome contains a 5′ long terminal repeat (LTR), 5′ leader sequence, gag, pol, env, and 3′ LTR. Transcription from proviral DNA begins from the R region of the 5′ LTR and ends at the polyadenylation signal located at the R region of the other end of the 3′ LTR. There is a 5′ splice site in the 5′ leader sequence and a 3′ splice site at the 3′ end of the pol region. Both full-length unspliced mRNAs and a singly spliced mRNA (env-mRNA) are produced in MLV-infected cells. The MLV Env protein plays important roles both in viral adsorption to host cells and in neuropathogenic disease in MLV-infected mice and rats. Understanding the regulatory mechanisms controlling Env expression is important for determining the functions of the Env protein. We have previously shown that splicing increases env-mRNA stability and translation efficiency. Generally, mRNA polysome formation correlates with translation efficiency. Therefore, here we investigated the effects of env-mRNA splicing on polysome formation to identify mechanisms for Env up-regulation due to splicing. We performed polysome profile analyses using Env-expression plasmids producing spliced or unspliced env-mRNA and showed that the former formed polysomes more efficiently than the latter. Thus, splicing of env-mRNA facilitated polysome formation, suggesting that this contributes to up-regulation of Env expression. We replaced the env region of the expression plasmids with a luciferase (luc) gene, and found that in this case both unspliced and spliced luc-mRNA formed polysomes to a similar extent. Thus, we conclude that whether mRNA polysome formation is affected by splicing depends on the structure of gene in question. PMID:26909075

  16. Genomic stability of murine leukemia viruses containing insertions at the Env-3' untranslated region boundary.

    PubMed

    Logg, C R; Logg, A; Tai, C K; Cannon, P M; Kasahara, N

    2001-08-01

    Retroviruses containing inserts of exogenous sequences frequently eliminate the inserted sequences upon spread in susceptible cells. We have constructed replication-competent murine leukemia virus (MLV) vectors containing internal ribosome entry site (IRES)-transgene cassettes at the env-3' untranslated region boundary in order to examine the effects of insert sequence and size on the loss of inserts during viral replication. A virus containing an insertion of 1.6 kb replicated with greatly attenuated kinetics relative to wild-type virus and lost the inserted sequences in a single infection cycle. In contrast, MLVs containing inserts of 1.15 to 1.30 kb replicated with kinetics only slightly attenuated compared to wild-type MLV and exhibited much greater stability, maintaining their genomic integrity over multiple serial infection cycles. Eventually, multiple species of deletion mutants were detected simultaneously in later infection cycles; once detected, these variants rapidly dominated the population and thereafter appeared to be maintained at a relative equilibrium. Sequence analysis of these variants identified preferred sites of recombination in the parental viruses, including both short direct repeats and inverted repeats. One instance of insert deletion through recombination with an endogenous retrovirus was also observed. When specific sequences involved in these recombination events were eliminated, deletion variants still arose with the same kinetics upon virus passage and by apparently similar mechanisms, although at different locations in the vectors. Our results suggest that while lengthened, insert-containing genomes can be maintained over multiple replication cycles, preferential deletions resulting in loss of the inserted sequences confer a strong selective advantage. PMID:11435579

  17. No Evidence of Murine Leukemia Virus-Related Viruses in Live Attenuated Human Vaccines

    PubMed Central

    Switzer, William M.; Zheng, HaoQiang; Simmons, Graham; Zhou, Yanchen; Tang, Shaohua; Shankar, Anupama; Kapusinszky, Beatrix; Delwart, Eric L.; Heneine, Walid

    2011-01-01

    Background The association of xenotropic murine leukemia virus (MLV)-related virus (XMRV) in prostate cancer and chronic fatigue syndrome reported in previous studies remains controversial as these results have been questioned by recent data. Nonetheless, concerns have been raised regarding contamination of human vaccines as a possible source of introduction of XMRV and MLV into human populations. To address this possibility, we tested eight live attenuated human vaccines using generic PCR for XMRV and MLV sequences. Viral metagenomics using deep sequencing was also done to identify the possibility of other adventitious agents. Results All eight live attenuated vaccines, including Japanese encephalitis virus (JEV) (SA-14-14-2), varicella (Varivax), measles, mumps, and rubella (MMR-II), measles (Attenuvax), rubella (Meruvax-II), rotavirus (Rotateq and Rotarix), and yellow fever virus were negative for XMRV and highly related MLV sequences. However, residual hamster DNA, but not RNA, containing novel endogenous gammaretrovirus sequences was detected in the JEV vaccine using PCR. Metagenomics analysis did not detect any adventitious viral sequences of public health concern. Intracisternal A particle sequences closest to those present in Syrian hamsters and not mice were also detected in the JEV SA-14-14-2 vaccine. Combined, these results are consistent with the production of the JEV vaccine in Syrian hamster cells. Conclusions We found no evidence of XMRV and MLV in eight live attenuated human vaccines further supporting the safety of these vaccines. Our findings suggest that vaccines are an unlikely source of XMRV and MLV exposure in humans and are consistent with the mounting evidence on the absence of these viruses in humans. PMID:22216219

  18. Crystal Structure of the Moloney Murine Leukemia Virus RNase H Domain

    SciTech Connect

    Lim,D.; Gregorio, G.; Bingman, C.; Martinez-Hackert, E.; Hendrickson, W.; Goff, S.

    2006-01-01

    A crystallographic study of the Moloney murine leukemia virus (Mo-MLV) RNase H domain was performed to provide information about its structure and mechanism of action. These efforts resulted in the crystallization of a mutant Mo-MLV RNase H lacking the putative helix C ({Delta}C). The 1.6-{angstrom} resolution structure resembles the known structures of the human immunodeficiency virus type 1 (HIV-1) and Escherichia coli RNase H. The structure revealed the coordination of a magnesium ion within the catalytic core comprised of the highly conserved acidic residues D524, E562, and D583. Surface charge mapping of the Mo-MLV structure revealed a high density of basic charges on one side of the enzyme. Using a model of the Mo-MLV structure superimposed upon a structure of HIV-1 reverse transcriptase bound to an RNA/DNA hybrid substrate, Mo-MLV RNase H secondary structures and individual amino acids were examined for their potential roles in binding substrate. Identified regions included Mo-MLV RNase H {beta}1-{beta}2, {alpha}A, and {alpha}B and residues from {alpha}B to {alpha}D and its following loop. Most of the identified substrate-binding residues corresponded with residues directly binding nucleotides in an RNase H from Bacillus halodurans as observed in a cocrystal structure with RNA/DNA. Finally, superimposition of RNases H of Mo-MLV, E. coli, and HIV-1 revealed that a loop of the HIV-1 connection domain resides within the same region of the Mo-MLV and E. coli C-helix. The HIV-1 connection domain may serve to recognize and bind the RNA/DNA substrate major groove.

  19. Basis for receptor specificity of nonecotropic murine leukemia virus surface glycoprotein gp70SU.

    PubMed Central

    Ott, D; Rein, A

    1992-01-01

    Murine leukemia viruses (MuLVs) initiate infection of NIH 3T3 cells by binding of the viral envelope (Env) protein to a cell surface receptor. Interference assays have shown that MuLVs can be divided into four groups, each using a distinct receptor: ecotropic, polytropic, amphotropic, and 10A1. In this study, we have attempted to map the determinants within viral Env proteins by constructing chimeric env genes. Chimeras were made in all six pairwise combinations between Moloney MCF (a polytropic MuLV), amphotropic MuLV, and 10A1, using a conserved EcoRI site in the middle of the Env coding region. The receptor specificity of each chimera was determined by using an interference assay. We found that amphotropic receptor specificity of each chimera was determined by using an interference assay. We found that amphotropic receptor specificity seems to map to the N-terminal portion of surface glycoprotein gp70SU. The difference between amphotropic and 10A1 receptor specificity can be attributed to one or more of only six amino acid differences in this region. Nearly all other cases showed evidence of interaction between Env domains in the generation of receptor specificity. Thus, a chimera composed exclusively of MCF and amphotropic sequences was found to exhibit 10A1 receptor specificity. None of the chimeras were able to infect cells by using the MCF receptor; however, two chimeras containing the C-terminal portion of MCF gp70SU could bind to this receptor, while they were able to infect cells via the amphotropic receptor. This result raises the possibility that receptor binding maps to the C-terminal portion of MCF gp70SU but requires MCF N-terminal sequences for a functional interaction with the MCF receptor. Images PMID:1321266

  20. Splicing of Friend Murine Leukemia Virus env-mRNA Enhances Its Ability to Form Polysomes.

    PubMed

    Machinaga, Akihito; Ishihara, Syuhei; Shirai, Akiko; Takase-Yoden, Sayaka

    2016-01-01

    Friend murine leukemia virus (MLV) belongs to the gamma retroviruses of the Retroviridae family. The positive-sense RNA of its genome contains a 5' long terminal repeat (LTR), 5' leader sequence, gag, pol, env, and 3' LTR. Transcription from proviral DNA begins from the R region of the 5' LTR and ends at the polyadenylation signal located at the R region of the other end of the 3' LTR. There is a 5' splice site in the 5' leader sequence and a 3' splice site at the 3' end of the pol region. Both full-length unspliced mRNAs and a singly spliced mRNA (env-mRNA) are produced in MLV-infected cells. The MLV Env protein plays important roles both in viral adsorption to host cells and in neuropathogenic disease in MLV-infected mice and rats. Understanding the regulatory mechanisms controlling Env expression is important for determining the functions of the Env protein. We have previously shown that splicing increases env-mRNA stability and translation efficiency. Generally, mRNA polysome formation correlates with translation efficiency. Therefore, here we investigated the effects of env-mRNA splicing on polysome formation to identify mechanisms for Env up-regulation due to splicing. We performed polysome profile analyses using Env-expression plasmids producing spliced or unspliced env-mRNA and showed that the former formed polysomes more efficiently than the latter. Thus, splicing of env-mRNA facilitated polysome formation, suggesting that this contributes to up-regulation of Env expression. We replaced the env region of the expression plasmids with a luciferase (luc) gene, and found that in this case both unspliced and spliced luc-mRNA formed polysomes to a similar extent. Thus, we conclude that whether mRNA polysome formation is affected by splicing depends on the structure of gene in question. PMID:26909075

  1. Preparation and characterization of the RNase H domain of Moloney murine leukemia virus reverse transcriptase.

    PubMed

    Nishimura, Kosaku; Yokokawa, Kanta; Hisayoshi, Tetsuro; Fukatsu, Kosuke; Kuze, Ikumi; Konishi, Atsushi; Mikami, Bunzo; Kojima, Kenji; Yasukawa, Kiyoshi

    2015-09-01

    Moloney murine leukemia virus reverse transcriptase (MMLV RT) contains fingers, palm, thumb, and connection subdomains as well as an RNase H domain. The DNA polymerase active site resides in the palm subdomain, and the RNase H active site is located in the RNase H domain. The RNase H domain contains a positively charged α-helix called the C helix (H(594)GEIYRRR(601)), that is thought to be involved in substrate recognition. In this study, we expressed three versions of the RNase H domain in Escherichia coli, the wild-type domain (WT) (residues Ile498-Leu671) and two variants that lack the regions containing the C helix (Ile593-Leu603 and Gly595-Thr605, which we called ΔC1 and ΔC2, respectively) with a strep-tag at the N-terminus and a deca-histidine tag at the C-terminus. These peptides were purified from the cells by anion-exchange, Ni(2+) affinity, and Strep-Tactin affinity column chromatography, and then the tags were removed by proteolysis. In an RNase H assay using a 25-bp RNA-DNA heteroduplex, WT, ΔC1, and ΔC2 produced RNA fragments ranging from 7 to 16 nucleotides (nt) whereas the full-length MMLV RT (Thr24-Leu671) produced 14-20-nt RNA fragments, suggesting that elimination of the fingers, palm, thumb, and connection subdomains affects the binding of the RNase H domain to the RNA-DNA heteroduplex. The activity levels of WT, ΔC1, and ΔC2 were estimated to be 1%, 0.01%, and 0.01% of full-length MMLV RT activity, indicating that the C helix is important, but not critical, for the activity of the isolated RNase H domain. PMID:25959458

  2. Characterization of a novel murine leukemia virus-related subgroup within mammals.

    PubMed Central

    Tristem, M; Kabat, P; Lieberman, L; Linde, S; Karpas, A; Hill, F

    1996-01-01

    The murine leukemia virus (MuLV)-related retroviruses are one of seven genera which together constitute the family Retroviridae. They are widespread as both endogenous and exogenous agents within vertebrates and have been associated with a variety of malignancies and other disorders. We isolated and characterized 12 endogenous representatives of this genus from a number of mammalian hosts. Subsequent sequence analysis revealed that the isolated viruses cluster into two clearly distinct groups. All of the exogenous MuLV-related retroviruses which have been isolated to date, as well as several endogenous examples, fall into the first group, whereas the second group is represented solely by endogenous representatives, including human endogenous retrovirus type E (HERV.E). The two groups are widespread within mammals, with both often present within one animal species. Despite this, there is no evidence to date that recombination between members of the different groups has occurred. Genetic distances and several other properties of the HERV.E genome suggest that if exogenous members of this subgroup exist, they are likely to have biological properties different from those of the other exogenous viruses of this genus. Several of these viruses are known to have been integrated within their hosts' genomes for a long period of time, and a most recent divergence date for the MuLV and HERV.E subgroups can thus be proposed. This date, approximately 30 million years ago, is the most recent date possible, and it is probable that the actual period of time since their divergence is significantly longer. PMID:8892961

  3. Physical properties of moloney murine leukemia virus high-molecular-weight RNA: a two subunit structure.

    PubMed Central

    Riggin, C H; Bondurant, M; Mitchell, W M

    1975-01-01

    The high-molecular-weight RNA of Moloney murine leukemia virus (MuLV) was analyzed by sedimentation equilibrium ultracentrifugation. Molecular weights of 7.2 x 10(6) and 3.4 x 10(6) were found for the native and subunit forms, respectively, indicating that the native structure is a dimer. S20,w and frictional coefficients were determined for MuLV RNA by analytical velocity centrifugation as a function of ionic strength. The apparent S20,w of native MuLV RNA was 47.3, 57.4, and 66.5 in 0.01, 0.1, and 0.20 M Na+, respectively; the corresponding frictional coefficients were 5.44, 4.48, and 3.87. Native RNA was estimated by circular dichroism to be 85% helical, whereas denatured RNA was 54% helical. Thermal denaturation profiles were obtained from uv absorbance scans. Melting temperatures of 57 and 68 C were obtained for high-molecular-weight RNA in 0.01 M Na+ and 0.122 M Na+, 1mM Mg2+, respectively. van't Hoff plots of the thermal denaturation data gave enthalpies for the helix-coil transition of 21,600 cal (ca. 90,500 J) per mol of cooperatively melting unit in high salt and 19,600 cal (ca. 82,100 J) per mol in low salt, consistent with both base stacking and pairing. The melting of Mu LV RNA occurred over a broad temprange and van't Hoff plots were linear over most of the melting range, indicating a noncooperative process of helix stabilization. PMID:1202247

  4. Comparative efficacies of amphotericin B lipid complex and amphotericin B deoxycholate suspension against murine blastomycosis.

    PubMed Central

    Clemons, K V; Stevens, D A

    1991-01-01

    Amphotericin B as a lipid complex and as a deoxycholate suspension (Fungizone) was tested against murine blastomycosis. All doses of each form prolonged survival (P less than 0.05 to 0.001). Fungizone was more effective than lipid complex at doses of 0.8 mg/kg of body weight. However, lipid complex at 12.8 mg/kg was not toxic and superior in efficacy (P less than 0.001) to 2.0 mg of Fungizone per kg (a toxic dose), and it cleared all animals of infection. Lipid complex is an effective therapy for murine blastomycosis. PMID:1759840

  5. Generation of mink cell focus-forming viruses by Friend murine leukemia virus: recombination with specific endogenous proviral sequences.

    PubMed Central

    Evans, L H; Cloyd, M W

    1984-01-01

    A family of recombinant mink cell focus-forming viruses (MCF) was derived by inoculation of NFS mice with a Friend murine leukemia virus, and their genomes were analyzed by RNase T1-resistant oligonucleotide fingerprinting. The viruses were obtained from the thymuses and spleens of preleukemic and leukemic animals and were evaluated for dualtropism and oncogenicity. All these isolates induced cytopathic foci on mink cells but could be classified into two groups based on their relative infectivities for SC-1 (mouse) or mink (ATCC CCL64) cells. One group of Friend MCFs (F-MCFs) (group I) exhibited approximately equal infectivities for SC-1 and mink cells, whereas a second group (group II) infected mink cells 1,000- to 10,000-fold more efficiently than SC-1 cells. Structural analyses of the F-MCFs revealed that group I and group II viruses correlated with recombination of Friend murine leukemia virus with two distinct, but closely related, endogenous NFS proviral sequences. No correlation was found between the type of F-MCF and the tissue of origin or the disease state of the animal. Furthermore, none of the F-MCF isolates were found to be oncogenic in NFS/N or AKR/J mice. F-MCFs of both groups underwent extensive substitution of ecotropic sequences, involving much of the gag and env genes of group I F-MCFs and most of the gag, pol, and env genes of group II F-MCFs. All F-MCF isolates retained the 3' terminal U3 region of Friend murine leukemia virus. Comparison of the RNAs of the F-MCFs with RNAs of MCFs derived from NFS.Akv-1 or NFS.Akv-2 mice indicated that the F-MCFs were derived from NFS proviral sequences which are distinct from the sequences contained in NFS.Akv MCF isolates. This result suggested that recombination with particular endogenous proviral sequences to generate MCFs may be highly specific for a given murine leukemia virus. Images PMID:6422051

  6. Leukemia.

    PubMed

    Juliusson, Gunnar; Hough, Rachael

    2016-01-01

    Leukemias are a group of life threatening malignant disorders of the blood and bone marrow. In the adolescent and young adult (AYA) population, the acute leukemias are most prevalent, with chronic myeloid leukemia being infrequently seen. Factors associated with more aggressive disease biology tend to increase in frequency with increasing age, whilst tolerability of treatment strategies decreases. There are also challenges regarding the effective delivery of therapy specific to the AYA group, consequences on the unique psychosocial needs of this age group, including compliance. This chapter reviews the current status of epidemiology, pathophysiology, treatment strategies and outcomes of AYA leukemia, with a focus on acute lymphoblastic leukemia and acute myeloid leukemia. PMID:27595359

  7. Identification of homeodomain proteins, PBX1 and PREP1, involved in the transcription of murine leukemia virus.

    PubMed

    Chao, Sheng-Hao; Walker, John R; Chanda, Sumit K; Gray, Nathanael S; Caldwell, Jeremy S

    2003-02-01

    Cyclin-dependent kinase inhibitors (CDKIs) have been shown to block human immunodeficiency virus and herpes simplex virus. It is hypothesized that CDKIs block viral replication by inhibiting transcription of specific cellular genes. Here we find that three CDKIs, flavopiridol, purvalanol A, and methoxy-roscovitine, block Moloney murine leukemia virus (MLV) transcription events. Using gene expression microarray technology to examine the inhibitory effects of CDKIs, we observed a cellular gene, the pre-B-cell leukemia transcription factor 1 (Pbx1) gene, down-regulated by CDKI treatment. The PBX consensus element (PCE), TGATTGAC, is conserved in the long terminal repeats of several murine retroviruses, including Moloney MLV. Mutations in the PCE completely inhibited viral transcription whereas overexpression of PBX1 and a PBX1-associated protein, PREP1, enhanced viral transcription. The interaction between the PCE and PBX1-PREP1 proteins was confirmed by gel shift experiments. Blocking PBX1 protein synthesis resulted in a significant decrease in viral transcription. Collectively, our results represent the first work demonstrating that the homeodomain proteins PBX1 and PREP1 are cellular factors involved in Moloney MLV transcription regulation. PMID:12529389

  8. Androgen-independent proliferation of LNCaP prostate cancer cells infected by xenotropic murine leukemia virus-related virus

    SciTech Connect

    Kakoki, Katsura; Kamiyama, Haruka; Izumida, Mai; Yashima, Yuka; Hayashi, Hideki; Yamamoto, Naoki; Matsuyama, Toshifumi; Igawa, Tsukasa; Sakai, Hideki; Kubo, Yoshinao

    2014-04-25

    Highlights: • XMRV infection induces androgen-independent growth in LNCaP cells. • XMRV infection reduces expression of androgen receptor. • XMRV promotes appearance of androgen blocker-resistant prostate cancer cells. - Abstract: Xenotropic murine leukemia virus-related virus (XMRV) is a novel gammaretrovirus that was originally isolated from human prostate cancer. It is now believed that XMRV is not the etiologic agent of prostate cancer. An analysis of murine leukemia virus (MLV) infection in various human cell lines revealed that prostate cancer cell lines are preferentially infected by XMRV, and this suggested that XMRV infection may confer some sort of growth advantage to prostate cancer cell lines. To examine this hypothesis, androgen-dependent LNCaP cells were infected with XMRV and tested for changes in certain cell growth properties. We found that XMRV-infected LNCaP cells can proliferate in the absence of the androgen dihydrotestosterone. Moreover, androgen receptor expression is significantly reduced in XMRV-infected LNCaP cells. Such alterations were not observed in uninfected and amphotropic MLV-infected LNCaP cells. This finding explains why prostate cancer cell lines are preferentially infected with XMRV.

  9. Leukemia

    MedlinePlus

    Leukemia is cancer of the white blood cells. White blood cells help your body fight infection. Your blood cells form in your bone marrow. In leukemia, the bone marrow produces abnormal white blood cells. ...

  10. Survivin suppressor (YM155) enhances chemotherapeutic efficacy against canine histiocytic sarcoma in murine transplantation models.

    PubMed

    Yamazaki, Hiroki; Takagi, Satoshi; Hosoya, Kenji; Okumura, Masahiro

    2015-04-01

    Histiocytic sarcoma (HS) in dogs exhibits aggressive clinical and biological behavior. Currently, no effective treatments are available for dogs with HS. Survivin, a member of a family of apoptosis protein inhibitors, could serve as a potential therapeutic target in several canine cancers. Sepantronium bromide (YM155) has recently been established as a novel survivin-targeting agent. The aim of this study was to use YM155 as a tool for evaluating survivin-targeted therapies against dogs with HS, and to investigate how YM155 treatment affects antitumor and chemotherapeutic efficacies in murine xenograft models using canine HS cells. The results showed that in HS cells with lomustine (CCNU) resistance, YM155 treatment suppressed both the cell-growth potential and cell resistance to CCNU, which essentially increases the chemotherapy efficacy in the murine models. The evidence presented here supports the favorable preclinical evaluation that survivin-targeted therapies might be effective against HS in dogs. PMID:25744435

  11. Infectious Entry by Amphotropic as well as Ecotropic Murine Leukemia Viruses Occurs through an Endocytic Pathway

    PubMed Central

    Katen, Louis J.; Januszeski, Michael M.; Anderson, W. French; Hasenkrug, Kim J.; Evans, Leonard H.

    2001-01-01

    Infectious entry of enveloped viruses is thought to proceed by one of two mechanisms. pH-dependent viruses enter the cells by receptor-mediated endocytosis and are inhibited by transient treatment with agents that prevent acidification of vesicles in the endocytic pathway, while pH-independent viruses are not inhibited by such agents and are thought to enter the cell by direct fusion with the plasma membrane. Nearly all retroviruses, including amphotropic murine leukemia virus (MuLV) and human immunodeficiency virus type 1, are classified as pH independent. However, ecotropic MuLV is considered to be a pH-dependent virus. We have examined the infectious entry of ecotropic and amphotropic MuLVs and found that they were equally inhibited by NH4Cl and bafilomycin A. These agents inhibited both viruses only partially over the course of the experiments. Agents that block the acidification of endocytic vesicles also arrest vesicular trafficking. Thus, partial inhibition of the MuLVs could be the result of virus inactivation during arrest in this pathway. In support of this contention, we found that that the loss of infectivity of the MuLVs during treatment of target cells with the drugs closely corresponded to the loss of activity due to spontaneous inactivation at 37°C in the same period of time. Furthermore, the drugs had no effect on the efficiency of infection under conditions in which the duration of infection was held to a very short period to minimize the effects of spontaneous inactivation. These results indicate that the infectious processes of both ecotropic and amphotropic MuLVs were arrested rather than aborted by transient treatment of the cells with the drugs. We also found that infectious viruses were efficiently internalized during treatment. This indicated that the arrest occurred in an intracellular compartment and that the infectious process of both the amphotropic and ecotropic MuLVs very likely involved endocytosis. An important aspect of this study

  12. Efficacy of lasalocid against murine Pneumocystis carinii pneumonitis.

    PubMed Central

    OZ, H S; Hughes, W T; Rehg, J E

    1997-01-01

    The efficacy of the ionophore lasalocid against Pneumocystis carinii pneumonitis in corticosteroid-immunosuppressed Sprague-Dawley rats was investigated. Lasalocid was effective in the prevention of the pneumonitis in a dose-dependent manner. At dosages of 0, 5, 10, and 20 mg/kg/day, P. carinii infection rates were 92, 60, 20, and 0%, respectively, during dexamethasone immunosuppression. Also, lasalocid compared favorably with other drugs known to have anti-P. carinii activity, including trimethoprim-sulfamethoxazole, atovaquone, and dapsone-trimethoprim. PMID:8980779

  13. Mutational analysis of the putative receptor-binding domain of Moloney murine leukemia virus glycoprotein gp70.

    PubMed

    Panda, B R; Kingsman, S M; Kingsman, A J

    2000-07-20

    The entry of Moloney murine leukemia virus (MoMuLV) to murine cells is mediated by the binding of its envelope glycoprotein gp70 to its receptor, the cationic amino acid transporter MCAT-1. The binding property of the envelope protein lies mainly in the N-terminal half of the protein. To identify essential residues involved in the binding of gp70 to its receptor, we have mutated amino acids within the putative receptor-binding domain of MoMuLV gp70. Changes in the residues P94 and W100 resulted in lower viral titers in comparison to the wild-type virions. Single, double, or triple point mutations involving the residue W100 make the envelope protein severely defective in binding to its receptor. Binding studies and cell fusion experiments with murine XC cells suggested that the residue W100 might play an important role in the process of infection by making contact between gp70 and its receptor. PMID:10891411

  14. Expression of murine leukemia viruses in RFM mice with host versus graft disease after perinatal inoculation of (T6 X RFM)F1 lymphohemopoietic cells.

    PubMed Central

    Cross, S S; Brede, G; Tucker, H S; Maloney, M; Montour, J L; Hard, R C

    1983-01-01

    Host versus graft disease is the fatal syndrome of altered immunity that follows the perinatal inoculation of related F1 hybrid spleen cells to susceptible strains of inbred mice. The allogenic reaction results in severe depletion of T-lymphocytes, but causes hyperplasia and hypersecretion of B-cells. Among the long-term survivors of acute host versus graft reactions, there is a high incidence of nonthymic lymphomas associated with ecotropic murine leukemia virus that may be of donor F1 origin. The present studies were done to determine whether ecotropic murine leukemia virus played any role in the pathogenesis of acute host versus graft disease in RFM mice perinatally inoculated with (T6 X RFM)F1 spleen cells. In RFM/(T6 X RFM)F1 chimeras, N-tropic murine leukemia virus can be detected as early as 3 days. The progression of the disease was accompanied by increasing viral expression. The inoculation of N-tropic virus of F1 donor origin into RFM neonates failed to induce disease, although the virus proliferated. Detection of progressively rising titers of antibody to murine leukemia virus linked the virus to the development of hyperimmunoglobulinemia by virtue of its ability to serve as a replicating source of antigens. These and other studies provided evidence that the seemingly paradoxical appearance of hyperimmunoglobulinemia in T-cell-deficient mice with the host versus graft syndrome is due, at least in part, to the stimulation of presensitized F1 donor B-cells, which are not destroyed in the allogenic reaction, as are the T-cells. Another unusual finding was the detection of polytropic murine leukemia virus in 25-day-old RFM/(T6 X RFM)F1 chimeras. It is suggested that the allogenic host versus graft reaction favored the formation of recombinants. PMID:6135664

  15. Combination angiostatin and endostatin gene transfer induces synergistic antiangiogenic activity in vitro and antitumor efficacy in leukemia and solid tumors in mice.

    PubMed

    Scappaticci, F A; Smith, R; Pathak, A; Schloss, D; Lum, B; Cao, Y; Johnson, F; Engleman, E G; Nolan, G P

    2001-02-01

    Angiostatin and endostatin are potent endothelial cell growth inhibitors that have been shown to inhibit angiogenesis in vivo and tumor growth in mice. However, tumor shrinkage requires chronic delivery of large doses of these proteins. Here we report synergistic antitumor activity and survival of animals when these factors are delivered in combination to tumors by retroviral gene transfer. We have demonstrated this efficacy in both murine leukemia and melanoma models. Complete loss of tumorigenicity was seen in 40% of the animals receiving tumors transduced by the combination of angiostatin and endostatin in the leukemia model. The synergy was also demonstrated in vitro on human umbilical vein endothelial cell differentiation and this antiangiogenic activity may suggest a mechanism for the antitumor activity in vivo. These findings imply separate pathways by which angiostatin and endostatin mediate their antiangiogenic effects. Together, these data suggest that a combination of antiangiogenic factors delivered by retroviral gene transfer may produce synergistic antitumor effects in both leukemia and solid tumors, thus avoiding long-term administration of recombinant proteins. The data also suggest that novel combinations of antiangiogenic factors delivered into tumors require further investigation as therapeutic modalities. PMID:11237675

  16. Diet-induced obesity accelerates acute lymphoblastic leukemia progression in two murine models*

    PubMed Central

    Yun, Jason P.; Behan, James W.; Heisterkamp, Nora; Butturini, Anna; Klemm, Lars; Ji, Lingyun; Groffen, John; Müschen, Markus; Mittelman, Steven D.

    2010-01-01

    Obesity is associated with an increased incidence of many cancers, including leukemia, though it is unknown whether leukemia incidence is increased directly by obesity, or rather by associated genetic, lifestyle, health, or socio-economic factors. We developed animal models of obesity and leukemia to test whether obesity could directly accelerate acute lymphoblastic leukemia (ALL) using BCR/ABL transgenic and AKR/J mice weaned onto a high-fat diet. Mice were observed until development of progressive ALL. Although obese and control BCR/ABL mice had similar median survival, older obese mice had accelerated ALL onset, implying a time-dependent effect of obesity on ALL. Obese AKR mice developed ALL significantly earlier than controls. The effect of obesity was not explained by WBC count, thymus/spleen weight, or ALL phenotype. However, obese AKR mice had higher leptin, insulin, and IL-6 levels than controls, and these obesity-related hormones all have potential roles in leukemia pathogenesis. In conclusion, obesity directly accelerates presentation of ALL, likely by increasing the risk of an early event in leukemogenesis. This is the first study to demonstrate that obesity can directly accelerate the progression of ALL. Thus, the observed associations between obesity and leukemia incidence are likely to be directly related to biological effects of obesity. PMID:20823291

  17. Capsid is an important determinant for functional complementation of murine leukemia virus and spleen necrosis virus Gag proteins.

    PubMed

    Lee, Sook-Kyung; Boyko, Vitaly; Hu, Wei-Shau

    2007-04-10

    In this report, we examined the abilities and requirements of heterologous Gag proteins to functionally complement each other to support viral replication. Two distantly related gammaretroviruses, murine leukemia virus (MLV) and spleen necrosis virus (SNV), were used as a model system because SNV proteins can support MLV vector replication. Using chimeric or mutant Gag proteins that could not efficiently support MLV vector replication, we determined that a homologous capsid (CA) domain was necessary for the functional complementation of MLV and SNV Gag proteins. Findings from the bimolecular fluorescence complementation assay revealed that MLV and SNV Gag proteins were capable of colocalizing and interacting in cells. Taken together, our results indicated that MLV and SNV Gag proteins can interact in cells; however, a homologous CA domain is needed for functional complementation of MLV and SNV Gag proteins to complete virus replication. This requirement of homologous Gag most likely occurs at a postassembly step(s) of the viral replication. PMID:17156810

  18. Expression of Moloney Murine Leukemia Virus RNase H Rescues the Growth Defect of an Escherichia coli Mutant

    PubMed Central

    Campbell, Andrew G.

    2001-01-01

    A 157-amino-acid fragment of Moloney murine leukemia virus reverse transcriptase encoding RNase H is shown to rescue the growth-defective phenotype of an Escherichia coli mutant. In vitro assays of the recombinant wild-type protein purified from the conditionally defective mutant confirm that it is catalytically active. Mutagenesis of one of the presumptive RNase H-catalytic residues results in production of a protein variant incapable of rescue and which lacks activity in vitro. Analyses of additional active site mutants demonstrate that their encoded variant proteins lack robust activity yet are able to rescue the bacterial mutant. These results suggest that genetic complementation may be useful for in vivo screening of mutant viral RNase H gene fragments and in evaluating their function under conditions that more closely mimic physiological conditions. The rescue system may also be useful in verifying the functional outcomes of mutations based on protein structural predictions and modeling. PMID:11390625

  19. Atomic resolution structure of Moloney murine leukemia virus matrix protein and its relationship to other retroviral matrix proteins.

    PubMed

    Riffel, Nico; Harlos, Karl; Iourin, Oleg; Rao, Zihe; Kingsman, Alan; Stuart, David; Fry, Elizabeth

    2002-12-01

    Matrix proteins associated with the viral membrane are important in the formation of the viral particle and in virus maturation. The 1.0 A crystal structure of the ecotropic Gammaretrovirus Moloney murine leukemia virus (M-MuLV) matrix protein reveals the conserved topology of other retroviral matrix proteins, despite undetectable sequence similarity. The N terminus (normally myristylated) is exposed and adjacent to a basic surface patch, features likely to contribute to membrane binding. The four proteins in the asymmetric unit make varied contacts. The M-MuLV matrix structure is intermediate, between those of the lentiviruses and other retroviruses. The protein fold appears to be maintained, in part, by the conservation of side chain packing, which may provide a useful tool for searching for weak distant similarities in proteins. PMID:12467570

  20. Virus-Specific Messenger RNA and Nascent Polypeptides in Polyribosomes of Cells Replicating Murine Sarcoma-Leukemia Viruses

    PubMed Central

    Vecchio, G.; Tsuchida, N.; Shanmugam, G.; Green, M.

