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Sample records for phosphotyrosine peptidomimetic prodrugs

  1. Design and Synthesis of Phosphotyrosine Peptidomimetic Prodrugs

    PubMed Central

    Garrido-Hernandez, Hugo; Moon, Kyung D.; Geahlen, Robert L.; Borch, Richard F.

    2008-01-01

    A novel approach to the intracellular delivery of aryl phosphates has been developed that utilizes a phosphoramidate-based prodrug approach. The prodrugs contain an ester group that undergoes reductive activation intracellularly with concomitant expulsion of a phosphoramidate anion. This anion undergoes intramolecular cyclization and hydrolysis to generate aryl phosphate exclusively with a t1/2 = ∼ 20 min. Phosphoramidate prodrugs (8-10) of phosphate-containing peptidomimetics that target the SH2 domain were synthesized. Evaluation of these peptidomimetic prodrugs in a growth inhibition assay and, in a cell-based transcriptional assay, demonstrated that the prodrugs had IC50 values in the low micromolar range. Synthesis of phosphorodiamidate analogs containing a P-NH-Ar linker (16 – 18) was also carried out in the hope that the phosphoramidates released might be phosphatase-resistant. Comparable activation rates and cell-based activities were observed for these prodrugs, but the intermediate phosphoramidate dianion underwent spontaneous hydrolysis with a t1/2 = ∼ 30 min. PMID:16722656

  2. Bifunctional peptidomimetic prodrugs of didanosine for improved intestinal permeability and enhanced acidic stability: synthesis, transepithelial transport, chemical stability and pharmacokinetics.

    PubMed

    Yan, Zhongtian; Sun, Jin; Chang, Yannan; Liu, Yanhua; Fu, Qiang; Xu, Youjun; Sun, Yongbing; Pu, Xiaohui; Zhang, Youxi; Jing, Yongkui; Yin, Shiliang; Zhu, Meng; Wang, Yongjun; He, Zhonggui

    2011-04-01

    Five peptidomimetic prodrugs of didanosine (DDI) were synthesized and designed to improve bioavailability of DDI following oral administration via targeting intestinal oligopeptide transporter (PepT1) and enhancing chemical stability. The permeability of prodrugs was screened in Caco-2 cells grown on permeable supports. 5'-O-L-valyl ester prodrug of DDI (compound 4a) demonstrated the highest membrane permeability and was selected as the optimal target prodrug for further studies. The uptake of glycylsarcosine (Gly-Sar, a typical substrate of PepT1) by Caco-2 cells could be inhibited by compound 4a in a concentration-dependent manner. The Caco-2 cells were treated with 0.2 nM leptin for enhanced PepT1 expression. The uptake of compound 4a was markedly increased in the leptin-treated Caco-2 cells compared with the control Caco-2 cells, both of which were obviously inhibited by 20 mM Gly-Sar. The K(m) and V(max) values of kinetic study of compound 4a transported by PepT1 in Caco-2 cells were 0.91 mM and 11.94 nmol/mg of protein/10 min, respectively. The chemical stability studies were performed in simulated gastric fluid (SGF), phosphate buffers under various pH conditions, rat tissue homogenates and plasma at 37 °C. The concentrations of DDI could not be detected in the two minutes in SGF. But compound 4a could significantly increase DDI acidic stability, and its t(½) was extended to as long as 36 min in SGF. Compound 4a was stable in pH 6.0 phosphate buffer but could be quickly transformed into DDI in plasma and tissue homogenates. The oral absolute bioavailability of DDI was 47.2% and 7.9% after compound 4a and DDI were orally administered to rats at a dose of 15 mg/kg, respectively. The coadministration with antiacid agent could also suggest that compound 4a was more stable under harsh acidic conditions compared with DDI. Compound 4a bioavailability in rats was reduced to 33.9% when orally co-administered with Gly-Sar (100 mg/kg). The In Vivo bioactivation

  3. Dendrimer Prodrugs.

    PubMed

    da Silva Santos, Soraya; Igne Ferreira, Elizabeth; Giarolla, Jeanine

    2016-01-01

    The main objective of this review is to describe the importance of dendrimer prodrugs in the design of new drugs, presenting numerous applications of these nanocomposites in the pharmaceutical field. Therefore, the use of dendrimer prodrugs as carrier for drug delivery, to improve pharmacokinetic properties of prototype, to promote drug sustained-release, to increase selectivity and, consequently, to decrease toxicity, are just some examples of topics that have been extensively reported in the literature, especially in the last decade. The examples discussed here give a panel of the growing interest dendrimer prodrugs have been evoking in the scientific community. PMID:27258239

  4. [Research progress of small peptidomimetics].

    PubMed

    Wang, Jia; Sun, De-qun

    2015-08-01

    The study of peptide drugs has been an important direction in research and development of new drugs. However, lots of natural macromolecular peptides are limited in clinical use by their metabolic instability and low bioavailability. In recent years, the active small peptidomimetics open up a new hotspot of peptide drug development with the characteristics of low molecular weight, high bioactivity and structural modification. Many peptidomimetics are on the market or on the clinical study. This paper elaborated the small peptidomimetics approved by American Food and Drug Administration (FDA) from 2005 to 2014, and reviewed their researching status with source, synthetic method, chemical structure, marketing time, indication, clinical efficacy and safety. Research prospects in this field were discussed. PMID:26668991

  5. Towards Quantitative Phosphotyrosine Profiling In Vivo

    PubMed Central

    Johnson, Hannah; White, Forest M.

    2012-01-01

    Tyrosine phosphorylation is a dynamic reversible post-translational modification that regulates many aspects of cell biology. To understand how this modification controls biological function, it is necessary to not only identify the specific sites of phosphorylation, but also to quantify how phosphorylation levels on these sites may be altered under specific physiological conditions. Due to its sensitivity and accuracy, mass spectrometry (MS) has widely been applied to the identification and characterization of phosphotyrosine signaling across biological systems. In this review we highlight the advances in both MS and phosphotyrosine enrichment methods that have been developed to enable the identification of low level tyrosine phosphorylation events. Computational and manual approaches to ensure confident identification of phosphopeptide sequence and determination of phosphorylation site localization are discussed along with methods that have been applied to the relative quantification of large numbers of phosphorylation sites. Finally, we provide an overview of the challenges ahead as we extend these technologies to the characterization of tyrosine phosphorylation signaling in vivo. With these latest developments in analytical and computational techniques, it is now possible to derive biological insight from quantitative MS-based analysis of signaling networks in vitro and in vivo. Application of these approaches to a wide variety of biological systems will define how signal transduction regulates cellular physiology in health and disease. PMID:22677333

  6. Expanding the genetic code of Escherichia coli with phosphotyrosine.

    PubMed

    Fan, Chenguang; Ip, Kevan; Söll, Dieter

    2016-09-01

    Protein phosphorylation is one of the most important post-translational modifications in nature. However, the site-specific incorporation of O-phosphotyrosine into proteins in vivo has not yet been reported. Endogenous phosphatases present in cells can dephosphorylate phosphotyrosine as a free amino acid or as a protein residue. Therefore, we deleted the genes of five phosphatases from the genome of Escherichia coli with the aim of stabilizing phosphotyrosine. Together with an engineered aminoacyl-tRNA synthetase (derived from Methanocaldococcus jannaschii tyrosyl-tRNA synthetase) and an elongation factor Tu variant, we were able to cotranslationally incorporate O-phosphotyrosine into the superfolder green fluorescent protein at a desired position in vivo. This system will facilitate future studies of tyrosine phosphorylation. PMID:27477338

  7. Quantitative Proteomic Approaches for Studying Phosphotyrosine Signaling

    SciTech Connect

    Ding, Shi-Jian; Qian, Weijun; Smith, Richard D.

    2007-02-01

    Protein tyrosine phosphorylation is a fundamental mechanism for controlling many aspects of cellular processes, as well as aspects of human health and diseases. Compared to phosphoserine (pSer) and phosphothreonine (pThr), phosphotyrosine (pTyr) signaling is more tightly regulated, but often more challenging to characterize due to significantly lower level of tyrosine phosphorylation (a relative abundance of 1800:200:1 was estimated for pSer/pThr/pTyr in vertebrate cells[1]). In this review, we outline the recent advances in analytical methodologies for enrichment, identification, and accurate quantitation of tyrosine phosphorylated proteins and peptides using antibody-based technologies, capillary liquid chromatography (LC) coupled with mass spectrometry (MS), and various stable isotope labeling strategies, as well as non-MS-based methods such as protein or peptide array methods. These proteomic technological advances provide powerful tools for potentially understanding signal transduction at the system level and provide a basis for discovering novel drug targets for human diseases. [1] Hunter, T. (1998) The Croonian Lecture 1997. The phosphorylation of proteins on tyrosine: its role in cell growth and disease. Philos. Trans. R. Soc. Lond. B Biol. Sci. 353, 583–605

  8. A phosphotyrosine switch regulates organic cation transporters

    PubMed Central

    Sprowl, Jason A.; Ong, Su Sien; Gibson, Alice A.; Hu, Shuiying; Du, Guoqing; Lin, Wenwei; Li, Lie; Bharill, Shashank; Ness, Rachel A.; Stecula, Adrian; Offer, Steven M.; Diasio, Robert B.; Nies, Anne T.; Schwab, Matthias; Cavaletti, Guido; Schlatter, Eberhard; Ciarimboli, Giuliano; Schellens, Jan H. M.; Isacoff, Ehud Y.; Sali, Andrej; Chen, Taosheng; Baker, Sharyn D.; Sparreboom, Alex; Pabla, Navjotsingh

    2016-01-01

    Membrane transporters are key determinants of therapeutic outcomes. They regulate systemic and cellular drug levels influencing efficacy as well as toxicities. Here we report a unique phosphorylation-dependent interaction between drug transporters and tyrosine kinase inhibitors (TKIs), which has uncovered widespread phosphotyrosine-mediated regulation of drug transporters. We initially found that organic cation transporters (OCTs), uptake carriers of metformin and oxaliplatin, were inhibited by several clinically used TKIs. Mechanistic studies showed that these TKIs inhibit the Src family kinase Yes1, which was found to be essential for OCT2 tyrosine phosphorylation and function. Yes1 inhibition in vivo diminished OCT2 activity, significantly mitigating oxaliplatin-induced acute sensory neuropathy. Along with OCT2, other SLC-family drug transporters are potentially part of an extensive ‘transporter-phosphoproteome' with unique susceptibility to TKIs. On the basis of these findings we propose that TKIs, an important and rapidly expanding class of therapeutics, can functionally modulate pharmacologically important proteins by inhibiting protein kinases essential for their post-translational regulation. PMID:26979622

  9. Peptides and Peptidomimetics for Antimicrobial Drug Design

    PubMed Central

    Mojsoska, Biljana; Jenssen, Håvard

    2015-01-01

    The purpose of this paper is to introduce and highlight a few classes of traditional antimicrobial peptides with a focus on structure-activity relationship studies. After first dissecting the important physiochemical properties that influence the antimicrobial and toxic properties of antimicrobial peptides, the contributions of individual amino acids with respect to the peptides antibacterial properties are presented. A brief discussion of the mechanisms of action of different antimicrobials as well as the development of bacterial resistance towards antimicrobial peptides follows. Finally, current efforts on novel design strategies and peptidomimetics are introduced to illustrate the importance of antimicrobial peptide research in the development of future antibiotics. PMID:26184232

  10. Diversity-oriented synthesis of macrocyclic peptidomimetics

    PubMed Central

    Isidro-Llobet, Albert; Murillo, Tiffanie; Bello, Paula; Cilibrizzi, Agostino; Hodgkinson, James T.; Galloway, Warren R. J. D.; Bender, Andreas; Welch, Martin; Spring, David R.

    2011-01-01

    Structurally diverse libraries of novel small molecules represent important sources of biologically active agents. In this paper we report the development of a diversity-oriented synthesis strategy for the generation of diverse small molecules based around a common macrocyclic peptidomimetic framework, containing structural motifs present in many naturally occurring bioactive compounds. Macrocyclic peptidomimetics are largely underrepresented in current small-molecule screening collections owing primarily to synthetic intractability; thus novel molecules based around these structures represent targets of significant interest, both from a biological and a synthetic perspective. In a proof-of-concept study, the synthesis of a library of 14 such compounds was achieved. Analysis of chemical space coverage confirmed that the compound structures indeed occupy underrepresented areas of chemistry in screening collections. Crucial to the success of this approach was the development of novel methodologies for the macrocyclic ring closure of chiral α-azido acids and for the synthesis of diketopiperazines using solid-supported N methylmorpholine. Owing to their robust and flexible natures, it is envisaged that both new methodologies will prove to be valuable in a wider synthetic context. PMID:21383137

  11. Pyruvate kinase M2 is a phosphotyrosine-binding protein

    SciTech Connect

    Christofk, H.R.; Vander Heiden, M.G.; Wu, N.; Asara, J.M.; Cantley, L.C.

    2008-06-03

    Growth factors stimulate cells to take up excess nutrients and to use them for anabolic processes. The biochemical mechanism by which this is accomplished is not fully understood but it is initiated by phosphorylation of signalling proteins on tyrosine residues. Using a novel proteomic screen for phosphotyrosine-binding proteins, we have made the observation that an enzyme involved in glycolysis, the human M2 (fetal) isoform of pyruvate kinase (PKM2), binds directly and selectively to tyrosine-phosphorylated peptides. We show that binding of phosphotyrosine peptides to PKM2 results in release of the allosteric activator fructose-1,6-bisphosphate, leading to inhibition of PKM2 enzymatic activity. We also provide evidence that this regulation of PKM2 by phosphotyrosine signalling diverts glucose metabolites from energy production to anabolic processes when cells are stimulated by certain growth factors. Collectively, our results indicate that expression of this phosphotyrosine-binding form of pyruvate kinase is critical for rapid growth in cancer cells.