    1973-01-01

    We present evidence that virus-specific RNA is present in polyribosomes of transformed cells replicating the murine sarcoma-leukemia virus complex and that it serves as messenger RNA for the synthesis of viral-coded proteins. Both virus-specific RNA (detected by hybridization with the [3H]DNA product of the viral RNA-directed DNA polymerase) and nascent viral polypeptides (measured by precipitation with antiserum to purified virus) were found in membrane-bound and free polyribosomes. Membrane-bound polyribosomes contained a higher content of both virus-specific RNA and nascent viral polypeptides. From 60 to 70% of viral RNA sequences were released from polyribosomes with EDTA, consistent with a function as messenger RNA. Maximum amounts of both virus-specific RNA and nascent viral polypeptides were found in the polyribosome region sedimenting at about 350 S. PMID:4352969

  1. Murine leukemia virus mutant with a frameshift in the reverse transcriptase coding region: implications for pol gene structure.

    PubMed Central

    Levin, J G; Hu, S C; Rein, A; Messer, L I; Gerwin, B I

    1984-01-01

    The molecular defect in the nonconditional B-tropic MuLV pol mutant, clone 23 (Gerwin et al., J. Virol. 31:741-751, 1979), has been characterized by recombinant DNA technology. The entire mutant genome was cloned from an EcoRI digest of integrated cellular DNA into bacteriophage lambda Charon 4A and then subcloned at the EcoRI site of pBR322. NIH-3T3 cells transfected with the plasmid clone, termed pRTM (RTM, reverse transcriptase mutant), reproduced the properties of clone 23 virus-infected cells. In vivo ligation experiments involving cotransfection of subclones of pRTM and wild-type murine leukemia virus localized the defect in the clone 23 genome to an approximately 400-base-pair region in the pol gene between the SalI and XhoI sites. Sequence analysis of this region in the wild-type and mutant genomes revealed that the mutant has one additional C residue located 231 bases downstream of the last base of the SalI recognition site. This 1-base insertion brings three TGA termination codons into phase. Thus, the mutation in clone 23 leads to premature termination of translation, explaining the presence in clone 23 virions of a truncated polymerase with low levels of enzymatic activity. It was previously shown that the gag precursor is cleaved normally in clone 23-infected cells; therefore, if a virus-coded protease is involved in this cleavage, it must be encoded by sequences upstream of the reverse transcriptase region of the pol gene. This consideration, coupled with the observed molecular weight of the mutant polymerase and our precise determination of its C terminus, have led to a proposal for the genetic organization of the murine leukemia virus pol gene. Images PMID:6205170

  2. Etomidate induces cytotoxic effects and gene expression in a murine leukemia macrophage cell line (RAW264.7).

    PubMed

    Wu, Rick Sai-Chuen; Wu, King-Chuen; Yang, Jai-Sing; Chiou, Shang-Ming; Yu, Chun-Shu; Chang, Shu-Jen; Chueh, Fu-Shin; Chung, Jing-Gung

    2011-06-01

    Etomidate is an important tool in the arsenal of the emergency physician, and it has been used in a variety of scenarios for both intubation and procedural sedation. In the present study, we investigated the cytotoxicity of etomidate including induction of apoptosis, and levels of protein and gene expressions associated with apoptotic cell death in murine leukemia RAW264.7 cells in vitro. Cytotoxic and apoptotic responses to etomidate of RAW264.7 cells, including cell morphological changes and cell viability were examined and measured by phase-contrast microscopy and flow cytometric assay, respectively. Results indicated that etomidate increased apoptotic cell morphological changes and reduced cell viability in RAW264.7 cells. 4',6-Diamidino-2-phenylindole (DAPI) staining also showed that etomidate induced the formation of apoptotic bodies, a characteristic of apoptosis. Results from Western blotting indicated that etomidate enhanced the levels of cytochrome c, apoptosis-inducing factor (AIF), endonuclease G (Endo G), caspase-9, caspase-3 active form and Bax proteins, but it inhibited the expression of Bcl-xl, leading to apoptosis. DNA microarray assay indicated that etomidate increased the expression of 17 genes (LOC676175; Gm14636; 2810021G02Rik; Iltifb; Olfr1167; Ttc30b; Olfr766; Gas5; Rgs1; LOC280487; V1rd4; Hist1h2bc; V1rj3; Gm10366; Olfr192; Gm10002 and Cspp1) and reduced the expression of 15 genes: (Gm10152; Gm5334; Olfr216; Lcn9; Gm10683; Gm5100; Tdgf1; Cypt2; Gm5595; 1700018F24Rik; Gm10417; Maml2; Olfr591; Trdn and Apol7c). In conclusion, etomidate induced cytotoxic and apoptotic effects the in murine leukemia RAW264.7 cells in vitro. PMID:21737642

  3. Structure of glycosylated and unglycosylated gag and gag-pol precursor proteins of Moloney murine leukemia virus.

    PubMed Central

    Saris, C J; van Eenbergen, J; Liskamp, R M; Bloemers, H P

    1983-01-01

    Precursor polyproteins containing translational products of the gag gene of Moloney murine leukemia virus were purified by gel electrophoresis and cleaved into large fragments by hydroxylamine, mild acid hydrolysis, or cyanogen bromide. The hydroxylamine cleavage method (specific for asparagine-glycine bonds) was adapted especially for this study. The electrophoretic mobility and antigenicity of the fragments and, in some cases, the presence or absence of [35S]methionine revealed detailed information on the structure of Pr65gag, gPr80gag, and Pr75gag (the unglycosylated variant of gPr80gag formed in vivo in the presence of tunicamycin or in vitro in a reticulocyte cell-free system). When compared with Pr65gag, gPr80gag contains 7,000 daltons of additional amino acids, presumably as, or as part of, a leader sequence at or very close to its N terminus. We present evidence that this leader may have replaced part of the p15 sequence. Furthermore, gPr80gag contains three separate carbohydrate groups. One is attached to the presumed leader sequence or to the p15 domain, and two are attached to the p30 domain. Each of the Moloney murine leukemia virus gag precursor proteins Pr65gag, gPr80gag, and Pr75gag corresponds with a read-through product into the pol gene. We designated these products Pr180gag-pol, gPr200gag-pol, and Pr190gag-pol (the unglycosylated variant of gPr200gag-pol), respectively. gPr200gag-pol contains all of the extra amino acids and carbohydrate groups present in gPr80gag and at least one carbohydrate group in its pol sequences. Images PMID:6602220

  4. No Evidence for Xenotropic Murine Leukemia-Related Virus Infection in Sweden Using Internally Controlled Multiepitope Suspension Array Serology

    PubMed Central

    Blomberg, Fredrik; Sjösten, Anna; Sheikholvaezin, Ali; Bölin-Wiener, Agnes; Elfaitouri, Amal; Hessel, Sanna; Gottfries, Carl-Gerhard; Zachrisson, Olof; Öhrmalm, Christina; Jobs, Magnus; Pipkorn, Rüdiger

    2012-01-01

    Many syndromes have a large number of differential diagnoses, a situation which calls for multiplex diagnostic systems. Myalgic encephalomyelitis (ME), also named chronic fatigue syndrome (CFS), is a common disease of unknown etiology. A mouse retrovirus, xenotropic murine leukemia-related virus (XMRV), was found in ME/CFS patients and blood donors, but this was not corroborated. However, the paucity of serological investigations on XMRV in humans prompted us to develop a serological assay which cover many aspects of XMRV antigenicity. It is a novel suspension array method, using a multiplex IgG assay with nine recombinant proteins from the env and gag genes of XMRV and 38 peptides based on known epitopes of vertebrate gammaretroviruses. IgG antibodies were sought in 520 blood donors and 85 ME/CFS patients and in positive- and negative-control sera from animals. We found no differences in seroreactivity between blood donors and ME/CFS patients for any of the antigens. This did not support an association between ME/CFS and XMRV infection. The multiplex serological system had several advantages: (i) biotinylated protein G allowed us to run both human and animal sera, which is essential because of a lack of XMRV-positive humans; (ii) a novel quality control was a pan-peptide positive-control rabbit serum; and (iii) synthetic XMRV Gag peptides with degenerate positions covering most of the variation of murine leukemia-like viruses did not give higher background than nondegenerate analogs. The principle may be used for creation of variant tolerant peptide serologies. Thus, our system allows rational large-scale serological assays with built-in quality control. PMID:22787191

  5. Selection of functional tRNA primers and primer binding site sequences from a retroviral combinatorial library: identification of new functional tRNA primers in murine leukemia virus replication

    PubMed Central

    Lund, Anders H.; Duch, Mogens; Pedersen, Finn Skou

    2000-01-01

    Retroviral reverse transcription is initiated from a cellular tRNA molecule and all known exogenous isolates of murine leukemia virus utilise a tRNAPro molecule. While several studies suggest flexibility in murine leukemia virus primer utilisation, studies on human immunodeficiency virus and avian retroviruses have revealed evidence of molecular adaptation towards the specific tRNA isoacceptor used as replication primer. In this study, murine leukemia virus tRNA utilisation is investigated by in vivo screening of a retroviral vector combinatorial library with randomised primer binding sites. While most of the selected primer binding sites are complementary to the 3′-end of tRNAPro, we also retrieved PBS sequences matching four other tRNA molecules and demonstrate that Akv murine leukemia virus vectors may efficiently replicate using tRNAArg(CCU), tRNAPhe(GAA) and a hitherto unknown human tRNASer(CGA). PMID:10637332

  6. Monoclonal antibody to the amino-terminal L sequence of murine leukemia virus glycosylated gag polyproteins demonstrates their unusual orientation in the cell membrane.

    PubMed Central

    Pillemer, E A; Kooistra, D A; Witte, O N; Weissman, I L

    1986-01-01

    To analyze cell surface murine leukemia virus gag protein expression, we have prepared monoclonal antibodies against the spontaneous AKR T lymphoma KKT-2. One of these antibodies, 43-13, detects an AKR-specific viral p12 determinant. A second monoclonal antibody, 43-17, detects a novel murine leukemia virus-related antigen found on glycosylated gag polyproteins (gp95gag, gp85gag, and gp55gag) on the surface of cells infected with and producing ecotropic endogenous viruses, but does not detect antigens within these virions. The 43-17 antibody immunoprecipitates the precursor of the cell surface gag protein whether in its glycosylated or unglycosylated state, but does not detect the cytoplasmic precursor of the virion gag proteins (Pr65gag). Based on these findings, we have localized the 43-17 determinant to the unique amino-terminal part of the glycosylated gag polyprotein (the L domain). We have determined that gp95gag contains L-p15-p12-p30-p10 determinants, whereas gp85gag lacks the carboxyterminal p10 determinant, and gp55gag lacks both p30 and p10 carboxy terminal determinants. Analysis of cell surface gag expression with the 43-17 antibody leads us to propose that the L domain plays a crucial role in (i) the insertion and orientation of murine leukemia virus gag polyproteins in the cell membrane and (ii) the relative abundance of expression of AKR leukemia virus versus Moloney murine leukemia virus glycosylated gag polyproteins in infected cells. Images PMID:2418213

  7. Immunovirotherapy with vesicular stomatitis virus and PD-L1 blockade enhances therapeutic outcome in murine acute myeloid leukemia.

    PubMed

    Shen, Weiwei; Patnaik, Mrinal M; Ruiz, Autumn; Russell, Stephen J; Peng, Kah-Whye

    2016-03-17

    Patients with relapsed acute myeloid leukemia (AML) have limited therapeutic options. Vesicular stomatitis virus (VSV)-interferon β (IFNβ)-sodium iodide symporter (NIS) is an oncolytic VSV encoding IFNβ and the NIS reporter. Syngeneic AML C1498 tumors responded to IV therapy with VSV-murine IFNβ (mIFNβ)-NIS in a dose-dependent manner. Imaging for NIS expression showed robust virus infection within the tumors. Virus infection did not increase programmed death ligand 1 (PD-L1) on tumor cells. Combining VSV-mIFNβ-NIS with anti-PD-L1 antibody (Ab) therapy enhanced antitumor activity compared with treatment with virus alone or Ab alone; this enhancement was not significant at higher VSV-mIFNβ-NIS doses. Systemic VSV therapy reduced systemic C1498-green fluorescent protein (GFP) tumor burden in the blood, bone marrow, spleen, and liver of mice with AML. Combination VSV-mIFNβ-NIS and anti-PD-L1 Ab therapy significantly enhanced the survival of these mice with no evidence of toxicity, compared with isotype control, anti-PD-L1, or virus alone. There was an increase in tumor-infiltrating CD4 and CD8 cells. Single-agent VSV-mIFNβ-NIS virotherapy induced both VSV-specific and GFP-specific CD8 T cells as determined by IFN-γ enzyme-linked immunospot, pentamer, and intracellular IFN-γ staining assays. Both of these responses were further enhanced by addition of anti-PD-L1 Ab. Depletion of CD8 or natural killer cells, but not CD4 cells, resulted in loss of antitumor activity in the VSV/anti-PD-L1 group. Clinical samples from chronic myelomonocytic leukemia and acute myelomonocytic leukemia appear to be especially susceptible to VSV. Overall, our studies show that oncolytic virotherapy combined with immune checkpoint blockade is a promising approach to AML therapy. PMID:26712908

  8. Immunomodulatory and Antioxidant Effects of Purple Sweet Potato Extract in LP-BM5 Murine Leukemia Virus-Induced Murine Acquired Immune Deficiency Syndrome.

    PubMed

    Kim, Ok-Kyung; Nam, Da-Eun; Yoon, Ho-Geun; Baek, Sun Jung; Jun, Woojin; Lee, Jeongmin

    2015-08-01

    The immunomodulatory effects of a dietary supplement of purple sweet potato extract (PSPE) in LP-BM5 murine leukemia virus (MuLV)-induced immune-deficient mice were investigated. Mice were divided into six groups: normal control, infected control (LP-BM5 MuLV infection), positive control (LP-BM5 MuLV infection+dietary supplement of red ginseng 300 mg/kg), purple sweet potato water extract (PSPWE) (LP-BM5 MuLV infection+dietary supplement of PSPE 300 mg/kg), PSP10EE (LP-BM5 MuLV infection+dietary supplement of 10% ethanol PSPE 300 mg/kg), and PSP80EE (LP-BM5 MuLV infection+dietary supplement of 80% ethanol PSPE 300 mg/kg). Dietary supplementation began on the day of LP-BM5 MuLV infection and continued for 12 weeks. Dietary supplementation of PSPE inhibited LP-BM5 MuLV-induced splenomegaly and lymphadenopathy and attenuated the suppression of T- and B-cell proliferation and T helper 1/T helper 2 cytokine imbalance in LP-BM5 MuLV-infected mice. Dietary supplement of PSPE increased the activity of the antioxidant enzymes, superoxide dismutase and glutathione peroxidase. The data suggest that PSPE may ameliorate immune dysfunction due to LP-BM5 MuLV infection by modulating antioxidant defense systems. PMID:26076116

  9. Efficacy of Posaconazole in a Murine Model of Systemic Infection by Saprochaete capitata

    PubMed Central

    Thomson, Pamela; Guarro, Josep; Mayayo, Emilio

    2015-01-01

    The fungus Saprochaete capitata causes opportunistic human infections, mainly in immunocompromised patients with hematological malignancies. The best therapy for this severe infection is still unknown. We evaluated the in vitro killing activity and the in vivo efficacy of posaconazole at 5, 10, or 20 mg/kg twice a day (BID) in a murine neutropenic model of systemic infection with S. capitata by testing a set of six clinical isolates. Posaconazole showed fungistatic activity against all of the isolates tested. The different doses of the drug, especially the highest one, showed good efficacy, measured by prolonged survival, reduction of (1-3)-β-d-glucan levels in serum, tissue burden reduction, and histopathology. PMID:26392490

  10. Murine leukemia virus-based Tat-inducible long terminal repeat replacement vectors: a new system for anti-human immunodeficiency virus gene therapy.

    PubMed Central

    Cannon, P M; Kim, N; Kingsman, S M; Kingsman, A J

    1996-01-01

    We have constructed new murine leukemia virus (MLV)-based vectors (TIN vectors) which, following integration, contain human immunodeficiency virus (HIV) type 1 U3 and R sequences in place of the MLV U3 and R regions. This provides, for the first time, single transcriptional unit retroviral vectors under the control of Tat. TIN vectors have several advantages for anti-HIV gene therapy applications. PMID:8892960

  11. Murine leukemia virus-based Tat-inducible long terminal repeat replacement vectors: a new system for anti-human immunodeficiency virus gene therapy.

    PubMed

    Cannon, P M; Kim, N; Kingsman, S M; Kingsman, A J

    1996-11-01

    We have constructed new murine leukemia virus (MLV)-based vectors (TIN vectors) which, following integration, contain human immunodeficiency virus (HIV) type 1 U3 and R sequences in place of the MLV U3 and R regions. This provides, for the first time, single transcriptional unit retroviral vectors under the control of Tat. TIN vectors have several advantages for anti-HIV gene therapy applications. PMID:8892960

  12. Efficacy of ponatinib against ABL tyrosine kinase inhibitor-resistant leukemia cells

    SciTech Connect

    Okabe, Seiichi Tauchi, Tetsuzo; Tanaka, Yuko; Ohyashiki, Kazuma

    2013-06-07

    Highlights: •Efficacy of ponatinib against ABL tyrosine kinase inhibitor-resistant leukemia cells okabe et al. •Imatinib or nilotinib resistance was involved Src family kinase. •The BCR-ABL point mutation (E334V) was highly resistant to imatinib or nilotinib. •Ponatinib was a powerful strategy against imatinib or nilotinib resistant Ph-positive cells. -- Abstract: Because a substantial number of patients with chronic myeloid leukemia acquire resistance to ABL tyrosine kinase inhibitors (TKIs), their management remains a challenge. Ponatinib, also known as AP24534, is an oral multi-targeted TKI. Ponatinib is currently being investigated in a pivotal phase 2 clinical trial. In the present study, we analyzed the molecular and functional consequences of ponatinib against imatinib- or nilotinib-resistant (R) K562 and Ba/F3 cells. The proliferation of imatinib- or nilotinib-resistant K562 cells did not decrease after treatment with imatinib or nilotinib. Src family kinase Lyn was activated. Point mutation Ba/F3 cells (E334 V) were also highly resistant to imatinib and nilotinib. Treatment with ponatinib for 72 h inhibited the growth of imatinib- and nilotinib-resistant cells. The phosphorylation of BCR-ABL, Lyn, and Crk-L was reduced. This study demonstrates that ponatinib has an anti-leukemia effect by reducing ABL and Lyn kinase activity and this information may be of therapeutic relevance.

  13. FLT3 mutations confer enhanced proliferation and survival properties to multipotent progenitors in a murine model of chronic myelomonocytic leukemia

    PubMed Central

    Lee, Benjamin H.; Tothova, Zuzana; Levine, Ross L.; Anderson, Kristina; Buza-Vidas, Natalija; Cullen, Dana E.; McDowell, Elizabeth P.; Adelsperger, Jennifer; Fröhling, Stefan; Huntly, Brian J.P.; Beran, Miloslav; Jacobsen, Sten Eirik; Gilliland, D. Gary

    2007-01-01

    SUMMARY Despite their known transforming properties, the effects of leukemogenic FLT3-ITD mutations on hematopoietic stem and multipotent progenitor cells and on hematopoietic differentiation are not well understood. We report a mouse model harboring an ITD in the murine Flt3 locus that develops myeloproliferative disease resembling CMML and further identified FLT3-ITD mutations in a subset of human CMML. These findings correlated with an increase in number, cell cycling and survival of multipotent stem and progenitor cells in an ITD dose-dependent manner in animals that exhibited alterations within their myeloid progenitor compartments and a block in normal B-cell development. This model provides insights into the consequences of constitutive signaling by an oncogenic tyrosine kinase on hematopoietic progenitor quiescence, function, and cell fate. SIGNIFICANCE Activating FLT3 mutations are among the most common genetic events in AML and confer a poor clinical prognosis. Essential to our understanding of how these lesions contribute to myeloid leukemia is the development of a Flt3-ITD ‘knock-in’ murine model that has allowed examination of the consequences of constitutive FLT3 signaling on primitive hematopoietic progenitors when expressed at appropriate physiologic levels. These animals informed us to the existence of FLT3-ITD-positive human CMML, which has clinical importance given the availability of FLT3 small molecule inhibitors. This model will not only serve as a powerful biological tool to identify mutations that cooperate with FLT3 in leukemogenesis, but also to assess molecular therapies that target either FLT3 or components of its signaling pathways. PMID:17936561

  14. Effective treatment of a murine model of adult T-cell leukemia using 211At-7G7/B6 and its combination with unmodified anti-Tac (daclizumab) directed toward CD25.

    PubMed

    Zhang, Zhuo; Zhang, Meili; Garmestani, Kayhan; Talanov, Vladimir S; Plascjak, Paul S; Beck, Barbara; Goldman, Carolyn; Brechbiel, Martin W; Waldmann, Thomas A

    2006-08-01

    Adult T-cell leukemia (ATL) consists of an overabundance of T cells, which express CD25. Therapeutic efficacy of astatine-211 ((211)At)-labeled murine monoclonal antibody 7G7/B6 alone and in combination with daclizumab was evaluated in nonobese diabetic/severe combined immunodeficiency (NOD/SCID) mice given injections of MET-1 human T-cell leukemia cells. Daclizumab and 7G7/B6 are directed toward different epitopes of CD25. Either a single dose of 12 microCi (0.444 MBq) (211)At-7G7/B6 per mouse given intravenously or receptor-saturating doses of daclizumab given at 100 microg weekly for 4 weeks intravenously inhibited tumor growth as monitored by serum levels of human beta-2 microglobulin (beta(2)mu) and by prolonged survival of leukemia-bearing mice compared with the control groups (P < .001). The combination of 2 agents enhanced the antitumor effect when compared with groups treated with 12 microCi (0.444 MBq) of (211)At-7G7/B6 (P < .05) or daclizumab alone (P < .05). The median survival duration of the PBS group was 62.6 days and 61.5 days in the radiolabeled nonspecific antibody (211)At-11F11-treated group. In contrast, 91% of mice in the combination group survived through day 94. These results that demonstrate a significantly improved therapeutic efficacy by combining (211)At-7G7/B6 with daclizumab support a clinical trial of this regimen in patients with ATL. PMID:16569769

  15. Tandemization of a Subregion of the Enhancer Sequences from SRS 19-6 Murine Leukemia Virus Associated with T-Lymphoid but Not Other Leukemias

    PubMed Central

    Granger, Steven W.; Bundy, Linda M.; Fan, Hung

    1999-01-01

    Most simple retroviruses induce tumors of a single cell type when infected into susceptible hosts. The SRS 19-6 murine leukemia virus (MuLV), which originated in mainland China, induces leukemias of multiple cellular origins. Indeed, infected mice often harbor more than one tumor type. Since the enhancers of many MuLVs are major determinants of tumor specificity, we tested the role of the SRS 19-6 MuLV enhancers in its broad disease specificity. The enhancer elements of the Moloney MuLV (M-MuLV) were replaced by the 170-bp enhancers of SRS 19-6 MuLV, yielding the recombinants ΔMo+SRS+ and ΔMo+SRS− M-MuLV. M-MuLV normally induces T-lymphoid tumors in all infected mice. Surprisingly, when neonatal mice were inoculated with ΔMo+SRS+ or ΔMo+SRS− M-MuLV, all tumors were of T-lymphoid origin, typical of M-MuLV rather than SRS 19-6 MuLV. Thus, the SRS 19-6 MuLV enhancers did not confer the broad disease specificity of SRS 19-6 MuLV to M-MuLV. However, all tumors contained ΔMo+SRS M-MuLV proviruses with common enhancer alterations. These alterations consisted of tandem multimerization of a subregion of the SRS 19-6 enhancers, encompassing the conserved LVb and core sites and adjacent sequences. Moreover, when tumors induced by the parental SRS 19-6 MuLV were analyzed, most of the T-lymphoid tumors had similar enhancer alterations in the same region whereas tumors of other lineages retained the parental SRS 19-6 MuLV enhancers. These results emphasize the importance of a subregion of the SRS 19-6 MuLV enhancer in induction of T-cell lymphoma. The relevant sequences were consistent with crucial sequences for T-cell lymphomagenesis identified for other MuLVs such as M-MuLV and SL3-3 MuLV. These results also suggest that other regions of the SRS 19-6 MuLV genome contribute to its broad leukemogenic spectrum. PMID:10438804

  16. Variable regions A and B in the envelope glycoproteins of feline leukemia virus subgroup B and amphotropic murine leukemia virus interact with discrete receptor domains.

    PubMed Central

    Tailor, C S; Kabat, D

    1997-01-01

    The surface (SU) envelope glycoproteins of feline leukemia virus subgroup B (FeLV-B) and amphotropic murine leukemia virus (A-MLV) are highly related, even in the variable regions VRA and VRB that have been shown to be required for receptor recognition. However, FeLV-B and A-MLV use different sodium-dependent phosphate symporters, Pit1 and Pit2, respectively, as receptors for infection. Pit1 and Pit2 are predicted to have 10 membrane-spanning domains and five extracellular loops. The close relationship of the retroviral envelopes enabled us to generate pseudotype virions carrying chimeric FeLV-B/A-MLV envelope glycoproteins. We found that some of the pseudotype viruses could not use Pit1 or Pit2 proteins but could efficiently utilize specific chimeric Pit1/Pit2 proteins as receptors. By studying Mus dunni tail fibroblasts expressing chimeric Pit1/Pit2 proteins and pseudotype virions carrying chimeric FeLV-B/A-MLV envelopes, we show that FeLV-B and A-MLV VRA and VRB interact in a modular manner with specific receptor domains. Our results suggest that FeLV-B VRA interacts with Pit1 extracellular loops 4 and 5 and that residues Phe-60 and Pro-61 of FeLV-B VRA are essential for receptor choice. However, this interaction is insufficient for infection, and an additional interaction between FeLV-B VRB and Pit1 loop 2 is essential. Similarly, A-MLV infection requires interaction of A-MLV VRA with Pit2 loops 4 and 5 and VRB with Pit2 loop 2, with residues Tyr-60 and Val-61 of A-MLV VRA being critical for receptor recognition. Together, our results suggest that FeLV-B and A-MLV infections require two major discrete interactions between the viral SU envelope glycoproteins and their respective receptors. We propose a common two-step mechanism for interaction between retroviral envelope glycoproteins and cell surface receptors. PMID:9371598

  17. Induction of donor-type chimerism in murine recipients of bone marrow allografts by different radiation regimens currently used in treatment of leukemia patients

    SciTech Connect

    Salomon, O.; Lapidot, T.; Terenzi, A.; Lubin, I.; Rabi, I.; Reisner, Y. )

    1990-11-01

    Three radiation protocols currently used in treatment of leukemia patients before bone marrow transplantation (BMT) were investigated in a murine model (C57BL/6----C3H/HeJ) for BM allograft rejection. These include (a) a single dose of total body irradiation (8.5 Gy TBI delivered at a dose rate of 0.2 Gy/min), (b) fractionated TBI 12 Gy administered in six fractions, 2 Gy twice a day in 3 days, delivered at a dose rate of 0.1 Gy/min, and (c) hyperfractionated TBI (14.4 Gy administered in 12 fractions, 1.2 Gy three times a day in 3 days, delivered at a dose rate of 0.1 Gy/min). Donor-type chimerism 6 to 8 weeks after BMT and hematologic reconstitution on day 12 after BMT found in these groups were compared with results obtained in mice conditioned with 8 Gy TBI delivered at a dose rate of 0.67 Gy/min, routinely used in this murine model. The results in both parameters showed a marked advantage for the single dose 8.5 Gy TBI over all the other treatments. This advantage was found to be equivalent to three- to fourfold increment in the BM inoculum when compared with hyperfractionated radiation, which afforded the least favorable conditions for development of donor-type chimerism. The fractionated radiation protocol was equivalent in its efficacy to results obtained in mice irradiated by single-dose 8 Gy TBI, both of which afforded a smaller but not significant advantage over the hyperfractionated protocol. This model was also used to test the effect of radiation dose rate on the development of donor-type chimerism. A significant enhancement was found after an increase in dose rate from 0.1 to 0.7 Gy/min. Further enhancement could be achieved when the dose rate was increased to 1.3 Gy/min, but survival at this high dose rate was reduced.

  18. DNA methyltransferase inhibition increases efficacy of adoptive cellular immunotherapy of murine breast cancer.

    PubMed

    Terracina, Krista P; Graham, Laura J; Payne, Kyle K; Manjili, Masoud H; Baek, Annabel; Damle, Sheela R; Bear, Harry D

    2016-09-01

    Adoptive T cell immunotherapy is a promising approach to cancer treatment that currently has limited clinical applications. DNA methyltransferase inhibitors (DNAMTi) have known potential to affect the immune system through multiple mechanisms that could enhance the cytotoxic T cell responses, including: upregulation of tumor antigen expression, increased MHC class I expression, and blunting of myeloid derived suppressor cells (MDSCs) expansion. In this study, we have investigated the effect of combining the DNAMTi, decitabine, with adoptive T cell immunotherapy in the murine 4T1 mammary carcinoma model. We found that expression of neu, MHC class I molecules, and several murine cancer testis antigens (CTA) was increased by decitabine treatment of 4T1 cells in vitro. Decitabine also increased expression of multiple CTA in two human breast cancer cell lines. Decitabine-treated 4T1 cells stimulated greater IFN-gamma release from tumor-sensitized lymphocytes, implying increased immunogenicity. Expansion of CD11b + Gr1 + MDSC in 4T1 tumor-bearing mice was significantly diminished by decitabine treatment. Decitabine treatment improved the efficacy of adoptive T cell immunotherapy in mice with established 4T1 tumors, with greater inhibition of tumor growth and an increased cure rate. Decitabine may have a role in combination with existing and emerging immunotherapies for breast cancer. PMID:27416831

  19. Murine Model Imitating Chronic Wound Infections for Evaluation of Antimicrobial Photodynamic Therapy Efficacy.

    PubMed

    Fila, Grzegorz; Kasimova, Kamola; Arenas, Yaxal; Nakonieczna, Joanna; Grinholc, Mariusz; Bielawski, Krzysztof P; Lilge, Lothar

    2016-01-01

    It is generally acknowledged that the age of antibiotics could come to an end, due to their widespread, and inappropriate use. Particularly for chronic wounds alternatives are being thought. Antimicrobial Photodynamic Therapy (APDT) is a potential candidate, and while approved for some indications, such as periodontitis, chronic sinusitis and other niche indications, its use in chronic wounds is not established. To further facilitate the development of APDT in chronic wounds we present an easy to use animal model exhibiting the key hallmarks of chronic wounds, based on full-thickness skin wounds paired with an optically transparent cover. The moisture-retaining wound exhibited rapid expansion of pathogen colonies up to 8 days while not jeopardizing the host survival. Use of two bioluminescent pathogens; methicillin resistant Staphylococcus aureus (MRSA) and Pseudomonas aeruginosa permits real time monitoring of the pathogens. The murine model was employed to evaluate the performance of four different photosensitizers as mediators in Photodynamic Therapy. While all four photosensitizers, Rose Bengal, porphyrin TMPyP, New Methylene Blue, and TLD1411 demonstrated good to excellent antimicrobial efficacy in planktonic solutions at 1 to 50 μM concentrations, whereas in in vivo the growth delay was limited with 24-48 h delay in pathogen expansion for MRSA, and we noticed longer growth suppression of P. aeruginosa with TLD1411 mediated Photodynamic Therapy. The murine model will enable developing new strategies for enhancement of APDT for chronic wound infections. PMID:27555843

  20. Murine Model Imitating Chronic Wound Infections for Evaluation of Antimicrobial Photodynamic Therapy Efficacy

    PubMed Central

    Fila, Grzegorz; Kasimova, Kamola; Arenas, Yaxal; Nakonieczna, Joanna; Grinholc, Mariusz; Bielawski, Krzysztof P.; Lilge, Lothar

    2016-01-01

    It is generally acknowledged that the age of antibiotics could come to an end, due to their widespread, and inappropriate use. Particularly for chronic wounds alternatives are being thought. Antimicrobial Photodynamic Therapy (APDT) is a potential candidate, and while approved for some indications, such as periodontitis, chronic sinusitis and other niche indications, its use in chronic wounds is not established. To further facilitate the development of APDT in chronic wounds we present an easy to use animal model exhibiting the key hallmarks of chronic wounds, based on full-thickness skin wounds paired with an optically transparent cover. The moisture-retaining wound exhibited rapid expansion of pathogen colonies up to 8 days while not jeopardizing the host survival. Use of two bioluminescent pathogens; methicillin resistant Staphylococcus aureus (MRSA) and Pseudomonas aeruginosa permits real time monitoring of the pathogens. The murine model was employed to evaluate the performance of four different photosensitizers as mediators in Photodynamic Therapy. While all four photosensitizers, Rose Bengal, porphyrin TMPyP, New Methylene Blue, and TLD1411 demonstrated good to excellent antimicrobial efficacy in planktonic solutions at 1 to 50 μM concentrations, whereas in in vivo the growth delay was limited with 24–48 h delay in pathogen expansion for MRSA, and we noticed longer growth suppression of P. aeruginosa with TLD1411 mediated Photodynamic Therapy. The murine model will enable developing new strategies for enhancement of APDT for chronic wound infections. PMID:27555843

  1. Effects of 3′ Untranslated Region Mutations on Plus-Strand Priming during Moloney Murine Leukemia Virus Replication

    PubMed Central

    Robson, Nicole D.; Telesnitsky, Alice

    1999-01-01

    A conserved purine-rich motif located near the 3′ end of retroviral genomes is involved in the initiation of plus-strand DNA synthesis. We mutated sequences both within and flanking the Moloney murine leukemia virus polypurine tract (PPT) and determined the effects of these alterations on viral DNA synthesis and replication. Our results demonstrated that both changes in highly conserved PPT positions and a mutation that left only the cleavage-proximal half of the PPT intact led to delayed replication and reduced the colony-forming titer of replication defective retroviral vectors. A mutation that altered the cleavage proximal half of the PPT and certain 3′ untranslated region mutations upstream of the PPT were incompatible with or severely impaired viral replication. To distinguish defects in plus-strand priming from other replication defects and to assess the relative use of mutant and wild-type PPTs, we examined plus-strand priming from an ectopic, secondary PPT inserted in U3. The results demonstrated that the analyzed mutations within the PPT primarily affected plus-strand priming whereas mutations upstream of the PPT appeared to affect both plus-strand priming and other stages of viral replication. PMID:9882295

  2. The "putative" leucine zipper region of murine leukemia virus transmembrane protein (P15e) is essential for viral infectivity.