  12. Structural and evolutionary division of phosphotyrosine binding (PTB) domains.

    PubMed

    Uhlik, Mark T; Temple, Brenda; Bencharit, Sompop; Kimple, Adam J; Siderovski, David P; Johnson, Gary L

    2005-01-01

    Proteins encoding phosphotyrosine binding (PTB) domains function as adaptors or scaffolds to organize the signaling complexes involved in wide-ranging physiological processes including neural development, immunity, tissue homeostasis and cell growth. There are more than 200 proteins in eukaryotes and nearly 60 human proteins having PTB domains. Six PTB domain encoded proteins have been found to have mutations that contribute to inherited human diseases including familial stroke, hypercholesteremia, coronary artery disease, Alzheimer's disease and diabetes, demonstrating the importance of PTB scaffold proteins in organizing critical signaling complexes. PTB domains bind both peptides and headgroups of phosphatidylinositides, utilizing two distinct binding motifs to mediate spatial organization and localization within cells. The structure of PTB domains confers specificity for binding peptides having a NPXY motif with differing requirements for phosphorylation of the tyrosine within this recognition sequence. In this review, we use structural, evolutionary and functional analysis to divide PTB domains into three groups represented by phosphotyrosine-dependent Shc-like, phosphotyrosine-dependent IRS-like and phosphotyrosine-independent Dab-like PTBs, with the Dab-like PTB domains representing nearly 75% of proteins encoding PTB domains. In addition, we further define the binding characteristics of the cognate ligands for each group of PTB domains. The signaling complexes organized by PTB domain encoded proteins are largely unknown and represents an important challenge in systems biology for the future. PMID:15567406

  13. Similarity analysis, synthesis, and bioassay of antibacterial cyclic peptidomimetics

    PubMed Central

    Berhanu, Workalemahu M; Ibrahim, Mohamed A; Pillai, Girinath G; Oliferenko, Alexander A; Khelashvili, Levan; Jabeen, Farukh; Mirza, Bushra; Ansari, Farzana Latif; ul-Haq, Ihsan; El-Feky, Said A

    2012-01-01

    Summary The chemical similarity of antibacterial cyclic peptides and peptidomimetics was studied in order to identify new promising cyclic scaffolds. A large descriptor space coupled with cluster analysis was employed to digitize known antibacterial structures and to gauge the potential of new peptidomimetic macrocycles, which were conveniently synthesized by acylbenzotriazole methodology. Some of the synthesized compounds were tested against an array of microorganisms and showed antibacterial activity against Bordetella bronchistepica, Micrococcus luteus, and Salmonella typhimurium. PMID:23019443

  14. Prodrug Strategies for Paclitaxel.

    PubMed

    Meng, Ziyuan; Lv, Quanxia; Lu, Jun; Yao, Houzong; Lv, Xiaoqing; Jiang, Feng; Lu, Aiping; Zhang, Ge

    2016-01-01

    Paclitaxel is an anti-tumor agent with remarkable anti-tumor activity and wide clinical uses. However, it is also faced with various challenges especially for its poor water solubility and low selectivity for the target. To overcome these disadvantages of paclitaxel, approaches using small molecule modifications and macromolecule modifications have been developed by many research groups from all over the world. In this review, we discuss the different strategies especially prodrug strategies that are currently used to make paclitaxel more effective. PMID:27223283

  15. Prodrug Strategies for Paclitaxel

    PubMed Central

    Meng, Ziyuan; Lv, Quanxia; Lu, Jun; Yao, Houzong; Lv, Xiaoqing; Jiang, Feng; Lu, Aiping; Zhang, Ge

    2016-01-01

    Paclitaxel is an anti-tumor agent with remarkable anti-tumor activity and wide clinical uses. However, it is also faced with various challenges especially for its poor water solubility and low selectivity for the target. To overcome these disadvantages of paclitaxel, approaches using small molecule modifications and macromolecule modifications have been developed by many research groups from all over the world. In this review, we discuss the different strategies especially prodrug strategies that are currently used to make paclitaxel more effective. PMID:27223283

  16. Adding Diverse Noncanonical Backbones to Rosetta: Enabling Peptidomimetic Design

    PubMed Central

    Craven, Timothy W.; Butterfoss, Glenn L.; Chou, Fang-Chieh; Lyskov, Sergey; Bullock, Brooke N.; Watkins, Andrew; Labonte, Jason W.; Pacella, Michael; Kilambi, Krishna Praneeth; Leaver-Fay, Andrew; Kuhlman, Brian; Gray, Jeffrey J.; Bradley, Philip; Kirshenbaum, Kent; Arora, Paramjit S.; Das, Rhiju; Bonneau, Richard

    2013-01-01

    Peptidomimetics are classes of molecules that mimic structural and functional attributes of polypeptides. Peptidomimetic oligomers can frequently be synthesized using efficient solid phase synthesis procedures similar to peptide synthesis. Conformationally ordered peptidomimetic oligomers are finding broad applications for molecular recognition and for inhibiting protein-protein interactions. One critical limitation is the limited set of design tools for identifying oligomer sequences that can adopt desired conformations. Here, we present expansions to the ROSETTA platform that enable structure prediction and design of five non-peptidic oligomer scaffolds (noncanonical backbones), oligooxopiperazines, oligo-peptoids, -peptides, hydrogen bond surrogate helices and oligosaccharides. This work is complementary to prior additions to model noncanonical protein side chains in ROSETTA. The main purpose of our manuscript is to give a detailed description to current and future developers of how each of these noncanonical backbones was implemented. Furthermore, we provide a general outline for implementation of new backbone types not discussed here. To illustrate the utility of this approach, we describe the first tests of the ROSETTA molecular mechanics energy function in the context of oligooxopiperazines, using quantum mechanical calculations as comparison points, scanning through backbone and side chain torsion angles for a model peptidomimetic. Finally, as an example of a novel design application, we describe the automated design of an oligooxopiperazine that inhibits the p53-MDM2 protein-protein interaction. For the general biological and bioengineering community, several noncanonical backbones have been incorporated into web applications that allow users to freely and rapidly test the presented protocols (http://rosie.rosettacommons.org). This work helps address the peptidomimetic community's need for an automated and expandable modeling tool for noncanonical

  17. Isocyanide-based multicomponent reactions towards cyclic constrained peptidomimetics

    PubMed Central

    Koopmanschap, Gijs; Ruijter, Eelco

    2014-01-01

    Summary In the recent past, the design and synthesis of peptide mimics (peptidomimetics) has received much attention. This because they have shown in many cases enhanced pharmacological properties over their natural peptide analogues. In particular, the incorporation of cyclic constructs into peptides is of high interest as they reduce the flexibility of the peptide enhancing often affinity for a certain receptor. Moreover, these cyclic mimics force the molecule into a well-defined secondary structure. Constraint structural and conformational features are often found in biological active peptides. For the synthesis of cyclic constrained peptidomimetics usually a sequence of multiple reactions has been applied, which makes it difficult to easily introduce structural diversity necessary for fine tuning the biological activity. A promising approach to tackle this problem is the use of multicomponent reactions (MCRs), because they can introduce both structural diversity and molecular complexity in only one step. Among the MCRs, the isocyanide-based multicomponent reactions (IMCRs) are most relevant for the synthesis of peptidomimetics because they provide peptide-like products. However, these IMCRs usually give linear products and in order to obtain cyclic constrained peptidomimetics, the acyclic products have to be cyclized via additional cyclization strategies. This is possible via incorporation of bifunctional substrates into the initial IMCR. Examples of such bifunctional groups are N-protected amino acids, convertible isocyanides or MCR-components that bear an additional alkene, alkyne or azide moiety and can be cyclized via either a deprotection–cyclization strategy, a ring-closing metathesis, a 1,3-dipolar cycloaddition or even via a sequence of multiple multicomponent reactions. The sequential IMCR-cyclization reactions can afford small cyclic peptide mimics (ranging from four- to seven-membered rings), medium-sized cyclic constructs or peptidic macrocycles

  18. Prodrugs of reverse fosmidomycin analogues.

    PubMed

    Brücher, Karin; Gräwert, Tobias; Konzuch, Sarah; Held, Jana; Lienau, Claudia; Behrendt, Christoph; Illarionov, Boris; Maes, Louis; Bacher, Adelbert; Wittlin, Sergio; Mordmüller, Benjamin; Fischer, Markus; Kurz, Thomas

    2015-02-26

    Fosmidomycin inhibits IspC (Dxr, 1-deoxy-d-xylulose 5-phosphate reductoisomerase), a key enzyme in nonmevalonate isoprenoid biosynthesis that is essential in Plasmodium falciparum. The drug has been used successfully to treat malaria patients in clinical studies, thus validating IspC as an antimalarial target. However, improvement of the drug's pharmacodynamics and pharmacokinetics is desirable. Here, we show that the conversion of the phosphonate moiety into acyloxymethyl and alkoxycarbonyloxymethyl groups can increase the in vitro activity against asexual blood stages of P. falciparum by more than 1 order of magnitude. We also synthesized double prodrugs by additional esterification of the hydroxamate moiety. Prodrugs with modified hydroxamate moieties are subject to bioactivation in vitro. All prodrugs demonstrated improved antiplasmodial in vitro activity. Selected prodrugs and parent compounds were also tested for their cytotoxicity toward HeLa cells and in vivo in a Plasmodium berghei malaria model as well as in the SCID mouse P. falciparum model. PMID:25633870

  19. Potent inhibition of protein-tyrosine phosphatase by phosphotyrosine-mimic containing cyclic peptides.

    PubMed

    Akamatsu, M; Roller, P P; Chen, L; Zhang, Z Y; Ye, B; Burke, T R

    1997-01-01

    In an effort to derive potent and bioavailable protein-tyrosine phosphatase inhibitors, we have previously reported hexameric peptides based on the epidermal growth factor receptor sequence EGFR988-993 (Asp-Ala-Asp-Glu-Xxx-Leu, where Xxx = Tyr), in which the tyrosyl residue has been replaced by the non-hydrolyzable phosphotyrosyl mimics phosphonomethylphenylalanine (Pmp), difluorophosphonomethylphenylalanine (F2Pmp) and O-malonyltyrosine (OMT). Inhibitory potencies (IC50 values) of these peptides against the tyrosine phosphatase PTP IB were 200, 0.2 and 10 microM, respectively. Since cellular penetration of peptides containing highly charged phosphonate residues is compromised, and good bioreversible protection strategies for the F2Pmp residue have not yet been reported, the OMT residue is of particular interest in that it affords potential new prodrug approaches. In the current study we have prepared cyclized versions of the OMT-containing EGFR988-993 peptide in order to increase its proteolytic stability and restrain conformational flexibility. Three different cyclic analogues were synthesized. Two of these were cyclized through the peptide backbone ('head to tail') using in one case a single glycine spacer (heptamer peptide) and in the second instance, two glycines (octamer peptide). In a PTPI-based assay the cyclic heptamer experienced a two-fold loss of potency (Ki = 25.2 +/- 3.9 microM) relative to the linear hexamer parent (Ki = 13 +/- 0.9 microM), while the cyclic octamer demonstrated a live-fold increase in potency (Ki = 2.60 +/- 0.11 microM). The third peptide was cyclized by means of a sulfide bridge between the side chain of a C-terminally added cysteine residue and the beta-carbon of a N-terminal acetyl residue. Although the overall size of this ring was identical to that exhibited by the preceding backbone-cyclized octamer, it displayed a three-fold enhancement in potency (Ki = 0.73 +/- 0.03 microM). The structural basis for the observed results are

  20. Advances in peptidic and peptidomimetic-based approaches to inhibit STAT signaling in human diseases.

    PubMed

    Szelag, Malgorzata; Wesoly, Joanna; Bluyssen, Hans A R

    2016-01-01

    STATs promote fundamental cellular processes, marking them as convergence points of many oncogenic and inflammatory pathways. Therefore, aberrant activation of STAT signaling is implicated in a plethora of human diseases, like cancer, inflammation and auto-immunity. Identification of STAT-specific inhibitors is the topic of great practical importance, and various inhibitory strategies are being pursued. An interesting approach includes peptides and peptide-like biopolymers, because they allow the manipulation of STAT signaling without the transfer of genetic material. Phosphopeptides and peptidomimetics directly target STATs by inhibiting dimerization. Despite that a large number of efficient peptide- based STAT3-specific inhibitors have been reported to date, none of them was able to meet the pharmacological requirements to serve as a potent anti-cancer drug. The existing limitations, like metabolic instability and poor cell permeability during in vivo tests, excluded these macromolecules from further clinical development. To overcome these liabilities, in the last five years many advances have been made to develop next generation STAT-specific inhibitors. Here we discuss the pitfalls of current STAT inhibitory strategies and review the progress on the development of peptide-like prodrugs directly targeting STATs. Novel strategies involve screening of high-complexity libraries of random peptides, as specific STAT3 or STAT5 DNA-binding inhibitors, to construct cell permeable peptide aptamers and aptides for cancer therapy. Another new direction is synthesis of negative dominant α-helical mimetics of the STAT3 N-domain, preventing oligomerization on DNA. Moreover, construction of phosphopeptide conjugates with molecules mediating cellular uptake offers new therapeutic possibilities in treatment of cancer, asthma and allergy. PMID:26521960

  1. Rational Design of Calpain Inhibitors Based on Calpastatin Peptidomimetics.

    PubMed

    Low, Kristin E; Ler, Spencer; Chen, Kevin J; Campbell, Robert L; Hickey, Jennifer L; Tan, Joanne; Scully, Conor C G; Davies, Peter L; Yudin, Andrei K; Zaretsky, Serge

    2016-06-01

    Our previously reported structures of calpain bound to its endogenous inhibitor calpastatin have motivated the use of aziridine aldehyde-mediated peptide macrocyclization toward the design of cyclic peptides and peptidomimetics as calpain inhibitors. Inspired by nature's hint that a β-turn loop within calpastatin forms a broad interaction around calpain's active site cysteine, we have constructed and tested a library of 45 peptidic compounds based on this loop sequence. Four molecules have shown reproducibly low micromolar inhibition of calpain-2. Further systematic sequence changes led to the development of probes that displayed increased potency and specificity of inhibition against calpain over other cysteine proteases. Calculated Ki values were in the low micromolar range, rivaling other peptidomimetic calpain inhibitors and presenting an improved selectivity profile against other therapeutically relevant proteases. Competitive and mixed inhibition against calpain-2 was observed, and an allosteric inhibition site on the enzyme was identified for a noncompetitive inhibitor. PMID:27148623

  2. Rationally Designed Peptidomimetic Modulators of Aβ Toxicity in Alzheimer's Disease

    NASA Astrophysics Data System (ADS)

    Rajasekhar, K.; Suresh, S. N.; Manjithaya, Ravi; Govindaraju, T.