    PubMed

    Ramsdale, E E; Kingsman, S M; Kingsman, A J

    1996-06-01

    In order to determine the role of the putative leucine zipper region of murine leukemia virus (MLV) transmembrane protein p15E, nine mutations in this region were introduced by site-directed mutagenesis. None of these mutations affected the expression or transport of the envelope protein or incorporation into virions. The mutants were analyzed for their ability to infect NIH3T3 cells and to induce cell fusion in a rat XC cell fusion assay. Mutations removing the charge of the hydrophilic residues reduced infectivity in NIH3T3 cells but had either no effect or a minor effect on envelope-induced XC cell fusion. Six mutations of hydrophobic residues of the putative leucine zipper region were constructed; four completely abolished the ability to infect NIH3T3 cells and these mutant envelopes were also unable to induce cell fusion in the XC cell fusion assay. These data demonstrate the absolute requirement for the putative leucine zipper region for both fusion and infection of MLV. PMID:8659102

  3. Mutagenesis analysis of the murine leukemia virus matrix protein: identification of regions important for membrane localization and intracellular transport.

    PubMed

    Soneoka, Y; Kingsman, S M; Kingsman, A J

    1997-07-01

    We have created two sets of substitution mutations in the Moloney murine leukemia virus (Mo-MuLV) matrix protein in order to identify domains involved in association with the plasma membrane and in incorporation of the viral envelope glycoproteins into virus particles. The first set of mutations was targeted at putative membrane-associating regions similar to those of the human immunodeficiency virus type 1 matrix protein, which include a polybasic region at the N terminus of the Mo-MuLV matrix protein and two regions predicted to form beta strands. The second set of mutations was created within hydrophobic residues to test for the production of virus particles lacking envelope proteins, with the speculation of an involvement of the membrane-spanning region of the envelope protein in incorporation into virus particles. We have found that mutation of the N-terminal polybasic region redirected virus assembly to the cytoplasm, and we show that tryptophan residues may also play a significant role in the intracellular transport of the matrix protein. In total, 21 mutants of the Mo-MuLV matrix protein were produced, but we did not observe any mutant virus particles lacking the envelope glycoproteins, suggesting that a direct interaction between the Mo-MuLV matrix protein and envelope proteins either may not exist or may occur through multiple redundant interactions. PMID:9188629

  4. Basic Residues in the Matrix Domain and Multimerization Target Murine Leukemia Virus Gag to the Virological Synapse

    PubMed Central

    Li, Fei; Jin, Jing; Herrmann, Christin

    2013-01-01

    Murine leukemia virus (MLV) can efficiently spread in tissue cultures by polarizing assembly to virological synapses. The viral envelope glycoprotein (Env) establishes cell-cell contacts and subsequently recruits Gag by a process that depends on its cytoplasmic tail. MLV Gag is recruited to virological synapses through the matrix domain (MA) (J. Jin, F. Li, and W. Mothes, J. Virol. 85:7672–7682, 2011). However, how MA targets Gag to sites of cell-cell contact remains unknown. Here we report that basic residues within MA are critical for directing MLV Gag to virological synapses. Alternative membrane targeting domains (MTDs) containing multiple basic residues can efficiently substitute MA to direct polarized assembly. Similarly, mutations in the polybasic cluster of MA that disrupt Gag polarization can be rescued by N-terminal addition of MTDs containing basic residues. MTDs containing basic residues alone fail to be targeted to the virological synapse. Systematic deletion experiments reveal that domains within Gag known to mediate Gag multimerization are also required. Thus, our data predict the existence of a specific “acidic” interface at virological synapses that mediates the recruitment of MLV Gag via the basic cluster of MA and Gag multimerization. PMID:23616653

  5. Alpha-phellandrene-induced DNA damage and affect DNA repair protein expression in WEHI-3 murine leukemia cells in vitro.

    PubMed

    Lin, Jen-Jyh; Wu, Chih-Chung; Hsu, Shu-Chun; Weng, Shu-Wen; Ma, Yi-Shih; Huang, Yi-Ping; Lin, Jaung-Geng; Chung, Jing-Gung

    2015-11-01

    Although there are few reports regarding α-phellandrene (α-PA), a natural compound from Schinus molle L. essential oil, there is no report to show that α-PA induced DNA damage and affected DNA repair associated protein expression. Herein, we investigated the effects of α-PA on DNA damage and repair associated protein expression in murine leukemia cells. Flow cytometric assay was used to measure the effects of α-PA on total cell viability and the results indicated that α-PA induced cell death. Comet assay and 4,6-diamidino-2-phenylindole dihydrochloride staining were used for measuring DNA damage and condensation, respectively, and the results indicated that α-PA induced DNA damage and condensation in a concentration-dependent manner. DNA gel electrophoresis was used to examine the DNA damage and the results showed that α-PA induced DNA damage in WEHI-3 cells. Western blotting assay was used to measure the changes of DNA damage and repair associated protein expression and the results indicated that α-PA increased p-p53, p-H2A.X, 14-3-3-σ, and MDC1 protein expression but inhibited the protein of p53, MGMT, DNA-PK, and BRCA-1. PMID:24861204

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

    SciTech Connect

    Zhou, Dongwen; Chung, Suhman; Miller, Maria; Le Grice, Stuart F.J.; Wlodawer, Alexander

    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 intact 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.

  7. Immunoprophylactic potential of wheat grass extract on benzene-induced leukemia: An in vivo study on murine model

    PubMed Central

    Khan, Neelofar; Ganeshpurkar, Aditya; Dubey, Nazneen; Bansal, Divya

    2015-01-01

    Objectives: Wheat grass (Triticum aestivum) is a gift of nature given to mankind. A number of scientific research on wheatgrass establishes its anticancer and antioxidant potential. Current work was focused to determine antileukemic effect of wheat grass. Materials and Methods: The commercial wheatgrass powder was extracted with 95% of methanol. Methanol extract of wheat grass was studied for acute oral toxicity as per revised Organization for Economic Cooperation and Development Guidelines number 423. Leukemia was successfully induced in Wister rats by intravenous injection of benzene. The blood was collected and analyzed for hematological parameters. Phagocytotic activity of the extract was determined. Results: Phytochemical screening revealed the presence of flavonoids, phenolics, carbohydrates, and amino acids. From acute toxicity studies, it was found that the methanol extract of wheatgrass was safe up to a dose level of 2000 mg/kg of body weight. Outcomes of hematological parameters in various experimental groups of murine model demonstrated antileukemic effect of extract. Methanol extract of wheatgrass aroused the process of phagocytosis of killed Candida albicans and also demonstrated a significant chemotactic activity at all tested concentrations. Conclusion: In the current work, methanol extract of wheat grass demonstrated antileukemic potential that might be due to the presence of flavonoids and polyphenolics in it. Further isolation, structural characterization of active constituents is necessary to extrapolate the mechanism of action. PMID:26288471

  8. Retroviral reverse transcriptase inhibitory activity in Thai herbs and spices: screening with Moloney murine leukemia viral enzyme.

    PubMed

    Suthienkul, O; Miyazaki, O; Chulasiri, M; Kositanont, U; Oishi, K

    1993-12-01

    Fifty-seven Thai herbs and spices were examined for their retroviral reverse transcriptase inhibitory activity. All herbs and spices were extracted with hot-water and methanol. Reverse transcriptase inhibitory activity of the extracts was determined by using Moloney Murine Leukemia Virus reverse transcriptase (M-MuLV-RT) reacted with 3H-dTTP and radioactivity measured with a scintillation counter. Eighty-one per cent (46/57) of hot-water extracts and 54% (31/57) of methanol extracts showed inhibitory activities. At a concentration of 125 micrograms/ml, 13% (6/46) of hot-water extracts, namely Eugenia caryophyllus Bullock et Harrison, Phyllanthus urinaria Linn., Terminalia belerica Roxb., Nelumbo nucifera Gaertn., Psidium guajava Linn. and Lawsonia inermis Linn., had a relative inhibitory ratio (IR) over 50%. They showed ratios of 100%, 91%, 75%, 74%, 61% and 60%, respectively. For methanol extracts, only 10% (3/31) had IR values over 50%. They were T. belerica, E. caryophyllus and N. nucifera which exhibited IR values of 83%, 54% and 54%, respectively. PMID:7524165

  9. Proviral Integration Site for Moloney Murine Leukemia Virus (PIM) Kinases Promote Human T Helper 1 Cell Differentiation*

    PubMed Central

    Tahvanainen, Johanna; Kyläniemi, Minna K.; Kanduri, Kartiek; Gupta, Bhawna; Lähteenmäki, Hanna; Kallonen, Teemu; Rajavuori, Anna; Rasool, Omid; Koskinen, Päivi J.; Rao, Kanury V. S.; Lähdesmäki, Harri; Lahesmaa, Riitta

    2013-01-01

    The differentiation of human primary T helper 1 (Th1) cells from naïve precursor cells is regulated by a complex, interrelated signaling network. The identification of factors regulating the early steps of Th1 cell polarization can provide important insight in the development of therapeutics for many inflammatory and autoimmune diseases. The serine/threonine-specific proviral integration site for Moloney murine leukemia virus (PIM) kinases PIM1 and PIM2 have been implicated in the cytokine-dependent proliferation and survival of lymphocytes. We have established that the third member of this family, PIM3, is also expressed in human primary Th cells and identified a new function for the entire PIM kinase family in T lymphocytes. Although PIM kinases are expressed more in Th1 than Th2 cells, we demonstrate here that these kinases positively influence Th1 cell differentiation. Our RNA interference results from human primary Th cells also suggest that PIM kinases promote the production of IFNγ, the hallmark cytokine produced by Th1 cells. Consistent with this, they also seem to be important for the up-regulation of the critical Th1-driving factor, T box expressed in T cells (T-BET), and the IL-12/STAT4 signaling pathway during the early Th1 differentiation process. In summary, we have identified PIM kinases as new regulators of human primary Th1 cell differentiation, thus providing new insights into the mechanisms controlling the selective development of human Th cell subsets. PMID:23209281

  10. Tyrosine phosphorylation of HSC70 and its interaction with RFC mediates methotrexate resistance in murine L1210 leukemia cells.

    PubMed

    Liu, Tuoen; Singh, Ratan; Rios, Zechary; Bhushan, Alok; Li, Mengxiong; Sheridan, Peter P; Bearden, Shawn E; Lai, James C K; Agbenowu, Senyo; Cao, Shousong; Daniels, Christopher K

    2015-02-01

    We previously identified and characterized a 66-68 kDa membrane-associated, tyrosine phosphorylated protein in murine leukemia L1210 cells as HSC70 which is a methotrexate (MTX)-binding protein. In order to further characterize the functional role of HSC70 in regulating MTX resistance in L1210 cells, we first showed that HSC70 colocalizes and interacts with reduced folate carrier (RFC) in L1210 cells by confocal laser scanning microscopy and Duolink in situ proximity ligation assay. The tyrosine phosphorylation status of HSC70 found in the membrane fraction was different from the parental L1210/0 and cisplatin (CDDP)-MTX cross resistant L1210/DDP cells. In MTX-binding assays, HSC70 from L1210/DDP cells showed less affinity for MTX-agarose beads than that of L1210/0 cells. In addition, genistein (a tyrosine phosphorylation inhibitor) significantly enhanced the resistance of L1210/0 cells to MTX. Moreover, site-directed mutation studies indicated the importance of tyrosine phosphorylation of HSC70 in regulating its binding to MTX. These findings suggest that tyrosine phosphorylation of HSC70 regulates the transportation of MTX into the cells via the HSC70-RFC system and contributes to MTX resistance in L1210 cells. PMID:25444929

  11. Multiparameter analyses of spontaneous nonthymic lymphomas occurring in NFS/N mice congenic for ecotropic murine leukemia viruses.

    PubMed Central

    Fredrickson, T. N.; Morse, H. C.; Yetter, R. A.; Rowe, W. P.; Hartley, J. W.; Pattengale, P. K.

    1985-01-01

    Mouse strains congenic for ecotropic retrovirus genes have a much higher frequency of spontaneous lymphomas than the background NFS/N strain. In this study, most of these lymphomas have been identified as B-cell in origin by morphologic features, identification of immunoglobulin class, and cell-surface antigens. The classification suggested by Pattengale and Taylor proved to be applicable to the lymphomas studied. Most were of large follicular center cells and are considered typical of the type formerly designated as "reticulum cell sarcoma, type B." Many lymphomas contained a large proportion of nonneoplastic cells which partially obscured their neoplastic component. The role of ecotropic murine leukemia viruses as etiologic agents for B-cell lymphomas remains equivocal. However, because the only difference between the NFS/N and congenic mice is the expression of viruses in the latter, it appears that these viruses are somehow involved in induction of B-cell lymphomas. Images Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Figure 8 Figure 9 Figure 10 PMID:2998195

  12. Sodium-Dependent myo-Inositol Transporter 1 Is a Cellular Receptor for Mus cervicolor M813 Murine Leukemia Virus

    PubMed Central

    Hein, Sibyll; Prassolov, Vladimir; Zhang, Yuanming; Ivanov, Dmitry; Löhler, Jürgen; Ross, Susan R.; Stocking, Carol

    2003-01-01

    Retrovirus infection is initiated by binding of the surface (SU) portion of the viral envelope glycoprotein (Env) to specific receptors on cells. This binding triggers conformational changes in the transmembrane portion of Env, leading to membrane fusion and cell entry, and is thus a major determinant of retrovirus tissue and species tropism. The M813 murine leukemia virus (MuLV) is a highly fusogenic gammaretrovirus, isolated from Mus cervicolor, whose host range is limited to mouse cells. To delineate the molecular mechanisms of its restricted host range and its high fusogenic potential, we initiated studies to characterize the cell surface protein that mediates M813 infection. Screening of the T31 mouse-hamster radiation hybrid panel for M813 infectivity localized the receptor gene to the distal end of mouse chromosome 16. Expression of one of the likely candidate genes (slc5a3) within this region in human cells conferred susceptibility to both M813 infection and M813-induced fusogenicity. slc5a3 encodes sodium myo-inositol transporter 1 (SMIT1), thus adding another sodium-dependent transporter to the growing list of proteins used by MuLVs for cell entry. Characterization of SMIT1 orthologues in different species identified several amino acid variations within two extracellular loops that may restrict susceptibility to M813 infection. PMID:12719585

  13. Pbx3 and Meis1 cooperate through multiple mechanisms to support Hox-induced murine leukemia

    PubMed Central

    Garcia-Cuellar, Maria-Paz; Steger, Julia; Füller, Elisa; Hetzner, Katrin; Slany, Robert K.

    2015-01-01

    Hox homeobox transcription factors drive leukemogenesis efficiently only in the presence of Meis or Pbx proteins. Here we show that Pbx3 and Meis1 need to dimerize to support Hox-induced leukemia and we analyze the molecular details of this cooperation. In the absence of Pbx3, Meis1 was highly unstable. As shown by a deletion analysis Meis1 degradation was contingent on a motif coinciding with the Pbx-binding domain. Either deletion of this sequence or binding to Pbx3 prolonged the half-life of Meis1 by preventing its ubiquitination. Meis1 break-down could also be blocked by inhibition of the ubiquitin proteasome system, indicating tight post-transcriptional control. In addition, Meis1 and Pbx3 cooperated genetically as overexpression of Pbx3 induced endogenous Meis1 transcription. These functional interactions translated into in vivo activity. Blocking Meis1/Pbx3 dimerization abrogated the ability to enhance proliferation and colony-forming cell numbers in primary cells transformed by Hoxa9. Furthermore, expression of Meis1 target genes Flt3 and Trib2 was dependent on Pbx3/Meis1 dimerization. This correlated with the requirement of Meis1 to bind Pbx3 in order to form high affinity DNA/Hoxa9/Meis1/Pbx3 complexes in vitro. Finally, kinetics and severity of disease in transplantation assays indicated that Pbx3/Meis1 dimers are rate-limiting factors for Hoxa9-induced leukemia. PMID:25911551

  14. New Mouse Models to Investigate the Efficacy of Drug Combinations in Human Chronic Myeloid Leukemia.

    PubMed

    Lin, Hanyang; Woolfson, Adrian; Jiang, Xiaoyan

    2016-01-01

    Chronic myeloid leukemia (CML) comprises a simple and effective paradigm for generating new insights into the cellular origin, pathogenesis, and treatment of many types of human cancer. In particular, mouse models of CML have greatly facilitated the understanding of the underlying molecular mechanisms and pathogenesis of this disease and have led to the identification of new drug targets that in some cases offer the possibility of functional cure. There are currently three established CML mouse models: the BCR-ABL transgenic model, the BCR-ABL retroviral transduction/transplantation model, and the xenotransplant immunodeficient model. Each has its own unique advantages and disadvantages. Depending on the question of interest, some models may be more appropriate than others. In this chapter, we describe a newly developed xenotransplant mouse model to determine the efficacy of novel therapeutic agents, either alone or in combination. The model facilitates the evaluation of the frequency of leukemic stem cells with long-term leukemia-initiating activity, a critical subcellular population that causes disease relapse and progression, through the utilization of primary CD34(+) CML stem/progenitor cells obtained from CML patients at diagnosis and prior to drug treatment. We have also investigated the effectiveness of new combination treatment strategies designed to prevent the development of leukemia in vivo using BCR-ABL (+) blast crisis cells as a model system. These types of in vivo studies are important for the prediction of individual patient responses to drug therapy, and have the potential to facilitate the design of personalized combination therapy strategies. PMID:27581149

  15. β2 Agonists enhance the efficacy of simultaneous enzyme replacement therapy in murine Pompe disease.

    PubMed

    Koeberl, Dwight D; Li, Songtao; Dai, Jian; Thurberg, Beth L; Bali, Deeksha; Kishnani, Priya S

    2012-02-01

    Enzyme replacement therapy (ERT) with recombinant human acid α-glucosidase (rhGAA) has improved clinical outcomes in patients with Pompe disease; however, the response of skeletal muscle and the central nervous system to ERT has been attenuated. The poor response of skeletal muscle to ERT has been attributed to the low abundance of the cation-independent mannose-6-phosphate receptor (CI-MPR), which mediates receptor-mediated uptake of rhGAA. Hence the ability of adjunctive therapy with β2-agonists to increase CI-MPR expression in skeletal muscle was evaluated during ERT in murine Pompe disease with regard to reversal of neuromuscular involvement. Mice with Pompe disease were treated with weekly rhGAA injections (20 mg/kg) and a selective β2-agonist, either albuterol (30 mg/l in drinking water) or low-dose clenbuterol (6 mg/l in drinking water). Biochemical correction was enhanced by β2-agonist treatment in both muscle and the cerebellum, indicating that adjunctive therapy could enhance efficacy from ERT in Pompe disease with regard to neuromuscular involvement. Intriguingly, clenbuterol slightly reduced muscle glycogen content independent of CI-MPR expression, as demonstrated in CI-MPR knockout/GAA knockout mice that were otherwise resistant to ERT. Thus, adjunctive therapy with β2 agonists might improve the efficacy of ERT in Pompe disease and possibly other lysosomal storage disorders through enhancing receptor-mediated uptake of recombinant lysosomal enzymes. PMID:22154081

  16. β2 Agonists Enhance the Efficacy of Simultaneous Enzyme Replacement Therapy in Murine Pompe Disease

    PubMed Central

    Koeberl, Dwight D.; Li, Songtao; Dai, Jian; Thurberg, Beth L.; Bali, Deeksha; Kishnani, Priya S.

    2011-01-01

    Enzyme replacement therapy (ERT) with recombinant human acid α-glucosidase (rhGAA) has improved clinical outcomes in patients with Pompe disease; however, the response of skeletal muscle and the central nervous system to ERT has been attenuated. The poor response of skeletal muscle to ERT has been attributed to the low abundance of the cation-independent mannose-6-phosphate receptor (CI-MPR), which mediates receptor-mediated uptake of rhGAA. Hence the ability of adjunctive therapy with β2-agonists to increase CI-MPR expression in skeletal muscle was evaluated during ERT in murine Pompe disease with regard to reversal of neuromuscular involvement. Mice with Pompe disease were treated with weekly rhGAA injections (20 mg/kg) and a selective β2-agonist, either albuterol (30 mg/l in drinking water) or low-dose clenbuterol (6 mg/l in drinking water). Biochemical correction was enhanced by β2-agonist treatment in both muscle and the cerebellum, indicating that adjunctive therapy could enhance efficacy from ERT in Pompe disease with regard to neuromuscular involvement. Intriguingly, clenbuterol slightly reduced muscle glycogen content independent of CI-MPR expression, as demonstrated in CI-MPR knockout/GAA knockout mice that were otherwise resistant to ERT. Thus, adjunctive therapy with β2 agonists might improve the efficacy of ERT in Pompe disease and possibly other lysosomal storage disorders through enhancing receptor-mediated uptake of recombinant lysosomal enzymes. PMID:22154081

  17. Efficacy of oxidizing disinfectants at inactivating murine norovirus on ready-to-eat foods.

    PubMed

    Girard, Maryline; Mattison, Kirsten; Fliss, Ismail; Jean, Julie

    2016-02-16

    Noroviruses are the leading cause of foodborne illness, and ready-to-eat foods are frequent vehicles of their transmission. Studies of the disinfection of fruits and vegetables are becoming numerous. It has been shown that strong oxidizing agents are more effective than other chemical disinfectants for inactivating enteric viruses. The aim of this study was to evaluate the efficacy of oxidizing disinfectants (sodium hypochlorite, chloride dioxide and peracetic acid) at inactivating noroviruses on fruits and vegetables, using a norovirus surrogate, namely murine norovirus 3, which replicates in cell culture. Based on plaque assay, solutions of peracetic acid (85 ppm) and chlorine dioxide (20 ppm) reduced the infectivity of the virus in suspension by at least 3 log10 units after 1 min, while sodium hypochlorite at 50 ppm produced a 2-log reduction. On the surface of blueberries, strawberries and lettuce, chlorine dioxide was less effective than peracetic acid and sodium hypochlorite, which reduced viral titers by approximately 4 logs. A surprising increase in the efficacy of sodium hypochlorite on surfaces fouled with artificial feces was noted. PMID:26686597

  18. Vaccination of adult and newborn mice of a resistant strain (C57BL/6J) against challenge with leukemias induced by Moloney murine leukemia virus

    SciTech Connect

    Reif, A.E.

    1985-01-01

    Adult or newborn C57BL/6J mice were immunized with isogenic Moloney strain MuLV-induced leukemia cells irradiated with 10,000 rads or treated with low concentrations of formalin. Groups of immunized and control mice were challenged with a range of doses of viable leukemia cells, and tumor deaths were recorded for 90 days after challenge. Then, the doses of challenge cells which produced 50% tumor deaths were calculated for immunized and control mice. The logarithm of their ratio quantified the degree of protection provided by immunization. For adult C57BL/6J mice, a single immunization with MuLV-induced leukemia cells was not effective; either cells plus Bacillus Calmette-Guerin or Corynebacterium parvum, or else two immunizations with irradiated leukemia cells were needed to produce statistically significant increases in the values of the doses of challenge cells which produced 50% tumor deaths. Cross-protection was obtained by immunization with other isogenic MuLV-induced leukemias, but not by immunization with isogenic carcinogen-induced tumors or with an isogenic spontaneous leukemia. For newborn mice, a single injection of irradiated leukemia cells provided 1.3 to 1.5 logs of protection, and admixture of B. Calmette-Guerin or C. parvum increased this protection to 2.4 to 2.7 logs. Since irradiated and frozen-thawed MuLV-induced leukemia cells contained viable MuLV, leukemia cells treated with 0.5 or 1.0% formalin were tested as an alternative. A single injection of formalin-treated isogenic leukemia cells admixed with C. parvum provided between 1.7 and 2.8 logs of protection. These results demonstrate that a single vaccination of newborn animals against a highly antigenic virally induced leukemia produces strong protection against a subsequent challenge with viable leukemia cells.

  19. Host proteins interacting with the Moloney murine leukemia virus integrase: Multiple transcriptional regulators and chromatin binding factors

    PubMed Central

    Studamire, Barbara; Goff, Stephen P

    2008-01-01

    Background A critical step for retroviral replication is the stable integration of the provirus into the genome of its host. The viral integrase protein is key in this essential step of the retroviral life cycle. Although the basic mechanism of integration by mammalian retroviruses has been well characterized, the factors determining how viral integration events are targeted to particular regions of the genome or to regions of a particular DNA structure remain poorly defined. Significant questions remain regarding the influence of host proteins on the selection of target sites, on the repair of integration intermediates, and on the efficiency of integration. Results We describe the results of a yeast two-hybrid screen using Moloney murine leukemia virus integrase as bait to screen murine cDNA libraries for host proteins that interact with the integrase. We identified 27 proteins that interacted with different integrase fusion proteins. The identified proteins include chromatin remodeling, DNA repair and transcription factors (13 proteins); translational regulation factors, helicases, splicing factors and other RNA binding proteins (10 proteins); and transporters or miscellaneous factors (4 proteins). We confirmed the interaction of these proteins with integrase by testing them in the context of other yeast strains with GAL4-DNA binding domain-integrase fusions, and by in vitro binding assays between recombinant proteins. Subsequent analyses revealed that a number of the proteins identified as Mo-MLV integrase interactors also interact with HIV-1 integrase both in yeast and in vitro. Conclusion We identify several proteins interacting directly with both MoMLV and HIV-1 integrases that may be common to the integration reaction pathways of both viruses. Many of the proteins identified in the screen are logical interaction partners for integrase, and the validity of a number of the interactions are supported by other studies. In addition, we observe that some of the

  20. Effect of Host Modification and Age on Airway Epithelial Gene Transfer Mediated by a Murine Leukemia Virus-Derived Vector

    PubMed Central

    Johnson, Larry G.; Mewshaw, Jennifer P.; Ni, Hong; Friedmann, Theodore; Boucher, Richard C.; Olsen, John C.

    1998-01-01

    To study retroviral gene transfer to airway epithelia, we used a transient transfection technique to generate high titers (∼109 infectious units/ml after concentration) of murine leukemia virus (MuLV)-derived vectors pseudotyped with the vesicular stomatitis virus envelope glycoprotein (VSV-G). Transformed (CFT1) and primary airway epithelial cells were efficiently transduced by a VSV-G-pseudotyped lacZ vector (HIT-LZ) in vitro. CFT1 cells and primary cystic fibrosis (CF) airway cell monolayers infected with a vector (HIT-LCFSN) containing human CF transmembrane conductance regulator (CFTR) in the absence of selection expressed CFTR, as assessed by Western blot analysis, and exhibited functional correction of CFTR-mediated Cl− secretion. In vitro studies of persistence suggested that pseudotransduction was not a significant problem with our vector preparations. In a sulfur dioxide (SO2) inhalational injury model, bromodeoxyuridine (BrdU) incorporation rates were measured and found to exceed 50% in SO2-injured murine tracheal epithelium. HIT-LZ vector (multiplicity of infection of ∼10) instilled into the SO2-injured tracheas of anesthetized mice transduced 6.1% ± 1.3% of superficial airway cells in tracheas of weanling mice (3 to 4 weeks old; n = 10), compared to 1.4 ± 0.9% in mice 5 weeks of age (n = 4) and 0.2% in mice older than 6 weeks (n = 15). No evidence for gene transfer following delivery of HIT-LZ to tracheas of either weanling or older mice not injured with SO2 was detected. Because only a small fraction of BrdU-labeled airway cells were transduced, we examined the stability of the vector. No significant loss of vector infectivity over intervals (2 h) paralleling those of in vivo protocols was detected in in vitro assays using CFT1 cells. In summary, high-titer vectors permitted complementation of defective CFTR-mediated Cl− transport in CF airway cells in vitro without selection and demonstrated that the age of the animal appeared to be a major

  1. Comparison of the efficacies of various formulations of amphotericin B against murine visceral leishmaniasis.

    PubMed Central

    Mullen, A B; Carter, K C; Baillie, A J

    1997-01-01

    The antileishmanial efficacies of four proprietary amphotericin B (AmB) formulations (Fungizone, AmBisome, Abelcet, and Amphocil) and an experimental nonionic surfactant vesicle (NIV) formulation were compared in a murine model of acute visceral leishmaniasis. By a multiple-dosing regimen, groups of Leishmania donovani-infected BALB/c mice were treated (2.5 mg of AmB per kg of body weight) on days 7 to 11 postinfection with one of the AmB formulations, and parasite burdens were determined on day 18 postinfection. All of the formulations caused significant suppression parasite burdens in spleens (P < 0.01 to 0.0005) and livers (P < 0.0005) compared with those in the spleens and livers of the controls. In addition, a significant suppression of parasite burdens in bone marrow (P < 0.0005) compared to the burdens in the bone marrow of the controls was obtained for all the formulations except Abelcet, which was inactive at this site. On the basis of their overall efficacies (activity against liver, spleen, and bone marrow parasites), the formulations could be ranked as follows: Amphocil = AmBisome > AmB-NIV > Abelcet >> Fungizone. On the basis of spectrophotometric measurements, AmB was shown to exist in a predominantly aggregated state in all of the formulations. Although incubation in 50% serum altered the degree of aggregation, the AmB remained predominantly aggregated, indicating that the AMB-lipid complex in all of the formulations was physically stable. The results of the study showed that antiparasitic efficacy is associated positively with the degree of AmB aggregation in the presence of serum. PMID:9333030

  2. Dual effect of LPS on murine myeloid leukemia cells: Pro-proliferation and anti-proliferation.

    PubMed

    Yu, Lingling; Zhao, Yingmin; Gu, Xin; Wang, Jijun; Pang, Lei; Zhang, Yanqing; Li, Yaoyao; Jia, Xiaoqin; Wang, Xin; Gu, Jian; Yu, Duonan

    2016-06-10

    Modification of the bone marrow microenvironment is considered as a promising strategy to control leukemic cell proliferation, diseases progression and relapse after treatment. However, due to the diversity and complexity of the cellular and molecular compartments in the leukemic microenvironment, it is extremely difficult to dissect the role of each individual molecule or cell type in vivo. Here we established an in vitro system to dissect the role of lipopolysaccharide (LPS), stromal cells and endothelial cells in the growth of mouse myeloid tumor cells and B-lymphoma cells. We found that either LPS or bone marrow stromal cells as a feeder layer in culture is required for the proliferation of myeloid tumor cells. Surprisingly, the growth of myeloid leukemic cells on stromal cells is strongly inhibited when coupled with LPS in culture. This opposing effect of LPS, a complete switch from pro-proliferation to antitumor growth is due, at least in part, to the rapidly increased production of interleukin 12, Fas ligand and tissue inhibitor of metalloproteinases-2 from stromal cells stimulated by LPS. These results demonstrate that LPS can either facilitate or attenuate tumor cell proliferation, thus changing the disease course of myeloid leukemias through its direct effect or modulation of the tumor microenvironment. PMID:26968634

  3. Benzoyl peroxide increases UVA-induced plasma membrane damage and lipid oxidation in murine leukemia L1210 cells.

    PubMed

    Ibbotson, S H; Lambert, C R; Moran, M N; Lynch, M C; Kochevar, I E

    1998-01-01

    Ultraviolet A radiation induces oxidative stress and cell damage. The purpose of this investigation was to examine whether ultraviolet A-induced cell injury was amplified by the presence of a non-ultraviolet A absorbing molecule capable of generating free radicals. Benzoyl peroxide was used as a lipid soluble potential radical-generating agent. Plasma membrane permeability assessed by trypan blue uptake was used to measure cell damage in murine leukemia L1210 cells. Cells were irradiated with a pulsed Nd/YAG laser at 355 nm using 0-160 J per cm2. The ratio of the fluence-response slope in the presence of 40 microM benzoyl peroxide to that of irradiated controls was 4.3 +/- 2.6. Benzoyl peroxide alone or benzoyl peroxide added after irradiation did not cause increased trypan blue uptake. The ratio of the fluence-response slopes in the presence of 40 microM benzoyl peroxide to that of irradiated controls was 4.7 +/- 1.4 when cells were irradiated (0-43 J per cm2) with a xenon lamp, filtered to remove wavelengths <320 nm. The increased trypan blue uptake in 355 nm-irradiated cells in the presence of benzoyl peroxide was inhibited in a concentration-dependent manner by butylated hydroxytoluene, vitamin E, and trolox, a water-soluble vitamin E derivative. Lipid oxidation, assessed as thiobarbituric acid reactive substances, was significantly increased in samples irradiated with ultraviolet A in the presence of benzoyl peroxide at fluences >34 J per cm2. The increased trypan blue uptake and thiobarbituric acid reactive substances were inhibited by butylated hydroxytoluene. These results suggest that agents not absorbing ultraviolet A radiation may enhance ultraviolet A-initiated oxidative stress in cells. PMID:9424093

  4. Purification and characterization of the DNA polymerase and RNase H activities in Moloney murine sarcoma-leukemia virus.

    PubMed Central

    Gerard, G F; Grandgenett, D P

    1975-01-01

    Two RNase H (RNA-DNA hybrid ribonucleotidohydrolase, EC 3.1.4.34) activities separable by Sephadex G-100 gel filtration were identified in lysates of Moloney murine sarcoma-leukemia virus (MSV). The larger enzyme, which we have called RNase H-I, represented about 10% of the RNase H activity in the virion. RNase H-I (i) copurified with RNA-directed DNA polymerase from the virus, (ii) had a sedimentation coefficient of 4.4S (corresponds to an apparent mol wt of 70,000), (iii) required Mn-2+ (2 mM optimum) for activity with a [3-h]poly(A)-poly(dT) substrate, (iv) eluted from phosphocellulose at 0.2 M KC1, and (v) degraded [3-H]poly(A)-poly(dT) and [3-H]poly(C)-poly(dG) at approximately equal rates. The smaller enzyme, designated RNase H-II, which represented the majority of the RNase H activity in the virus preparation, was shown to be different since it (i) had no detectable, associated DNA polymerase activity, (ii) had a sedmimentation coefficient of 2.6S (corresponds to an apparent mol wt of 30,000), (iii) preferred Mg-2+ (10 to 15 mM optimum) over Mn-2+ (5 to 10 mM optimum) 2.5-fold for the degradation of [3-H]poly(A)-poly(dT), and (iv) degraded [3-H]poly(A)-poly(dT) 6 and 60 times faster than [3-H]poly(C)-poly(dG) in the presence of Mn-2+ and Mg-2+, respectively. Moloney MSV DNA polymerase (RNase H-I), purified by Sephadex G-100 gel filtration followed by phosphocellulose, poly(A)-oligo(dT)-cellulose, and DEAE-cellulose chromatography, transcribed heteropolymeric regions of avian myeloblastosis virus 70S RNA at a rate comparable to avian myeloblastosis virus DNA polymerase purified by the same procedure. PMID:46924

  5. Oxygen radical detoxification enzymes in doxorubicin-sensitive and -resistant P388 murine leukemia cells

    SciTech Connect

    Ramu, A.; Cohen, L.; Glaubiger, D.

    1984-05-01

    One of the proposed mechanisms for the cytotoxic effects of anthracycline compounds suggests that the effect is mediated through the formation of intracellular superoxide radicals. It is therefore possible that doxorubicin resistance is associated with increased intracellular enzyme capacity to convert these superoxide radicals to inactive metabolites. We have measured the relative activities of superoxide dismutase, glutathione peroxidase, and catalase in P388 mouse leukemia cells and in a doxorubicin-resistant subline. Since oxygen-reactive metabolites also play a role in mediating the cytotoxicity of ionizing radiation, the radiosensitivity of both cell lines was also studied. No significant differences in superoxide dismutase activity between these cell lines was observed, indicating that they have a similar capacity to convert superoxide anion radicals to hydrogen peroxide. P388 cells that are resistant to doxorubicin have 1.5 times the glutathione content and 1.5 times the activity of glutathione peroxidase measured in drug-sensitive P388 cells. However, incubation with 1-chloro-2,4-dinitrobenzene, which covalently binds glutathione, had no effect on the sensitivity of either cell line to doxorubicin. Measured catalase activity in drug-resistant P388 cells was one-third of the activity measured in doxorubicin-sensitive P388 cells. The activity of this enzyme was much higher than that of glutathione peroxidase in terms of H/sub 2/O/sub 2/ deactivation in both cell lines. It is therefore unlikely that doxorubicin-resistant P388 cells have an increased ability to detoxify reactive oxygen metabolites when compared to drug-sensitive cells. Doxorubicin-resistant P388 cells were significantly more sensitive to X-irradiation than were drug-sensitive P388 cells. These observations suggest that the difference in catalase activity in these cell lines may be associated with the observed differences in radiosensitivity.

  6. Comparative Virucidal Efficacy of Seven Disinfectants Against Murine Norovirus and Feline Calicivirus, Surrogates of Human Norovirus.

    PubMed

    Zonta, William; Mauroy, Axel; Farnir, Frederic; Thiry, Etienne

    2016-03-01

    Human noroviruses (HuNoV) are the leading cause of acute non-bacterial gastroenteritis in humans and can be transmitted either by person-to-person contact or by consumption of contaminated food. A knowledge of an efficient disinfection for both hands and food-contact surfaces is helpful for the food sector and provides precious information for public health. The aim of this study was to evaluate the effect of seven disinfectants belonging to different groups of biocides (alcohol, halogen, oxidizing agents, quaternary ammonium compounds, aldehyde and biguanide) on infectious viral titre and on genomic copy number. Due to the absence of a cell culture system for HuNoV, two HuNoV surrogates, such as murine norovirus and feline calicivirus, were used and the tests were performed in suspension, on gloves and on stainless steel discs. When, as criteria of efficacy, a log reduction >3 of the infectious viral titre on both surrogates and in the three tests is used, the most efficacious disinfectants in this study appear to be biocidal products B, C and D, representing the halogens, the oxidizing agents group and a mix of QAC, alcohol and aldehyde, respectively. In addition, these three disinfectants also elicited a significant effect on genomic copy number for both surrogate viruses and in all three tests. The results of this study demonstrate that a halogen compound, oxidizing agents and a mix of QAC, alcohol and aldehyde are advisable for HuNoV disinfection of either potentially contaminated surfaces or materials in contact with foodstuffs. PMID:26445948

  7. Ibrutinib enhances chimeric antigen receptor T-cell engraftment and efficacy in leukemia.

    PubMed

    Fraietta, Joseph A; Beckwith, Kyle A; Patel, Prachi R; Ruella, Marco; Zheng, Zhaohui; Barrett, David M; Lacey, Simon F; Melenhorst, Jan Joseph; McGettigan, Shannon E; Cook, Danielle R; Zhang, Changfeng; Xu, Jun; Do, Priscilla; Hulitt, Jessica; Kudchodkar, Sagar B; Cogdill, Alexandria P; Gill, Saar; Porter, David L; Woyach, Jennifer A; Long, Meixiao; Johnson, Amy J; Maddocks, Kami; Muthusamy, Natarajan; Levine, Bruce L; June, Carl H; Byrd, John C; Maus, Marcela V

    2016-03-01

    Anti-CD19 chimeric antigen receptor (CAR) T-cell therapy is highly promising but requires robust T-cell expansion and engraftment. A T-cell defect in chronic lymphocytic leukemia (CLL) due to disease and/or therapy impairs ex vivo expansion and response to CAR T cells. To evaluate the effect of ibrutinib treatment on the T-cell compartment in CLL as it relates to CAR T-cell generation, we examined the phenotype and function of T cells in a cohort of CLL patients during their course of treatment with ibrutinib. We found that ≥5 cycles of ibrutinib therapy improved the expansion of CD19-directed CAR T cells (CTL019), in association with decreased expression of the immunosuppressive molecule programmed cell death 1 on T cells and of CD200 on B-CLL cells. In support of these findings, we observed that 3 CLL patients who had been treated with ibrutinib for ≥1 year at the time of T-cell collection had improved ex vivo and in vivo CTL019 expansion, which correlated positively together and with clinical response. Lastly, we show that ibrutinib exposure does not impair CAR T-cell function in vitro but does improve CAR T-cell engraftment, tumor clearance, and survival in human xenograft models of resistant acute lymphocytic leukemia and CLL when administered concurrently. Our collective findings indicate that ibrutinib enhances CAR T-cell function and suggest that clinical trials with combination therapy are warranted. Our studies demonstrate that improved T-cell function may also contribute to the efficacy of ibrutinib in CLL. These trials were registered at www.clinicaltrials.gov as #NCT01747486, #NCT01105247, and #NCT01217749. PMID:26813675

  8. Dexamethasone-loaded Block Copolymer Nanoparticles Induce Leukemia Cell Death and Enhances Therapeutic Efficacy: A Novel Application in Pediatric Nanomedicine

    PubMed Central

    Krishnan, Vinu; Xu, Xian; Barwe, Sonali P.; Yang, Xiaowei; Czymmek, Kirk; Waldman, Scott A.; Mason, Robert W.; Jia, Xinqiao; Rajasekaran, Ayyappan K.

    2014-01-01

    Nanotechnology approaches have tremendous potential for enhancing treatment efficacy with lower doses of chemotherapeutics. Nanoparticle-based drug delivery approaches are poorly developed for childhood leukemia. Dexamethasone (Dex) is one of the most common chemotherapeutic drugs used in the treatment of childhood leukemia. In this study, we encapsulated Dex in polymeric nanoparticles and validated their anti-leukemic potential in vitro and in vivo. Nanoparticles (NPs) with an average diameter of 110 nm were assembled from amphiphilic block copolymers poly (ethylene glycol) (PEG) and poly (ε-caprolactone) (PCL) bearing pendant cyclic ketals. The blank nanoparticles were non-toxic to cultured cells in vitro and to mice in vivo. Encapsulation of Dex into the nanoparticles (Dex-NP) did not compromise the bioactivity of the drug. Dex-NPs induced glucocorticoid phosphorylation and showed cytotoxicity similar to the free Dex in leukemic cells. Studies using nanoparticles labeled with fluorescent dyes revealed leukemic cell surface binding and internalization. In vivo biodistribution studies showed NP accumulation in the liver and spleen with subsequent clearance of the particles with time. In a pre-clinical model of leukemia, Dex-NPs significantly improved the quality of life and survival of mice compared to the free drug. To our knowledge, this is the first report showing the efficacy of polymeric nanoparticles to deliver Dex to potentially treat childhood leukemia and reveals that low dose of Dex should be sufficient for inducing cell death and improve survival. PMID:23194373

  9. Safrole suppresses murine myelomonocytic leukemia WEHI-3 cells in vivo, and stimulates macrophage phagocytosis and natural killer cell cytotoxicity in leukemic mice.

    PubMed

    Yu, Fu-Shun; Yang, Jai-Sing; Yu, Chun-Shu; Chiang, Jo-Hua; Lu, Chi-Cheng; Chung, Hsiung-Kwang; Yu, Chien-Chih; Wu, Chih-Chung; Ho, Heng-Chien; Chung, Jing-Gung

    2013-11-01

    Many anticancer drugs are obtained from phytochemicals and natural products. However, some phytochemicals have mutagenic effects. Safrole, a component of Piper betle inflorescence, has been reported to be a carcinogen. We have previously reported that safrole induced apoptosis in human oral cancer cells in vitro and inhibited the human oral tumor xenograft growth in vivo. Until now, there is no information addressing if safrole promotes immune responses in vivo. To evaluate whether safrole modulated immune function, BALB/c mice were intraperitoneally injected with murine myelomonocytic WEHI-3 leukemia cells to establish leukemia and then were treated with or without safrole at 4 and 16 mg/kg. Animals were sacrificed after 2 weeks post-treatment with safrole for examining the immune cell populations, phagocytosis of macrophages and the natural killer (NK) cells' cytotoxicity. Results indicated that safrole increased the body weight, and decreased the weights of spleen and liver in leukemic mice. Furthermore, safrole promoted the activities of macrophages phagocytosis and NK cells' cytotoxicity in leukemic mice when compared with untreated leukemic mice. After determining the cell marker population, we found that safrole promoted the levels of CD3 (T cells), CD19 (B cells) and Mac-3 (macrophages), but it did not affect CD11b (monocytes) in leukemic mice. In conclusion, safrole altered the immune modulation and inhibited the leukemia WEHI-3 cells in vivo. PMID:24150866

  10. The Icsbp locus is a common proviral insertion site in mature B-cell lymphomas/plasmacytomas induced by exogenous murine leukemia virus

    SciTech Connect

    Ma Shiliang; Sorensen, Annette Balle; Kunder, Sandra; Sorensen, Karina Dalsgaard; Quintanilla-Martinez, Leticia; Morris, David W.; Schmidt, Joerg; Pedersen, Finn Skou . E-mail: fsp@mb.au.dk

    2006-09-01

    ICSBP (interferon consensus sequence binding protein)/IRF8 (interferon regulatory factor 8) is an interferon gamma-inducible transcription factor expressed predominantly in hematopoietic cells, and down-regulation of this factor has been observed in chronic myelogenous leukemia and acute myeloid leukemia in man. By screening about 1200 murine leukemia virus (MLV)-induced lymphomas, we found proviral insertions at the Icsbp locus in 14 tumors, 13 of which were mature B-cell lymphomas or plasmacytomas. Only one was a T-cell lymphoma, although such tumors constituted about half of the samples screened. This indicates that the Icsbp locus can play a specific role in the development of mature B-lineage malignancies. Two proviral insertions in the last Icsbp exon were found to act by a poly(A)-insertion mechanism. The remaining insertions were found within or outside Icsbp. Since our results showed expression of Icsbp RNA and protein in all end-stage tumor samples, a simple tumor suppressor function of ICSBP is not likely. Interestingly, proviral insertions at Icsbp have not been reported from previous extensive screenings of mature B-cell lymphomas induced by endogenous MLVs. We propose that ICSBP might be involved in an early modulation of an immune response to exogenous MLVs that might also play a role in proliferation of the mature B-cell lymphomas.

  11. Efficacy and Mechanisms of Murine Norovirus Inhibition by Pulsed-Light Technology

    PubMed Central

    Vimont, Allison; Fliss, Ismaïl

    2015-01-01

    Pulsed light is a nonthermal processing technology recognized by the FDA for killing microorganisms on food surfaces, with cumulative fluences up to 12 J cm−2. In this study, we investigated its efficacy for inactivating murine norovirus 1 (MNV-1) as a human norovirus surrogate in phosphate-buffered saline, hard water, mineral water, turbid water, and sewage treatment effluent and on food contact surfaces, including high-density polyethylene, polyvinyl chloride, and stainless steel, free or in an alginate matrix. The pulsed-light device emitted a broadband spectrum (200 to 1,000 nm) at a fluence of 0.67 J cm−2 per pulse, with 2% UV at 8 cm beneath the lamp. Reductions in viral infectivity exceeded 3 log10 in less than 3 s (5 pulses; 3.45 J cm−2) in clear suspensions and on clean surfaces, even in the presence of alginate, and in 6 s (11 pulses; 7.60 J cm−2) on fouled surfaces except for stainless steel (2.6 log10). The presence of protein or bentonite interfered with viral inactivation. Analysis of the morphology, the viral proteins, and the RNA integrity of treated MNV-1 allowed us to elucidate the mechanisms involved in the antiviral activity of pulsed light. Pulsed light appeared to disrupt MNV-1 structure and degrade viral protein and RNA. The results suggest that pulsed-light technology could provide an effective alternative means of inactivating noroviruses in wastewaters, in clear beverages, in drinking water, or on food-handling surfaces in the presence or absence of biofilms. PMID:25681193

  12. Efficacy and mechanisms of murine norovirus inhibition by pulsed-light technology.

    PubMed

    Vimont, Allison; Fliss, Ismaïl; Jean, Julie

    2015-04-01

    Pulsed light is a nonthermal processing technology recognized by the FDA for killing microorganisms on food surfaces, with cumulative fluences up to 12 J cm(-2). In this study, we investigated its efficacy for inactivating murine norovirus 1 (MNV-1) as a human norovirus surrogate in phosphate-buffered saline, hard water, mineral water, turbid water, and sewage treatment effluent and on food contact surfaces, including high-density polyethylene, polyvinyl chloride, and stainless steel, free or in an alginate matrix. The pulsed-light device emitted a broadband spectrum (200 to 1,000 nm) at a fluence of 0.67 J cm(-2) per pulse, with 2% UV at 8 cm beneath the lamp. Reductions in viral infectivity exceeded 3 log10 in less than 3 s (5 pulses; 3.45 J cm(-2)) in clear suspensions and on clean surfaces, even in the presence of alginate, and in 6 s (11 pulses; 7.60 J cm(-2)) on fouled surfaces except for stainless steel (2.6 log10). The presence of protein or bentonite interfered with viral inactivation. Analysis of the morphology, the viral proteins, and the RNA integrity of treated MNV-1 allowed us to elucidate the mechanisms involved in the antiviral activity of pulsed light. Pulsed light appeared to disrupt MNV-1 structure and degrade viral protein and RNA. The results suggest that pulsed-light technology could provide an effective alternative means of inactivating noroviruses in wastewaters, in clear beverages, in drinking water, or on food-handling surfaces in the presence or absence of biofilms. PMID:25681193

  13. Virucidal efficacy of treatment with photodynamically activated curcumin on murine norovirus bio-accumulated in oysters.

    PubMed

    Wu, Juan; Hou, Wei; Cao, Binbin; Zuo, Tao; Xue, Changhu; Leung, Albert Wingnang; Xu, Chuanshan; Tang, Qing-Juan

    2015-09-01

    Norovirus (NoV) is one of the most important seafood- and water-borne viruses, and is a major cause of acute gastroenteritis outbreaks. In the present study we investigated the effect of curcumin as a sensitizer to photodynamic treatment both in buffer and in oysters against murine norovirus 1 (MNV-1), a surrogate of NoV. MNV-1 cultured in buffer and MNV-1 bio-accumulated in oysters were irradiated with a novel LED light source with a wavelength of 470nm and an energy of 3.6J/cm(2). Inactivation of MNV-1 was investigated by plaque assays. After virus was extracted from the gut of oysters treated over a range of curcumin concentrations, the ultrastructural morphology of the virus was observed using electron microscopy, and the integrity of viral nucleic acids and stability of viral capsid proteins were also determined. Results showed that the infectivity of MNV-1 was significantly inhibited by 1-3logPFU/ml, with significant damage to viral nucleic acids in a curcumin dose-dependent manner after photodynamic activation. Virus morphology was altered after the photodynamic treatment with curcumin, presumably due to the change of the viral capsid protein structures. The data suggest that treatment of oysters with photodynamic activation of curcumin is a potentially efficacious and cost-effective method to inactivate food-borne NoV. Further studies are necessary to evaluate the toxicology of this approach in detail and perform sensory evaluation of the treated product. PMID:26117199

  14. Efficacy of Exercise Interventions in Patients with Acute Leukemia: A Meta-Analysis

    PubMed Central

    Zhu, Jinjie; Gu, Zejuan; Yin, Xiangguang

    2016-01-01

    Background Decreased physical performance and impaired physiological and psychological fitness have been reported in patients with acute leukemia (AL). We performed a meta-analysis to assess the efficacy of exercise in patients with AL. Methods In this meta-analysis, the electronic databases MEDLINE, Embase, Cochrane, Web of Science, SPORTDiscus, CINAHL and PEDro were searched through November 2015. Three authors participated in the study selection, data extraction and quality assessment. The instrument used for quality assessment was derived from the Cochrane Handbook for Systematic Reviews of Interventions. Analyses were performed according to the recommendations of The Cochrane Collaboration using Review Manager 5.3. Results Nine trials (8 randomized controlled trials and 1 quasi-experimental design trial) with 314 AL participants were included in this meta-analysis. The pooled standardized mean differences between the exercise and control groups were 0.45 (95% confidence interval (CI): 0.09 to 0.80, P value = 0.01, P for heterogeneity = 0.23, I2 = 28%) for cardiorespiratory fitness and 0.67 (95% CI: 0.28 to 1.06, P value = 0.0007, P for heterogeneity = 0.14, I2 = 43%) for muscle strength. Based on the data for fatigue, anxiety, and depression, there were no significant differences in these parameters between the exercise and control groups. Conclusions Exercise has beneficial effects on cardiorespiratory fitness, muscle strength and functional mobility; however, no significant improvements in fatigue, anxiety, depression or quality of life were observed. Further large-scale randomized trials are needed to assess the safety, feasibility and efficacy of exercise programs for AL patients. PMID:27463234

  15. CD19-Targeted Nanodelivery of Doxorubicin Enhances Therapeutic Efficacy in B-cell Acute Lymphoblastic Leukemia

    PubMed Central

    Krishnan, Vinu; Xu, Xian; Kelly, Dakota; Snook, Adam; Waldman, Scott A.; Mason, Robert W.; Jia, Xinqiao; Rajasekaran, Ayyappan K.

    2015-01-01

    Nanomedicine has advanced to clinical trials for adult cancer therapy. However, the field is still in its infancy for treatment of childhood malignancies such as acute lymphoblastic leukemia (ALL). Nanotherapy offers multiple advantages over conventional therapy. It facilitates targeted delivery and enables controlled release of drugs to reduce treatment-related side effects. Here, we demonstrate, that doxorubicin (DOX) encapsulated in polymeric nanoparticles (NPs) modified with targeting ligands against CD19 (CD19-DOX-NPs) can be delivered in a CD19-specific manner to leukemic cells. The CD19-DOX-NPs were internalized via receptor-mediated endocytosis and imparted cytotoxicity in a CD19-dependent manner in CD19 positive ALL cells. Leukemic mice treated with CD19-DOX-NPs survived significantly longer and manifested a higher degree of agility indicating reduced apparent systemic toxicity during treatment compared to mice treated with free DOX. We suggest that targeted delivery of drugs used in childhood cancer treatment should improve therapeutic efficacy and reduce treatment-related side effects in children. PMID:25898125

  16. New hope for chronic myelogenous leukemia patients: dasatinib offers better efficacy with shorter treatment

    PubMed Central

    Ichinohe, Tatsuo

    2016-01-01

    Although the discovery of tyrosine kinase inhibitors (TKIs) has dramatically improved the prognoses of chronic myelogenous leukemia (CML) patients, a cure has remained elusive. Unanswered questions include how long must a patient continue on TKI therapy, and how does a patient know when he/she can safely stop or finish this therapy? Imagawa et al. have carefully addressed these questions of safety and efficacy using a stop study of the second-generation TKI dasatinib. The results of a multicenter phase II trial termed the “dasatinib discontinuation” (DADI) trial indicated that 48% (30/63) of CML patients who had maintained a deep molecular response (DMR) to second-line or subsequent dasatinib therapy for at least for 1 year did not show any signs of disease relapse. Thus, even after it is stopped, dasatinib treatment may decrease the chance of disease relapse and provide a curative benefit to CML patients. This work by Imagawa et al. strongly supports the clinical utility of the second-generation TKI dasatinib for CML treatment. PMID:27488943

  17. CD19-Targeted Nanodelivery of Doxorubicin Enhances Therapeutic Efficacy in B-Cell Acute Lymphoblastic Leukemia.

    PubMed

    Krishnan, Vinu; Xu, Xian; Kelly, Dakota; Snook, Adam; Waldman, Scott A; Mason, Robert W; Jia, Xinqiao; Rajasekaran, Ayyappan K

    2015-06-01

    Nanomedicine has advanced to clinical trials for adult cancer therapy. However, the field is still in its infancy for treatment of childhood malignancies such as acute lymphoblastic leukemia (ALL). Nanotherapy offers multiple advantages over conventional therapy. It facilitates targeted delivery and enables controlled release of drugs to reduce treatment-related side effects. Here, we demonstrate that doxorubicin (DOX) encapsulated in polymeric nanoparticles (NPs) modified with targeting ligands against CD19 (CD19-DOX-NPs) can be delivered in a CD19-specific manner to leukemic cells. The CD19-DOX-NPs were internalized via receptor-mediated endocytosis and imparted cytotoxicity in a CD19-dependent manner in CD19-positive ALL cells. Leukemic mice treated with CD19-DOX-NPs survived significantly longer and manifested a higher degree of agility, indicating reduced apparent systemic toxicity during treatment compared to mice treated with free DOX. We suggest that targeted delivery of drugs used in childhood cancer treatment should improve therapeutic efficacy and reduce treatment-related side effects in children. PMID:25898125

  18. Proviral activation of the c-myb proto-oncogene is detectable in preleukemic mice infected neonatally with Moloney murine leukemia virus but not in resulting end stage T lymphomas.

    PubMed

    Belli, B; Wolff, L; Nazarov, V; Fan, H

    1995-08-01

    Moloney murine leukemia virus induces myeloid leukemia when inoculated intravenously into pristane-primed adult BALB/c mice. One hundred percent of these tumors show insertional activation of the c-myb proto-oncogene, and reverse transcriptase PCR assays have shown that the c-myb activation could be detected soon after infection. We tested BALB/c and NIH Swiss mice that had been inoculated as newborns with Moloney murine leukemia virus, under which conditions they develop T lymphomas exclusively. Reverse transcriptase-PCR assays indicated that c-myb activations were detectable soon after neonatal infection. However, none of the resulting T lymphomas contained c-myb activations. The implications of these results to the timing of proto-oncogene activations in leukemogenesis and the specificity of proto-oncogene activations for different diseases are discussed. PMID:7609084

  19. Effects of diallyl trisulfide on induction of apoptotic death in murine leukemia WEHI-3 cells in vitro and alterations of the immune responses in normal and leukemic mice in vivo.

    PubMed

    Hung, Fang-Ming; Shang, Hung-Sheng; Tang, Nou-Ying; Lin, Jen-Jyh; Lu, Kung-Wen; Lin, Jing-Pin; Ko, Yang-Ching; Yu, Chien-Chih; Wang, Hai-Lung; Liao, Jung-Chi; Lu, Hsu-Feng; Chung, Jing-Gung

    2015-11-01

    Diallyl trisulfide (DATS), a chemopreventive dietary constituent and extracted from garlic, has been shown to against cultured many types of human cancer cell liens but the fate of apoptosis in murine leukemia cells in vitro and immune responses in leukemic mice remain elusive. Herein, we clarified the actions of DATS on growth inhibition of murine leukemia WEHI-3 cells in vitro and used WEHI-3 cells to generate leukemic mice in vivo, following to investigate the effects of DATS in animal model. In in vitro study, DATS induced apoptosis of WEHI-3 cells through the G0/G1 phase arrest and induction of caspase-3 activation. In in vivo study DATS decreased the weight of spleen of leukemia mice but did not affect the spleen weight of normal mice. DATS promoted the immune responses such as promotions of the macrophage phagocytosis and NK cell activities in WEHI-3 leukemic and normal mice. However, DATS only promotes NK cell activities in normal mice. DATS increases the surface markers of CD11b and Mac-3 in leukemia mice but only promoted CD3 in normal mice. In conclusion, the present study indicates that DATS induces cell death through induction of apoptosis in mice leukemia WHEI-3 cells. DATS also promotes immune responses in leukemia and normal mice in vivo. PMID:24890016

  20. No association found between the detection of either xenotropic murine leukemia virus-related virus or polytropic murine leukemia virus and chronic fatigue syndrome in a blinded, multi-site, prospective study by the establishment and use of the SolveCFS BioBank

    PubMed Central

    2014-01-01

    Background In 2009, a retrospective study reported the detection of xenotropic murine leukemia virus-related virus (XMRV) in clinical isolates derived from individuals with chronic fatigue syndrome or myalgic encephalomyelitis (CFS). While many efforts to confirm this observation failed, one report detected polytropic murine leukemia virus (pMLV), instead of XMRV. In both studies, Polymerase Chain Reaction (PCR)-based methods were employed which could provide the basis for the development of a practical diagnostic tool. To confirm these studies, we hypothesized that the ability to detect these viruses will not only depend upon the technical details of the methods employed but also on the criteria used to diagnose CFS and the availability of well characterized clinical isolates. Methods A repository of clinical isolates from geographically distinct sites was generated by the collection of fresh blood samples from well characterized CFS and healthy subjects. Molecular techniques were used to generate assay positive controls and to determine the lower limit of detection (LLOD) for murine retroviral and Intracisternal A particle (Cell 12(4):963-72, 1977) detection methods. Results We report the establishment of a repository of well-defined, clinical isolates from five, geographically distinct regions of the US, the comparative determination of the LLODs and validation efforts for the previously reported detection methods and the results of an effort to confirm the association of these retroviral signatures in isolates from individuals with CFS in a blinded, multi-site, prospective study. We detected various, murine retroviral DNA signatures but were unable to resolve a difference in the incidence of their detection between isolates from CFS (5/72; 6.7%) and healthy (2/37; 5.4%) subjects (Fisher’s Exact Test, p-value = 1). The observed sequences appeared to reflect the detection of endogenous murine retroviral DNA, which was not identical to either XMRV or p

  1. B-cell-specific Moloney murine leukemia virus integration site 1: potential stratification factor and therapeutic target for epithelial ovarian cancer.

    PubMed

    Zhao, Qianying; Gui, Ting; Qian, Qiuhong; Li, Lei; Shen, Keng

    2016-01-01

    Epithelial ovarian cancer, a vexing challenge for clinical management, still lacks biomarkers for early diagnosis, precise stratification, and prognostic evaluation of patients. B-cell-specific Moloney murine leukemia virus integration site 1 (BMI1), a member of the polycomb group of proteins, engages in diverse cellular processes, including proliferation, differentiation, senescence, and stem cell renewal. In addition, BMI1, as a cancer stem-cell marker, participates in tumorigenesis through various pathways. Rewardingly, recent studies have also revealed a relationship between BMI1 expression and the clinical grade/stage, therapy response, and survival outcome in a majority of human malignancies, including epithelial ovarian cancer. Therefore, BMI1 might serve as a potential stratification factor and treatment target for epithelial ovarian cancer, pending evidence from further investigations. PMID:27578986

  2. A human TRIM5alpha B30.2/SPRY domain mutant gains the ability to restrict and prematurely uncoat B-tropic murine leukemia virus.

    PubMed

    Diaz-Griffero, Felipe; Perron, Michel; McGee-Estrada, Kathleen; Hanna, Robert; Maillard, Pierre V; Trono, Didier; Sodroski, Joseph

    2008-09-01

    Human TRIM5alpha restricts N-tropic murine leukemia virus (N-MLV) but not B-tropic MLV (B-MLV) infection. Here we study B30.2/SPRY domain mutants of human TRIM5alpha that acquire the ability to inhibit B-MLV infection prior to reverse transcription without losing the ability to restrict N-MLV infection. Remarkably, these mutants gain the ability to decrease the amount of particulate B-MLV capsids in the cytosol of infected cells. In addition, these mutants gain the ability to restrict SIV(mac) and HIV-2 infection. B-MLV and SIV(mac) infections were blocked by the mutant TRIM5alpha proteins prior to reverse transcription. Thus, the range of retroviruses restricted by human TRIM5alpha can be increased by changes in the B30.2/SPRY domain, which also result in the ability to cause premature uncoating of the restricted retroviral capsid. PMID:18586294

  3. B-cell-specific Moloney murine leukemia virus integration site 1: potential stratification factor and therapeutic target for epithelial ovarian cancer

    PubMed Central

    Zhao, Qianying; Gui, Ting; Qian, Qiuhong; Li, Lei; Shen, Keng

    2016-01-01

    Epithelial ovarian cancer, a vexing challenge for clinical management, still lacks biomarkers for early diagnosis, precise stratification, and prognostic evaluation of patients. B-cell-specific Moloney murine leukemia virus integration site 1 (BMI1), a member of the polycomb group of proteins, engages in diverse cellular processes, including proliferation, differentiation, senescence, and stem cell renewal. In addition, BMI1, as a cancer stem-cell marker, participates in tumorigenesis through various pathways. Rewardingly, recent studies have also revealed a relationship between BMI1 expression and the clinical grade/stage, therapy response, and survival outcome in a majority of human malignancies, including epithelial ovarian cancer. Therefore, BMI1 might serve as a potential stratification factor and treatment target for epithelial ovarian cancer, pending evidence from further investigations. PMID:27578986

  4. [Efficacy of siRNA on feline leukemia virus replication in vitro].

    PubMed

    Lehmann, Melanie; Weber, Karin; Rauch, Gisep; Hofmann-Lehmann, Regina; Hosie, Margaret J; Meli, Marina L; Hartmann, Katrin

    2015-01-01

    Feline leukemia virus (FeLV) can lead to severe clinical signs in cats. Until now, there is no effective therapy for FeLV-infected cats. RNA interference-based antiviral therapy is a new concept. Specific small interfering RNA (siRNA) are designed complementary to the mRNA of a target region, and thus inhibit replication. Several studies have proven efficacy of siRNAs in inhibiting virus replication. The aim of this study was to evaluate the inhibitory potential of siRNAs against FeLV replication in vitro. siRNAs against the FeLV env gene and the host cell surface receptor (feTHTR1) which is used by FeLV-A for entry as well as siRNA that were not complementary to the FeLV or cat genome, were tested. Crandell feline kidney cells (CrFK cells) were transfected with FeLV-A/Glasgow-1. On day 13, infected cells were transfected with siRNAs. As control, cells were mock-transfected or treated with azidothymidine (AZT) (5 μg/ml). Culture supernatants were analyzed for FeLV RNA using quantitative real-time RT-PCR and for FeLV p27 by ELISA every 24 hours for five days. All siRNAs significantly reduced viral RNA and p27 production, starting after 48 hours. The fact that non-complementary siRNAs also inhibited virus replication may lead to the conclusion that unspecific mechanisms rather than specific binding lead to inhibition. PMID:26054227

  5. Efficacy of species-specific protein antibiotics in a murine model of acute Pseudomonas aeruginosa lung infection

    PubMed Central

    McCaughey, Laura C.; Ritchie, Neil. D.; Douce, Gillian R.; Evans, Thomas J.; Walker, Daniel

    2016-01-01

    Protein antibiotics, known as bacteriocins, are widely produced by bacteria for intraspecies competition. The potency and targeted action of bacteriocins suggests that they could be developed into clinically useful antibiotics against highly drug resistant Gram-negative pathogens for which there are few therapeutic options. Here we show that Pseudomonas aeruginosa specific bacteriocins, known as pyocins, show strong efficacy in a murine model of P. aeruginosa lung infection, with the concentration of pyocin S5 required to afford protection from a lethal infection at least 100-fold lower than the most commonly used inhaled antibiotic tobramycin. Additionally, pyocins are stable in the lung, poorly immunogenic at high concentrations and efficacy is maintained in the presence of pyocin specific antibodies after repeated pyocin administration. Bacteriocin encoding genes are frequently found in microbial genomes and could therefore offer a ready supply of highly targeted and potent antibiotics active against problematic Gram-negative pathogens. PMID:27444885

  6. Extract of Hedyotis diffusa Willd influences murine leukemia WEHI-3 cells in vivo as well as promoting T- and B-cell proliferation in leukemic mice.

    PubMed

    Lin, Chin-Chung; Kuo, Chao-Lin; Lee, Mau-Hva; Hsu, Shu-Chun; Huang, An-Cheng; Tang, Nou-Ying; Lin, Jing-Pin; Yang, Jai-Sing; Lu, Chi-Cheng; Chiang, Jo-Hua; Chueh, Fu-Shin; Chung, Jing-Gung

    2011-01-01

    Medicinal plants and herbs are widely used in the treatment of various types of cancer in Taiwan, China and many other countries. Hedyotis diffusa Willd (HDW) has been known as a traditional Chinese medicine for a long time, and possesses various bioactivities and anticancer activity. There is no available information on the effects of HDW extracts in leukemic mice and on immune responses in vivo. In this study, we established murine WEHI-3 leukemia in BALB/c mice and hypothesized that an aqueous HDW extract might have antileukemia effects on leukemic animals in vivo. The major characteristic of leukemic mice was an enlarged spleen after intraperitoneal injection with WEHI-3 cells. HDW extract reduced the weights of spleen and liver, but had no significant effect on body weight in WEHI-3 leukemic mice. HDW extract increased the percentage of CD11b cell surface marker (monocytes), but it reduced the percentage of CD3 (T-cell) and CD19 (B-cell) markers. However, HDW extract did not affect the level of Mac-3 and there was no influence on phagocytosis by macrophages from peripheral blood mononuclear cells and the peritoneal cavity in leukemic mice. The isolated splenocytes from HDW extract-treated leukemic mice demonstrated an increase of T- and B-cell proliferation in vivo. Based on these results, HDW extract would appear to have antileukemia activity in WEHI-3 cell-induced leukemia in vivo. PMID:21709007

  7. Efficacy of an Fc-modified anti-CD123 antibody (CSL362) combined with chemotherapy in xenograft models of acute myelogenous leukemia in immunodeficient mice

    PubMed Central

    Lee, Erwin M.; Yee, Dean; Busfield, Samantha J.; McManus, Julie F.; Cummings, Nik; Vairo, Gino; Wei, Andrew; Ramshaw, Hayley S.; Powell, Jason A.; Lopez, Angel F.; Lewis, Ian D.; McCall, Martin N.; Lock, Richard B.