    2015-01-01

    Alzheimer's disease is one of the devastating illnesses mankind is facing in the 21st century. The main pathogenic event in Alzheimer's disease is believed to be the aggregation of the β-amyloid (Aβ) peptides into toxic aggregates. Molecules that interfere with this process may act as therapeutic agents for the treatment of the disease. Use of recognition unit based peptidomimetics as inhibitors are a promising approach, as they exhibit greater protease stability compared to natural peptides. Here, we present peptidomimetic inhibitors of Aβ aggregation designed based on the KLVFF (P1) sequence that is known to bind Aβ aggregates. We improved inhibition efficiency of P1 by introducing multiple hydrogen bond donor-acceptor moieties (thymine/barbiturate) at the N-terminal (P2 and P3), and blood serum stability by modifying the backbone by incorporating sarcosine (N-methylglycine) units at alternate positions (P4 and P5). The peptidomimetics showed moderate to good activity in both inhibition and dissolution of Aβ aggregates as depicted by thioflavin assay, circular dichroism (CD) measurements and microscopy (TEM). The activity of P4 and P5 were studied in a yeast cell model showing Aβ toxicity. P4 and P5 could rescue yeast cells from Aβ toxicity and Aβ aggregates were cleared by the process of autophagy.

  3. Rationally Designed Peptidomimetic Modulators of Aβ Toxicity in Alzheimer's Disease

    PubMed Central

    Rajasekhar, K.; Suresh, S. N.; Manjithaya, Ravi; Govindaraju, T.

    2015-01-01

    Alzheimer's disease is one of the devastating illnesses mankind is facing in the 21st century. The main pathogenic event in Alzheimer's disease is believed to be the aggregation of the β-amyloid (Aβ) peptides into toxic aggregates. Molecules that interfere with this process may act as therapeutic agents for the treatment of the disease. Use of recognition unit based peptidomimetics as inhibitors are a promising approach, as they exhibit greater protease stability compared to natural peptides. Here, we present peptidomimetic inhibitors of Aβ aggregation designed based on the KLVFF (P1) sequence that is known to bind Aβ aggregates. We improved inhibition efficiency of P1 by introducing multiple hydrogen bond donor-acceptor moieties (thymine/barbiturate) at the N-terminal (P2 and P3), and blood serum stability by modifying the backbone by incorporating sarcosine (N-methylglycine) units at alternate positions (P4 and P5). The peptidomimetics showed moderate to good activity in both inhibition and dissolution of Aβ aggregates as depicted by thioflavin assay, circular dichroism (CD) measurements and microscopy (TEM). The activity of P4 and P5 were studied in a yeast cell model showing Aβ toxicity. P4 and P5 could rescue yeast cells from Aβ toxicity and Aβ aggregates were cleared by the process of autophagy. PMID:25633824

  4. Substrate Mediated Enzyme Prodrug Therapy

    PubMed Central

    Fejerskov, Betina; Zelikin, Alexander N.

    2012-01-01

    In this report, we detail Substrate Mediated Enzyme Prodrug Therapy (SMEPT) as a novel approach in drug delivery which relies on enzyme-functionalized cell culture substrates to achieve a localized conversion of benign prodrug(s) into active therapeutics with subsequent delivery to adhering cells or adjacent tissues. For proof-of-concept SMEPT, we use surface adhered micro-structured physical hydrogels based on poly(vinyl alcohol), β-glucuronidase enzyme and glucuronide prodrugs. We demonstrate enzymatic activity mediated by the assembled hydrogel samples and illustrate arms of control over rate of release of model fluorescent cargo. SMEPT was not impaired by adhering cells and afforded facile time - and dose – dependent uptake of the in situ generated fluorescent cargo by hepatic cells, HepG2. With the use of a glucuronide derivative of an anticancer drug, SN-38, SMEPT afforded a decrease in cell viability to a level similar to that achieved using parent drug. Finally, dose response was achieved using SMEPT and administration of judiciously chosen concentration of SN-38 glucuronide prodrug thus revealing external control over drug delivery using drug eluting surface. We believe that this highly adaptable concept will find use in diverse biomedical applications, specifically surface mediated drug delivery and tissue engineering. PMID:23152927

  5. Phospho-tyrosine dependent protein–protein interaction network

    PubMed Central

    Grossmann, Arndt; Benlasfer, Nouhad; Birth, Petra; Hegele, Anna; Wachsmuth, Franziska; Apelt, Luise; Stelzl, Ulrich

    2015-01-01

    Post-translational protein modifications, such as tyrosine phosphorylation, regulate protein–protein interactions (PPIs) critical for signal processing and cellular phenotypes. We extended an established yeast two-hybrid system employing human protein kinases for the analyses of phospho-tyrosine (pY)-dependent PPIs in a direct experimental, large-scale approach. We identified 292 mostly novel pY-dependent PPIs which showed high specificity with respect to kinases and interacting proteins and validated a large fraction in co-immunoprecipitation experiments from mammalian cells. About one-sixth of the interactions are mediated by known linear sequence binding motifs while the majority of pY-PPIs are mediated by other linear epitopes or governed by alternative recognition modes. Network analysis revealed that pY-mediated recognition events are tied to a highly connected protein module dedicated to signaling and cell growth pathways related to cancer. Using binding assays, protein complementation and phenotypic readouts to characterize the pY-dependent interactions of TSPAN2 (tetraspanin 2) and GRB2 or PIK3R3 (p55γ), we exemplarily provide evidence that the two pY-dependent PPIs dictate cellular cancer phenotypes. PMID:25814554

  6. Photosensitive Peptidomimetic for Light-Controlled, Reversible DNA Compaction.

    PubMed

    Schimka, Selina; Santer, Svetlana; Mujkić-Ninnemann, Nina M; Bléger, David; Hartmann, Laura; Wehle, Marko; Lipowsky, Reinhard; Santer, Mark

    2016-06-13

    Light-induced DNA compaction as part of nonviral gene delivery was investigated intensively in the past years, although the bridging between the artificial light switchable compacting agents and biocompatible light insensitive compacting agents was not achieved until now. In this paper, we report on light-induced compaction and decompaction of DNA molecules in the presence of a new type of agent, a multivalent cationic peptidomimetic molecule containing a photosensitive Azo-group as a branch (Azo-PM). Azo-PM is synthesized using a solid-phase procedure during which an azobenzene unit is attached as a side chain to an oligo(amidoamine) backbone. We show that within a certain range of concentrations and under illumination with light of appropriate wavelengths, these cationic molecules induce reversible DNA compaction/decompaction by photoisomerization of the incorporated azobenzene unit between a hydrophobic trans- and a hydrophilic cis-conformation, as characterized by dynamic light scattering and AFM measurements. In contrast to other molecular species used for invasive DNA compaction, such as widely used azobenzene containing cationic surfactant (Azo-TAB, C4-Azo-OCX-TMAB), the presented peptidomimetic agent appears to lead to different complexation/compaction mechanisms. An investigation of Azo-PM in close proximity to a DNA segment by means of a molecular dynamics simulation sustains a picture in which Azo-PM acts as a multivalent counterion, with its rather large cationic oligo(amidoamine) backbone dominating the interaction with the double helix, fine-tuned or assisted by the presence and isomerization state of the Azo-moiety. However, due to its peptidomimetic backbone, Azo-PM should be far less toxic than photosensitive surfactants and might represent a starting point for a conscious design of photoswitchable, biocompatible vectors for gene delivery. PMID:27030485

  7. A diversity-oriented synthesis strategy enabling the combinatorial-type variation of macrocyclic peptidomimetic scaffolds.

    PubMed

    Isidro-Llobet, Albert; Hadje Georgiou, Kathy; Galloway, Warren R J D; Giacomini, Elisa; Hansen, Mette R; Méndez-Abt, Gabriela; Tan, Yaw Sing; Carro, Laura; Sore, Hannah F; Spring, David R

    2015-04-21

    Macrocyclic peptidomimetics are associated with a broad range of biological activities. However, despite such potentially valuable properties, the macrocyclic peptidomimetic structural class is generally considered as being poorly explored within drug discovery. This has been attributed to the lack of general methods for producing collections of macrocyclic peptidomimetics with high levels of structural, and thus shape, diversity. In particular, there is a lack of scaffold diversity in current macrocyclic peptidomimetic libraries; indeed, the efficient construction of diverse molecular scaffolds presents a formidable general challenge to the synthetic chemist. Herein we describe a new, advanced strategy for the diversity-oriented synthesis (DOS) of macrocyclic peptidomimetics that enables the combinatorial variation of molecular scaffolds (core macrocyclic ring architectures). The generality and robustness of this DOS strategy is demonstrated by the step-efficient synthesis of a structurally diverse library of over 200 macrocyclic peptidomimetic compounds, each based around a distinct molecular scaffold and isolated in milligram quantities, from readily available building-blocks. To the best of our knowledge this represents an unprecedented level of scaffold diversity in a synthetically derived library of macrocyclic peptidomimetics. Cheminformatic analysis indicated that the library compounds access regions of chemical space that are distinct from those addressed by top-selling brand-name drugs and macrocyclic natural products, illustrating the value of our DOS approach to sample regions of chemical space underexploited in current drug discovery efforts. An analysis of three-dimensional molecular shapes illustrated that the DOS library has a relatively high level of shape diversity. PMID:25778821

  8. Phosphotyrosine recognition domains: the typical, the atypical and the versatile

    PubMed Central

    2012-01-01

    SH2 domains are long known prominent players in the field of phosphotyrosine recognition within signaling protein networks. However, over the years they have been joined by an increasing number of other protein domain families that can, at least with some of their members, also recognise pTyr residues in a sequence-specific context. This superfamily of pTyr recognition modules, which includes substantial fractions of the PTB domains, as well as much smaller, or even single member fractions like the HYB domain, the PKCδ and PKCθ C2 domains and RKIP, represents a fascinating, medically relevant and hence intensely studied part of the cellular signaling architecture of metazoans. Protein tyrosine phosphorylation clearly serves a plethora of functions and pTyr recognition domains are used in a similarly wide range of interaction modes, which encompass, for example, partner protein switching, tandem recognition functionalities and the interaction with catalytically active protein domains. If looked upon closely enough, virtually no pTyr recognition and regulation event is an exact mirror image of another one in the same cell. Thus, the more we learn about the biology and ultrastructural details of pTyr recognition domains, the more does it become apparent that nature cleverly combines and varies a few basic principles to generate a sheer endless number of sophisticated and highly effective recognition/regulation events that are, under normal conditions, elegantly orchestrated in time and space. This knowledge is also valuable when exploring pTyr reader domains as diagnostic tools, drug targets or therapeutic reagents to combat human diseases. PMID:23134684

  9. Global Impact of Oncogenic Src on a Phosphotyrosine Proteome

    PubMed Central

    Luo, Weifeng; Slebos, Robbert J.; Hill, Salisha; Li, Ming; Brábek, Jan; Amanchy, Ramars; Chaerkady, Raghothama; Pandey, Akhilesh; Ham, Amy-Joan L.; Hanks, Steven K.

    2008-01-01

    Elevated activity of Src, the first characterized protein-tyrosine kinase, is associated with progression of many human cancers, and Src has attracted interest as a therapeutic target. Src is known to act in various receptor signaling systems to impact cell behavior, yet it remains likely that the spectrum of Src protein substrates relevant to cancer is incompletely understood. To better understand the cellular impact of deregulated Src kinase activity, we extensively applied a mass spectrometry shotgun phosphotyrosine (pTyr) proteomics strategy to obtain global pTyr profiles of Src-transformed mouse fibroblasts as well as their nontransformed counterparts. A total of 867 peptides representing 563 distinct pTyr sites on 374 different proteins were identified from the Src-transformed cells, while 514 peptides representing 275 pTyr sites on 167 proteins were identified from nontransformed cells. Distinct characteristics of the two profiles were revealed by spectral counting, indicative of pTyr site relative abundance, and by complementary quantitative analysis using stable isotope labeling with amino acids in cell culture (SILAC). While both pTyr profiles are replete with sites on signaling and adhesion/cytoskeletal regulatory proteins, the Src-transformed profile is more diverse with enrichment in sites on metabolic enzymes and RNA and protein synthesis and processing machinery. Forty-three pTyr sites (32 proteins) are predicted as major biologically relevant Src targets on the basis of frequent identification in both cell populations. This select group, of particular interest as diagnostic biomarkers, includes well-established Src sites on signaling/adhesion/cytoskeletal proteins, but also uncharacterized sites of potential relevance to the transformed cell phenotype. PMID:18563927

  10. Development of Anti-EGF Receptor Peptidomimetics (AERP) as Tumor Imaging Agent

    PubMed Central

    Ponde, Datta E.; Su, ZiFen; Berezov, Alan; Zhang, Hongtao; Alavi, Abbas; Greene, Mark I.; Murali, Ramachandran

    2011-01-01

    EGFR is over-expressed in several solid tumors including breast, prostate, pancreas and lung cancers and is correlated to the metastasic potential of the tumor. Anti-EGFR receptor-binding peptidomimetics (AERP) were examined to assess the small molecule's potential use as tumor-specific imaging agents. The aim of this work was to design and characterize the binding specificity of the radiolabeled peptidomimetics to EGFR over-expressing cell lysate and to A431 xenograft tumors. Our newly designed peptidomimetic, AERP, was conjugated to DTPA and labeled with 99mTc. The in vivo tumor accumulation of [99mTc] DTPA-AERP-2 was 1.6 ± 0.1 %ID/g and tumor to muscle ratio was 5.5. Our studies suggest that this novel peptidomimetic, AERP-2, warrants further development as an EGFR-specific tumor-imaging agent. PMID:21392985

  11. Synthesis of sansalvamide A peptidomimetics: triazole, oxazole, thiazole, and pseudoproline containing compounds

    PubMed Central

    Davis, Melinda R.; Singh, Erinprit K.; Wahyudi, Hendra; Alexander, Leslie D.; Kunicki, Joseph B.; Nazarova, Lidia A.; Fairweather, Kelly A.; Giltrap, Andrew M.; Jolliffe, Katrina A.; McAlpine, Shelli R.

    2011-01-01

    Peptidomimetic-based macrocycles typically have improved pharmacokinetic properties over those observed with peptide analogs. Described are the syntheses of 13 peptidomimetic derivatives that are based on active Sansalvamide A structures, where these analogs incorporate heterocycles (triazoles, oxazoles, thiazoles, or pseudoprolines) along the macrocyclic backbone. The syntheses of these derivatives employ several approaches that can be applied to convert a macrocyclic peptide into its peptidomimetic counterpart. These approaches include peptide modifications to generate the alkyne and azide for click chemistry, a serine conversion into an oxazole, a Hantzsch reaction to generate the thiazole, and protected threonine to generate the pseudoproline derivatives. Furthermore, we show that two different peptidomimetic moieties, triazoles and thiazoles, can be incorporated into the macrocyclic backbone without reducing cytotoxicity: triazole and thiazole. PMID:22287031

  12. Adaptive Evolution of Escherichia coli to an α-Peptide/β-Peptoid Peptidomimetic Induces Stable Resistance

    PubMed Central

    Hein-Kristensen, Line; Franzyk, Henrik; Holch, Anne; Gram, Lone

    2013-01-01

    Antimicrobial peptides (AMPs) and synthetic analogues thereof target conserved structures of bacterial cell envelopes and hence, development of resistance has been considered an unlikely event. However, recently bacterial resistance to AMPs has been observed, and the aim of the present study was to determine whether bacterial resistance may also evolve against synthetic AMP analogues, e.g. α-peptide/β-peptoid peptidomimetics. E. coli ATCC 25922 was exposed to increasing concentrations of a peptidomimetic (10 lineages), polymyxin B (10 lineages), or MilliQ water (4 lineages) in a re-inoculation culturing setup covering approx. 500 generations. All 10 lineages exposed to the peptidomimetic adapted to 32×MIC while this occurred for 8 out of 10 of the polymyxin B-exposed lineages. All lineages exposed to 32×MIC of either the peptidomimetic or polymyxin B had a significantly increased MIC (16–32×) to the selection agent. Five transfers (∼35 generations) in unsupplemented media did not abolish resistance indicating that resistance was heritable. Single isolates from peptidomimetic-exposed lineage populations displayed MICs against the peptidomimetic from wild-type MIC to 32×MIC revealing heterogeneous populations. Resistant isolates showed no cross-resistance against a panel of membrane-active AMPs. These isolates were highly susceptible to blood plasma antibacterial activity and were killed when plasma concentrations exceeded ∼30%. Notably, MIC of the peptidomimetic against resistant isolates returned to wild-type level upon addition of 25% plasma. Whole-genome sequencing of twenty isolates from four resistant lineages revealed mutations, in murein transglycosylase D (mltD) and outer-membrane proteins, which were conserved within and between lineages. However, no common resistance-conferring mutation was identified. We hypothesise that alterations in cell envelope structure result in peptidomimetic resistance, and that this may occur via several distinct

  13. Poly-lysine peptidomimetics having potent antimicrobial activity without hemolytic activity.

    PubMed

    Ahn, Mija; Jacob, Binu; Gunasekaran, Pethaiah; Murugan, Ravichandran N; Ryu, Eun Kyoung; Lee, Ga-hyang; Hyun, Jae-Kyung; Cheong, Chaejoon; Kim, Nam-Hyung; Shin, Song Yub; Bang, Jeong Kyu

    2014-09-01

    Diversity of sequence and structure in naturally occurring antimicrobial peptides (AMPs) limits their intensive structure-activity relationship (SAR) study. In contrast, peptidomimetics have several advantages compared to naturally occurring peptide in terms of simple structure, convenient to analog synthesis, rapid elucidation of optimal physiochemical properties and low-cost synthesis. In search of short antimicrobial peptides using peptidomimetics, which provide facile access to identify the key factors involving in the destruction of pathogens through SAR study, a series of simple and short peptidomimetics consisting of multi-Lys residues and lipophilic moiety have been prepared and found to be active against several Gram-negative and Gram-positive bacteria containing methicillin-resistant Staphylococcus aureus (MRSA) without hemolytic activity. Based on the SAR studies, we found that hydrophobicity, +5 charges of multiple Lys residues, hydrocarbon tail lengths and cyclohexyl group were crucial for antimicrobial activity. Furthermore, membrane depolarization, dye leakage, inner membrane permeability and time-killing kinetics revealed that bacterial-killing mechanism of our peptidomimetics is different from the membrane-targeting AMPs (e. g. melittin and SMAP-29) and implied our peptidomimetics might kill bacteria via the intracellular-targeting mechanism as done by buforin-2. PMID:24961649

  14. Hydrogen sulfide prodrugs-a review.

    PubMed

    Zheng, Yueqin; Ji, Xingyue; Ji, Kaili; Wang, Binghe

    2015-09-01

    Hydrogen sulfide (H2S) is recognized as one of three gasotransmitters together with nitric oxide (NO) and carbon monoxide (CO). As a signaling molecule, H2S plays an important role in physiology and shows great potential in pharmaceutical applications. Along this line, there is a need for the development of H2S prodrugs for various reasons. In this review, we summarize different H2S prodrugs, their chemical properties, and some of their potential therapeutic applications. PMID:26579468

  15. Direct Photocontrol of Peptidomimetics: An Alternative to Oxygen-Dependent Photodynamic Cancer Therapy.

    PubMed

    Babii, Oleg; Afonin, Sergii; Garmanchuk, Liudmyla V; Nikulina, Viktoria V; Nikolaienko, Tetiana V; Storozhuk, Olha V; Shelest, Dmytro V; Dasyukevich, Olga I; Ostapchenko, Liudmyla I; Iurchenko, Volodymyr; Zozulya, Sergey; Ulrich, Anne S; Komarov, Igor V

    2016-04-25

    Conventional photodynamic treatment strategies are based on the principle of activating molecular oxygen in situ by light, mediated by a photosensitizer, which leads to the generation of reactive oxygen species and thereby causes cell death. A diarylethene-derived peptidomimetic is presented that is suitable for photodynamic cancer therapy without any involvement of oxygen. This light-sensitive molecule is not a mediator but is itself the cytotoxic agent. As a derivative of the cyclic amphiphilic peptide gramicidin S, the peptidomimetic exists in two thermally stable photoforms that are interconvertible by light of different wavelengths. The isomer generated by visible light shows much stronger toxicity against tumor cells than the UV-generated isomer. First in vivo applications are demonstrated on a tumor animal model to illustrate how the peptidomimetic can be administered in the less toxic form and then activated locally in a solid tumor by visible light. PMID:27028784

  16. Antibacterial Peptidomimetics: Polymeric Synthetic Mimics of Antimicrobial Peptides

    NASA Astrophysics Data System (ADS)

    Lienkamp, Karen; Madkour, Ahmad E.; Tew, Gregory N.

    Polymer-based peptidomimetics, or proteinomimetics, are a relatively young and dynamic field of research. The ability to successfully mimic the biochemical activity of antimicrobial peptides (AMPs) has been demonstrated by several groups. This has been accomplished by careful tuning of the molecule's hydrophobicity and charge density. At the same time, many important questions remain to be answered, including the role of backbone rigidity, details of membrane insertion, and the role of curvature in the self-assemblies between these novel peptidemimetics and phospholipids. As the biological properties of polymeric synthetic mimics of AMPs (SMAMPs) result from the interplay of many parameters, it is not yet possible to predict the exact properties of such molecules from their mere chemical structure. However, as demonstrated here, the effect of certain design features such as charge and hydrophobicity on the properties across a polymer series is understood. Compared to the mechanistic specifics that are known about the interactions of AMPs or small antibacterial molecules with membranes and cells, relatively little is known concerning the interaction of polymeric SMAMPs with membranes. Beyond SMAMPs, numerous opportunities exist and protein transduction domain mimics are an active area of research in the Tew laboratory. These two examples, one quite new and the other studied for almost a decade, demonstrate that it is possible to teach synthetic polymers to behave like peptides, despite their lack of sequence specificity and secondary structure.

  17. Identification of Glioblastoma Phosphotyrosine-Containing Proteins with Two-Dimensional Western Blotting and Tandem Mass Spectrometry

    PubMed Central

    Guo, Tianyao; Wang, Xiaowei; Li, Maoyu; Yang, Haiyan; Li, Ling; Peng, Fang

    2015-01-01

    To investigate the presence of, and the potential biological roles of, protein tyrosine phosphorylation in the glioblastoma pathogenesis, two-dimensional gel electrophoresis- (2DGE-) based Western blotting coupled with liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS) analysis was used to detect and identify the phosphotyrosine immunoreaction-positive proteins in a glioblastoma tissue. MS/MS and Mascot analyses were used to determine the phosphotyrosine sites of each phosphopeptide. Protein domain and motif analysis and systems pathway analysis were used to determine the protein domains/motifs that contained phosphotyrosine residue and signal pathway networks to clarify the potential biological functions of protein tyrosine phosphorylation. A total of 24 phosphotyrosine-containing proteins were identified. Each phosphotyrosine-containing protein contained at least one tyrosine kinase phosphorylation motif and a certain structural and functional domains. Those phosphotyrosine-containing proteins were involved in the multiple signal pathway systems such as oxidative stress, stress response, and cell migration. Those data show 2DGE-based Western blotting, MS/MS, and bioinformatics are a set of effective approaches to detect and identify glioblastoma tyrosine-phosphorylated proteome and to effectively rationalize the biological roles of tyrosine phosphorylation in the glioblastoma biological systems. It provides novel insights regarding tyrosine phosphorylation and its potential role in the molecular mechanism of a glioblastoma. PMID:26090378

  18. Evaluation of the Interaction between Phosphohistidine Analogues and Phosphotyrosine Binding Domains

    PubMed Central

    McAllister, Tom E; Horner, Katherine A; Webb, Michael E

    2014-01-01

    We have investigated the interaction of peptides containing phosphohistidine analogues and their homologues with the prototypical phosphotyrosine binding SH2 domain from the eukaryotic cell signalling protein Grb2 by using a combination of isothermal titration calorimetry and a fluorescence anisotropy competition assay. These investigations demonstrated that the triazole class of phosphohistidine analogues are capable of binding too, suggesting that phosphohistidine could potentially be detected by this class of proteins in vivo. PMID:24771713

  19. Immunostimulating activities of the novel peptidomimetic L-glutamyl-histamine

    PubMed Central

    Babizhayev, M A; Semiletov, Y A; Lul’kin, Y A; Sakina, N L; Savel’yeva, E L; Alimbarova, L M; Barinskii, I P

    2005-01-01

    An original representative of histamine-containing peptidomimetics L-glutamyl-histamine (L-Glu-Hist) was synthesized and characterized as a cytokine mimic leading to cellular responses of improved specificity. The energy-minimized 3-D conformations of L-Glu-Hist derived from its chemical structure resulted in stabilization for Fe2+ chelating complexes. L-Glu-Hist accelerated the decrease of ferrous iron in the ferrous sulphate solution in a concentration-dependent mode and showed the ferroxidase-like activity at concentrations less than 3 mm in the phenanthroline assay, whereas in the concentration range 3–20 mm L-Glu-Hist restricted the availability of Fe2+ to phenanthroline due to binding of ferrous ions in chelating complexes. L-Glu-Hist showed a stimulatory effect on phosphatidylcholine liposomal peroxidation (LPO) catalysed by the superoxide anion radical ()-generating system (Fe2++ ascorbate) at low (less or about 1 mm) L-Glu-Hist concentrations and both revealed the inhibitory effect on LPO in this system of high (∼ 10 mm) L-Glu-Hist concentration. L-Glu-Hist released in concentrations which stimulated [3H]-thymidine incorporation into DNA and proliferation of mouse spleen lymphocytes and mononuclear cells from human blood. The structural peptide-like analogues of L-Glu-Hist such as L-Glu-Trp, carcinine (β-alanylhistamine), but not L-Pro-Glu-Trp were active in stimulating thymidine incorporation and in inducing proliferation of mononuclear cells compared to mitogen concanavalin A at doses 2·5–25·0 µg/ml. Our data provide evidence that L-Glu-Hist may act as a very fast and sensitive trigger for lymphocyte proliferation and immunoregulation. PMID:15730390