    2015-01-01

    The prognosis of older patients with acute myelogenous leukemia is generally poor. The interleukin-3 receptor α-chain (CD123) is highly expressed on the surface of acute leukemia cells compared with normal hematopoietic stem cells. CSL362 is a fully humanized, CD123-neutralizing monoclonal antibody containing a modified Fc structure, which enhances human natural killer cell antibody-dependent cell-mediated cytotoxicity. Six continuous acute myelogenous leukemia xenografts established from patient explants and characterized by cell and molecular criteria, produced progressively lethal disease 42-202 days after transplantation. CSL362 alone reduced engraftment of one of four and three of four acute myelogenous leukemia xenografts in the bone marrow and peripheral organs, respectively. A cytarabine and daunorubicin regimen was optimized using this model to identify potentially synergistic interactions with CSL362. Cytarabine/daunorubicin improved the survival of mice engrafted with four of four acute myelogenous leukemia xenografts by 31–41 days. Moreover, CSL362 extended the survival of cytarabine/daunorubicin-treated mice for two of two acute myelogenous leukemia xenografts, while augmentation of natural killer cell-deficient NSG mice with adoptively transferred human natural killer cells improved survival against a single xenograft. Interestingly, this enhanced CSL362 efficacy was lost in the absence of chemotherapy. This study shows that acute myelogenous leukemia xenografts provide a platform for the evaluation of new therapeutics, simulating complex in vivo interactions, and that the in vivo efficacy of CSL362 supports continued clinical development of this drug. PMID:26130514

  8. Efficacy of Humanized Carbapenem and Ceftazidime Regimens against Enterobacteriaceae Producing OXA-48 Carbapenemase in a Murine Infection Model

    PubMed Central

    Wiskirchen, Dora E.; Nordmann, Patrice; Crandon, Jared L.

    2014-01-01

    Enterobacteriaceae producing the OXA-48 carbapenemase are emerging worldwide, leaving few treatment options. Efficacy has been demonstrated in vivo with ceftazidime against a ceftazidime-susceptible OXA-48 isolate but not with imipenem despite maintaining susceptibility. The relationship between phenotype and in vivo efficacy was assessed for OXA-48 producers using humanized regimens of 2 g doripenem every 8 h (q8h; 4 h infusion), 1 g ertapenem q24h, 2 g ceftazidime q8h (2 h inf), and 500 mg levofloxacin q24h. Each regimen was evaluated over 24 h against an isogenic pair (wild-type and OXA-48 Klebsiella pneumoniae strains) and six clinical OXA-48 isolates with and without other extended-spectrum β-lactamases in immunocompetent and neutropenic murine thigh infection models. Efficacy was determined using the change in bacterial density versus 24-h growth controls in immunocompetent studies and 0-h controls in neutropenic studies. Bacterial reductions of ≥1 log CFU were observed with all agents for the wild-type strain. Consistent with low MICs, ceftazidime and levofloxacin exhibited efficacy against the isogenic OXA-48 strain, whereas doripenem did not, despite having a susceptible MIC; no activity was observed with ertapenem, consistent with a resistant MIC. Similar trends were observed for the clinical isolates evaluated. Ceftazidime, levofloxacin, and ertapenem efficacy against isogenic and clinical OXA-48-producing strains correlated well with phenotypic profiles and pharmacodynamic targets, whereas efficacy with doripenem was variable over the MIC range studied. These data suggest that carbapenems may not be a reliable treatment for treating OXA-48 producers and add to previous observations with KPC and NDM-1 suggesting that genotype may better predict activity of the carbapenems than the phenotypic profile. PMID:24379200

  9. Further evidence for the existence of 'homing' receptors on murine leukemia cells which mediate adherence to normal bone marrow stromal cells.

    PubMed

    Kamenov, B; Longenecker, B M

    1985-01-01

    A significant proportion of 131IUDR-labelled cells from murine leukemia cell lines L1210 and P388, but not the L5178Y lymphoma cell line, are retained in the bone marrow (B.M.) following i.v. injection into syngeneic mice. Following this, L1210 and P388 cells grow and rapidly replace the normal hematopoietic cells of the B.M. L1210 and P388 cells, but not several lymphoma cell lines, also bind avidly to monolayers of B.M. stromal cells (Dexter cultures) and soon overgrow the cultures following rapid cell proliferation. P388 cells bound equally well to confluent monolayers of B.M., whole mouse embryo and newborn mouse kidney while L1210 cells bound well to B.M. and whole mouse embryo but showed little binding to newborn kidney monolayers. The accumulation of the two leukemia cell lines in the B.M. was constant and indistinguishable over a 48-h period. In contrast, in both spleen and liver the number of L1210 cells decreased during the same period while P388 cells were retained at a constant level. Generally there was a lack of correlation of B.M. metastasis of a cell line and its metastasis to other organs although P388 cells, but not L1210 cells, demonstrated a tremendous capacity for metastatic growth in both spleen and liver. Normal B.M. cells were fused with the syngeneic SP2/0 murine myeloma fusor line and 10 hybridomas plus the SP2/0 parent were tested for in-vitro adherence to B.M. monolayers and in-vivo metastatic behavior. The same 3 (out of 10) hybridomas showed a high level of adherence to B.M. monolayers, high levels of retention of cells in the B.M. following i.v. injection, and rapid growth and takeover of the normal B.M. In marked contrast, neither the SP2/0 parent nor the remaining 7 hybridomas show significant adherence, B.M. retention or growth in the B.M. A distinct lack of correlation of B.M. vs liver or spleen metastasis was once again noted for the hybridomas although all of the hybridomas showed much less metastatic growth in the liver than

  10. Equivalent chemotherapy efficacy against leukemia in mice treated with topical vasoconstrictors to prevent cancer therapy side effects.

    PubMed

    Graul-Conroy, Amanda; Hicks, Emily J; Fahl, William E

    2016-06-15

    Topically applied vasoconstrictor is a new strategy to prevent oral mucositis and alopecia, two complications of chemotherapy and stem-cell transplant. We sought to determine whether mice treated with topical vasoconstrictor minutes before chemotherapy to suppress L1210 leukemia would develop a vasoconstrictor-induced L1210 cell sanctuary, and with it, significantly worse survival outcomes. B6D2F1 mice received 10(4) mouse L1210 leukemia cells via retro-orbital intravenous injection and were then divided into treatment groups, which included: (i) no further treatment, (ii) a single, sub-curative, intraperitoneal dose of cyclophosphamide (90 µg/gm bw) 24 hr after L1210 cell inoculation, (iii) topical epinephrine (25-400 mM) to clipped dorsal backs 20 min before cyclophosphamide or (iv) orotopical phenylephrine (16-130 mM), epinephrine (10 mM) or norepinephrine (25 mM) 20 min before cyclophosphamide. All mice were then followed until day of death. Differences in median survival time and percent survival between mice receiving cyclophosphamide alone and mice treated with either orotopical phenylephrine, epinephrine or norepinephrine; or topical epinephrine before cyclophosphamide were not significantly different. A discernible leukemia sanctuary was not created by topical vasoconstrictor treatment prior to chemotherapy; there was no significant difference in leukemia progression between untreated mice and those treated with either orotopical or topical vasoconstrictor before chemotherapy. We have opened a Phase I/IIa dose escalation trial to evaluate the safety and efficacy of orotopical phenylephrine in preventing oral mucositis in subjects undergoing hematopoietic stem cell transplant conditioning with cyclophosphamide plus total body irradiation. This could provide a cost-effective and convenient method to prevent oral mucositis. PMID:26860340

  11. Influence of WR 2721 on the efficacy of radiotherapy and chemotherapy of murine tumors

    SciTech Connect

    Clement, J.J.; Johnson, R.K.

    1982-03-01

    The effect of WR2721 on the response of tumors to radiation, antineoplastic alkylating drugs, and DNA binding agents was evaluated and compared to the degree of normal tissue protection provided by WR 2721 against these agents. WR 2721 administered to mice bearing P388 leukemia or Lewis lung carcinoma was found to reduce the radiosensitivity of the leukemia and lung tumor by dose modifying factors of 1.4 and 1.3, respectively. WR 2721 protected bone marow, intestine, and skin from radiation by factors of 1.9. 1.4 and 1.8. WR 2721 protected mice from the lethality of cyclophosphamide by a factor of only 1.2 whereas protection from melphalan toxicity was more dramatic with a dose modifying factor of 1.6. In chemotherapy studies of established M5076 ovarian tumor, the combination of WR 2721 plus cyclophosphamide was equivalent in activity to cyclophosphamide alone. WR 2721 did not modify the antitumor activity of melphalan in early Lewis lung carcinoma did not decrease the antileukemic effects of this agent by a factor of 2.6 indicating tumor protection greater than host protection in the leukemia. The antitumor activity of the DNA binding agents etoposide (VP16-213) and mitoxantrone against systemic P388 leukemia was not diminished by WR 2721, while a substantial increase in host toxicity was noted for the combinations. The protective effects of WR 2721 against radiation and drug damage were, therefore, not entirely selective for normal tissues. In some cases the degree of tumor protection can be similar to, or greater than, normal tissue protection.

  12. The efficacy of X-ray does on murine norovirus-1 (MNV-1) in pure culture, half-shell oyster, salmon sushi, and tuna salad

    Technology Transfer Automated Retrieval System (TEKTRAN)

    In this investigation, we determined the efficacy of X-ray doses on reducing a human norovirus (HuNoV) surrogate [murine norovirus-1 (MNV-1)] in pure culture, half-shell oyster, salmon sushi and tuna salad. The pure culture (phosphate-buffer saline, pH 7.4), half-shell oyster, salmon sushi and tuna ...

  13. Improving anticancer efficacy of (–)-epigallocatechin-3-gallate gold nanoparticles in murine B16F10 melanoma cells

    PubMed Central

    Chen, Cheng-Cheung; Hsieh, Dar-Shih; Huang, Kao-Jean; Chan, Yi-Lin; Hong, Po-Da; Yeh, Ming-Kung; Wu, Chang-Jer

    2014-01-01

    (–)-Epigallocatechin-3-gallate (EGCG), the major bioactive constituent in green tea, has been reported to effectively inhibit the formation and development of tumors. To maximize the effectiveness of EGCG, we attached it to nanogold particles (EGCG-pNG) in various ratios to examine in vitro cytotoxicity and in vivo anti-cancer activity. EGCG-pNG showed improved anti-cancer efficacy in B16F10 murine melanoma cells; the cytotoxic effect in the melanoma cells treated with EGCG-pNG was 4.91 times higher than those treated with EGCG. The enhancement is achieved through mitochondrial pathway-mediated apoptosis as determined by annexin V assay, JC-10 staining, and caspase-3, -8, -9 activity assay. Moreover, EGCG-pNG was 1.66 times more potent than EGCG for inhibition of tumor growth in a murine melanoma model. In the hemolysis assay, the pNG surface conjugated with EGCG is most likely the key factor that contributes to the decreased release of hemoglobin from human red blood cells. PMID:24855338

  14. In vivo localization of ⁹⁰Y and ¹⁷⁷Lu radioimmunoconjugates using Cerenkov luminescence imaging in a disseminated murine leukemia model.

    PubMed

    Balkin, Ethan R; Kenoyer, Aimee; Orozco, Johnnie J; Hernandez, Alexandra; Shadman, Mazyar; Fisher, Darrell R; Green, Damian J; Hylarides, Mark D; Press, Oliver W; Wilbur, D Scott; Pagel, John M

    2014-10-15

    Cerenkov radiation generated by positron-emitting radionuclides can be exploited for a molecular imaging technique known as Cerenkov luminescence imaging (CLI). Data have been limited, however, on the use of medium- to high-energy β-emitting radionuclides of interest for cancer imaging and treatment. We assessed the use of CLI as an adjunct to determine localization of radioimmunoconjugates to hematolymphoid tissues. Radiolabeled (177)Lu- or (90)Y-anti-CD45 antibody (Ab; DOTA-30F11) was administered by tail vein injection to athymic mice bearing disseminated murine myeloid leukemia, with CLI images acquired at times afterward. Gamma counting of individual organs showed preferential uptake in CD45(+) tissues with significant retention of radiolabeled Ab in sites of leukemia (spleen and bone marrow). This result was confirmed in CLI images with 1.35 × 10(5) ± 2.2 × 10(4) p/s/cm(2)/sr and 3.45 × 10(3) ± 7.0 × 10(2) p/s/cm(2)/sr for (90)Y-DOTA-30F11 and (177)Lu-DOTA-30F11, respectively, compared with undetectable signal for both radionuclides using the nonbinding control Ab. Results showed that CLI allows for in vivo visualization of localized β-emissions. Pixel intensity variability resulted from differences in absorbed doses of the associated energies of the β-emitting radionuclide. Overall, our findings offer a preclinical proof of concept for the use of CLI techniques in tandem with currently available clinical diagnostic tools. PMID:25261237

  15. New class of leukemogenic ecotropic recombinant murine leukemia virus isolated from radiation-induced thymomas of C57BL/6 mice

    SciTech Connect

    Rassart, E.; Sankar-Mistry, P.; Lemay, G.; DesGroseillers, L.; Jalicoeur, P.

    1983-02-01

    We previously reported the establishment of several lymphoid cell lines from X-ray-induced thymomas of C57BL/Ka mice, and all, except one, produce retroviruses. Biological characterization of five of these new primary radiation leukemia viruses (RadLVs) indicated that they had a B-tropic, fibrotropic, and ecotropic host range and were leukemogenic when reinjected into C57BL/Ka newborn mice. The leukemogenic potential of one isolate (G/sub 6/T/sub 2/) was further assessed and shown to be retained after prolonged passaging on fibroblasts in vitro. Restriction endonuclease analysis of the DNA of four of our new RadLV isolates (G/sub 6/T/sub 2/, Ti-7, Ti-8, and Ti-9) revealed that G/sub 6/T/sub 2/ and Ti-7 murine leukemia virus (MuLV) genomes had identical restriction maps, whereas Ti-8 and Ti-9 genomes were different from each other and from the G/sub 6/T/sub 2/ and Ti-7 genomes. The physical maps of these genomes were similar to that of known ecotropic MuLV genomes (including the C57BL/Ka endogenous ecotropic MuLV) within their long terminal repeats, env, the right portion of pol, and the left portion of gag. However, a region covering the end of gag and the beginning of pol was different and showed several similarities with xenotropic MuLV genomes of BALB/c, AKR, and C58 mice previously mapped. Our results suggest that these primary RadLV genomes are recombinants between the parental ecotropic MuLV genome and a nonecotropic (xenotropic) sequence. To further study the leukemic potential of these RadLVs, the genome of one of them (G/sub 6/T/sub 2/) was cloned in Charon 21A as an infectious molecule.

  16. In vivo Localization of 90Y- and 177Lu-Radioimmunoconjugates Using Cerenkov Luminescence Imaging in a Disseminated Murine Leukemia Model

    PubMed Central

    Balkin, Ethan R.; Kenoyer, Aimee; Orozco, Johnnie J.; Hernandez, Alexandra; Shadman, Mazyar; Fisher, Darrell R.; Green, Damian J.; Hylarides, Mark D.; Press, Oliver W.; Wilbur, D. Scott; Pagel, John M.

    2014-01-01

    Cerenkov radiation generated by positron-emitting radionuclides can be exploited for a molecular imaging technique known as Cerenkov Luminescence Imaging (CLI). Data have been limited, however, on the use of medium-to-high energy β-emitting radionuclides of interest for cancer imaging and treatment. We assessed the use of CLI as an adjunct to determine localization of radioimmunoconjugates to hematolymphoid tissues. Radiolabeled 177Lu- or 90Y-anti-CD45 antibody (Ab; DOTA-30F11) was administered by tail vein injection to athymic mice bearing disseminated murine myeloid leukemia with CLI images acquired at times afterward. Gamma counting of individual organs showed preferential uptake in CD45+ tissues with significant retention of radiolabeled Ab in sites of leukemia (spleen and bone marrow). This result was confirmed in CLI images with 1.35 × 105 ± 2.2 × 104 p/sec/cm2/sr and 3.45 × 103 ± 7.0 × 102 p/sec/cm2/sr for 90Y-DOTA-30F11 and 177Lu-DOTA-30F11, respectively, compared to undetectable signal for both radionuclides using the non-binding control Ab. Results showed that CLI allows for in vivo visualization of localized β-emissions. Pixel intensity variability resulted from differences in absorbed doses of the associated energies of the β-emitting radionuclide. Overall, our findings offer a preclinical proof of concept for the use of CLI techniques in tandem with currently available clinical diagnostic tools. PMID:25261237

  17. Novel Quinazolinone MJ-29 Triggers Endoplasmic Reticulum Stress and Intrinsic Apoptosis in Murine Leukemia WEHI-3 Cells and Inhibits Leukemic Mice

    PubMed Central

    Lu, Chi-Cheng; Yang, Jai-Sing; Chiang, Jo-Hua; Hour, Mann-Jen; Lin, Kuei-Li; Lin, Jen-Jyh; Huang, Wen-Wen; Tsuzuki, Minoru

    2012-01-01

    The present study was to explore the biological responses of the newly compound, MJ-29 in murine myelomonocytic leukemia WEHI-3 cells in vitro and in vivo fates. We focused on the in vitro effects of MJ-29 on ER stress and mitochondria-dependent apoptotic death in WEHI-3 cells, and to hypothesize that MJ-29 might fully impair the orthotopic leukemic mice. Our results indicated that a concentration-dependent decrease of cell viability was shown in MJ-29-treated cells. DNA content was examined utilizing flow cytometry, whereas apoptotic populations were determined using annexin V/PI, DAPI staining and TUNEL assay. Increasing vital factors of mitochondrial dysfunction by MJ-29 were further investigated. Thus, MJ-29-provaked apoptosis of WEHI-3 cells is mediated through the intrinsic pathway. Importantly, intracellular Ca2+ release and ER stress-associated signaling also contributed to MJ-29-triggered cell apoptosis. We found that MJ-29 stimulated the protein levels of calpain 1, CHOP and p-eIF2α pathways in WEHI-3 cells. In in vivo experiments, intraperitoneal administration of MJ-29 significantly improved the total survival rate, enhanced body weight and attenuated enlarged spleen and liver tissues in leukemic mice. The infiltration of immature myeloblastic cells into splenic red pulp was reduced in MJ-29-treated leukemic mice. Moreover, MJ-29 increased the differentiations of T and B cells but decreased that of macrophages and monocytes. Additionally, MJ-29-stimulated immune responses might be involved in anti-leukemic activity in vivo. Based on these observations, MJ-29 suppresses WEHI-3 cells in vitro and in vivo, and it is proposed that this potent and selective agent could be a new chemotherapeutic candidate for anti-leukemia in the future. PMID:22662126

  18. Efficacy of Liposomal Amphotericin B in Treatment of Systemic Murine Fusariosis

    PubMed Central

    Ortoneda, Montserrat; Capilla, Javier; Pastor, Francisco J.; Pujol, Isabel; Guarro, Josep

    2002-01-01

    We have compared the activities of liposomal amphotericin B (LAMB) at 3, 5, 10, and 20 mg/kg/day and amphotericin B deoxycholate (AMB) at 1.5 and 2.5 mg/kg/day in a murine systemic infection by Fusarium verticillioides. Survival was improved by all treatments except AMB at 1.5 mg/kg/day. The tissue burden in liver was reduced by LAMB at all dosages and by AMB at 2.5 mg/kg/day. The two highest dosages of LAMB showed significant reductions in the spleen. PMID:12069988

  19. Nucleotide Sequence of the Envelope Gene of Gardner-Arnstein Feline Leukemia Virus B Reveals Unique Sequence Homologies with a Murine Mink Cell Focus-Forming Virus †

    PubMed Central

    Elder, John H.; Mullins, James I.

    1983-01-01

    The nucleotide sequence of the envelope gene and the adjacent 3′ long terminal repeat (LTR) of Gardner-Arnstein feline leukemia virus of subgroup B (GA-FeLV-B) has been determined. Comparison of the derived amino acid sequence of the gp70-p15E polyprotein to those of several previously reported murine retroviruses revealed striking homologies between GA-FeLV-B gp70 and the gp70 of a Moloney virus-derived mink cell focus-forming virus. These homologies were located within the substituted (presumably xenotropic) portion of the mink cell focus-forming virus envelope gene and comprised amino acid sequences not present in three ecotropic virus gp70s. In addition, areas of insertions and deletions, in general, were the same between GA-FeLV-B and Moloney mink cell focus-forming virus, although the sizes of the insertions and deletions differed. Homologies between GA-FeLV-B and mink cell focus-forming virus gp70s is functionally significant in that they both possess expanded host ranges, a property dictated by gp70. The amino acid sequence of FeLV-B contains 12 Asn-X-Ser/Thr sequences, indicating 12 possible sites of N-linked glycosylation as compared with 7 or 8 for its murine counterparts. Comparison of the 3′ LTR of GA-FeLV-B to AKR and Moloney virus LTRs revealed extensive conservation in several regions including the “CCAAT” and Goldberg-Hogness (TATA) boxes thought to be involved in promotion of transcription and in the repeat region of the LTR. The inverted repeats that flanked the LTR of GA-FeLV-B were identical to the murine inverted repeats, but were one base longer than the latter. The region of U3 corresponding to the approximately 75-nucleotide “enhancer sequence” is present in GA-FeLV-B, but contains deletions relative to AKR and Moloney virus and is not repeated. An interesting pallindrome in the repeat region immediately 3′ to the U3 region was noted in all the LTRs, but was particularly pronounced in GA-FeLV-B. Possible roles for this

  20. Efficacy and effects of palifermin for the treatment of oral mucositis in patients affected by acute lymphoblastic leukemia.

    PubMed

    Lucchese, Alessandra; Matarese, Giovanni; Ghislanzoni, Luis Huanca; Gastaldi, Giorgio; Manuelli, Maurizio; Gherlone, Enrico

    2016-01-01

    This randomized-controlled trial studied the efficacy of palifermin, administered as a dose during hematopoietic stem cell transplant (HSCT) therapy, as primary prophylaxis on pediatric patients with acute lymphoblastic leukemia in order to reduce oral mucositis (OM). Patients in the palifermin group were randomly assigned to receive palifermin, 60 μg/kg, intravenously as a single dose 3 days before and 0, +1, and +2 post autologous HSCT infusion. The patients in the control group received only a placebo treatment. OM-related assessments were the WHO oral-toxicity scale and the patient-reported outcomes. There was a statistically significant reduction in the incidence of OM grade 3 and 4 in the palifermin group compared to the control group. There was also a reduction in the degree of severity of OM in the palifermin group (1.65 grade respect to 2.33 in the control group). Palifermin could prevent the recurrence of severe OM and improve the quality of life in patients with acute lymphoblastic leukemia (ALL). PMID:26428409

  1. Oncocin Onc72 is efficacious against antibiotic-susceptible Klebsiella pneumoniae ATCC 43816 in a murine thigh infection model.

    PubMed

    Knappe, Daniel; Adermann, Knut; Hoffmann, Ralf

    2015-11-01

    Oncocins and apidaecins are short proline-rich antimicrobial peptides (PrAMPs) representing novel antibiotic drug lead compounds that kill bacteria after internalization and inhibition of intracellular targets (e.g. 70S ribosome and DnaK). Oncocin Onc72 is highly active against Gram-negative bacteria in vitro and in vivo protecting mice in systemic infection models with Escherichia coli and KPC-producing Klebsiella pneumoniae. Here we studied its efficacy in a murine thigh infection model using meropenem as antibiotic comparator that had a 44-fold higher molar in vitro activity than Onc72. Male CD1 mice were rendered neutropenic using cyclophosphamide for four days before intramuscular infection with K. pneumoniae ATCC 43816. After 75 min oncocin Onc72 or the antibiotic comparator meropenem were administered subcutaneously with 100 mg (43 µmol) and 25 mg (65 µmol) per kg of body weight, respectively, six times every 75 min. Onc72 and meropenem administered subcutaneously reduced the thigh tissue burden of K. pneumoniae ATCC 43816 in neutropenic mice significantly by 4.14 and 4.65 a log10 cfu/g, respectively. The bacterial counts were ∼0.5 and ∼1 log10 below the pre-treatment burden, respectively, indicating bactericidal effects for both compounds. Thus, Onc72 was as efficacious as meropenem in vivo despite its much lower in vitro activity determined according to CLSI standard antimicrobial activity tests. PMID:25968331

  2. The prognostic value of polycomb group protein B-cell-specific moloney murine leukemia virus insertion site 1 in stage II colon cancer patients.

    PubMed

    Espersen, Maiken L M; Linnemann, Dorte; Christensen, Ib J; Alamili, Mahdi; Troelsen, Jesper T; Høgdall, Estrid

    2016-07-01

    The aim of this study was to investigate the prognostic value of B-cell-specific moloney murine leukemia virus insertion site 1 (BMI1) protein expression in primary tumors of stage II colon cancer patients. BMI1 protein expression was assessed by immunohistochemistry in a retrospective patient cohort consisting of 144 stage II colon cancer patients. BMI1 expression at the invasive front of the primary tumors correlated with mismatch repair status of the tumors. Furthermore, BMI1 expression at the luminal surface correlated with T-stage, tumor location, and the histological subtypes of the tumors. In a univariate Cox proportional hazard analysis, no statistical significant association between risk of relapse and BMI1 protein expression at the invasive front (HR: 1.12; 95% CI 0.78-1.60; p = 0.53) or at the luminal surface of the tumor (HR: 1.06; 95% CI 0.75-1.48; p = 0.70) was found. Likewise, there was no association between 5-year overall survival and BMI1 expression at the invasive front (HR: 1.12; 95% CI 0.80-1.56; p = 0.46) or at the luminal surface of the tumor (HR: 1.16; 95% CI 0.86-1.60; p = 0.33). In conclusion, BMI1 expression in primary tumors of stage II colon cancer patients could not predict relapse or overall survival of the patients, thus having a limited prognostic value in stage II colon cancer patients. PMID:27102362

  3. Phosphorylation of mouse SAMHD1 regulates its restriction of human immunodeficiency virus type 1 infection, but not murine leukemia virus infection.

    PubMed

    Wang, Feifei; St Gelais, Corine; de Silva, Suresh; Zhang, Hong; Geng, Yu; Shepard, Caitlin; Kim, Baek; Yount, Jacob S; Wu, Li

    2016-01-01

    Human SAMHD1 (hSAMHD1) restricts HIV-1 infection in non-dividing cells by depleting intracellular dNTPs to limit viral reverse transcription. Phosphorylation of hSAMHD1 at threonine (T) 592 by cyclin-dependent kinase (CDK) 1 and CDK2 negatively regulates HIV-1 restriction. Mouse SAMHD1 (mSAMHD1) restricts HIV-1 infection in non-dividing cells, but whether its phosphorylation regulates retroviral restriction is unknown. Here we identified six phospho-sites of mSAMHD1, including T634 that is homologous to T592 of hSAMHD1 and phosphorylated by CDK1 and CDK2. We found that wild-type (WT) mSAMHD1 and a phospho-ablative mutant, but not a phospho-mimetic mutant, restricted HIV-1 infection in differentiated U937 cells. Murine leukemia virus (MLV) infection of dividing NIH3T3 cells was modestly restricted by mSAMHD1 WT and phospho-mutants, but not by a dNTPase-defective mutant. Our results suggest that phosphorylation of mSAMHD1 at T634 by CDK1/2 negatively regulates its HIV-1 restriction in differentiated cells, but does not affect its MLV restriction in dividing cells. PMID:26580513

  4. Early Detection of a Two-Long-Terminal-Repeat Junction Molecule in the Cytoplasm of Recombinant Murine Leukemia Virus-Infected Cells

    PubMed Central

    Serhan, Fatima; Penaud, Magalie; Petit, Caroline; Leste-Lasserre, Thierry; Trajcevski, Stéphane; Klatzmann, David; Duisit, Ghislaine; Sonigo, Pierre; Moullier, Philippe

    2004-01-01

    We showed that a U5-U3 junction was reproducibly detected by a PCR assay as early as 1 to 2 h postinfection with a DNase-treated murine leukemia virus (MLV)-containing supernatant in aphidicolin-arrested NIH 3T3 cells, as well as in nonarrested cells. Such detection is azidothymidine sensitive and corresponded to neosynthesized products of the reverse transcriptase. This observation was confirmed in two additional human cell lines, TE671 and ARPE-19. Using cell fractionation combined with careful controls, we found that a two-long-terminal-repeat (two-LTR) junction molecule was detectable in the cytoplasm as early as 2 h post virus entry. Altogether, our data indicated that the neosynthesized retroviral DNA led to the early formation of structures including true two-LTR junctions in the cytoplasm of MLV-infected cells. Thus, the classical assumption that two-LTR circles are a mitosis-dependent dead-end product accumulating in the nucleus must be reconsidered. MLV-derived products containing a two-LTR junction can no longer be used as an exclusive surrogate for the preintegration complex nuclear translocation event. PMID:15163712

  5. Serologic and PCR testing of persons with chronic fatigue syndrome in the United States shows no association with xenotropic or polytropic murine leukemia virus-related viruses

    PubMed Central

    2011-01-01

    In 2009, a newly discovered human retrovirus, xenotropic murine leukemia virus (MuLV)-related virus (XMRV), was reported by Lombardi et al. in 67% of persons from the US with chronic fatigue syndrome (CFS) by PCR detection of gag sequences. Although six subsequent studies have been negative for XMRV, CFS was defined more broadly using only the CDC or Oxford criteria and samples from the US were limited in geographic diversity, both potentially reducing the chances of identifying XMRV positive CFS cases. A seventh study recently found polytropic MuLV sequences, but not XMRV, in a high proportion of persons with CFS. Here we tested blood specimens from 45 CFS cases and 42 persons without CFS from over 20 states in the United States for both XMRV and MuLV. The CFS patients all had a minimum of 6 months of post-exertional malaise and a high degree of disability, the same key symptoms described in the Lombardi et al. study. Using highly sensitive and generic DNA and RNA PCR tests, and a new Western blot assay employing purified whole XMRV as antigen, we found no evidence of XMRV or MuLV in all 45 CFS cases and in the 42 persons without CFS. Our findings, together with previous negative reports, do not suggest an association of XMRV or MuLV in the majority of CFS cases. PMID:21342521

  6. Molecular cloning of two paralytogenic, temperature-sensitive mutants, ts1 and ts7, and the parental wild-type Moloney murine leukemia virus.

    PubMed Central

    Yuen, P H; Malehorn, D; Nau, C; Soong, M M; Wong, P K

    1985-01-01

    ts1 and ts7, the paralytogenic, temperature-sensitive mutants of Moloney murine leukemia virus (MoMuLV), together with their wild-type parent, MoMuLV-TB, were molecularly cloned. ts1-19, ts7-22, and wt-25, the infectious viruses obtained on transfection to NIH/3T3 cells of the lambda Charon 21A recombinants of ts1, ts7, and wt, were found to have retained the characteristics of their non-molecularly cloned parents. In contrast to the wt virus, ts1-19 and ts7-22 are temperature-sensitive, inefficient in the intracellular processing of Pr80env at the restrictive temperature, and able to induce paralysis in CFW/D mice. Like the non-molecularly cloned ts7, the ts7-22 virion was also shown to be heat labile. The heat lability of the ts7 virion distinguishes it from ts1. Endonuclease restriction mapping with 11 endonucleases demonstrated that the base composition of MoMuLV-TB differs from that of the standard MoMuLV, but no difference was detected between the molecularly cloned ts1 and ts7 genomes. However, ts1 and ts7 differ from MoMuLV in the loss or acquisition of four different restriction sites, whereas they differ from MoMuLV-TB in the loss or acquisition of three different restriction sites. Images PMID:2983112

  7. Nuclear Import of Moloney Murine Leukemia Virus DNA Mediated by Adenovirus Preterminal Protein Is Not Sufficient for Efficient Retroviral Transduction in Nondividing Cells

    PubMed Central

    Lieber, André; Kay, Mark A.; Li, Zong-Yi

    2000-01-01

    Moloney murine leukemia virus (MoMLV)-derived vectors require cell division for efficient transduction, which may be related to an inability of the viral DNA-protein complex to cross the nuclear membrane. In contrast, adenoviruses (Ad) can efficiently infect nondividing cells. This property may be due to the presence of multiple nuclear translocation signals in a number of Ad proteins, which are associated with the incoming viral genomes. Of particular interest is the Ad preterminal protein (pTP), which binds alone or in complex with the Ad polymerase to specific sequences in the Ad inverted terminal repeat. The goal of this study was to test whether coexpression of pTP with retroviral DNA carrying pTP-binding sites would facilitate nuclear import of the viral preintegration complex and transduction of quiescent cells. In preliminary experiments, we demonstrated that the karyophylic pTP can coimport plasmid DNA into the nuclei of growth-arrested cells. Retroviral transduction studies were performed with G1/S-arrested LTA cells or stationary-phase human primary fibroblasts. These studies demonstrated that pTP or pTP-Ad polymerase conferred nuclear import of retroviral DNA upon arrested cells when the retrovirus vector contained the corresponding binding motifs. However, pTP-mediated nuclear translocation of MoMLV DNA in nondividing cells was not sufficient for stable transduction. Additional cellular factors activated during S phase or DNA repair synthesis were required for efficient retroviral integration. PMID:10623734

  8. Highly Efficient Transfer of Chromosomes to a Broad Range of Target Cells Using Chinese Hamster Ovary Cells Expressing Murine Leukemia Virus-Derived Envelope Proteins.