  20. Amino Acid Carbamates As Prodrugs Of Resveratrol.

    PubMed

    Mattarei, Andrea; Azzolini, Michele; La Spina, Martina; Zoratti, Mario; Paradisi, Cristina; Biasutto, Lucia

    2015-01-01

    Resveratrol (3, 5, 4'-trihydroxy-trans-stilbene), a plant polyphenol, has important drug-like properties, but its pharmacological exploitation in vivo is hindered by its rapid transformation via phase II conjugative metabolism. One approach to bypass this problem relies on prodrugs. We report here the synthesis, characterization, stability and in vivo pharmacokinetic behaviour of prodrugs of resveratrol in which the OH groups are engaged in an N-monosubstituted carbamate ester (-OC(O)NHR) linkage with a natural amino acid (Leu, Ile, Phe, Thr) to prevent conjugation and modulate the physicochemical properties of the molecule. We also report a convenient, high-yield protocol to obtain derivatives of this type. The new carbamate ester derivatives are stable at pH 1, while they undergo slow hydrolysis at physiological pH and hydrolyse with kinetics suitable for use in prodrugs in whole blood. After administration to rats by oral gavage the isoleucine-containing prodrug was significantly absorbed, and was present in the bloodstream as non-metabolized unaltered or partially deprotected species, demonstrating effective shielding from first-pass metabolism. We conclude that prodrugs based on the N-monosubstituted carbamate ester bond have the appropriate stability profile for the systemic delivery of phenolic compounds. PMID:26463125

  1. Amino Acid Carbamates As Prodrugs Of Resveratrol

    PubMed Central

    Mattarei, Andrea; Azzolini, Michele; La Spina, Martina; Zoratti, Mario; Paradisi, Cristina; Biasutto, Lucia

    2015-01-01

    Resveratrol (3, 5, 4′-trihydroxy-trans-stilbene), a plant polyphenol, has important drug-like properties, but its pharmacological exploitation in vivo is hindered by its rapid transformation via phase II conjugative metabolism. One approach to bypass this problem relies on prodrugs. We report here the synthesis, characterization, stability and in vivo pharmacokinetic behaviour of prodrugs of resveratrol in which the OH groups are engaged in an N-monosubstituted carbamate ester (-OC(O)NHR) linkage with a natural amino acid (Leu, Ile, Phe, Thr) to prevent conjugation and modulate the physicochemical properties of the molecule. We also report a convenient, high-yield protocol to obtain derivatives of this type. The new carbamate ester derivatives are stable at pH 1, while they undergo slow hydrolysis at physiological pH and hydrolyse with kinetics suitable for use in prodrugs in whole blood. After administration to rats by oral gavage the isoleucine-containing prodrug was significantly absorbed, and was present in the bloodstream as non-metabolized unaltered or partially deprotected species, demonstrating effective shielding from first-pass metabolism. We conclude that prodrugs based on the N-monosubstituted carbamate ester bond have the appropriate stability profile for the systemic delivery of phenolic compounds. PMID:26463125

  2. Selective Water-Soluble Gelatinase Inhibitor Prodrugs

    PubMed Central

    Gooyit, Major; Lee, Mijoon; Schroeder, Valerie A.; Ikejiri, Masahiro; Suckow, Mark A.; Mobashery, Shahriar; Chang, Mayland

    2011-01-01

    SB-3CT (1), a selective and potent thiirane-based gelatinase inhibitor, is effective in animal models of cancer metastasis and stroke; however, it is limited by poor aqueous solubility and extensive metabolism. We addressed these issues by blocking the primary site of metabolism and capitalizing on a prodrug strategy to achieve >5000-fold increased solubility. The amide prodrugs were quantitatively hydrolyzed in human blood to a potent gelatinase inhibitor, ND-322 (3). The arginyl amide prodrug (ND-478, 5d) was metabolically stable in mouse, rat, and human liver microsomes. Both 5d and 3 were non-mutagenic in the Ames II mutagenicity assay. The prodrug 5d showed moderate clearance of 0.0582 L/min/kg, remained mostly in the extracellular fluid compartment (Vd = 0.0978 L/kg), and had a terminal half-life of >4 h. The prodrug 5d had superior pharmacokinetic properties than 3, making the thiirane class of selective gelatinase inhibitors suitable for intravenous administration in treatment of acute gelatinase-dependent diseases. PMID:21866961

  3. Rapid Multistep Synthesis of a Bioactive Peptidomimetic Oligomer for the Undergraduate Laboratory

    ERIC Educational Resources Information Center

    Utku, Yeliz; Rohatgi, Abhinav; Yoo, Barney; Kirshenbaum, Kent; Zuckermann, Ronald N.; Pohl, Nicola L.

    2010-01-01

    Peptidomimetic compounds are increasingly important in drug-discovery applications. We introduce the synthesis of an N-substituted glycine oligomer, a bioactive "peptoid" trimer. The six-step protocol is conducted on solid-phase resin, enabling the synthesis to be performed by undergraduate organic chemistry students. This synthesis lab was…

  4. [Radiation Anticarcinogenesis by Thiazolidine Pro-drug

    NASA Technical Reports Server (NTRS)

    Warters, Raymond L.; Roberts, Jeanette C.; Fain, Heidi

    1999-01-01

    The original goal of this work was to determine the capacity of selected aminothiols to modulate radiation induced cytotoxicity, mutagenesis and carcinogenesis in a human mammary epithelial cell line. The conclusions from this work are that WR-1065 is the "gold standard" for protection against radiation induced cytotoxicity, mutagenesis and carcinogenesis. While a potent radiation protector, WR-1065 is cytotoxic in vitro and in vivo. Our rationale for a study of the thiazolidine pro-drugs was that these compounds are neither toxic in vitro or in vivo. The results obtained during this funding period indicate that the thiazolidine pro-drugs are as potent as WR-1065 as protectors against radiation induced mutation induction, and thus presumably against radiation induced carcinogenesis. Our results indicate that the thiazolidine prodrugs are excellent candidates to test as non-toxic anticarcinogens for protecting astronauts from cancer induction during space travel.

  5. Photoactivatable Prodrugs of Antimelanoma Agent Vemurafenib.

    PubMed

    Horbert, Rebecca; Pinchuk, Boris; Davies, Paul; Alessi, Dario; Peifer, Christian

    2015-09-18

    In this study, we report on novel photoactivatable caged prodrugs of vemurafenib. This kinase inhibitor was the first approved drug for the personalized treatment of BRAF-mutated melanoma and showed impressive results in clinical studies. However, the occurrence of severe side effects and drug resistance illustrates the urgent need for innovative therapeutic approaches. To conquer these limitations, we implemented photoremovable protecting groups into vemurafenib. In general, this caging concept provides spatial and temporal control over the activation of molecules triggered by ultraviolet light. Thus, higher inhibitor concentrations in tumor tissues might be reached with less systemic effects. Our study describes the first development of caged vemurafenib prodrugs useful as pharmacological tools. We investigated their photochemical characteristics and photoactivation. In vitro evaluation proved the intended loss-of-function and the light-dependent recovery of efficacy in kinase and cellular assays. The reported vemurafenib photo prodrugs represent a powerful biological tool for novel pharmacological approaches in cancer research. PMID:26061392

  6. Virtual Screening of Peptide and Peptidomimetic Fragments Targeted to Inhibit Bacterial Dithiol Oxidase DsbA.

    PubMed

    Duprez, Wilko; Bachu, Prabhakar; Stoermer, Martin J; Tay, Stephanie; McMahon, Róisín M; Fairlie, David P; Martin, Jennifer L

    2015-01-01

    Antibacterial drugs with novel scaffolds and new mechanisms of action are desperately needed to address the growing problem of antibiotic resistance. The periplasmic oxidative folding system in Gram-negative bacteria represents a possible target for anti-virulence antibacterials. By targeting virulence rather than viability, development of resistance and side effects (through killing host native microbiota) might be minimized. Here, we undertook the design of peptidomimetic inhibitors targeting the interaction between the two key enzymes of oxidative folding, DsbA and DsbB, with the ultimate goal of preventing virulence factor assembly. Structures of DsbB--or peptides--complexed with DsbA revealed key interactions with the DsbA active site cysteine, and with a hydrophobic groove adjacent to the active site. The present work aimed to discover peptidomimetics that target the hydrophobic groove to generate non-covalent DsbA inhibitors. The previously reported structure of a Proteus mirabilis DsbA active site cysteine mutant, in a non-covalent complex with the heptapeptide PWATCDS, was used as an in silico template for virtual screening of a peptidomimetic fragment library. The highest scoring fragment compound and nine derivatives were synthesized and evaluated for DsbA binding and inhibition. These experiments discovered peptidomimetic fragments with inhibitory activity at millimolar concentrations. Although only weakly potent relative to larger covalent peptide inhibitors that interact through the active site cysteine, these fragments offer new opportunities as templates to build non-covalent inhibitors. The results suggest that non-covalent peptidomimetics may need to interact with sites beyond the hydrophobic groove in order to produce potent DsbA inhibitors. PMID:26225423

  7. Virtual Screening of Peptide and Peptidomimetic Fragments Targeted to Inhibit Bacterial Dithiol Oxidase DsbA

    PubMed Central

    Stoermer, Martin J.; Tay, Stephanie; McMahon, Róisín M.; Fairlie, David P.; Martin, Jennifer L.

    2015-01-01

    Antibacterial drugs with novel scaffolds and new mechanisms of action are desperately needed to address the growing problem of antibiotic resistance. The periplasmic oxidative folding system in Gram-negative bacteria represents a possible target for anti-virulence antibacterials. By targeting virulence rather than viability, development of resistance and side effects (through killing host native microbiota) might be minimized. Here, we undertook the design of peptidomimetic inhibitors targeting the interaction between the two key enzymes of oxidative folding, DsbA and DsbB, with the ultimate goal of preventing virulence factor assembly. Structures of DsbB - or peptides - complexed with DsbA revealed key interactions with the DsbA active site cysteine, and with a hydrophobic groove adjacent to the active site. The present work aimed to discover peptidomimetics that target the hydrophobic groove to generate non-covalent DsbA inhibitors. The previously reported structure of a Proteus mirabilis DsbA active site cysteine mutant, in a non-covalent complex with the heptapeptide PWATCDS, was used as an in silico template for virtual screening of a peptidomimetic fragment library. The highest scoring fragment compound and nine derivatives were synthesized and evaluated for DsbA binding and inhibition. These experiments discovered peptidomimetic fragments with inhibitory activity at millimolar concentrations. Although only weakly potent relative to larger covalent peptide inhibitors that interact through the active site cysteine, these fragments offer new opportunities as templates to build non-covalent inhibitors. The results suggest that non-covalent peptidomimetics may need to interact with sites beyond the hydrophobic groove in order to produce potent DsbA inhibitors. PMID:26225423

  8. Vipegitide: a folded peptidomimetic partial antagonist of α2β1 integrin with antiplatelet aggregation activity

    PubMed Central

    Momic, Tatjana; Katzhendler, Jehoshua; Shai, Ela; Noy, Efrat; Senderowitz, Hanoch; Eble, Johannes A; Marcinkiewicz, Cezary; Varon, David; Lazarovici, Philip

    2015-01-01

    Linear peptides containing the sequence WKTSRTSHY were used as lead compounds to synthesize a novel peptidomimetic antagonist of α2β1 integrin, with platelet aggregation-inhibiting activity, named Vipegitide. Vipegitide is a 13-amino acid, folded peptidomimetic molecule, containing two α-aminoisobutyric acid residues at positions 6 and 8 and not stable in human serum. Substitution of glycine and tryptophan residues at positions 1 and 2, respectively, with a unit of two polyethylene glycol (PEG) molecules yielded peptidomimetic Vipegitide-PEG2, stable in human serum for over 3 hours. Vipegitide and Vipegitide-PEG2 showed high potency (7×10−10 M and 1.5×10−10 M, respectively) and intermediate efficacy (40% and 35%, respectively) as well as selectivity toward α2 integrin in inhibition of adhesion of α1/α2 integrin overexpressing cells toward respective collagens. Interaction of both peptidomimetics with extracellular active domain of α2 integrin was confirmed in cell-free binding assay with recombinant α2 A-domain. Integrin α2β1 receptor is found on the platelet membrane and triggers collagen-induced platelet aggregation. Vipegitide and Vipegitide-PEG2 inhibited α2β1 integrin-mediated adhesion of human and murine platelets under the flow condition, by 50%. They efficiently blocked adenosine diphosphate- and collagen I-induced platelet aggregation in platelet rich plasma and whole human blood. Higher potency of Vipegitide than Vipegitide-PEG2 is consistent with results of computer modeling of the molecules in water. These peptidomimetic molecules were acutely tolerated in mice upon intravenous bolus injection of 50 mg/kg. These results underline the potency of Vipegitide and Vipegitide-PEG2 molecules as platelet aggregation-inhibiting drug lead compounds in antithrombotic therapy. PMID:25609915

  9. Exploration of the P3 region of PEXEL peptidomimetics leads to a potent inhibitor of the Plasmodium protease, plasmepsin V.