    PubMed

    Suzuki, Teruhiko; Kazuki, Yasuhiro; Oshimura, Mitsuo; Hara, Takahiko

    2016-01-01

    Microcell-mediated chromosome transfer (MMCT) is an essential step for introducing chromosomes from donor cells to recipient cells. MMCT allows not only for genetic/epigenetic analysis of specific chromosomes, but also for utilization of human and mouse artificial chromosomes (HACs/MACs) as gene delivery vectors. Although the scientific demand for genome scale analyses is increasing, the poor transfer efficiency of the current method has hampered the application of chromosome engineering technology. Here, we developed a highly efficient chromosome transfer method, called retro-MMCT, which is based on Chinese hamster ovary cells expressing envelope proteins derived from ecotropic or amphotropic murine leukemia viruses. Using this method, we transferred MACs to NIH3T3 cells with 26.5 times greater efficiency than that obtained using the conventional MMCT method. Retro-MMCT was applicable to a variety of recipient cells, including embryonic stem cells. Moreover, retro-MMCT enabled efficient transfer of MAC to recipient cells derived from humans, monkeys, mice, rats, and rabbits. These results demonstrate the utility of retro-MMCT for the efficient transfer of chromosomes to various types of target cell. PMID:27271046

  9. The conserved His8 of the Moloney murine leukemia virus Env SU subunit directs the activity of the SU-TM disulphide bond isomerase

    SciTech Connect

    Li Kejun; Zhang, Shujing; Kronqvist, Malin; Ekstroem, Maria; Wallin, Michael; Garoff, Henrik . E-mail: henrik.garoff@cbt.ki.se

    2007-04-25

    Murine leukemia virus (MLV) fusion is controlled by isomerization of the disulphide bond between the receptor-binding surface (SU) and fusion-active transmembrane subunits of the Env-complex. The bond is in SU linked to a CXXC motif. This carries a free thiol that upon receptor binding can be activated (ionized) to attack the disulphide and rearrange it into a disulphide isomer within the motif. To find out whether His8 in the conserved SPHQ sequence of Env directs thiol activation, we analyzed its ionization in MLV vectors with wtEnv and Env with His8 deleted or substituted for Tyr or Arg, which partially or completely arrests fusion. The ionization was monitored by following the pH effect on isomerization in vitro by Ca{sup 2+} depletion or in vivo by receptor binding. We found that wtEnv isomerized optimally at slightly basic pH whereas the partially active mutant required higher and the inactive mutants still higher pH. This suggests that His8 directs the ionization of the CXXC thiol.

  10. Retrovirus gene expression during the cell cycle. I. Virus production, synthesis, and expression of viral proteins in Rauscher murine leukemia virus-infected mouse cells.

    PubMed Central

    Balazs, I; Caldarella, J

    1981-01-01

    Synchronized mouse cells (JLS-V9) chronically infected with Rauscher murine leukemia virus were used to study virus production, the synthesis of gag and env precursor proteins, and the expression of env protein on the cell surface during the cell cycle. The amount of virus released into the medium by synchronized cells during a 30-min interval was determined by using the XC plaque assay and by measuring reverse transcriptase activity. The results show that virus production occurs during mitosis. Labeling of the cell surface of synchronized cells with 125I or with fluorescein-conjugated antiserum shows that the amount of gp 70env on the cell surface parallels cellular growth. Therefore, the cell cycle-dependent release of virus is not accompanied by similar variations in the amount of viral envelope protein on the cell surface. Immunoprecipitation of cells labeled with [35S]methionine, followed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, was used to measure viral protein synthesis during the cell cycle. The rate of synthesis of gag precursor proteins show three maximums corresponding to the G1, middle S, and late S to G2 phases of the cell cycle. The rate of synthesis of env precursor proteins does not change, suggesting that in these cells the synthesis of these two gene products is controlled separately. Images PMID:7288918

  11. trans-dominant interference with virus infection at two different stages by a mutant envelope protein of Friend murine leukemia virus.

    PubMed Central

    Matano, T; Odawara, T; Ohshima, M; Yoshikura, H; Iwamoto, A

    1993-01-01

    A dominant negative mutant Friend murine leukemia virus (FMLV) env gene was cloned from an immunoselected Friend erythroleukemia cell. The mutant env had a point mutation which resulted in a Cys-to-Arg substitution at the 361st amino acid in the FMLV envelope protein (Env). The mutant Env was retained in the endoplasmic reticulum (ER) and accumulated because of its slow degradation. The NIH 3T3 cells expressing the mutant env were resistant to ecotropic Moloney MLV (MoMLV) penetration, suggesting that the mutant Env traps the ecotropic MLV receptors in the ER. When the mutant env gene was transfected into and expressed in the cells persistently infected with MoMLV, the wild-type Env was trapped in the ER, and the MoMLV production was suppressed. Thus, the mutant Env accumulating in the ER trans-dominantly and efficiently interfered with the ecotropic MLV infection at both the early and the late stages. Images PMID:8445721

  12. Highly Efficient Transfer of Chromosomes to a Broad Range of Target Cells Using Chinese Hamster Ovary Cells Expressing Murine Leukemia Virus-Derived Envelope Proteins

    PubMed Central

    Kazuki, Yasuhiro; Oshimura, Mitsuo; Hara, Takahiko

    2016-01-01

    Microcell-mediated chromosome transfer (MMCT) is an essential step for introducing chromosomes from donor cells to recipient cells. MMCT allows not only for genetic/epigenetic analysis of specific chromosomes, but also for utilization of human and mouse artificial chromosomes (HACs/MACs) as gene delivery vectors. Although the scientific demand for genome scale analyses is increasing, the poor transfer efficiency of the current method has hampered the application of chromosome engineering technology. Here, we developed a highly efficient chromosome transfer method, called retro-MMCT, which is based on Chinese hamster ovary cells expressing envelope proteins derived from ecotropic or amphotropic murine leukemia viruses. Using this method, we transferred MACs to NIH3T3 cells with 26.5 times greater efficiency than that obtained using the conventional MMCT method. Retro-MMCT was applicable to a variety of recipient cells, including embryonic stem cells. Moreover, retro-MMCT enabled efficient transfer of MAC to recipient cells derived from humans, monkeys, mice, rats, and rabbits. These results demonstrate the utility of retro-MMCT for the efficient transfer of chromosomes to various types of target cell. PMID:27271046

  13. Enhancer mutations of Akv murine leukemia virus inhibit the induction of mature B-cell lymphomas and shift disease specificity towards the more differentiated plasma cell stage

    SciTech Connect

    Sorensen, Karina Dalsgaard; Kunder, Sandra; Quintanilla-Martinez, Leticia; Sorensen, Jonna; Schmidt, Joerg; Pedersen, Finn Skou . E-mail: fsp@mb.au.dk

    2007-05-25

    This study investigates the role of the proviral transcriptional enhancer for B-lymphoma induction by exogenous Akv murine leukemia virus. Infection of newborn inbred NMRI mice with Akv induced 35% plasma cell proliferations (PCPs) (consistent with plasmacytoma), 33% diffuse large B-cell lymphomas, 25% follicular B-cell lymphomas and few splenic marginal zone and small B-cell lymphomas. Deleting one copy of the 99-bp proviral enhancer sequence still allowed induction of multiple B-cell tumor types, although PCPs dominated (77%). Additional mutation of binding sites for the glucocorticoid receptor, Ets, Runx, or basic helix-loop-helix transcription factors in the proviral U3 region, however, shifted disease induction to almost exclusively PCPs, but had no major influence on tumor latency periods. Southern analysis of immunoglobulin rearrangements and ecotropic provirus integration patterns showed that many of the tumors/cell proliferations induced by each virus were polyclonal. Our results indicate that enhancer mutations weaken the ability of Akv to induce mature B-cell lymphomas prior to the plasma cell stage, whereas development of plasma cell proliferations is less dependent of viral enhancer strength.

  14. A. cantoniensis inhibits the proliferation of murine leukemia WEHI-3 cells in vivo and promotes immunoresponses in vivo.

    PubMed

    Tan, Tzu-Wei; Lin, Yuh-Tzy; Yang, Jai-Sing; Lu, Chi-Cheng; Chiang, Jo-Hua; Wu, Chang-Lin; Lin, Jing-Pin; Tang, Nou-Ying; Yeh, Chin-Chung; Fan, Ming-Jen; Chung, Jing-Gung

    2009-01-01

    Ampelopsis cantoniensis (AC) has been used as a folk medicine for reducing pain in the Taiwanese population. Our previous studies have shown that the crude extract of AC induced apoptosis in human promyelocytic leukemia HL-60 cells. In this study, the in vivo effects of AC on leukemia WEHI-3 cells and immune responses such as phagocytosis and natural killer (NK) cell activity were investigated. The weights of the livers and spleens were decreased in the AC-treated groups compared to the control groups. The AC treatment increased the percentage of CD3 and CD19 marker cells in WEHI-3-injected mice, indicating that the precursors of T and B cells were inhibited. The AC treatment promoted the activity of macrophage phagocytosis in the peripheral blood mononuclear cells (PBMC) and peritoneal cells. It was found that the NK cells from mice after treatment with AC can kill the YAC-1 target cells. Therefore, the AC treatment increased NK cell activity. In conclusion, AC can affect WEHI-3 cells in vivo and promote macrophage and NK cell activities. PMID:19567391

  15. Efficacy of the Mineral Oil and Hyaluronic Acid Mixture Eye Drops in Murine Dry Eye

    PubMed Central

    Choi, Jung Han; Kim, Jung Han; Li, Zhengri; Oh, Han Jin; Ahn, Kyu Youn

    2015-01-01

    Purpose To investigate the therapeutic effects of mineral oil (MO) and hyaluronic acid (HA) mixture eye drops on the tear film and ocular surface in a mouse model of experimental dry eye (EDE). Methods Eye drops consisting of 0.1% HA alone or mixed with 0.1%, 0.5%, or 5.0% MO were applied to desiccating stress-induced murine dry eyes. Tear volume, corneal irregularity score, tear film break-up time (TBUT), and corneal fluorescein staining scores were measured at 5 and 10 days after treatment. Ten days after treatment, goblet cells in the conjunctiva were counted after Periodic acid-Schiff staining. Results There was no significant difference in the tear volume between desiccating stress-induced groups. The corneal irregularity score was lower in the 0.5% MO group compared with the EDE and HA groups. The 0.5% and 5.0% MO groups showed a significant improvement in TBUT compared with the EDE group. Mice treated with 0.1% and 0.5% MO mixture eye drops showed a significant improvement in fluorescein staining scores compared with the EDE group and the HA group. The conjunctival goblet cell count was higher in the 0.5% MO group compared with the EDE group and HA group. Conclusions The MO and HA mixture eye drops had a beneficial effect on the tear films and ocular surface of murine dry eye. The application of 0.5% MO and 0.1% HA mixture eye drops could improve corneal irregularity, the corneal fluorescein staining score, and conjunctival goblet cell count compared with 0.1% HA eye drops in the treatment of EDE. PMID:25829831

  16. In vivo efficacy and pharmacokinetics of biapenem in a murine model of ventilator-associated pneumonia with Pseudomonas aeruginosa.

    PubMed

    Yamada, Koichi; Yamamoto, Yoshihiro; Yanagihara, Katsunori; Araki, Nobuko; Harada, Yosuke; Morinaga, Yoshitomo; Izumikawa, Koichi; Kakeya, Hiroshi; Hasegawa, Hiroo; Kohno, Shigeru; Kamihira, Shimeru

    2012-08-01

    Biapenem (BIPM) has high bactericidal activity against Pseudomonas aeruginosa and similar activity in vitro as meropenem (MEPM). We used a murine model to examine the efficacy of biapenem against ventilator-associated pneumonia (VAP) caused by P. aeruginosa. Mice were treated by intraperitoneal injection with 100 mg/kg BIPM or MEPM every 12 h beginning 12 h after inoculation with P. aeruginosa. Survival was evaluated for 7 days, and 24 h after infection, lung histopathology was analyzed and the number of viable bacteria in the lungs and blood was counted. In addition, the pharmacokinetics of BIPM and MEPM were analyzed after the initial treatment. BIPM and MEPM significantly prolonged survival compared to control (P < 0.05). The lungs of mice treated with BIPM or MEPM had significantly fewer viable bacteria (3.54 ± 0.28 vs. 3.77 ± 0.14 log(10) CFU/ml) than in the lungs of control mice (6.65 ± 0.57 log(10) CFU/ml) (P < 0.05). Furthermore, viable bacteria were not detected in the blood of mice treated with BIPM or MEPM (control 2.85 ± 0.85 log(10) CFU/ml) (P < 0.05). Histopathological examination of lung specimens indicated that BIPM and MEPM prevent the progression of lung inflammation, including alveolar neutrophil infiltration and hemorrhage. The % time above MIC for BIPM and MEPM was 15.4% and 18.3% in plasma and 19.8% and 19.8% in lungs, respectively. These results show that BIPM and MEPM significantly prolongs survival and reduces the number of viable bacteria in a murine model of VAP caused by P. aeruginosa. Therefore, BIPM might be a potent and effective treatment for VAP caused by this bacterium. PMID:22215228

  17. Efficacy of Lysophosphatidylcholine in Combination with Antimicrobial Agents against Acinetobacter baumannii in Experimental Murine Peritoneal Sepsis and Pneumonia Models.

    PubMed

    Parra Millán, R; Jiménez Mejías, M E; Sánchez Encinales, V; Ayerbe Algaba, R; Gutiérrez Valencia, A; Pachón Ibáñez, M E; Díaz, C; Pérez Del Palacio, J; López Cortés, L F; Pachón, J; Smani, Y

    2016-08-01

    Immune response stimulation to prevent infection progression may be an adjuvant to antimicrobial treatment. Lysophosphatidylcholine (LPC) is an immunomodulator involved in immune cell recruitment and activation. In this study, we aimed to evaluate the efficacy of LPC in combination with colistin, tigecycline, or imipenem in experimental murine models of peritoneal sepsis and pneumonia. We used Acinetobacter baumannii strain Ab9, which is susceptible to colistin, tigecycline, and imipenem, and multidrug-resistant strain Ab186, which is susceptible to colistin and resistant to tigecycline and imipenem. Pharmacokinetic and pharmacodynamic parameters for colistin, tigecycline, and imipenem and the 100% minimal lethal dose (MLD100) were determined for both strains. The therapeutic efficacies of LPC, colistin (60 mg/kg of body weight/day), tigecycline (10 mg/kg/day), and imipenem (180 mg/kg/day), alone or in combination, were assessed against Ab9 and Ab186 at the MLD100 in murine peritoneal sepsis and pneumonia models. The levels of pro- and anti-inflammatory cytokines, i.e., tumor necrosis factor alpha (TNF-α) and interleukin-10 (IL-10), were determined by enzyme-linked immunosorbent assay (ELISA) for the same experimental models after inoculating mice with the MLD of both strains. LPC in combination with colistin, tigecycline, or imipenem markedly enhanced the bacterial clearance of Ab9 and Ab186 from the spleen and lungs and reduced bacteremia and mouse mortality rates (P < 0.05) compared with those for colistin, tigecycline, and imipenem monotherapies. Moreover, at 4 h post-bacterial infection, Ab9 induced higher TNF-α and lower IL-10 levels than those with Ab186 (4 μg/ml versus 3 μg/ml [P < 0.05] and 2 μg/ml versus 3.4 μg/ml [P < 0.05], respectively). LPC treatment combined with colistin, tigecycline, or imipenem modestly reduced the severity of infection by A. baumannii strains with different resistance phenotypes compared to LPC monotherapy in both

  18. Efficacy of a Novel Tricyclic Topoisomerase Inhibitor in a Murine Model of Neisseria gonorrhoeae Infection.

    PubMed

    Savage, Victoria J; Charrier, Cédric; Salisbury, Anne-Marie; Box, Helen; Chaffer-Malam, Nathan; Huxley, Anthony; Kirk, Ralph; Noonan, Gary M; Mohmed, Sarfraz; Craighead, Mark W; Ratcliffe, Andrew J; Best, Stuart A; Stokes, Neil R

    2016-09-01

    There is an urgent need for new antibiotics to treat multidrug-resistant Neisseria gonorrhoeae In this report, the microbiology, in vivo pharmacokinetics, and efficacy of REDX05931, a representative novel tricyclic topoisomerase inhibitor, were evaluated. REDX05931 demonstrated high oral bioavailability in mice and reduced N. gonorrhoeae infection after a single dose in a mouse model of gonorrhea. These data support the potential of this series of small molecules as a new treatment for drug-resistant gonorrheal infections. PMID:27324777

  19. Efficacy of quercetin against chemically induced murine oral squamous cell carcinoma

    PubMed Central

    DROGUETT, DANIEL; CASTILLO, CHRISTIAN; LEIVA, ELBA; THEODULOZ, CRISTINA; SCHMEDA-HIRSCHMANN, GUILLERMO; KEMMERLING, ULRIKE

    2015-01-01

    Oral squamous cell carcinoma (OSCC) is the most common form of head and neck cancer, and oxidative damage is associated with the development of OSCCs. Antioxidants have therefore been proposed for use as chemoprotective agents against different types of cancer. In the present study, the effect of the antioxidant quercetin, administered at doses of 10 and 100 mg/kg/day, was investigated in an experimental murine model of 4-nitroquinoline 1-oxide (4-NQO)-induced carcinogenesis. The survival of the treated animals, the plasmatic levels of reduced glutathione and the type and severity of lesions (according the International Histological Classification of Tumors and Bryne's Multifactorial Grading System for the Invasive Tumor Front) were assessed. Additionally, the organization of the extracellular matrix was analyzed by carbohydrate and collagen histochemistry, and immunohistochemistry was used to assess the expression of the tumor markers proliferating cell nuclear antigen and mutated p53. The results indicate that, despite the promising effect of quercetin in other studies, this drug is ineffective as a chemoprotective agent against 4-NQO-induced OSCC in mice at the assayed doses. PMID:26622865

  20. Preclinical efficacy of maternal embryonic leucine-zipper kinase (MELK) inhibition in acute myeloid leukemia.

    PubMed

    Alachkar, Houda; Mutonga, Martin B G; Metzeler, Klaus H; Fulton, Noreen; Malnassy, Gregory; Herold, Tobias; Spiekermann, Karsten; Bohlander, Stefan K; Hiddemann, Wolfgang; Matsuo, Yo; Stock, Wendy; Nakamura, Yusuke

    2014-12-15

    Maternal embryonic leucine-zipper kinase (MELK), which was reported to be frequently up-regulated in various types of solid cancer, plays critical roles in formation and maintenance of cancer stem cells. However, little is known about the relevance of this kinase in hematologic malignancies. Here we report characterization of possible roles of MELK in acute myeloid leukemia (AML). MELK is expressed in AML cell lines and AML blasts with higher levels in less differentiated cells. MELK is frequently upregulated in AML with complex karyotypes and is associated with worse clinical outcome. MELK knockdown resulted in growth inhibition and apoptosis of leukemic cells. Hence, we investigated the potent anti-leukemia activity of OTS167, a small molecule MELK kinase inhibitor, in AML, and found that the compound induced cell differentiation and apoptosis as well as decreased migration of AML cells. MELK expression was positively correlated with the expression of FOXM1 as well as its downstream target genes. Furthermore, MELK inhibition resulted in downregulation of FOXM1 activity and the expression of its downstream targets. Taken together, and given that OTS167 is undergoing a phase I clinical trial in solid cancer, our study warrants clinical evaluation of this compound as a novel targeted therapy for AML patients. PMID:25365263

  1. New Class of Leukemogenic Ecotropic Recombinant Murine Leukemia Virus Isolated from Radiation-Induced Thymomas of C57BL/6 Mice

    PubMed Central

    Rassart, E.; Sankar-Mistry, P.; Lemay, G.; DesGroseillers, L.; Jolicoeur, P.

    1983-01-01

    We previously reported the establishment of several lymphoid cell lines from X-ray-induced thymomas of C57BL/Ka mice, and all, except one, produce retroviruses (P. Sankar-Mistry and P. Jolicoeur, J. Virol.35:270-275, 1980). Biological characterization of five of these new primary radiation leukemia viruses (RadLVs) indicated that they had a B-tropic, fibrotropic, and ecotropic host range and were leukemogenic when reinjected into C57BL/Ka newborn mice. The leukemogenic potential of one isolate (G6T2) was further assessed and shown to be retained after prolonged passaging on fibroblasts in vitro. Restriction endonuclease analysis of the DNA of four of our new RadLV isolates (G6T2, Ti-7, Ti-8, and Ti-9) revealed that G6T2 and Ti-7 murine leukemia virus (MuLV) genomes had identical restriction maps, whereas Ti-8 and Ti-9 genomes were different from each other and from the G6T2 and Ti-7 genomes. The physical maps of these genomes were similar to that of known ecotropic MuLV genomes (including the C57BL/Ka endogenous ecotropic MuLV) within their long terminal repeats, env, the right portion of pol, and the left portion of gag. However, a region covering the end of gag and the beginning of pol was different and showed several similarities with xenotropic MuLV genomes of BALB/c, AKR, and C58 mice previously mapped. Our results suggest that these primary RadLV genomes are recombinants between the parental ecotropic MuLV genome and a nonecotropic (xenotropic) sequence. This nonecotropic gag-pol region might be important in conferring the leukemogenic potential to these isolates. Therefore, these RadLVs appear to form a new class of leukemogenic recombinant MuLVs recovered from leukemic tissues of mice. They appear to be distinct from the recombinant AKR mink cell focus-inducing MuLVs which have a dual-tropic host range and harbor xenotropic env sequences. To further study the leukemogenic potential of these RadLVs, the genome of one of them (G6T2) was cloned in Charon 21A

  2. Efficacy of Astaxanthin for the Treatment of Atopic Dermatitis in a Murine Model.

    PubMed

    Yoshihisa, Yoko; Andoh, Tsugunobu; Matsunaga, Kenji; Rehman, Mati Ur; Maoka, Takashi; Shimizu, Tadamichi

    2016-01-01

    Atopic dermatitis (AD) is a common chronic inflammatory skin disease associated with various factors, including immunological abnormalities and exposure to allergens. Astaxanthin (AST) is a xanthophyll carotenoid that has recently been demonstrated to have anti-inflammatory effects and to regulate the expression of inflammatory cytokines. Thus, we investigated whether AST could improve the dermatitis and pruritus in a murine model of AD using NC/Nga mice. In addition to a behavioral evaluation, the effects of AST on the AD were determined by the clinical skin severity score, serum IgE level, histological analyses of skin, and by reverse transcription-PCR and Western blotting analyses for the expression of inflammation-related factors. AST (100 mg/kg) or vehicle (olive oil) was orally administered once day and three times a week for 26 days. When compared with vehicle-treated group, the administration of AST significantly reduced the clinical skin severity score. In addition, the spontaneous scratching in AD model mice was reduced by AST administration. Moreover, the serum IgE level was markedly decreased by the oral administration of AST compared to that in vehicle-treated mice. The number of eosinophils, total and degranulated mast cells all significantly decreased in the skin of AST-treated mice compared with vehicle-treated mice. The mRNA and protein levels of eotaxin, MIF, IL-4, IL-5 and L-histidine decarboxylase were significantly decreased in the skin of AST-treated mice compared with vehicle-treated mice. These results suggest that AST improves the dermatitis and pruritus in AD via the regulation of the inflammatory effects and the expression of inflammatory cytokines. PMID:27023003

  3. PASylation of Murine Leptin Leads to Extended Plasma Half-Life and Enhanced in Vivo Efficacy.

    PubMed

    Morath, Volker; Bolze, Florian; Schlapschy, Martin; Schneider, Sarah; Sedlmayer, Ferdinand; Seyfarth, Katrin; Klingenspor, Martin; Skerra, Arne

    2015-05-01

    Leptin plays a central role in the control of energy homeostasis and appetite and, thus, has attracted attention for therapeutic approaches in spite of its limited pharmacological activity owing to the very short circulation in the body. To improve drug delivery and prolong plasma half-life, we have fused murine leptin with Pro/Ala/Ser (PAS) polypeptides of up to 600 residues, which adopt random coil conformation with expanded hydrodynamic volume in solution and, consequently, retard kidney filtration in a similar manner as polyethylene glycol (PEG). Relative to unmodified leptin, size exclusion chromatography and dynamic light scattering revealed an approximately 21-fold increase in apparent size and a much larger molecular diameter of around 18 nm for PAS(600)-leptin. High receptor-binding activity for all PASylated leptin versions was confirmed in BIAcore measurements and cell-based dual-luciferase assays. Pharmacokinetic studies in mice revealed a much extended plasma half-life after ip injection, from 26 min for the unmodified leptin to 19.6 h for the PAS(600) fusion. In vivo activity was investigated after single ip injection of equimolar doses of each leptin version. Strongly increased and prolonged hypothalamic STAT3 phosphorylation was detected for PAS(600)-leptin. Also, a reduction in daily food intake by up to 60% as well as loss in body weight of >10% lasting for >5 days was observed, whereas unmodified leptin was merely effective for 1 day. Notably, application of a PASylated superactive mouse leptin antagonist (SMLA) led to the opposite effects. Thus, PASylated leptin not only provides a promising reagent to study its physiological role in vivo but also may offer a superior drug candidate for clinical therapy. PMID:25811325

  4. Efficacy of Astaxanthin for the Treatment of Atopic Dermatitis in a Murine Model

    PubMed Central

    Yoshihisa, Yoko; Andoh, Tsugunobu; Matsunaga, Kenji; Rehman, Mati Ur; Maoka, Takashi; Shimizu, Tadamichi

    2016-01-01

    Atopic dermatitis (AD) is a common chronic inflammatory skin disease associated with various factors, including immunological abnormalities and exposure to allergens. Astaxanthin (AST) is a xanthophyll carotenoid that has recently been demonstrated to have anti-inflammatory effects and to regulate the expression of inflammatory cytokines. Thus, we investigated whether AST could improve the dermatitis and pruritus in a murine model of AD using NC/Nga mice. In addition to a behavioral evaluation, the effects of AST on the AD were determined by the clinical skin severity score, serum IgE level, histological analyses of skin, and by reverse transcription-PCR and Western blotting analyses for the expression of inflammation-related factors. AST (100 mg/kg) or vehicle (olive oil) was orally administered once day and three times a week for 26 days. When compared with vehicle-treated group, the administration of AST significantly reduced the clinical skin severity score. In addition, the spontaneous scratching in AD model mice was reduced by AST administration. Moreover, the serum IgE level was markedly decreased by the oral administration of AST compared to that in vehicle-treated mice. The number of eosinophils, total and degranulated mast cells all significantly decreased in the skin of AST-treated mice compared with vehicle-treated mice. The mRNA and protein levels of eotaxin, MIF, IL-4, IL-5 and L-histidine decarboxylase were significantly decreased in the skin of AST-treated mice compared with vehicle-treated mice. These results suggest that AST improves the dermatitis and pruritus in AD via the regulation of the inflammatory effects and the expression of inflammatory cytokines. PMID:27023003

  5. Synergistic Efficacy from Gene Therapy with Coreceptor Blockade and a β2-Agonist in Murine Pompe Disease.

    PubMed

    Han, Sang-oh; Li, Songtao; Bird, Andrew; Koeberl, Dwight

    2015-11-01

    Pompe disease (glycogen storage disease type II; acid maltase deficiency) is a devastating myopathy resulting from acid α-glucosidase (GAA) deficiency in striated and smooth muscle. Despite the availability of enzyme replacement therapy (ERT) with recombinant human GAA (rhGAA), the limitations of ERT have prompted the preclinical development of gene therapy. Gene therapy has the advantage of continuously producing GAA, in contrast to ERT, which requires frequent injections of rhGAA. An adeno-associated viral (AAV) vector containing a muscle-specific promoter, AAV-MHCK7hGAApA, achieved high GAA expression in heart and skeletal muscle in mice with Pompe disease. However, elevated GAA activity was not sufficient to completely clear accumulated glycogen in skeletal muscle. The process of glycogen clearance from lysosomes might require improved trafficking of GAA to the lysosomes in skeletal muscle, previously achieved with the β(2)-agonist clenbuterol that enhanced glycogen clearance in skeletal muscle without increasing GAA activity. Glycogen clearance was clearly enhanced by treatment with a nondepleting anti-CD4 monoclonal antibody (anti-CD4 mAb) along with muscle-specific GAA expression in cardiac muscle, but that treatment was not effective in skeletal muscle. Furthermore, anti-CD4 mAb treatment along with clenbuterol achieved synergistic therapeutic efficacy in both cardiac and skeletal muscle. This triple therapy increased both muscle strength and weight gain. Overall, triple therapy to enhance GAA trafficking and to suppress immune responses significantly improved the efficacy of muscle-targeted gene therapy in murine Pompe disease. PMID:26417913

  6. Efficacy of Oritavancin in a Murine Model of Bacillus anthracis Spore Inhalation Anthrax ▿

    PubMed Central

    Heine, H. S.; Bassett, J.; Miller, L.; Bassett, A.; Ivins, B. E.; Lehoux, D.; Arhin, F. F.; Parr, T. R.; Moeck, G.

    2008-01-01

    The inhaled form of Bacillus anthracis infection may be fatal to humans. The current standard of care for inhalational anthrax postexposure prophylaxis is ciprofloxacin therapy twice daily for 60 days. The potent in vitro activity of oritavancin, a semisynthetic lipoglycopeptide, against B. anthracis (MIC against Ames strain, 0.015 μg/ml) prompted us to test its efficacy in a mouse aerosol-anthrax model. In postexposure prophylaxis dose-ranging studies, a single intravenous (i.v.) dose of oritavancin of 5, 15, or 50 mg/kg 24 h after a challenge with 50 to 75 times the median lethal dose of Ames strain spores provided 40, 70, and 100% proportional survival, respectively, at 30 days postchallenge. Untreated animals died within 4 days of challenge, whereas 90% of control animals receiving ciprofloxacin at 30 mg/kg intraperitoneally twice daily for 14 days starting 24 h after challenge survived. Oritavancin demonstrated significant activity post symptom development; a single i.v. dose of 50 mg/kg administered 42 h after challenge provided 56% proportional survival at 30 days. In a preexposure prophylaxis study, a single i.v. oritavancin dose of 50 mg/kg administered 1, 7, 14, or 28 days before lethal challenge protected 90, 100, 100, and 20% of mice at 30 days; mice treated with ciprofloxacin 24 h or 24 and 12 h before challenge all died within 5 days. Efficacy in pre- and postexposure models of inhalation anthrax, together with a demonstrated low propensity to engender resistance, promotes further study of oritavancin pharmacokinetics and efficacy in nonhuman primate models. PMID:18606841

  7. Atomic force microscopy investigation of fibroblasts infected with wild-type and mutant murine leukemia virus (MuLV).

    PubMed Central

    Kuznetsov, Yurii G; Datta, Shoibal; Kothari, Natantara H; Greenwood, Aaron; Fan, Hung; McPherson, Alexander

    2002-01-01

    NIH 3T3 cells were infected in culture with the oncogenic retrovirus, mouse leukemia virus (MuLV), and studied using atomic force microscopy (AFM). Cells fixed with glutaraldehyde alone, and those postfixed with osmium tetroxide, were imaged under ethanol according to procedures that largely preserved their structures. With glutaraldehyde fixation alone, the lipid bilayer was removed and maturing virions were seen emerging from the cytoskeletal matrix. With osmium tetroxide postfixation, the lipid bilayer was maintained and virions were observable still attached to the cell surfaces. The virions on the cell surfaces were imaged at high resolution and considerable detail of the arrangement of protein assemblies on their surfaces was evident. Infected cells were also labeled with primary antibodies against the virus env surface protein, followed by secondary antibodies conjugated with colloidal gold particles. Other 3T3 cells in culture were infected with MuLV containing a mutation in the gPr80(gag) gene. Those cells were observed by AFM not to produce normal MuLV on their surfaces, or at best, only at very low levels. The cell surfaces, however, became covered with tubelike structures that appear to result from a failure of the virions to properly undergo morphogenesis, and to fail in budding completely from the cell's surfaces. PMID:12496133

  8. Improved BM212 MmpL3 Inhibitor Analogue Shows Efficacy in Acute Murine Model of Tuberculosis Infection

    PubMed Central

    Alfonso, Salvatore; Cocozza, Martina; Porretta, Giulio Cesare; Ballell, Lluís; Rullas, Joaquin; Ortega, Fátima; De Logu, Alessandro; Agus, Emanuela; La Rosa, Valentina; Pasca, Maria Rosalia; De Rossi, Edda; Wae, Baojie; Franzblau, Scott G.; Manetti, Fabrizio; Botta, Maurizio; Biava, Mariangela

    2013-01-01

    1,5-Diphenyl pyrroles were previously identified as a class of compounds endowed with high in vitro efficacy against M. tuberculosis. To improve the physical chemical properties and drug-like parameters of this class of compounds, a medicinal chemistry effort was undertaken. By selecting the optimal substitution patterns for the phenyl rings at N1 and C5 and by replacing the thiomorpholine moiety with a morpholine one, a new series of compounds was produced. The replacement of the sulfur with oxygen gave compounds with lower lipophilicity and improved in vitro microsomal stability. Moreover, since the parent compound of this family has been shown to target MmpL3, mycobacterial mutants resistant to two compounds have been isolated and characterized by sequencing the mmpL3 gene; all the mutants showed point mutations in this gene. The best compound identified to date was progressed to dose-response studies in an acute murine TB infection model. The resulting ED99 of 49 mg/Kg is within the range of commonly employed tuberculosis drugs, demonstrating the potential of this chemical series. The in vitro and in vivo target validation evidence presented here adds further weight to MmpL3 as a druggable target of interest for anti-tubercular drug discovery. PMID:23437287

  9. Sanitizer Efficacy against Murine Norovirus, a Surrogate for Human Norovirus, on Stainless Steel Surfaces when Using Three Application Methods

    PubMed Central

    Kotwal, Grishma; Harrison, Mark A.; Law, S. Edward; Harrison, Judy A.