    PubMed

    Gazdik, Michelle; Jarman, Kate E; O'Neill, Matthew T; Hodder, Anthony N; Lowes, Kym N; Jousset Sabroux, Helene; Cowman, Alan F; Boddey, Justin A; Sleebs, Brad E

    2016-05-01

    The use of arginine isosteres is a known strategy to overcome poor membrane permeability commonly associated with peptides or peptidomimetics that possess this highly polar amino acid. Here, we apply this strategy to peptidomimetics that are potent inhibitors of the malarial protease, plasmepsin V, with the aim of enhancing their activity against Plasmodium parasites, and exploring the structure-activity relationship of the P3 arginine within the S3 pocket of plasmepsin V. Of the arginine isosteres trialled in the P3 position, we discovered that canavanine was the ideal and that this peptidomimetic potently inhibits plasmepsin V, efficiently blocks protein export and inhibits parasite growth. Structure studies of the peptidomimetics bound to plasmepsin V provided insight into the structural basis for the enzyme activity observed in vitro and provides further evidence why plasmepsin V is highly sensitive to substrate modification. PMID:27021426

  10. Modern prodrug design for targeted oral drug delivery.

    PubMed

    Dahan, Arik; Zimmermann, Ellen M; Ben-Shabat, Shimon

    2014-01-01

    The molecular information that became available over the past two decades significantly influenced the field of drug design and delivery at large, and the prodrug approach in particular. While the traditional prodrug approach was aimed at altering various physiochemical parameters, e.g., lipophilicity and charge state, the modern approach to prodrug design considers molecular/cellular factors, e.g., membrane influx/efflux transporters and cellular protein expression and distribution. This novel targeted-prodrug approach is aimed to exploit carrier-mediated transport for enhanced intestinal permeability, as well as specific enzymes to promote activation of the prodrug and liberation of the free parent drug. The purpose of this article is to provide a concise overview of this modern prodrug approach, with useful successful examples for its utilization. In the past the prodrug approach used to be viewed as a last option strategy, after all other possible solutions were exhausted; nowadays this is no longer the case, and in fact, the prodrug approach should be considered already in the very earliest development stages. Indeed, the prodrug approach becomes more and more popular and successful. A mechanistic prodrug design that aims to allow intestinal permeability by specific transporters, as well as activation by specific enzymes, may greatly improve the prodrug efficiency, and allow for novel oral treatment options. PMID:25317578

  11. Novel prodrugs with a spontaneous cleavable guanidine moiety.

    PubMed

    Hamada, Yoshio

    2016-04-01

    Water-soluble prodrug strategy is a practical alternative for improving the drug bioavailability of sparingly-soluble drugs with reduced drug efficacy. Many water-soluble prodrugs of sparingly-soluble drugs, such as the phosphate ester of a drug, have been reported. Recently, we described a novel water-soluble prodrug based on O-N intramolecular acyl migration. However, these prodrug approaches require a hydroxy group in the structure of their drugs, and other prodrug approaches are often restricted by the structure of the parent drugs. To develop prodrugs with no restriction in the structure, we focused on a decomposition reaction of arginine methyl ester. This reaction proceeds at room temperature under neutral conditions, and we applied this reaction to the prodrug strategy for drugs with an amino group. We designed and synthesized novel prodrugs of representative sparingly soluble drugs phenytoin and sulfathiazole. Phenytoin and sulfathiazole were obtained as stable salt that were converted to parent drugs under physiological conditions. Phenytoin prodrug 3 showed a short half-life (t1/2) of 13min, whereas sulfathiazole prodrug 7 had a moderate t1/2 of 40min. Prodrugs 3 and 7 appear to be suitable for use as an injectable formulation and orally administered drug, respectively. PMID:26923694

  12. Involvement of SH2-containing phosphotyrosine phosphatase Syp in erythropoietin receptor signal transduction pathways.

    PubMed

    Tauchi, T; Feng, G S; Shen, R; Hoatlin, M; Bagby, G C; Kabat, D; Lu, L; Broxmeyer, H E

    1995-03-10

    Erythropoietin (Epo) regulates the proliferation and differentiation of erythroid precursors. The phosphorylation of proteins at tyrosine residues is critical in the growth signaling induced by Epo. This mechanism is regulated by the activities of both protein-tyrosine kinases and protein tyrosine phosphatases. The discovery of phosphotyrosine phosphatases that contain SH2 domains suggests roles for these molecules in growth factor signaling pathways. We found that Syp, a phosphotyrosine phosphatase, widely expressed in all tissues in mammals became phosphorylated on tyrosine after stimulation with Epo in M07ER cells engineered to express high levels of human EpoR. Syp was complexed with Grb2 in Epo-stimulated M07ER cells. Direct binding between Syp and Grb2 was also observed in vitro. Furthermore, Syp appeared to bind directly to tyrosine-phosphorylated EpoR in M07ER cells. Both NH2-terminal and COOH-terminal SH2 domains of Syp, made as glutathione S-transferase fusion proteins, were able to bind to the tyrosine-phosphorylated EpoR in vitro. These results suggest that Syp may be an important signaling component downstream of the EpoR and may regulate the proliferation and differentiation of hematopoietic cells. PMID:7534299

  13. Estradiol receptor: phosphorylation on tyrosine in uterus and interaction with anti-phosphotyrosine antibody.

    PubMed Central

    Migliaccio, A; Rotondi, A; Auricchio, F

    1986-01-01

    Estradiol receptor from rat uteri incubated with [32P] orthophosphate has been purified by diethylstilbestrol--Sepharose followed by heparin--Sepharose chromatography. The purified receptor, analyzed by centrifugation through sucrose gradients after incubation with monoclonal antibodies against purified estradiol receptor, appears to be labeled with 32P. The receptor preparation has been further purified by immunoaffinity chromatography and submitted to SDS--poly-acrylamide gel electrophoresis. A heavily 32P-labeled 68 kd protein and a very lightly 32P-labeled 48 kd protein, probably a proteolytic product of the 68 kd protein, were detected. Phosphoamino acid analysis of the receptor eluted from the immunoaffinity column shows that its 32P-labeling occurs exclusively on tyrosine. This is the first report on phosphorylation on tyrosine of a steroid receptor in tissue. It is consistent with our previous finding that a uterus estradiol receptor-kinase, which confers hormone binding ability to the estradiol receptor, in vitro phosphorylates this receptor exclusively on tyrosine. Calf uterus receptor binds with high specificity and affinity to monoclonal anti-phosphotyrosine antibodies covalently bound to Sepharose (Kd = 0.28 nM). Dephosphorylation of the receptor by nuclei containing the calf uterus nuclear phosphatase abolishes the interaction with antibodies. These results suggest that also in calf uterus, estradiol receptor is phosphorylated on tyrosine. Anti-phosphotyrosine antibodies bound to Sepharose have been used to partially purify the estradiol receptor from calf uterus. Images Fig. 2. Fig. 3. PMID:2431901

  14. Thin-layer chromatography can resolve phosphotyrosine, phosphoserine, and phosphothreonine in a protein hydrolyzate

    SciTech Connect

    Neufeld, E.; Goren, H.J.; Boland, D. )

    1989-02-15

    A solution of propionic acid, 1 M ammonium hydroxide, and isopropyl alcohol (45/17.5/17.5, v/v) was the ascending solvent in the separation of phosphotyrosine, phosphothreonine, and phosphoserine by thin-layer chromatography. The immobile phase was cellulose. The relative migrations were 0.44, 0.38, and 0.2, respectively. A previously described thin-layer system consisting of isobutyric acid and 0.5 M ammonium hydroxide (50/30, v/v) gave very similar relative migrations. To determine the usefulness of thin-layer chromatography in phosphoamino acid analysis, the propionic acid/ammonium hydroxide/isopropyl alcohol solution was used to characterize phosphorylated residues in a plasma membrane protein which is a substrate for the insulin receptor kinase, in insulin receptor phosphorylated histone H2B, and in an in vivo phosphorylated 90000-Da protein from IM9 cells. {sup 32}P-labeled proteins were separated by dodecyl sulfate-gel electrophoresis, digested with trypsin, and then hydrolyzed with 6 N HCl, 2 h, 110 degrees C. Following thin-layer chromatography of the hydrolyzates and autoradiography, phosphotyrosine was detected in insulin receptor substrates, and phosphoserine and phosphothreonine were found in the in vivo-phosphorylated protein. This study supports previous reports about the practicality of thin-layer chromatography in phosphoamino acid analysis and it demonstrates that a propionic acid, ammonium hydroxide, isoprophyl alcohol solution may be a useful ascending solvent mixture for this purpose.

  15. Antibody-based fluorescent and fluorescent ratiometric indicators for detection of phosphotyrosine.

    PubMed

    Huynh Nhat, Kim Phuong; Watanabe, Takayoshi; Yoshikoshi, Kensuke; Hohsaka, Takahiro

    2016-08-01

    Fluorescent indicators for protein phosphorylation are very important in not only fundamental biology but also biomedical applications. In this study, we developed novel fluorescent and fluorescent ratiometric indicators for detection of phosphotyrosine (pTyr) derivatives. A single-chain antibody variable fragment (scFv) against phosphotyrosine was fluorescent-labeled by incorporation of tetramethylrhodamine (TAMRA)-linked nonnatural amino acid at the N- or C-terminus. The TAMRA-labeled scFv showed fluorescence enhancement upon addition of pTyr-containing peptides based on antigen-dependent fluorescence quenching effect on TAMRA. The TAMRA-labeled scFv was further fused with enhanced green fluorescent protein (EGFP) to generate a double-labeled scFv for pTyr. In the absence of antigen, fluorescence resonance energy transfer (FRET) occurred from EGFP to TAMRA but TAMRA was quenched. The antigen-binding removed the quenching of TAMRA while FRET occurred without altering its efficiency. As a result of the FRET and antigen-dependent fluorescence quenching effect, the double-labeled scFv exhibited fluorescence ratio enhancement upon the antigen-binding. The fluorescent and fluorescent ratiometric indicators obtained in this study will become a novel tool for analysis of protein phosphorylation. Moreover, this strategy utilizes antibody derivatives, and therefore, can be easily applied to other antigen-antibody pairs to generate fluorescent ratiometric indicators for various target molecules. PMID:26896314

  16. A duplex "Gemini" prodrug of naltrexone for transdermal delivery.

    PubMed

    Hammell, Dana C; Hamad, Mohamed; Vaddi, Haranath K; Crooks, Peter A; Stinchcomb, Audra L

    2004-06-18

    Transdermal naltrexone delivery is desirable in the treatment of narcotic dependence and alcoholism. The purpose of this study was to increase the delivery rate of naltrexone (NTX) across human skin by using a novel prodrug. A duplex "gemini" prodrug of naltrexone was synthesized and evaluated. In vitro human skin permeation rates of naltrexone and prodrug were measured using a flow-through diffusion cell system. Drug concentrations in the skin were quantitated at the end of the diffusion experiment. The prodrug was hydrolyzed on passing through the skin and appeared mainly as naltrexone in the receiver compartment. The prodrug provided a significantly higher naltrexone equivalent flux across human skin in vitro than naltrexone base. The naltrexone equivalent solubilities of naltrexone and the prodrug in the donor solution were not significantly different. No significant increase in drug concentration in the skin after prodrug treatment, as compared to naltrexone, was observed. The naltrexone equivalent permeability from the prodrug exceeded the permeability of naltrexone base by two-fold. Due to the design of this prodrug, toxicities associated with this compound should be nonexistent, because only naltrexone and carbon dioxide (carbonic acid) are released when the prodrug is cleaved. PMID:15196755

  17. Peptidomimetic β-Secretase Inhibitors Comprising a Sequence of Amyloid-β Peptide for Alzheimer's Disease.

    PubMed

    Vila-Real, Helder; Coelho, Helena; Rocha, João; Fernandes, Adelaide; Ventura, M Rita; Maycock, Christopher D; Iranzo, Olga; Simplício, Ana L

    2015-07-23

    Alzheimer's disease is a grave social problem in an aging population. A major problem is the passage of drugs through the blood-brain barrier. This work tests the hypothesis that the conjugation of peptidomimetic β-secretase inhibitors with a fragment of amyloid-β peptide facilitates entrance into the central nervous system. HVR-3 (compound 4), one of the conjugation products, was found to be as potent as OM00-3, a known peptidomimetic inhibitor, 4-fold more selective toward β-secretase 1 in relation to β-secretase 2 and 3-fold more resistant to in vitro metabolization in human serum. Its intravenous administration to mice and Wistar rats generated an active metabolite recovered from the rodent's brains. PMID:26061085

  18. Peptide and peptidomimetic ligands for CXC chemokine receptor 4 (CXCR4).

    PubMed

    Oishi, Shinya; Fujii, Nobutaka

    2012-08-14

    The development of novel peptide and peptidomimetic ligands for the CXC chemokine receptor 4 (CXCR4) as therapeutic agents for HIV-1 infection, cancer, and immune system diseases has grown over the last decade. In this perspective article, the design of CXCR4 agonists and antagonists from endogenous stromal cell-derived factor-1 (SDF-1)/CXCL12 and horseshoe crab-derived antimicrobial peptides and their therapeutic and diagnostic applications are described. PMID:22517031

  19. Synthesis of Cyclic Peptidomimetics via a Pd-Catalyzed Macroamination Reaction.

    PubMed

    Hopkins, Brett A; Smith, Graham F; Sciammetta, Nunzio

    2016-08-19

    A new method to access cyclic peptidomimetics via a Pd-catalyzed macroamination reaction is presented. Natural amino acid amines are revealed as proficient coupling partners in these transformations. With a commercially available CPhos G3 catalyst system and substrates bearing diverse amino acid and aryl halide backbones, the unique head to side-chain (or side-chain mimic) macrocycles are afforded with ring sizes from 11 to 23 members in yields up to 84%. PMID:27508926

  20. The Determination of Protonation Constants of Peptidomimetic Cyclophanes in Binary Methanol-Water Mixtures

    PubMed Central

    Tomczyk, Danuta; Andrijewski, Grzegorz

    2016-01-01

    The protonation constants of new group of peptidomimetic cyclophanes with valine or phenylalanine moieties incorporated into the macrocyclic skeleton as well as their linear analogues were determined by potentiometric measurements in solutions of methanol-water mixtures at 25°C and constant ionic strength. The influence of cavity size, location of protonation sites, and attached substituents of the macrocyclic ligands on the protonation constants were discussed on the basis of potentiometric measurement as well as H1-NMR results. PMID:27516918

  1. The Determination of Protonation Constants of Peptidomimetic Cyclophanes in Binary Methanol-Water Mixtures.

    PubMed

    Seliger, Piotr; Tomczyk, Danuta; Andrijewski, Grzegorz; Tomal, Ewa

    2016-01-01

    The protonation constants of new group of peptidomimetic cyclophanes with valine or phenylalanine moieties incorporated into the macrocyclic skeleton as well as their linear analogues were determined by potentiometric measurements in solutions of methanol-water mixtures at 25°C and constant ionic strength. The influence of cavity size, location of protonation sites, and attached substituents of the macrocyclic ligands on the protonation constants were discussed on the basis of potentiometric measurement as well as H(1)-NMR results. PMID:27516918

  2. Discovery of Fluoromethylketone-Based Peptidomimetics as Covalent ATG4B (Autophagin-1) Inhibitors.

    PubMed

    Qiu, Zongxing; Kuhn, Bernd; Aebi, Johannes; Lin, Xianfeng; Ding, Haiyuan; Zhou, Zheng; Xu, Zhiheng; Xu, Danqing; Han, Li; Liu, Cheng; Qiu, Hongxia; Zhang, Yuxia; Haap, Wolfgang; Riemer, Claus; Stahl, Martin; Qin, Ning; Shen, Hong C; Tang, Guozhi

    2016-08-11

    ATG4B or autophagin-1 is a cysteine protease that cleaves ATG8 family proteins. ATG4B plays essential roles in the autophagosome formation and the autophagy pathway. Herein we disclose the design and structural modifications of a series of fluoromethylketone (FMK)-based peptidomimetics as highly potent ATG4B inhibitors. Their structure-activity relationship (SAR) and protease selectivity are also discussed. PMID:27563406

  3. Cyclic alpha-conotoxin peptidomimetic chimeras as potent GLP-1R agonists.

    PubMed

    Swedberg, Joakim E; Schroeder, Christina I; Mitchell, Justin M; Durek, Thomas; Fairlie, David P; Edmonds, David J; Griffith, David A; Ruggeri, Roger B; Derksen, David R; Loria, Paula M; Liras, Spiros; Price, David A; Craik, David J

    2015-10-20

    Type 2 diabetes mellitus (T2DM) results from compromised pancreatic β-cell function, reduced insulin production, and lowered insulin sensitivity in target organs resulting in hyperglycemia. The GLP-1 hormone has two biologically active forms, GLP-1-(7-37) and GLP-1-(7-36)amide, which are equipotent at the glucagon-like peptide-1 receptor (GLP-1R). These peptides are central both to normal glucose metabolism and dysregulation in T2DM. Several structurally modified GLP-1 analogues are now approved drugs, and a number of other analogues are in clinical trials. None of these compounds is orally bioavailable and all require parenteral delivery. Recently, a number of smaller peptidomimetics containing 11-12 natural and unnatural amino acids have been identified that have similar insulin regulating profiles as GLP-1. The α-conotoxins are a class of disulfide rich peptide venoms isolated from cone snails, and are known for their highly constrained structures and resistance to enzymatic degradation. In this study, we examined whether 11-residue peptidomimetics incorporated into α-conotoxin scaffolds, forming monocyclic or bicyclic compounds constrained by disulfide bonds and/or backbone cyclization, could activate the GLP-1 receptor (GLP-1R). Several compounds showed potent (nanomolar) agonist activity at GLP-1R, as evaluated via cAMP signaling. In addition, HPLC retention times and in silico calculations suggested that mono- and bicyclic compounds had more favorable n-octanol/water partition coefficients according to the virtual partition coefficient model (vLogP), while maintaining a smaller radius of gyration compared to corresponding uncyclized peptidomimetics. Our findings suggest that cyclic peptidomimetics provide a potential avenue for future design of potent, compact ligands targeting GLP-1R and possessing improved physicochemical properties. PMID:26352676

  4. Rationally designed peptidomimetic modulators of aβ toxicity in Alzheimer's disease.

    PubMed

    Rajasekhar, K; Suresh, S N; Manjithaya, Ravi; Govindaraju, T

    2015-01-01

    Alzheimer's disease is one of the devastating illnesses mankind is facing in the 21st century. The main pathogenic event in Alzheimer's disease is believed to be the aggregation of the β-amyloid (Aβ) peptides into toxic aggregates. Molecules that interfere with this process may act as therapeutic agents for the treatment of the disease. Use of recognition unit based peptidomimetics as inhibitors are a promising approach, as they exhibit greater protease stability compared to natural peptides. Here, we present peptidomimetic inhibitors of Aβ aggregation designed based on the KLVFF (P1) sequence that is known to bind Aβ aggregates. We improved inhibition efficiency of P1 by introducing multiple hydrogen bond donor-acceptor moieties (thymine/barbiturate) at the N-terminal (P2 and P3), and blood serum stability by modifying the backbone by incorporating sarcosine (N-methylglycine) units at alternate positions (P4 and P5). The peptidomimetics showed moderate to good activity in both inhibition and dissolution of Aβ aggregates as depicted by thioflavin assay, circular dichroism (CD) measurements and microscopy (TEM). The activity of P4 and P5 were studied in a yeast cell model showing Aβ toxicity. P4 and P5 could rescue yeast cells from Aβ toxicity and Aβ aggregates were cleared by the process of autophagy. PMID:25633824

  5. Prodrugs for transdermal drug delivery - trends and challenges.

    PubMed

    Ita, Kevin B

    2016-09-01

    Prodrugs continue to attract significant interest in the transdermal drug delivery field. These moieties can confer favorable physicochemical properties on transdermal drug delivery candidates. Alkyl chain lengthening, pegylation are some of the strategies used for prodrug synthesis. It is usually important to optimize partition coefficient, water and oil solubilities of drugs. In this review, progress made in the field of prodrugs for percutaneous penetration is highlighted and the challenges discussed. PMID:26878159

  6. Synthesis and characterization of novel dipeptide ester prodrugs of acyclovir

    NASA Astrophysics Data System (ADS)

    Nashed, Yasser E.; Mitra, Ashim K.

    2003-07-01

    Four dipeptide (Gly-Gly, Gly-Val, Val-Val, Val-Gly) ester prodrugs of 9-[(2-hydroxyethoxy)methyl]guanine (acyclovir, ACV) were synthesized. LC/MS was used to characterize the new prodrugs. Both 1H NMR and 13C NMR spectra of the four prodrugs of ACV were measured and assigned based on spectral comparison with compounds of similar structures.

  7. [Fospropofol: A new prodrug of propofol].

    PubMed

    Telletxea, S; Lauzirika, Z; Etxebarria, A; Ortega, L F

    2012-11-01

    The development of new propofol formulations has advanced rapidly in the last ten years with the achievement of the marketing a new prodrug of propofol: fospropofol, pharmacologically different from the original compound. It is a water soluble compound that requires metabolism of the prodrug to propofol, which leads to a time delay between its administration and the appearance of its pharmacological effect. Its pharmacokinetic and pharmacodynamic characteristics are different to the original formula. Due to its formulation it does not cause pain on intravenous injection, does not lead to hyperlipidaemia or excess bacterial growth. Although it is currently unavailable in Spain, it has been approved by the FDA (American Food and Drug Administration) for sedation in controlled care in diagnostic and therapeutic procedures in adults. It must only be administered by personnel qualified to administer anaesthesia, and the patients must be monitored throughout the whole procedure. PMID:22748853

  8. Lipid prodrug nanocarriers in cancer therapy.

    PubMed

    Mura, Simona; Bui, Duc Trung; Couvreur, Patrick; Nicolas, Julien

    2015-06-28

    Application of nanotechnology in the medical field (i.e., nanomedicine) plays an important role in the development of novel drug delivery methods. Nanoscale drug delivery systems can indeed be customized with specific functionalities in order to improve the efficacy of the treatments. However, despite the progresses of the last decades, nanomedicines still face important obstacles related to: (i) the physico-chemical properties of the drug moieties which may reduce the total amount of loaded drug; (ii) the rapid and uncontrolled release (i.e., burst release) of the encapsulated drug after administration and (iii) the instability of the drug in biological media where a fast transformation into inactive metabolites can occur. As an alternative strategy to alleviate these drawbacks, the prodrug approach has found wide application. The covalent modification of a drug molecule into an inactive precursor from which the drug will be freed after administration offers several benefits such as: (i) a sustained drug release (mediated by chemical or enzymatic hydrolysis of the linkage between the drug-moiety and its promoiety); (ii) an increase of the drug chemical stability and solubility and, (iii) a reduced toxicity before the metabolization occurs. Lipids have been widely used as building blocks for the design of various prodrugs. Interestingly enough, these lipid-derivatized drugs can be delivered through a nanoparticulate form due to their ability to self-assemble and/or to be incorporated into lipid/polymer matrices. Among the several prodrugs developed so far, this review will focus on the main achievements in the field of lipid-based prodrug nanocarriers designed to improve the efficacy of anticancer drugs. Gemcitabine (Pubchem CID: 60750); 5-fluorouracil (Pubchem CID: 3385); Doxorubicin (Pubchem CID: 31703); Docetaxel (Pubchem CID: 148124); Methotrexate (Pubchem CID: 126941); Paclitaxel (Pubchem CID: 36314). PMID:25617724

  9. Phosphotyrosine enrichment identifies focal adhesion kinase and other tyrosine kinases for targeting in canine hemangiosarcoma.

    PubMed

    Marley, K; Maier, C S; Helfand, S C

    2012-09-01

    Canine hemangiosarcoma (HSA) is an endothelial cell malignancy driven, in part, by activating mutations in receptor and non-receptor tyrosine kinases. Proteomics, Western blots and a tyrosine kinase inhibitor were used to elucidate activating mechanisms in HSA cell lines. Phosphotyrosine peptides from focal adhesion kinase (FAK) STAT3, Lyn, Fyn and other signal transduction kinases were identified by mass spectrometry. FAK was constitutively activated at tyrosine 397, the autophosphorylation site, and this was reversible with high concentrations of a FAK inhibitor. FAK inhibitor-14 suppressed migration and phosphorylation of FAK tyrosine 397 and tyrosines 576/577 and was cytotoxic to HSA cells suggesting FAK signalling may be an important contributor to canine HSA survival. PMID:22487216

  10. Purification, crystallization and preliminary X-ray diffraction analysis of the Fyn SH2 domain and its complex with a phosphotyrosine peptide

    PubMed Central

    Huculeci, Radu; Buts, Lieven; Lenaerts, Tom; van Nuland, Nico A. J.; Garcia-Pino, Abel

    2012-01-01

    SH2 domains are widespread protein-binding modules that recognize phosphotyrosines and play central roles in intracellular signalling pathways. The SH2 domain of the human protein tyrosine kinase Fyn has been expressed, purified and crystallized in the unbound state and in complex with a high-affinity phosphotyrosine peptide. X-ray data were collected to a resolution of 2.00 Å for the unbound form and 1.40 Å for the protein in complex with the phosphotyrosine peptide. PMID:22442244

  11. Prodrugs of anthracyclines for use in antibody-directed enzyme prodrug therapy.

    PubMed

    Florent, J C; Dong, X; Gaudel, G; Mitaku, S; Monneret, C; Gesson, J P; Jacquesy, J C; Mondon, M; Renoux, B; Andrianomenjanahary, S; Michel, S; Koch, M; Tillequin, F; Gerken, M; Czech, J; Straub, R; Bosslet, K

    1998-09-10

    A series of new prodrugs of daunorubicin and doxorubicin which are candidates for antibody-directed enzyme prodrug therapy (ADEPT) is reported. These compounds (25a,b,c and 32a,b,c) have been designed to generate cytotoxic drugs after activation with beta-glucuronidase. As expected, recovery of the active drug was observed after enzymatic cleavage by Escherichia coli beta-glucuronidase as well as by a fusion protein which has been obtained from human beta-glucuronidase and humanized CEA-specific binding region. The six prodrugs are highly stable and are more than 100-fold less cytotoxic than doxorubicin against murine L1210 cell lines. The ortho-substituted phenyl carbamates 25a,b,c are better substrates for beta-glucuronidase than the corresponding para-substituted analogues. After taking into account additional factors such as stability in plasma and kinetics of enzymatic cleavage, we selected the o-nitro prodrug 25c for clinical trials. PMID:9733483

  12. Synthesis, hydrolysis kinetics and pharmacological evaluation of aceclofenac prodrugs.

    PubMed

    Dhokchawle, Bharat V; Bhandari, Anil B

    2015-01-01

    The mutual prodrugs of aceclofenac with various naturally available antioxidants; menthol, thymol, eugenol, guiacol and vanillin have been synthesized by the DCC coupling method, purified and characterized by spectral data, as well as, partition coefficient, solubility and hydrolytic studies. The title compounds have more lipophilic character as compared to the parent moieties and good stability in acidic environment, which is prerequisite for the oral absorption of the drug. Under gastric as well as intestinal pH conditions these prodrugs showed variable susceptibility towards hydrolysis. The synthesized derivatives were evaluated for antiinflammatory, analgesic activities and ulcerogenic potential. Prodrug showed improved solubility in organic solvents, which implies lipophilic character of ester prodrugs and were also found to be chemically stable in acidic environment. The aceclofenac mutual prodrugs showed improved analgesic and anti-inflammatory activities with reduced ulcerogenicity. PMID:25403255

  13. In silico studies, synthesis and binding evaluation of substituted 2-pyrrolidinones as peptidomimetics of RGD tripeptide sequence.