    2013-01-01

    Human noroviruses are major etiologic agents of epidemic gastroenteritis. Outbreaks are often accompanied by contamination of environmental surfaces, but since these viruses cannot be routinely propagated in laboratory cultures, their response to surface disinfectants is predicted by using surrogates, such as murine norovirus 1 (MNV-1). This study compared the virucidal efficacies of various liquid treatments (three sanitizer liquids, 5% levulinic acid plus 2% SDS [LEV/SDS], 200 ppm chlorine, and an isopropanol-based quaternary ammonium compound [Alpet D2], and two control liquids, sterile tap water and sterile tap water plus 2% SDS) when delivered to MNV-1-inoculated stainless steel surfaces by conventional hydraulic or air-assisted, induction-charged (AAIC) electrostatic spraying or by wiping with impregnated towelettes. For the spray treatments, LEV/SDS proved effective when applied with hydraulic and AAIC electrostatic spraying, providing virus reductions of 2.71 and 1.66 log PFU/ml, respectively. Alpet D2 provided a 2.23-log PFU/ml reduction with hydraulic spraying, outperforming chlorine (1.16-log PFU/ml reduction). Chlorine and LEV/SDS were equally effective as wipes, reducing the viral load by 7.05 log PFU/ml. Controls reduced the viral load by <1 log with spraying applications and by >3 log PFU/ml with wiping. Results indicated that both sanitizer type and application methods should be carefully considered when choosing a surface disinfectant to best prevent and control environmental contamination by noroviruses. PMID:23263949

  10. Efficacy of paracetamol on patent ductus arteriosus closure may be dose dependent: evidence from human and murine studies

    PubMed Central

    El-Khuffash, Afif; Jain, Amish; Corcoran, David; Shah, Prakesh S.; Hooper, Christopher W.; Brown, Naoko; Poole, Stanley D.; Shelton, Elaine L.; Milne, Ginger L.; Reese, Jeff

    2015-01-01

    BACKGROUND We evaluated the clinical effectiveness of variable courses of paracetamol on patent ductus arteriosus (PDA) closure and examined its effect on the in vitro term and preterm murine ductus arteriosus (DA). METHODS Neonates received one of the following three paracetamol regimens: short course of oral paracetamol (SCOP), long course of oral paracetamol (LCOP), and intravenous paracetamol (IVP) for 2–6 d. Pressure myography was used to examine changes in vasomotor tone of the preterm and term mouse DA in response to paracetamol or indomethacin. Their effect on prostaglandin synthesis by DA explants was measured by mass spectroscopy. RESULTS Twenty-one preterm infants were included. No changes in PDA hemodynamics were seen in SCOP infants (n = 5). The PDA became less significant and eventually closed in six LCOP infants (n = 7). PDA closure was achieved in eight IVP infants (n = 9). On pressure myograph, paracetamol induced a concentration-dependent constriction of the term mouse DA, up to 30% of baseline (P < 0.01), but required >1 μmol/l. Indomethacin induced greater DA constriction and suppression of prostaglandin synthesis (P < 0.05). CONCLUSION The clinical efficacy of paracetamol on PDA closure may depend on the duration of treatment and the mode of administration. Paracetamol is less potent than indomethacin for constriction of the mouse DA in vitro. PMID:24941212

  11. Comparative Efficacies of Tedizolid Phosphate, Linezolid, and Vancomycin in a Murine Model of Subcutaneous Catheter-Related Biofilm Infection Due to Methicillin-Susceptible and -Resistant Staphylococcus aureus.

    PubMed

    Bayer, Arnold S; Abdelhady, Wessam; Li, Liang; Gonzales, Rachelle; Xiong, Yan Q

    2016-08-01

    Tedizolid, a novel oxazolidinone, exhibits bacteriostatic activity through inhibition of protein synthesis. The efficacies of tedizolid, linezolid, and vancomycin were compared in a murine catheter-related biofilm infection caused by methicillin-susceptible and -resistant Staphylococcus aureus (MSSA and MRSA, respectively) strains engineered for bioluminescence. We observed significantly improved efficacy in terms of decreased S. aureus densities and bioluminescent signals in the tedizolid-treated group versus the linezolid- and vancomycin-treated groups in the model of infection caused by the MSSA and MRSA strains. PMID:27297485

  12. Open Label, Phase II Study to Evaluate Efficacy and Safety of Oral Nilotinib in Philadelphia Positive (Ph+) Chronic Myelogenous Leukemia (CML) Pediatric Patients.

    ClinicalTrials.gov

    2016-08-05

    Leukemia; Leukemia,Pediatric; Leukemia, Myleiod; Leukemia, Mylegenous, Chronic; Leukemia, Mylegenous, Accelerated; BCR-ABL Positive; Myeloproliferative Disorder; Bone Marrow Disease; Hematologic Diseases; Neoplastic Processes; Imatinib; Dasatinib; Enzyme Inhibitor; Protein Kinase Inhibitor

  13. Evaluation of infectivity and reverse transcriptase real-time polymerase chain reaction assays for detection of xenotropic murine leukemia virus used in virus clearance validation.

    PubMed

    Anwaruzzaman, Mohammad; Wang, Wensheng; Wang, Eunice; Erfe, Lolita; Lee, Janice; Liu, Shengjiang

    2015-07-01

    Infectivity and reverse transcriptase quantitative real-time polymerase chain reaction (qRT-PCR) assays have been optimized and validated for xenotropic murine leukemia virus (X-MuLV) detection. We have evaluated the assays systematically with regard to specificity, linearity, lower limit of detection (LLOD), lower limit of quantification (LLOQ), and precision. Both assays are specific for X-MuLV detection, with a linear detection range of 0.6-5.6 log(10) TCID(50)/mL for the infectivity assay, and 1.4-6.5 log(10) particles/mL for the qRT-PCR assay. The LLOD and LLOQ of the infectivity and the qRT-PCR assays are determined as 0.5 and 1.0 log(10)/mL, and 1.4 and 2.2 log(10)/mL. The inter-assay repeatability of qRT-PCR assay (4.2% coefficient of variation [CV]) is higher than the infectivity assay (7.9% CV). We have shown that both assays are closely correlated (r = 0.85, P < 0.05, n = 22). The particle/infectivity ratio is determined as 66. Both assays were applied to evaluate virus removal using virus clearance samples of chromatographic and filtration processes. Here, we have demonstrated that the qRT-PCR assay is much faster in testing and is approximately 8-fold more sensitive than the infectivity assay. Therefore, the qRT-PCR assay can replace the infectivity assay in many cases, but both assays are complementary in elucidating the mechanism of virus inactivation and removal in virus clearance validation. PMID:25997567

  14. The neurovirulent determinants of ts1, a paralytogenic mutant of Moloney murine leukemia virus TB, are localized in at least two functionally distinct regions of the genome.

    PubMed Central

    Yuen, P H; Tzeng, E; Knupp, C; Wong, P K

    1986-01-01

    To better understand the molecular mechanism involved in retrovirus ts1-induced paralytic disease in mice, we constructed a panel of recombinant viruses between ts1 and the wild-type viruses Moloney murine leukemia virus (MoMuLV) and MoMuLV-TB, a strain of MoMuLV. These recombinant viruses were constructed in an attempt to identify the sequence(s) in the genome of ts1 which contains the critical mutation(s) responsible for the neurovirulence of ts1. Two functionally distinct sequences in the genome of ts1 were found to be responsible for its paralytogenic ability. One of these sequences, the 0.77-kilobase-pair XbaI-BamHI (nucleotides 5765 to 6537) fragment which encodes the 5' half of gp70 and 11 base pairs upstream of the env gene coding sequence, determines the inability of ts1 to process Pr80env. The other sequence, the 2.30-kilobase-pair BamHI-PstI (nucleotides 538 to 8264 and 1 to 567) fragment, which comprises nearly two-thirds of the env gene, the long terminal repeat, and the 5' noncoding sequence, determines the enhanced neurotropism of ts1. Replacement of any one of these two regions with the homologous region from either one of the two wild-type viruses resulted in recombinant viruses which either totally failed to induce paralysis or induced a greatly attenuated form of paresis in some of the infected mice. Images PMID:3712556

  15. Susceptibility of muridae cell lines to ecotropic murine leukemia virus and the cationic amino acid transporter 1 viral receptor sequences: implications for evolution of the viral receptor.

    PubMed

    Kakoki, Katsura; Shinohara, Akio; Izumida, Mai; Koizumi, Yosuke; Honda, Eri; Kato, Goro; Igawa, Tsukasa; Sakai, Hideki; Hayashi, Hideki; Matsuyama, Toshifumi; Morita, Tetsuo; Koshimoto, Chihiro; Kubo, Yoshinao

    2014-06-01

    Ecotropic murine leukemia viruses (Eco-MLVs) infect mouse and rat, but not other mammalian cells, and gain access for infection through binding the cationic amino acid transporter 1 (CAT1). Glycosylation of the rat and hamster CAT1s inhibits Eco-MLV infection, and treatment of rat and hamster cells with a glycosylation inhibitor, tunicamycin, enhances Eco-MLV infection. Although the mouse CAT1 is also glycosylated, it does not inhibit Eco-MLV infection. Comparison of amino acid sequences between the rat and mouse CAT1s shows amino acid insertions in the rat protein near the Eco-MLV-binding motif. In addition to the insertion present in the rat CAT1, the hamster CAT1 has additional amino acid insertions. In contrast, tunicamycin treatment of mink and human cells does not elevate the infection, because their CAT1s do not have the Eco-MLV-binding motif. To define the evolutionary pathway of the Eco-MLV receptor, we analyzed CAT1 sequences and susceptibility to Eco-MLV infection of other several murinae animals, including the southern vole (Microtus rossiaemeridionalis), large Japanese field mouse (Apodemus speciosus), and Eurasian harvest mouse (Micromys minutus). Eco-MLV infection was enhanced by tunicamycin in these cells, and their CAT1 sequences have the insertions like the hamster CAT1. Phylogenetic analysis of mammalian CAT1s suggested that the ancestral CAT1 does not have the Eco-MLV-binding motif, like the human CAT1, and the mouse CAT1 is thought to be generated by the amino acid deletions in the third extracellular loop of CAT1. PMID:24469466

  16. A deletion mutation in the 5' part of the pol gene of Moloney murine leukemia virus blocks proteolytic processing of the gag and pol polyproteins.

    PubMed Central

    Crawford, S; Goff, S P

    1985-01-01

    Deletion mutations in the 5' part of the pol gene of Moloney murine leukemia virus were generated by restriction enzyme site-directed mutagenesis of cloned proviral DNA. DNA sequence analysis indicated that one such deletion was localized entirely within the 5' part of the pol gene, did not affect the region encoding reverse transcriptase, and preserved the translational reading frame downstream of the mutation. The major viral precursor polyproteins (Pr65gag, Pr200gag-pol, and gPr80env) were synthesized at wild-type levels in cell lines carrying the mutant genome. These cell lines assembled and released wild-type levels of virion particles into the medium. Cleavage of both Pr65gag and Pr200gag-pol precursors to the mature proteins was completely blocked in the mutant virions. Surprisingly, these virions contained high levels of active reverse transcriptase; examination of the endogenous reverse transcription products synthesized by the mutant virions revealed normal amounts of minus-strand strong-stop DNA, indicating that the RNA genome was packaged and that reverse transcription in detergent-permeabilized virions was not significantly impaired. Processing of gPr80env to gP70env and P15E was not affected by the mutation, but cleavage of P15E to P12E was not observed. The mutant particles were poorly infectious; analysis indicated that infection was blocked at an early stage. The data are consistent with the idea that the 5' part of the pol gene encodes a protease directly responsible for processing Pr65gag, and possibly Pr200gag-pol, to the structural virion proteins. It appears that cleavage of the gag gene product is not required for budding and release of virions and that complete processing of the pol gene product to the mature form of reverse transcriptase is not required for its functional activation. Images PMID:3882995

  17. The Homopolyadenylate and Adjacent Nucleotides at the 3′-Terminus of 30-40S RNA Subunits in the Genome of Murine Sarcoma-Leukemia Virus

    PubMed Central

    Rho, Hyune Mo; Green, Maurice

    1974-01-01

    Adenosine is the major 3′OH-terminal nucleoside of the 60-70S RNA genome of the murine sarcoma-leukemia virus, its 30-40S RNA subunits, and the poly(A) segments derived by RNase treatment of both RNA species, as determined by periodate oxidation-[3H]-borohydride reduction. The binding 30-40S RNA to oligo(dT)-cellulose suggests that most viral RNA subunits contain poly(A). The molecular weight of poly(A) derived from viral RNA by digestion with RNase and purified by affinity chromatography is 64,000-68,000, as determined by gel electrophoresis. From the size of poly(A) and the poly(A) content of viral RNA (1.6%), it is estimated that there is about one poly(A) segment for each viral 30-40S RNA subunit. The results of 3′-termini labeling with [3H]borohydride, in vivo labeling with [3H]adenosine, and base composition of [32P]poly(A) indicate that a homopoly(A) segment is located at the 3′-end of a 30-40S RNA subunit. The homogeneous poly(A) segments isolated from RNase T1 digests of 60-70S [32P]RNA consist of one cytidylate, one uridylate, and about 190 adenylate residues, while those isolated from RNase A digests consist exclusively of adenylate residues. These results indicate that -G(C,U)A190AOH is the 3′-terminal nucleotide sequence of the viral 30-40S RNA subunits. PMID:4366765

  18. Mouse Siglec-1 Mediates trans-Infection of Surface-bound Murine Leukemia Virus in a Sialic Acid N-Acyl Side Chain-dependent Manner.

    PubMed

    Erikson, Elina; Wratil, Paul R; Frank, Martin; Ambiel, Ina; Pahnke, Katharina; Pino, Maria; Azadi, Parastoo; Izquierdo-Useros, Nuria; Martinez-Picado, Javier; Meier, Chris; Schnaar, Ronald L; Crocker, Paul R; Reutter, Werner; Keppler, Oliver T

    2015-11-01

    Siglec-1 (sialoadhesin, CD169) is a surface receptor on human cells that mediates trans-enhancement of HIV-1 infection through recognition of sialic acid moieties in virus membrane gangliosides. Here, we demonstrate that mouse Siglec-1, expressed on the surface of primary macrophages in an interferon-α-responsive manner, captures murine leukemia virus (MLV) particles and mediates their transfer to proliferating lymphocytes. The MLV infection of primary B-cells was markedly more efficient than that of primary T-cells. The major structural protein of MLV particles, Gag, frequently co-localized with Siglec-1, and trans-infection, primarily of surface-bound MLV particles, efficiently occurred. To explore the role of sialic acid for MLV trans-infection at a submolecular level, we analyzed the potential of six sialic acid precursor analogs to modulate the sialylated ganglioside-dependent interaction of MLV particles with Siglec-1. Biosynthetically engineered sialic acids were detected in both the glycolipid and glycoprotein fractions of MLV producer cells. MLV released from cells carrying N-acyl-modified sialic acids displayed strikingly different capacities for Siglec-1-mediated capture and trans-infection; N-butanoyl, N-isobutanoyl, N-glycolyl, or N-pentanoyl side chain modifications resulted in up to 92 and 80% reduction of virus particle capture and trans-infection, respectively, whereas N-propanoyl or N-cyclopropylcarbamyl side chains had no effect. In agreement with these functional analyses, molecular modeling indicated reduced binding affinities for non-functional N-acyl modifications. Thus, Siglec-1 is a key receptor for macrophage/lymphocyte trans-infection of surface-bound virions, and the N-acyl side chain of sialic acid is a critical determinant for the Siglec-1/MLV interaction. PMID:26370074

  19. Adoptive transfer of T cells transduced with a chimeric antigen receptor to treat relapsed or refractory acute leukemia: efficacy and feasibility of immunotherapy approaches.

    PubMed

    Ding, Guoliang; Chen, Hu

    2016-07-01

    Treatment outcomes of acute leukemia (AL) have not improved over the past several decades and relapse rates remain high despite the availability of aggressive therapies. Conventional relapsed leukemia treatment includes second allogeneic hematopoietic stem cell transplantation (allo-HSCT) and donor lymphocyte infusion (DLI), which in most cases mediate, at best, a modest graft-versus-leukemia effect, although their clinical efficacy is still limited. Although allo-HSCT following myeloablative conditioning is a curative treatment option for younger patients with acute myeloid leukemia (AML) in a first complete remission (CR), allo-HSCT as a clinical treatment is usually limited because of treatment-related toxicity. The overall DLI remission rate is only 15%-42% and 2-year overall survival (OS) is approximately 15%-20%, with a high (40%-60%) incidence of DLI-related graft-versus-host disease (GVHD). Therefore, development of new, targeted treatment strategies for relapsed and refractory AL patients is ongoing. Adoptive transfer of T cells with genetically engineered chimeric antigen receptors (CARs) is an encouraging approach for treating hematological malignancies. These T cells are capable of selectively recognizing tumor-associated antigens and may overcome many limitations of conventional therapies, inducing remission in patients with chemotherapy-refractory or relapsed AL. In this review, we aimed to highlight the current understanding of this promising treatment modality, discussing its adverse effects and efficacy. PMID:27142351

  20. Efficacy of irreversible electroporation in human pancreatic adenocarcinoma: advanced murine model.

    PubMed

    Philips, Prejesh; Li, Yan; Li, Suping; St Hill, Charles R; Martin, Robert Cg

    2015-01-01

    Irreversible electroporation (IRE) is a promising cell membrane ablative modality for pancreatic cancer. There have been recent concerns regarding local recurrence and the potential use of IRE as a debulking (partial ablation) modality. We hypothesize that incomplete ablation leads to early recurrence and a more aggressive biology. We created the first ever heterotopic murine model by inoculating BALB/c nude mice in the hindlimb with a subcutaneous injection of Panc-1 cells, an immortalized human pancreatic adenocarcinoma cell line. Tumors were allowed to grow from 0.75 to 1.5 cm and then treated with the goal of complete ablation or partial ablation using standard IRE settings. Animals were recovered and survived for 2 days (n = 6), 7 (n = 6), 14 (n = 6), 21 (n = 6), 30 (n = 8), and 60 (n = 8) days. All 40 animals/tumors underwent successful IRE under general anesthesia with muscle paralysis. The mean tumor volume of the animals undergoing ablation was 1,447.6 mm(3) ± 884). Histologically, in the 14-, 21-, 30-, and 60-day survival groups the entire tumor was nonviable, with a persistent tumor nodule completely replaced fibrosis. In the group treated with partial ablation, incomplete electroporation/recurrences (N = 10 animals) were seen, of which 66% had confluent tumors and this was a significant predictor of recurrence (P < 0.001). Recurrent tumors were also significantly larger (mean 4,578 mm(3) ± SD 877 versus completed electroporated tumors 925.8 ± 277, P < 0.001). Recurrent tumors had a steeper growth curve (slope = 0.73) compared with primary tumors (0.60, P = 0.02). Recurrent tumors also had a significantly higher percentage of EpCAM expression, suggestive of stem cell activation. Tumors that recur after incomplete electroporation demonstrate a biologically aggressive tumor that could be more resistant to standard of care chemotherapy. Clinical correlation of this data is limited, but should be considered when IRE of pancreatic cancer is being

  1. Praziquantel in a Clay Nanoformulation Shows More Bioavailability and Higher Efficacy against Murine Schistosoma mansoni Infection

    PubMed Central

    Mohamed, Wael S.; Nasr, Hanaa E.; El-Lakkany, Naglaa M.; Seif el-Din, Sayed H.; Botros, Sanaa S.

    2015-01-01

    Consideration of existing compounds always simplifies and shortens the long and difficult process of discovering new drugs specifically for diseases of developing countries, an approach that may add to the significant potential cost savings. This study focused on improving the biological characteristics of the already-existing antischistosomal praziquantel (PZQ) by incorporating it into montmorillonite (MMT) clay as a delivery carrier to overcome its known bioavailability drawbacks. The oral bioavailability of a PZQ-MMT clay nanoformulation and its in vivo efficacy against Schistosoma mansoni were investigated. The PZQ-MMT clay nanoformulation provided a preparation with a controlled release rate, a decrease in crystallinity, and an appreciable reduction in particle size. Uninfected and infected mice treated with PZQ-MMT clay showed 3.61- and 1.96-fold and 2.16- and 1.94-fold increases, respectively, in area under the concentration-time curve from 0 to 8 h (AUC0–8) and maximum concentration of drug in serum (Cmax), with a decrease in elimination rate constant (kel) by 2.84- and 1.35-fold and increases in the absorption rate constant (ka) and half-life (t1/2e) by 2.11- and 1.51-fold and 2.86- and 1.34-fold, respectively, versus the corresponding conventional PZQ-treated groups. This improved bioavailability has been expressed in higher efficacy of the drug, where the dose necessary to kill 50% of the worms was reduced by >3-fold (PZQ 50% effective dose [ED50] was 20.25 mg/kg of body weight for PZQ-MMT clay compared to 74.07 mg/kg for conventional PZQ), with significant reduction in total tissue egg load and increase in total immature, mature, and dead eggs in most of the drug-treated groups. This formulation showed better bioavailability, enhanced antischistosomal efficacy, and a safer profile despite the longer period of residence in the systemic circulation. Although the conventional drug's toxicity was not examined, animal mortality rates were not different

  2. Efficacy of Lychnopholide Polymeric Nanocapsules after Oral and Intravenous Administration in Murine Experimental Chagas Disease.

    PubMed

    de Mello, Carlos Geraldo Campos; Branquinho, Renata Tupinambá; Oliveira, Maykon Tavares; Milagre, Matheus Marques; Saúde-Guimarães, Dênia Antunes; Mosqueira, Vanessa Carla Furtado; Lana, Marta de

    2016-09-01

    The etiological treatment of Chagas disease remains neglected. The compounds available show several limitations, mainly during the chronic phase. Lychnopholide encapsulated in polymeric nanocapsules (LYC-NC) was efficacious in mice infected with Trypanosoma cruzi and treated by intravenous administration during the acute phase (AP). As the oral route is preferred for treatment of chronic infections, such as Chagas disease, this study evaluated the use of oral LYC-NC in the AP and also compared it with LYC-NC administered to mice by the oral and intravenous routes during the chronic phase (CP). The therapeutic efficacy was evaluated by fresh blood examination, hemoculture, PCR, and enzyme-linked immunosorbent assay (ELISA). The cure rates in the AP and CP were 62.5% and 55.6%, respectively, upon oral administration of LYC-poly(d,l-lactide)-polyethylene glycol nanocapsules (LYC-PLA-PEG-NC) and 57.0% and 30.0%, respectively, with LYC-poly-ε-caprolactone nanocapsules (LYC-PCL-NC). These cure rates were significantly higher than that of free LYC, which did not cure any animals. LYC-NC formulations administered orally during the AP showed cure rates similar to that of benznidazole, but only LYC-NC cured mice in the CP. Similar results were achieved with intravenous treatment during the CP. The higher cure rates obtained with LYC loaded in PLA-PEG-NC may be due to the smaller particle size of these NC and the presence of PEG, which influence tissue diffusion and the controlled release of LYC. Furthermore, PLA-PEG-NC may improve the stability of the drug in the gastrointestinal tract. This work is the first report of cure of experimental Chagas disease via oral administration during the CP. These findings represent a new and important perspective for oral treatment of Chagas disease. PMID:27324760

  3. Comparative efficacy of amphotericin B colloidal dispersion and amphotericin B deoxycholate suspension in treatment of murine coccidioidomycosis.

    PubMed Central

    Clemons, K V; Stevens, D A

    1991-01-01

    The efficacy of a novel sterol-complexed preparation of amphotericin B, amphotericin B colloidal dispersion, was compared with that of deoxycholate-complexed amphotericin B in an acute murine model of systemic coccidioidomycosis. Mice (CD-1, female) were infected intravenously with 180 or 200 arthroconidia of Coccidioides immitis, and intravenous therapy was begun 3 days later. Six doses in various regimens of either preparation were given over 14 days, and deaths were tallied for an additional 35 days. All regimens that were not acutely lethal prolonged the survival of mice over that of controls (P less than 0.001). Quantitative determination of residual burdens of C. immitis in the spleen, liver, and lungs of survivors revealed that the colloidal dispersion was not as effective as the deoxycholate suspension on a milligram-per-kilogram basis. Deoxycholate suspension at 1.3 mg/kg cleared the organs in all mice, whereas colloidal dispersion at 5.0 mg/kg was the lowest dose that cleared organisms from all animals. Lower doses cleared organisms from fewer animals or cleared only selected organs. Deoxycholate suspension was more efficacious than colloidal dispersion in clearing C. immitis from the liver or lungs (P less than 0.05 to 0.01, dose and organ dependent) at identical doses. No overt toxicity was observed in mice treated with colloidal dispersion at 10 mg/kg. In contrast, deoxycholate suspension at 2.0 mg/kg was acutely toxic; 50% of the treated mice died after treatment. The two complexes were not equivalent on a milligram-per-kilogram basis; the deoxycholate suspension was three to four times more efficacious and also greater than 5- to greater than or equal to 8-fold more toxic. Thus, the therapeutic index of the colloidal dispersion complex is greater than that of the deoxycholate complex. The amount of amphotericin B per dose could also be increased when given as a colloidal dispersion to an optimally level. Amphotericin B colloidal dispersion shows

  4. Characterization of a Murine Pressure Ulcer Model to Assess Efficacy of Adipose-derived Stromal Cells

    PubMed Central

    Strong, Amy L.; Bowles, Annie C.; MacCrimmon, Connor P.; Lee, Stephen J.; Frazier, Trivia P.; Katz, Adam J.; Gawronska-Kozak, Barbara; Bunnell, Bruce A.

    2015-01-01

    Background: As the world’s population lives longer, the number of individuals at risk for pressure ulcers will increase considerably in the coming decades. In developed countries, up to 18% of nursing home residents suffer from pressure ulcers and the resulting hospital costs can account for up to 4% of a nation’s health care budget. Although full-thickness surgical skin wounds have been used as a model, preclinical rodent studies have demonstrated that repeated cycles of ischemia and reperfusion created by exposure to magnets most closely mimic the human pressure ulcer condition. Methods: This study uses in vivo and in vitro quantitative parameters to characterize the temporal kinetics and histology of pressure ulcers in young, female C57BL/6 mice exposed to 2 or 3 ischemia-reperfusion cycles. This pressure ulcer model was validated further in studies examining the efficacy of adipose-derived stromal/stem cell administration. Results: Optimal results were obtained with the 2-cycle model based on the wound size, histology, and gene expression profile of representative angiogenic and reparative messenger RNAs. When treated with adipose-derived stromal/stem cells, pressure ulcer wounds displayed a dose-dependent and significant acceleration in wound closure rates and improved tissue histology. Conclusion: These findings document the utility of this simplified preclinical model for the evaluation of novel tissue engineering and medical approaches to treat pressure ulcers in humans. PMID:25878945

  5. Targeted nanoparticle delivery overcomes off-target immunostimulatory effects of oligonucleotides and improves therapeutic efficacy in chronic lymphocytic leukemia

    PubMed Central

    Yu, Bo; Mao, Yicheng; Bai, Li-Yuan; Herman, Sarah E. M.; Wang, Xinmei; Ramanunni, Asha; Jin, Yan; Mo, Xiaokui; Cheney, Carolyn; Chan, Kenneth K.; Jarjoura, David; Marcucci, Guido; Lee, Robert J.; Byrd, John C.

    2013-01-01

    Several RNA-targeted therapeutics, including antisense oligonucleotides (ONs), small interfering RNAs, and miRNAs, constitute immunostimulatory CpG motifs as an integral part of their design. The limited success with free antisense ONs in hematologic malignancies in recent clinical trials has been attributed to the CpG motif–mediated, TLR-induced prosurvival effects and inefficient target modulation in desired cells. In an attempt to diminish their off-target prosurvival and proinflammatory effects and specific delivery, as a proof of principle, in the present study, we developed an Ab-targeted liposomal delivery strategy using a clinically relevant CD20 Ab (rituximab)–conjugated lipopolyplex nanoparticle (RIT-INP)– and Bcl-2–targeted antisense G3139 as archetypical antisense therapeutics. The adverse immunostimulatory responses were abrogated by selective B cell–targeted delivery and early endosomal compartmentalization of G3139-encapsulated RIT-INPs, resulting in reduced NF-κB activation, robust Bcl-2 down-regulation, and enhanced sensitivity to fludarabine-induced cytotoxicity. Furthermore, significant in vivo therapeutic efficacy was noted after RIT-INP–G3139 administration in a disseminated xenograft leukemia model. The results of the present study demonstrate that CD20-targeted delivery overcomes the immunostimulatory properties of CpG-containing ON therapeutics and improves efficient gene silencing and in vivo therapeutic efficacy for B-cell malignancies. The broader implications of similar approaches in overcoming immunostimulatory properties of RNA-directed therapeutics in hematologic malignancies are also discussed. PMID:23165478

  6. Low efficacy and high mortality associated with clofarabine treatment of relapsed/refractory acute myeloid leukemia and myelodysplastic syndromes.

    PubMed

    Roberts, Daniel A; Wadleigh, Martha; McDonnell, Anne M; DeAngelo, Daniel J; Stone, Richard M; Steensma, David P

    2015-02-01

    Clofarabine, a second-generation nucleoside analog, has clinical activity in relapsed or refractory acute myelogenous leukemia (AML) and higher-risk myelodysplastic syndromes (MDS). However, there are few data evaluating performance of clofarabine in populations of patients not enrolled in clinical trials. We reviewed outcomes for 84 patients treated with clofarabine for relapsed or refractory AML or MDS, either with clofarabine as monotherapy (n=19) or in combination with cytarabine (n=65). Using International Working Group (IWG) response criteria, the overall response rate (ORR) of all treated patients was 21%, with a complete response rate with either complete or incomplete hematopoietic recovery (CRR=CR+CRi) of 14%. For combination therapy, ORR was 22% with CRR of 18%, and monotherapy patients had an ORR of 21% with CRR of 11%. Although limited by small numbers, subgroup analysis did not reveal variation in response rates when comparing different risk factors. The 30-day mortality was 21% and median survival was 3 months; a subset of 12 patients who were able to go to transplant had an 18-month median survival. Clofarabine's efficacy in a "real-world" setting appears to be less than has been reported in clinical trials, and treatment is associated with a high early mortality rate. PMID:25554239

  7. Efficacy of helper-dependent adenovirus vector-mediated gene therapy in murine glycogen storage disease type Ia.

    PubMed

    Koeberl, Dwight D; Sun, B; Bird, A; Chen, Y T; Oka, K; Chan, L

    2007-07-01

    Genetic deficiency of glucose-6-phosphatase (G6Pase) underlies glycogen storage disease type Ia (GSD-Ia, also known as von Gierke disease; MIM 232200), an autosomal recessive disorder of metabolism associated with life-threatening hypoglycemia and growth retardation. We tested whether helper-dependent adenovirus (HDAd)-mediated hepatic delivery of G6Pase would lead to prolonged survival and sustained correction of the metabolic abnormalities in G6Pase knockout (KO) mice, a model for a severe form of GSD-Ia. An HDAd vector encoding G6Pase was administered intravenously (2 or 5 x 10(12)vector particles/kg) to 2-week-old (w.o.) G6Pase-KO mice. Following HDAd vector administration survival was prolonged to a median of 7 months, in contrast to untreated affected mice that did not survive past 3 weeks of age. G6Pase levels increased more than tenfold between 3 days and 28 weeks after HDAd injection (P < 0.03). The weights of untreated 2 w.o. G6Pase-KO mice were approximately half those of their unaffected littermates, and treatment stimulated their growth to the size of wild-type mice. Severe hypoglycemia and hypercholesterolemia, which are hallmarks of GSD-Ia both in humans and in mice, were also restored to normalcy by the treatment. Glycogen accumulation in the liver was markedly reduced. The efficacy of HDAd-G6Pase treatment in reversing the physiological and biochemical abnormalities associated with GSD-Ia in affected G6Pase-KO mice justifies further preclinical evaluation in murine and canine models of GSD-Ia. PMID:17505475

  8. Efficacy of Rifampin Plus Clofazimine in a Murine Model of Mycobacterium ulcerans Disease

    PubMed Central

    Converse, Paul J.; Tyagi, Sandeep; Xing, Yalan; Li, Si-Yang; Kishi, Yoshito; Adamson, John; Nuermberger, Eric L.; Grosset, Jacques H.

    2015-01-01

    Treatment of Buruli ulcer, or Mycobacterium ulcerans disease, has shifted from surgical excision and skin grafting to antibiotic therapy usually with 8 weeks of daily rifampin (RIF) and streptomycin (STR). Although the results have been highly favorable, administration of STR requires intramuscular injection and carries the risk of side effects, such as hearing loss. Therefore, an all-oral, potentially less toxic, treatment regimen has been sought and encouraged by the World Health Organization. A combination of RIF plus clarithromycin (CLR) has been successful in patients first administered RIF+STR for 2 or 4 weeks. Based on evidence of efficacy of clofazimine (CFZ) in humans and mice with tuberculosis, we hypothesized that the combination of RIF+CFZ would be effective against M. ulcerans in the mouse footpad model of M. ulcerans disease because CFZ has similar MIC against M. tuberculosis and M. ulcerans. For comparison, mice were also treated with the gold standard of RIF+STR, the proposed RIF+CLR alternative regimen, or CFZ alone. Treatment was initiated after development of footpad swelling, when the bacterial burden was 4.64±0.14log10 CFU. At week 2 of treatment, the CFU counts had increased in untreated mice, remained essentially unchanged in mice treated with CFZ alone, decreased modestly with either RIF+CLR or RIF+CFZ, and decreased substantially with RIF+STR. At week 4, on the basis of footpad CFU counts, the combination regimens were ranked as follows: RIF+STR>RIF+CLR>RIF+CFZ. At weeks 6 and 8, none of the mice treated with these regimens had detectable CFU. Footpad swelling declined comparably with all of the combination regimens, as did the levels of detectable mycolactone A/B. In mice treated for only 6 weeks and followed up for 24 weeks, there were no relapses in RIF+STR treated mice, one (5%) relapse in RIF+CFZ-treated mice, but >50% in RIF+CLR treated mice. On the basis of these results, RIF+CFZ has potential as a continuation phase regimen for

  9. Efficacy of Rifampin Plus Clofazimine in a Murine Model of Mycobacterium ulcerans Disease.

    PubMed

    Converse, Paul J; Tyagi, Sandeep; Xing, Yalan; Li, Si-Yang; Kishi, Yoshito; Adamson, John; Nuermberger, Eric L; Grosset, Jacques H

    2015-01-01

    Treatment of Buruli ulcer, or Mycobacterium ulcerans disease, has shifted from surgical excision and skin grafting to antibiotic therapy usually with 8 weeks of daily rifampin (RIF) and streptomycin (STR). Although the results have been highly favorable, administration of STR requires intramuscular injection and carries the risk of side effects, such as hearing loss. Therefore, an all-oral, potentially less toxic, treatment regimen has been sought and encouraged by the World Health Organization. A combination of RIF plus clarithromycin (CLR) has been successful in patients first administered RIF+STR for 2 or 4 weeks. Based on evidence of efficacy of clofazimine (CFZ) in humans and mice with tuberculosis, we hypothesized that the combination of RIF+CFZ would be effective against M. ulcerans in the mouse footpad model of M. ulcerans disease because CFZ has similar MIC against M. tuberculosis and M. ulcerans. For comparison, mice were also treated with the gold standard of RIF+STR, the proposed RIF+CLR alternative regimen, or CFZ alone. Treatment was initiated after development of footpad swelling, when the bacterial burden was 4.64±0.14log10 CFU. At week 2 of treatment, the CFU counts had increased in untreated mice, remained essentially unchanged in mice treated with CFZ alone, decreased modestly with either RIF+CLR or RIF+CFZ, and decreased substantially with RIF+STR. At week 4, on the basis of footpad CFU counts, the combination regimens were ranked as follows: RIF+STR>RIF+CLR>RIF+CFZ. At weeks 6 and 8, none of the mice treated with these regimens had detectable CFU. Footpad swelling declined comparably with all of the combination regimens, as did the levels of detectable mycolactone A/B. In mice treated for only 6 weeks and followed up for 24 weeks, there were no relapses in RIF+STR treated mice, one (5%) relapse in RIF+CFZ-treated mice, but >50% in RIF+CLR treated mice. On the basis of these results, RIF+CFZ has potential as a continuation phase regimen for

  10. Efficacy and Safety of Cladribine: Subcutaneous versus Intravenous Administration in Hairy Cell Leukemia Patients

    PubMed Central

    Khorshid, Ola; Namour, Alfred Elias; El-Gammal, Mosaad M; Mahmoud, Tarek Yakout; Fortpied, Catherine; Abdel-Malek, Raafat; Ramadan, Safaa

    2015-01-01

    Cladribine induces durable complete remission (CR) in approximately 85% of hairy cell leukemia (HCL) patients. In Egypt, cladribine is mainly used as IV continuous infusion at a dose of 0.1 mg/kg/day for 7 days and as SC bolus injection at a dose of 0.14 mg/kg/day for 5 days. We aimed to compare the outcome and toxicity between these two regimens. We retrospectively collected data from HCL patients treated at the National Cancer Institute and its affiliated center, Nasser Institute, Cairo, Egypt. Forty-nine patients were identified, 18 treated with the IV regimen (IV group) and 31 with the SC regimen (SC group). Forty-one patients were newly diagnosed. Patient characteristics were balanced across the two groups. The CR rates in the IV and the SC group were 94% and 97%, respectively. The main complications in the IV group and the SC were neutropenia G3–4 (67% vs. 87%), mucositis mainly G1–2 (67% vs 32%) and infections (mainly viral, 78% vs 34%). In the IV group, five patients died, three of progression and infection, one of unknown cause and one of late heart failure. In the SC group, one patient died of disease progression and one of second cancer. After 33.5 months, median follow-up, the 3-year event free survival was 60% and 96%, respectively (p=0.104). The 3-year overall survival was 81% and 100%, respectively (p=0.277). In conclusion, SC cladribine is an excellent alternative to the IV regimen for the treatment of HCL. PMID:26543527

  11. Hydrodynamic diameters of murine mammary, Rous sarcoma, and feline leukemia RNA tumor viruses: studies by laser beat frequency light-scattering spectroscopy and electron microscopy.

    PubMed Central

    Salmeen, I; Rimai, L; Luftig, R B; Libes, L; Retzel, E; Rich, M; McCormick, J J

    1976-01-01

    We have studied purified preparations of murine mammary tumor virus (MuMTV), Rous sarcoma virus (RSV; Prague strain), and feline leukemia virus (FeLV) by laser beat frequency light-scattering spectroscopy, ultra-centrifugation, and electron microscopy. The laser beat frequency light-scattering spectroscopy measurements yield the light-scattering intensity, weighted diffusion coefficients. The corresponding average hydrodynamic diameters, as calculated from the diffusion coefficients by the Stokes-Einstein equation for MuMTV, RSV, and FeLV, respectively, are: 144 +/- 6 nm, 147 +/- 7 nm, and 168 +/- 6 nm. Portions of the purified RSV and MuMTV preparations, from which light-scattering samples were obtained, and portions of the actual FeLV light-scattering samples were examined by negatively stained, catalase crystal-calibrated electron microscopy. The light-scattering intensity weighted averages of the electron micrograph size distributions were calculated by weighing each size by its theoretical relative scattering intensity, as obtained from published tables computed according to the Mie scattering theory. These averages and the experimentally observed hydrodynamic diameters agreed to within +/- 5%, which is the combined experimental error in the electron microscopic and light-scattering techniques. We conclude that the size distributions of singlet particles observed in the electron micrographs are statistically true representations of the sedimentation-purified solution size distributions. The sedimentation coefficients (S20, w) for MuMTV, RSV, and FeLV, respectively, are: 595 +/- 29S, 689 +/- 35S, and 880 +/- 44S. Virus partial specific volumes were taken as the reciprocals of the buoyant densities, determined in sucrose density gradients. The Svedberg equation was used to calculate particle weights from the measured diffusion and sedimentation coefficients. The particle weights for MuMTV, RSV, and FeLV, respectively, are: (3.17 +/- 0.32) x 10(8), (4.17 +/- 0

  12. Negative regulatory element associated with potentially functional promoter and enhancer elements in the long terminal repeats of endogenous murine leukemia virus-related proviral sequences.

    PubMed Central

    Ch'ang, L Y; Yang, W K; Myer, F E; Yang, D M

    1989-01-01

    Three series of recombinant DNA clones were constructed, with the bacterial chloramphenicol acetyltransferase (CAT) gene as a quantitative indicator, to examine the activities of promoter and enhancer sequence elements in the 5' long terminal repeat (LTR) of murine leukemia virus (MuLV)-related proviral sequences isolated from the mouse genome. Transient CAT expression was determined in mouse NIH 3T3, human HT1080, and mink CCL64 cultured cells transfected with the LTR-CAT constructs. The 700-base-pair (bp) LTRs of three polytropic MuLV-related proviral clones and the 750-bp LTRs of four modified polytropic proviral clones, in complete structures either with or without the adjacent downstream sequences, all showed very little or negligible activities for CAT expression, while ecotropic MuLV LTRs were highly active. The MuLV-related LTRs were divided into three portions and examined separately. The 3' portion of the MuLV-related LTRs that contains the CCAAC and TATAA boxes was found to be a functional promoter, being about one-half to one-third as active as the corresponding portion of ecotropic MuLV LTRs. A MboI-Bg/II fragment, representing the distinct 190- to 200-bp inserted segment in the middle, was found to be a potential enhancer, especially when examined in combination with the simian virus 40 promoter in CCL64 cells. A PstI-MboI fragment of the 5' portion, which contains the protein-binding motifs of the enhancer segment as well as the upstream LTR sequences, showed moderate enhancer activities in CCL6 cells but was virtually inactive in NIH 3T3 cells and HT1080 cells; addition of this fragment to the ecotropic LTR-CAT constructs depressed CAT expression. Further analyses using chimeric LTR constructs located the presence of a strong negative regulatory element within the region containing the 5' portion of the enhancer and the immediate upstream sequences in the MuLV-related LTRs. Images PMID:2542587

  13. Efficacy of Azacitidine in autoimmune and inflammatory disorders associated with myelodysplastic syndromes and chronic myelomonocytic leukemia.

    PubMed

    Fraison, Jean-Baptiste; Mekinian, Arsène; Grignano, Eric; Kahn, Jean-Emmanuel; Arlet, Jean-Benoit; Decaux, Olivier; Denis, Guillaume; Buchdahl, Anne-Laure; Omouri, Mohamed; Maigne, Gwenola; Aouba, Achille; Leon, Nathalie; Berthier, Sabine; Liozon, Eric; Park, Sophie; Gardin, Claude; Lortholary, Olivier; Rossignol, Julien; Fenaux, Pierre; Fain, Olivier; Braun, Thorsten

    2016-04-01

    This retrospective study describes efficacy of Azacitidine on autoimmune disorders (AID) associated with MDS/CMML in 22 patients. Response of AID to Azacitidine was observed in 19 patients (86%). Reduction or discontinuation of steroids and/or immunosuppressive therapy (IST) was possible in 16 cases (73%). Hematologic response was seen in 55% of the patients. MDS/CMML and AID evolution was concordant in 13 cases (59%): both favorable (n=11), both unfavorable (n=2), but AID improved while MDS/CMML worsened (n=8) and vice versa (n=1). Azacitidine frequently seems effective in controlling steroid-dependent AID associated with MDS/CMML, but prospective studies are necessary to confirm those findings. PMID:26922775

  14. Efficacy and tolerability of treatment with azacitidine for 5 days in elderly patients with acute myeloid leukemia

    PubMed Central

    Sadashiv, Santhosh K; Hilton, Christie; Khan, Cyrus; Rossetti, James M; Benjamin, Heather L; Fazal, Salman; Sahovic, Entezam; Shadduck, Richard K; Lister, John

    2014-01-01

    Acute myeloid leukemia (AML) patients aged ≥60 years tolerate standard induction chemotherapy poorly. Therapy with azacitidine at a dose of 75 mg/m2/day for 7 days appears to be better tolerated, and is approved by the Food and Drug Administration (FDA) for the treatment of elderly AML patients with bone marrow (BM) blast counts of 20–30%. Here, we report the results of a prospective, phase 2, open-label study that evaluated the tolerability and efficacy of a 5-day regimen of single-agent subcutaneous azacitidine 100 mg/m2/day administered every 28 days in 15 elderly patients with newly diagnosed AML, 14 of whom had BM blast counts >30%. The overall response rate was 47%. Complete remission, partial remission, and hematologic improvement were achieved by 20, 13, and 13% of patients, respectively. Median overall survival was 355 days for the entire cohort, and 532 days for responders. Median time to best response was 95 days, and median treatment duration was 198 days (range = 13–724 days). Grade 3–4 hematologic toxicities comprised predominantly febrile neutropenia (40%) and thrombocytopenia (20%). Febrile neutropenia was the most common cause of hospitalization. Nonhematologic toxicities, consisting of injection-site skin reactions and fatigue (Grades 1–2), occurred in 73% (n = 11) of patients. No treatment-related deaths occurred during the study. The dose and schedule of therapy remained constant in all but four patients. The findings of this study suggest that administration of subcutaneous azacitidine 100 mg/m2/day for 5 days every 28 days is a feasible, well-tolerated, and effective alternative to standard induction chemotherapy in elderly patients with AML. PMID:25132519

  15. Murine acute leukemia cell line with megakaryocytic differentiation (MK-8057) induced by whole-body irradiation in C sup 3 H/He mice: Cytological properties and kinetics of its leukemic stem cells

    SciTech Connect

    Hirabayashi, Y.; Inoue, T.; Yoshida, K.; Sasaki, H.; Kubo, S.; Kanisawa, M.; Seki, M. )

    1991-01-01

    Five cases of murine leukemia with megakaryocytic differentiation were observed among the 417 cases of radiation-induced leukemias which developed in 30% of C{sup 3}H/HeMs mice exposed at 8 to 10 weeks to 0.5 to 5 gy total body irradiation. Cells from individual leukemic colonies in the spleen of the irradiated mice, and cells from colonies in methylcellulose (MC) culture in vitro, derived from one of these leukemias, MK-8057, were injected into mice; both types of cells caused the deaths of the recipient mice by inducing the same type of leukemia. MK-8057 can be maintained in Dexter-type liquid culture with a feeder layer of irradiated bone marrow cells. There was a linear reciprocal relationship between the increasing number of MK-8057 cells injected versus the survival of the recipient mice. A reciprocal relationship also was seen between an increasing number of leukemic stem cells, corresponding to the number of MK-8057 cells, and the survival of mice injected with MK-8057. Giant nuclear megakaryocytes developed during the course of colony growth in the spleen as they did in the MC culture. Such megakaryocytes were acetylcholinesterase positive, whereas leukemic cells in the peripheral blood showed no sign of platelet production nor of a positive reaction to acetylcholinesterase. Cells maintained in culture were entirely positive in platelet glycoprotein IIb/IIIa when anti-human antibody was used. The larger cells in a splenic cell suspension derived from a moribund mouse were separated and enriched by velocity sedimentation using centrifugal elutriation (CE), and then subjected to flow cytometry using propidium iodide staining. Cells with up to 32N-DNA content were detected. After separating MK-8057 by counter-flow CE, the larger cell fraction produced more leukemic colonies when injected into irradiated mice than did the small cell fraction.

  16. CHK1 and WEE1 inhibition combine synergistically to enhance therapeutic efficacy in acute myeloid leukemia ex vivo

    PubMed Central

    Chaudhuri, Leena; Vincelette, Nicole D.; Koh, Brian D.; Naylor, Ryan M.; Flatten, Karen S.; Peterson, Kevin L.; McNally, Amanda; Gojo, Ivana; Karp, Judith E.; Mesa, Ruben A; Sproat, Lisa O.; Bogenberger, James M.; Kaufmann, Scott H.; Tibes, Raoul

    2014-01-01

    Novel combinations targeting new molecular vulnerabilities are needed to improve the outcome of patients with acute myeloid leukemia. We recently identified WEE1 kinase as a novel target in leukemias. To identify genes that are synthetically lethal with WEE1 inhibition, we performed a short interfering RNA screen directed against cell cycle and DNA repair genes during concurrent treatment with the WEE1 inhibitor MK1775. CHK1 and ATR, genes encoding two replication checkpoint kinases, were among the genes whose silencing enhanced the effects of WEE1 inhibition most, whereas CDK2 short interfering RNA antagonized MK1775 effects. Building on this observation, we examined the impact of combining MK1775 with selective small molecule inhibitors of CHK1, ATR and cyclin-dependent kinases. The CHK1 inhibitor MK8776 sensitized acute myeloid leukemia cell lines and primary leukemia specimens to MK1775 ex vivo, whereas smaller effects were observed with the MK1775/MK8776 combination in normal myeloid progenitors. The ATR inhibitor VE-821 likewise enhanced the antiproliferative effects of MK1775, whereas the cyclin-dependent kinase inhibitor roscovitine antagonized MK1775. Further studies showed that MK8776 enhanced MK1775-mediated activation of the ATR/CHK1 pathway in acute leukemia cell lines and ex vivo. These results indicate that combined cell cycle checkpoint interference with MK1775/MK8776 warrants further investigation as a potential treatment for acute myeloid leukemia. PMID:24179152

  17. Efficacy of continuous venovenous hemofiltration with chemotherapy in patients with Burkitt lymphoma and leukemia at high risk of tumor lysis syndrome.

    PubMed

    Choi, Kyung A; Lee, Jung Eun; Kim, Yoon-Goo; Kim, Dae Joong; Kim, Kihyun; Ko, Young Hyeh; Oh, Ha Young; Kim, Won Seog; Huh, Wooseong

    2009-07-01

    Tumor lysis syndrome (TLS) is a potentially fatal metabolic complication of chemotherapy for Burkitt lymphoma. It has not been established whether chemotherapy should be delayed in patients with spontaneous TLS, and several studies have shown poor prognoses in this group. This retrospective study evaluated the efficacy and safety of continuous venovenous hemofiltration (CVVH) with prephase chemotherapy using the modified LMB-89 regimen in patients with Burkitt lymphoma and leukemia (BL/L) at a high risk of developing TLS from February 1998 to February 2007. The chemotherapy regimen was followed by the modified LMB-89 protocol. CVVH was applied to all patients before prephase chemotherapy or within 2 h of chemotherapy. The median follow-up was 19.7 months (range 1-97.8). Eight patients had Burkitt lymphoma and three had Burkitt leukemia; their median age was 48 years. The international prognostic indices were >3 for all patients. Seven patients had spontaneous TLS and four patients were at a high risk of TLS. CVVH was continued for 109 h (range 70.5-157.5). No patient had fatal metabolic complications related to TLS. Renal function had recovered fully before induction chemotherapy in all but one patient. The 1-year event-free survival and overall survival rates were both 82%. In conclusion, chemotherapy combined with CVVH might be effective and safe in patients with advanced Burkitt lymphoma and leukemia at a high risk of developing TLS. PMID:19030857

  18. Evaluation of the Efficacies of Amphotericin B, Posaconazole, Voriconazole, and Anidulafungin in a Murine Disseminated Infection by the Emerging Opportunistic Fungus Sarocladium (Acremonium) kiliense

    PubMed Central

    Fernández-Silva, Fabiola; Capilla, Javier; Mayayo, Emilio; Sutton, Deanna A.; Hernández, Pilar

    2013-01-01

    We evaluated and compared the efficacies of different antifungal drugs against Sarocladium kiliense (formerly Acremonium kiliense), a clinically relevant opportunistic fungus, in a murine model of systemic infection. Three clinical strains of this fungus were tested, and the therapy administered was as follows: posaconazole at 20 mg/kg of body weight (twice daily), voriconazole at 40 mg/kg, anidulafungin at 10 mg/kg, or amphotericin B at 0.8 mg/kg. The efficacy was evaluated by prolonged animal survival, tissue burden reduction, and (1→3)-β-d-glucan serum levels. In general, the four antifungal drugs showed high MICs and poor in vitro activity. The efficacy of the different treatments was only modest, since survival rates were never higher than 40% and no drug was able to reduce fungal load in all the organs for the three strains tested. Posaconazole, in spite of its high MICs (≥16 μg/ml), showed the highest efficacy. The (1→3)-β-d-glucan serum levels were equally reduced by all drugs evaluated. PMID:24100490

  19. Genome structure of mink cell focus-forming murine leukemia virus in epithelial mink lung cells transformed vitro by iododeoxyuridine-induced C3H/MuLV cells.

    PubMed Central

    Rapp, U R; Birkenmeier, E; Bonner, T I; Gonda, M A; Gunnell, M

    1983-01-01

    We characterized mink cell focus-forming murine leukemia viruses that were isolated from C3H/MCA-5 cells after induction with 5-iododeoxyuridine in culture. Mink lung epithelial cells malignantly transformed in vitro by induced virus were the source of four molecular clones of mink cell focus-forming virus. CI-1, CI-2, CI-3, and CI-4. Three clones, CI-1, CI-2, and CI-3, had full-length mink cell focus-forming viral genomes, one of which (CI-3) was infectious. In addition, we obtained a defective viral genome (CI-4) which had a deletion in the envelope gene. A comparison between the envelope genes of CI-4 and those of spleen focus-forming virus by heteroduplex mapping showed close homology in the substitution region and defined the deletion as being identical to the p15E deletion of spleen focus-forming virus. The recombinant mink cell focus-forming genomes are not endogenous in C3H/MCA-5 cells and therefore must have been formed in culture after induction by 5-iododeoxyuridine. CI-3, the infectious clone of mink cell focus-forming murine leukemia virus, was dualtropic, and mink cells infected with CI-3 were altered in their response to epidermal growth factor. In the presence of epidermal growth factor at 10 ng/ml, uninfected mink cells retained their epithelial morphology in monolayer culture and did not form colonies in soft agar. In contrast, CI-3 virus-infected mink cells grew with fibroblastic morphology in monolayer culture and showed an increased growth rate in soft agar in the presence of epidermal growth factor. Images PMID:6300431

  20. Dose-reduced fludarabine, cyclophosphamide and rituximab is well tolerated in older patients with chronic lymphocytic leukemia and has preserved therapeutic efficacy.

    PubMed

    Lew, Thomas E; Cheah, Chan Y; Carney, Dennis A; Prince, H Miles; Wolf, Max; Bazargan, Ali; Januszewicz, E Henry; Filshie, Robin; Westerman, David; Seymour, John F; Tam, Constantine S

    2016-05-01

    Despite its efficacy in prospective trials, full dose fludarabine, cyclophosphamide and rituximab (FCR) may be too toxic for elderly patients with chronic lymphocytic leukemia (CLL) in clinical practice. We retrospectively reviewed the impact of dose reductions in FCR therapy on the outcomes of 42 consecutive patients aged 65-87 (median 72) years. Despite a median cumulative fludarabine dose reduction of 50% from full dose, the objective response and complete response rates were 86% and 38% respectively (frontline 94%/59%; previously treated 80%/24%). Dose reductions of 25-75% were not significantly associated with inferior progression free survival compared to minimal reductions (≤25%) (p = 0.49), and did not preclude deep responses, including six cases (14%) of minimal residual disease negativity. Although hematological and infectious toxicities were common, treatment limiting adverse effects were infrequent. Dose attenuated FCR appears to have preserved efficacy and may be a viable therapeutic option for elderly patients with CLL. PMID:26464106

  1. Efficacy and safety of intrathecal liposomal cytarabine for the treatment of meningeal relapse in acute lymphoblastic leukemia: experience of two pediatric institutions.

    PubMed

    Parasole, Rosanna; Menna, Giuseppe; Marra, Nicoletta; Petruzziello, Fara; Locatelli, Franco; Mangione, Argia; Misuraca, Aldo; Buffardi, Salvatore; Di Cesare-Merlone, Alessandra; Poggi, Vincenzo

    2008-08-01

    The treatment of meningeal relapse in acute lymphoblastic leukemia (ALL) remains a challenging clinical problem. Liposomal cytarabine (DepoCyte) permits to decrease frequency of lumbar punctures, without loss of efficacy, because intrathecal levels of the drug remain cytotoxic for up to 14 days. We investigated the efficacy and safety of intrathecal DepoCyte in six children with meningeal relapse, treated in two pediatric institutions. DepoCyte was well tolerated in all patients, who achieved complete clearance of blasts from the cerebrospinal fluid after the first three intrathecal drug administrations. Five of the six patients were concurrently treated with high-dose administration of systemic cytarabine, without additional neurological side effects. Our results suggest that DepoCyte is a valid option for children with ALL experiencing meningeal relapse; it deserves further investigation in intensive treatment regimens, taking into due consideration potential neurotoxicity. PMID:18766969

  2. Efficacy of the clinical agent VT-1161 against fluconazole-sensitive and -resistant Candida albicans in a murine model of vaginal candidiasis.

    PubMed

    Garvey, E P; Hoekstra, W J; Schotzinger, R J; Sobel, J D; Lilly, E A; Fidel, P L

    2015-09-01

    Vulvovaginal candidiasis (VVC) and recurrent VVC (RVVC) remain major health problems for women. VT-1161, a novel fungal CYP51 inhibitor which has potent antifungal activity against fluconazole-sensitive Candida albicans, retained its in vitro potency (MIC50 of ≤0.015 and MIC90 of 0.12 μg/ml) against 10 clinical isolates from VVC or RVVC patients resistant to fluconazole (MIC50 of 8 and MIC90 of 64 μg/ml). VT-1161 pharmacokinetics in mice displayed a high volume of distribution (1.4 liters/kg), high oral absorption (73%), and a long half-life (>48 h) and showed rapid penetration into vaginal tissue. In a murine model of vaginal candidiasis using fluconazole-sensitive yeast, oral doses as low as 4 mg/kg VT-1161 significantly reduced the fungal burden 1 and 4 days posttreatment (P < 0.0001). Similar VT-1161 efficacy was measured when an isolate highly resistant to fluconazole (MIC of 64 μg/ml) but fully sensitive in vitro to VT-1161 was used. When an isolate partially sensitive to VT-1161 (MIC of 0.12 μg/ml) and moderately resistant to fluconazole (MIC of 8 μg/ml) was used, VT-1161 remained efficacious, whereas fluconazole was efficacious on day 1 but did not sustain efficacy 4 days posttreatment. Both agents were inactive in treating an infection with an isolate that demonstrated weaker potency (MICs of 2 and 64 μg/ml for VT-1161 and fluconazole, respectively). Finally, the plasma concentrations of free VT-1161 were predictive of efficacy when in excess of the in vitro MIC values. These data support the clinical development of VT-1161 as a potentially more efficacious treatment for VVC and RVVC. PMID:26124165

  3. Efficacy of the Clinical Agent VT-1161 against Fluconazole-Sensitive and -Resistant Candida albicans in a Murine Model of Vaginal Candidiasis

    PubMed Central

    Hoekstra, W. J.; Schotzinger, R. J.; Sobel, J. D.; Lilly, E. A.; Fidel, P. L.

    2015-01-01

    Vulvovaginal candidiasis (VVC) and recurrent VVC (RVVC) remain major health problems for women. VT-1161, a novel fungal CYP51 inhibitor which has potent antifungal activity against fluconazole-sensitive Candida albicans, retained its in vitro potency (MIC50 of ≤0.015 and MIC90 of 0.12 μg/ml) against 10 clinical isolates from VVC or RVVC patients resistant to fluconazole (MIC50 of 8 and MIC90 of 64 μg/ml). VT-1161 pharmacokinetics in mice displayed a high volume of distribution (1.4 liters/kg), high oral absorption (73%), and a long half-life (>48 h) and showed rapid penetration into vaginal tissue. In a murine model of vaginal candidiasis using fluconazole-sensitive yeast, oral doses as low as 4 mg/kg VT-1161 significantly reduced the fungal burden 1 and 4 days posttreatment (P < 0.0001). Similar VT-1161 efficacy was measured when an isolate highly resistant to fluconazole (MIC of 64 μg/ml) but fully sensitive in vitro to VT-1161 was used. When an isolate partially sensitive to VT-1161 (MIC of 0.12 μg/ml) and moderately resistant to fluconazole (MIC of 8 μg/ml) was used, VT-1161 remained efficacious, whereas fluconazole was efficacious on day 1 but did not sustain efficacy 4 days posttreatment. Both agents were inactive in treating an infection with an isolate that demonstrated weaker potency (MICs of 2 and 64 μg/ml for VT-1161 and fluconazole, respectively). Finally, the plasma concentrations of free VT-1161 were predictive of efficacy when in excess of the in vitro MIC values. These data support the clinical development of VT-1161 as a potentially more efficacious treatment for VVC and RVVC. PMID:26124165

  4. Leukemia -- Eosinophilic

    MedlinePlus

    ... Leukemia - Eosinophilic: Overview Request Permissions Print to PDF Leukemia - Eosinophilic: Overview Approved by the Cancer.Net Editorial ... Platelets that help the blood to clot About leukemia Types of leukemia are named after the specific ...

  5. Expression of Interferon Consensus Sequence Binding Protein (ICSBP) Is Downregulated in Bcr-Abl-Induced Murine Chronic Myelogenous Leukemia-Like Disease, and Forced Coexpression of ICSBP Inhibits Bcr-Abl-Induced Myeloproliferative Disorder

    PubMed Central

    Hao, Sheryl X.; Ren, Ruibao

    2000-01-01

    Chronic myelogenous leukemia (CML) is a clonal myeloproliferative disorder resulting from the neoplastic transformation of a hematopoietic stem cell. The majority of cases of CML are associated with the (9;22) chromosome translocation that generates the bcr-abl chimeric gene. Alpha interferon (IFN-α) treatment induces hematological remission and prolongs life in 75% of CML patients in the chronic phase. It has been shown that mice deficient in interferon consensus sequence binding protein (ICSBP), a member of the interferon regulatory factor family, manifest a CML-like syndrome. We have shown that expression of Bcr-Abl in bone marrow (BM) cells from 5-fluorouracil (5-FU)-treated mice by retroviral transduction efficiently induces a myeloproliferative disease in mice resembling human CML. To directly test whether icsbp can function as a tumor suppressor gene, we examined the effect of ICSBP on Bcr-Abl-induced CML-like disease using this murine model for CML. We found that expression of the ICSBP protein was significantly decreased in Bcr-Abl-induced CML-like disease. Forced coexpression of ICSBP inhibited the Bcr-Abl-induced colony formation of BM cells from 5-FU-treated mice in vitro and Bcr-Abl-induced CML-like disease in vivo. Interestingly, coexpression of ICSBP and Bcr-Abl induced a transient B-lymphoproliferative disorder in the murine model of Bcr-Abl-induced CML-like disease. Overexpression of ICSBP consistently promotes rather than inhibits Bcr-Abl-induced B lymphoproliferation in a murine model where BM cells from non-5-FU-treated donors were used, indicating that ICSBP has a specific antitumor activity toward myeloid neoplasms. We also found that overexpression of ICSBP negatively regulated normal hematopoiesis. These data provide direct evidence that ICSBP can act as a tumor suppressor that regulates normal and neoplastic proliferation of hematopoietic cells. PMID:10648600

  6. Leukemia - B-Cell Prolymphocytic Leukemia and Hairy Cell Leukemia

    MedlinePlus

    ... Leukemia: Introduction Request Permissions Print to PDF Leukemia - B-cell Prolymphocytic Leukemia and Hairy Cell Leukemia: Introduction ... Research and Advocacy Survivorship Blog About Us Leukemia - B-cell Prolymphocytic Leukemia and Hairy Cell Leukemia Guide ...

  7. Antibody formation and mannose-6-phosphate receptor expression impact the efficacy of muscle-specific transgene expression in murine Pompe disease

    PubMed Central

    Sun, Baodong; Li, Songtao; Bird, Andrew; Yi, Haiqing; Kemper, Alex; Koeberl, Dwight D.

    2013-01-01

    BACKGROUND Lysosomal storage disorders such as Pompe disease can be more effectively treated, if immune tolerance to enzyme or gene replacement therapy can be achieved. Alternatively, immune responses against acid α-glucosidase (GAA) might be evaded in Pompe disease through muscle-specific expression of GAA with adeno-associated virus (AAV) vectors. METHODS An AAV vector containing the MHCK7 regulatory cassette to drive muscle-specific GAA expression was administered to GAA knockout (KO) mice, immune tolerant GAA-KO mice, and mannose-6-phosphate deficient GAA-KO mice. GAA activity and glycogen content were analyzed in striated muscle to determine biochemical efficacy. RESULTS The biochemical efficacy from GAA expression was slightly reduced in GAA-KO mice, as demonstrated by higher residual glycogen content in skeletal muscles. Next immune tolerance to GAA was induced in GAA-KO mice by co-administration of a second AAV vector encoding liver-specific GAA along with the AAV vector encoding muscle-specific GAA. Antibody formation was prevented by liver-specific GAA, and the biochemical efficacy of GAA expression was improved in absence of antibodies as evidenced by significantly reduced glycogen content in the diaphragm. Efficacy was reduced in old GAA-KO mice despite the absence of antibodies. The greatest impact upon gene therapy was observed in GAA-KO mice lacking the mannose-6-phosphate receptor in muscle. The clearance of stored glycogen was markedly impaired despite high GAA expression in receptor-deficient Pompe disease mice. CONCLUSIONS Overall, antibody formation had a subtle effect upon efficacy, while the absence of mannose-6-phosphate receptors markedly impaired muscle-targeted gene therapy in murine Pompe disease. PMID:20967919

  8. Effect of preexisting anti-herpes immunity on the efficacy of herpes simplex viral therapy in a murine intraperitoneal tumor model.

    PubMed

    Lambright, E S; Kang, E H; Force, S; Lanuti, M; Caparrelli, D; Kaiser, L R; Albelda, S M; Molnar-Kimber, K L

    2000-10-01

    HSV-1716, a replicating nonneurovirulent herpes simplex virus type 1, has shown efficacy in treating multiple types of human tumors in immunodeficient mice. Since the majority of the human population has been previously exposed to herpes simplex virus, the efficacy of HSV-based oncolytic therapy was investigated in an immunocompetent animal tumor model. EJ-6-2-Bam-6a, a tumor cell line derived from h-ras-transformed murine fibroblast, exhibit a diffuse growth pattern in the peritoneal cavity of BALB/c mice and replicate HSV-1716 to titers observed in human tumors. An established intraperitoneal (ip) tumor model of EJ-6-2-Bam-6a in naive and HSV-immunized mice was used to evaluate the efficacy of single or multiple ip administrations of HSV-1716 (4 x 10(6) pfu/treatment) or of carrier cells, which are irradiated, ex vivo virally infected EJ-6-2-Bam-6a cells that can amplify the viral load in situ. All treated groups significantly prolonged survival versus media control with an approximately 40% long-term survival rate (cure) in the multiply treated, HSV-naive animals. Prior immunization of the mice with HSV did not significantly decrease the median survival of the single or multiply treated HSV-1716 or the carrier cell-treated groups. These studies support the development of replication-selective herpes virus mutants for use in localized intraperitoneal malignancies. PMID:11020355

  9. Long-term efficacy and safety of bosutinib in patients with advanced leukemia following resistance/intolerance to imatinib and other tyrosine kinase inhibitors.

    PubMed

    Gambacorti-Passerini, Carlo; Kantarjian, Hagop M; Kim, Dong-Wook; Khoury, Hanna J; Turkina, Anna G; Brümmendorf, Tim H; Matczak, Ewa; Bardy-Bouxin, Nathalie; Shapiro, Mark; Turnbull, Kathleen; Leip, Eric; Cortes, Jorge E

    2015-09-01

    Long-term efficacy and safety of bosutinib (≥4 years follow-up from last enrolled patient) were evaluated in an ongoing phase 1/2 study in the advanced leukemia cohort with prior treatment failure (accelerated-phase [AP, n = 79] chronic myeloid leukemia [CML], blast-phase [BP, n = 64] CML, acute lymphoblastic leukemia [ALL, n = 24]). Fourteen AP, 2 BP, and 1 ALL patient remained on bosutinib at 4 years (vs. 38, 8, 1 at 1 year); median (range) treatment durations: 10.2 (0.1-88.6), 2.8 (0.03-55.9), 0.97 (0.3-89.2) months. Among AP and BP patients, 57% and 28% newly attained or maintained baseline overall hematologic response (OHR); 40% and 37% attained/maintained major cytogenetic response (MCyR) by 4 years (most by 12 months). In responders at 1 versus 4 years, Kaplan-Meier (KM) probabilities of maintaining OHR were 78% versus 49% (AP) and 28% versus 19% (BP); KM probabilities of maintaining MCyR were 65% versus 49% (AP) and 21% versus 21% (BP). Most common AEs (AP, BP) were gastrointestinal (96%; 83%), primarily diarrhea (85%; 64%), which was typically low grade (maximum grade 1/2: 81%; 59%) and transient; no patient discontinued due to diarrhea. Serious AEs occurred in 44 (56%) AP and 37 (58%) BP patients, most commonly pneumonia (n = 9) for AP and pyrexia (n = 6) for BP; 11 and 13 died within 30 days of last dose (2 considered bosutinib-related [AP] per investigator). Responses were durable in ∼50% AP responders at 4 years (∼25% BP patients responded at year 1, suggesting possible bridge-to-transplant role in BP patients); toxicity was manageable. PMID:26040495

  10. Efficacy of levocarnitine for tyrosine kinase inhibitor-induced painful muscle cramps in patients with chronic myelogenous leukemia.

    PubMed

    Yamada, Michiko; Kuroda, Hiroyuki; Shimoyama, Saori; Ito, Ryo; Sugama, Yusuke; Sato, Ken; Yamauchi, Natsumi; Horiguchi, Hiroto; Nakamura, Hajime; Hamaguchi, Kota; Abe, Tomoyuki; Fujii, Shigeyuki; Maeda, Masahiro; Kato, Junji

    2016-04-01

    Muscle cramps are side effects commonly associated with tyrosine kinase inhibitor (TKI) treatment. Patients suffering from muscle cramps are treated with various medications such as calcium, magnesium and vitamin supplements, but these therapies are often ineffective. We report two patients with chronic myelogenous leukemia who developed muscle cramps caused by TKI. These patients were treated successfully with levocarnitine. Both of our cases revealed the beneficial effects of levocarnitine treatment on TKI-induced muscle cramps. PMID:27169456