    PubMed

    Toum, Valérie; Bolley, Julie; Lalatonne, Yoann; Barbey, Carole; Motte, Laurence; Lecouvey, Marc; Royer, Jacques; Dupont, Nathalie; Pérard-Viret, Joëlle

    2015-03-26

    In silico optimisation, synthesis and binding evaluation of αvβ3 integrin's affinity for precursors of a new RGD peptidomimetics family are presented. The 2-pyrrolidinone building block was obtained by condensation of l-lysine with dimethoxydihydrofuran followed by reduction. The ring was functionalized with a carboxylic acid and a guanidinium appendage. On the pyrrolidinone heterocycle, the effects on affinity of position, length and relative geometry of the two acid or basic functionalized side chains introduced on the pyrrolidinone ring have been previously evaluated by docking studies. Peptidomimetics have finally been evaluated by competition binding assays for αvβ3 integrin's affinity using radio-ligands. PMID:25721024

  14. Development of macromolecular prodrug for rheumatoid arthritis☆

    PubMed Central

    Yuan, Fang; Quan, Ling-dong; Cui, Liao; Goldring, Steven R.; Wang, Dong

    2012-01-01

    Rheumatoid arthritis (RA) is a chronic autoimmune disease that is considered to be one of the major public health problems worldwide. The development of therapies that target tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6) and co-stimulatory pathways that regulate the immune system have revolutionized the care of patients with RA. Despite these advances, many patients continue to experience symptomatic and functional impairment. To address this issue, more recent therapies that have been developed are designed to target intracellular signaling pathways involved in immunoregulation. Though this approach has been encouraging, there have been major challenges with respect to off-target organ side effects and systemic toxicities related to the widespread distribution of these signaling pathways in multiple cell types and tissues. These limitations have led to an increasing interest in the development of strategies for the macromolecularization of anti-rheumatic drugs, which could target them to the inflamed joints. This approach enhances the efficacy of the therapeutic agent with respect to synovial inflammation, while markedly reducing non-target organ adverse side effects. In this manuscript, we provide a comprehensive overview of the rational design and optimization of macromolecular prodrugs for treatment of RA. The superior and the sustained efficacy of the prodrug may be partially attributed to their Extravasation through Leaky Vasculature and subsequent Inflammatory cell-mediated Sequestration (ELVIS) in the arthritic joints. This biologic process provides a plausible mechanism, by which macromolecular prodrugs preferentially target arthritic joints and illustrates the potential benefits of applying this therapeutic strategy to the treatment of other inflammatory diseases. PMID:22433784

  15. Structure-activity Relationships of Peptidomimetics that Inhibit PPI of HER2-HER3

    PubMed Central

    Kanthala, Shanthi; Gauthier, Ted; Satyanarayanajois, Seetharama

    2014-01-01

    Human epidermal growth factor receptor-2 (HER2) is a tyrosine kinase family protein receptor that is known to undergo heterodimerization with other members of the family of epidermal growth factor receptors (EGFR) for cell signaling. Overexpression of HER2 and deregulation of signaling has implications in breast, ovarian, and lung cancers. We have designed several peptidomimetics to block the HER2-mediated dimerization, resulting in antiproliferative activity for cancer cells. In the present work we have investigated the structure-activity relationships of peptidomimetic analogs of compound 5. Compound 5 was conformationally constrained by N- and C-terminal modification and cyclization as well as by substitution with D-amino acids at the N-and C-termini. Among the compounds studied in this work, a peptidomimetic compound 21 with D-amino acid substitution and its N- and C-termini capped with acetyl and amide functional groups and a reversed sequence compared to that of compound 5 exhibited better antiproliferative activity in HER2-overexpressed breast, ovarian, and lung cancer cell lines. Compound 21 was further evaluated for its protein-protein interaction (PPI) inhibition ability using enzyme fragment complementation (EFC) assay, proximity ligation assay (PLA), and Western blot analysis. Results suggested that compound 21 is able to block HER2:HER3 interaction and inhibit phosphorylation of the kinase domain of HER2. The mode of binding of compound 21 to HER2 protein was modeled using a docking method. Compound 21 seems to bind to domain IV of HER2 near the PPI site of EGFR:HER2 and HER:HER3 and inhibit PPI. PMID:24222531

  16. Polyethylene terephthalate membrane grafted with peptidomimetics: endothelial cell compatibility and retention under shear stress.

    PubMed

    Rémy, Murielle; Bareille, Reine; Rerat, Vincent; Bourget, Chantal; Marchand-Brynaert, Jacqueline; Bordenave, Laurence

    2013-01-01

    The present work aimed to treat a polyethylene terephthalate (PET) surface to make the biomaterial more 'attractive' in terms of attachment and shear stress response to endothelial cells with a view to possible applications in vascular grafting. A surface wet-chemistry protocol was applied to graft track-etched PET membranes with RGD peptidomimetics based on the tyrosine template and active at the nano-level vs. isolated human αvβ3 receptor, which was monitored by X-ray photoelectron spectroscopy, contact angle measurement and atomic force microscopy for characterization. A primary culture of human saphenous vein endothelial cells was used before and after sterilization of the membranes (heat treatment or γ-ray irradiation) to test the benefit of grafting. The optimal surface concentrations of grafted molecules were around 50 pmol/cm². Compared to GRGDS, the peptidomimetics promoted cell attachment with similar or slightly better performances. Endothelialized grafted supports were further exposed to 2 h of shear stress mimicking arterial conditions. Cells were lost on non-grafted PET whereas cells on grafted polymers sterilized by γ-ray irradiation withstood forces with no significant difference in focal contacts. At the mRNA level, cells on functionalized PET were able to respond to shear stress with NFkB upregulation. Thus, grafting of peptidomimetics as ligands of the αvβ3 integrin could be a relevant strategy to improve the adhesion of human endothelial cells and to obtain an efficient endothelialized PET for the surgery of small-diameter vascular prostheses. PMID:23565647

  17. A mechanistic study on the inhibition of α-chymotrypsin by a macrocyclic peptidomimetic aldehyde.

    PubMed

    Zhang, X; Bruning, J B; George, J H; Abell, A D

    2016-08-01

    Here we describe an NMR and X-ray crystallography-based characterisation of the mechanism by which a new class of macrocyclic peptidomimetic aldehyde inhibits α-chymotrypsin. In particular, a (13)C-labelled analogue of the inhibitor was prepared and used in NMR experiments to confirm formation of a hemiacetal intermediate on binding with α-chymotrypsin. Analysis of an X-ray crystallographic structure in complex with α-chymotrypsin reveals that the backbone adopts a stable β-strand conformation as per its design. Binding is further stabilised by interaction with the oxyanion hole near the S1 subsite and multiple hydrogen bonds. PMID:27349772

  18. The Phosphotyrosine Interactome of the Insulin Receptor Family and Its Substrates IRS-1 and IRS-2*S⃞

    PubMed Central

    Hanke, Stefan; Mann, Matthias

    2009-01-01

    The insulin signaling pathway is critical in regulating glucose levels and is associated with diabetes, obesity, and longevity. A tyrosine phosphorylation cascade creates docking sites for protein interactions, initiating subsequent propagation of the signal throughout the cell. The phosphotyrosine interactome of this medically important pathway has not yet been studied comprehensively. We therefore applied quantitative interaction proteomics to exhaustively profile all potential phosphotyrosine-dependent interaction sites in its key players. We targeted and compared insulin receptor substrates 1 and 2 (IRS-1 and IRS-2) as central distributors of the insulin signal, the insulin receptor, the insulin-like growth factor 1 receptor, and the insulin receptor-related receptor. Using the stable isotope labeling by amino acids in cell culture (SILAC) approach with phosphorylated versus non-phosphorylated bait peptides, we found phosphorylation-specific interaction partners for 52 out of 109 investigated sites. In addition, doubly and triply phosphorylated motifs provided insight into the combinatorial effects of phosphorylation events in close proximity to each other. Our results retrieve known interactions and substantially broaden the spectrum of potential interaction partners of IRS-1 and IRS-2. A large number of common interactors rationalize their extensive functional redundancy. However, several proteins involved in signaling and metabolism interact differentially with IRS-1 and IRS-2 and thus provide leads into their different physiological roles. Differences in interactions at the receptor level are reflected in multisite recruitment of SHP2 by the insulin-like growth factor 1 receptor and limited but exclusive interactions with the IRR. In common with other recent reports, our data furthermore hint at non-SH2 or phosphotyrosine-binding domain-mediated phosphotyrosine binding. PMID:19001411

  19. Insulin-induced tyrosine dephosphorylation of paxillin and focal adhesion kinase requires active phosphotyrosine phosphatase 1D.

    PubMed Central

    Ouwens, D M; Mikkers, H M; van der Zon, G C; Stein-Gerlach, M; Ullrich, A; Maassen, J A

    1996-01-01

    Insulin stimulation of fibroblasts rapidly induces the tyrosine dephosphorylation of proteins of 68 kDa and 125 kDa, in addition to the tyrosine phosphorylation of the insulin receptor beta-chain, insulin receptor substrates 1 and 2, and Shc. Using specific antibodies, the 68 kDa and 125 kDa proteins were identified as paxillin and focal adhesion kinase (pp125FAK) respectively. We have examined whether dephosphorylation of paxillin and pp125FAK requires interaction of the cells with the extracellular matrix. For this, cells were grown on poly(L-lysine) plates, and the tyrosine phosphorylation of pp125FAK and paxillin was increased by addition of lysophosphatidic acid. Under these conditions, insulin still induced the complete dephosphorylation of pp125FAK and paxillin, indicating that this process can occur independently of the interaction of integrins with extracellular matrix proteins. We also studied whether dephosphorylation of pp125FAK and paxillin results from the action of a phosphotyrosine phosphatase. It was found that phenylarsine oxide, a phosphotyrosine phosphatase inhibitor, prevented the insulin-induced dephosphorylation of pp125FAK and paxillin. Furthermore, this insulin-induced dephosphorylation was also impaired in cells expressing a dominant-negative mutant of phosphotyrosine phosphatase 1D (PTP 1D). Thus we have identified paxillin as a target for dephosphorylation by insulin. In addition, we have obtained evidence that the insulin-mediated dephosphorylation of paxillin and pp125FAK requires active PTP 1D. PMID:8809054

  20. Prodrugs of perzinfotel with improved oral bioavailability.

    PubMed

    Baudy, Reinhardt B; Butera, John A; Abou-Gharbia, Magid A; Chen, Hong; Harrison, Boyd; Jain, Uday; Magolda, Ronald; Sze, Jean Y; Brandt, Michael R; Cummons, Terri A; Kowal, Diane; Pangalos, Menelas N; Zupan, Bojana; Hoffmann, Matthew; May, Michael; Mugford, Cheryl; Kennedy, Jeffrey; Childers, Wayne E

    2009-02-12

    Previous studies with perzinfotel (1), a potent, selective, competitive NMDA receptor antagonist, showed it to be efficacious in inflammatory and neuropathic pain models. To increase the low oral bioavailability of 1 (3-5%), prodrug derivatives (3a-h) were synthesized and evaluated. The oxymethylene-spaced diphenyl analogue 3a demonstrated good stability at acidic and neutral pH, as well as in simulated gastric fluid. In rat plasma, 3a was rapidly converted to 1 via 2a. Pharmacokinetic studies indicated that the amount of systemic exposure of 1 produced by a 10 mg/kg oral dose of 3a was 2.5-fold greater than that produced by a 30 mg/kg oral dose of 1. Consistent with these results, 3a was significantly more potent and had a longer duration of activity than 1 following oral administration in a rodent model of inflammatory pain. Taken together, these results demonstrate that an oxymethylene-spaced prodrug approach increased the bioavailability of 1. PMID:19146418

  1. 10-Boronic acid substituted camptothecin as prodrug of SN-38.

    PubMed

    Wang, Lei; Xie, Shao; Ma, Longjun; Chen, Yi; Lu, Wei

    2016-06-30

    Malignant tumor cells have been found to have high levels of reactive oxygen species such as hydrogen peroxide (H2O2), supporting the hypothesis that a prodrug could be activated by intracellular H2O2 and lead to a potential antitumor therapy. In this study, the 7-ethyl-10-boronic acid camptothecin (B1) was synthesized for the first time as prodrug of SN-38, by linking a cleavable aryl carbon-boron bond to the SN-38. Prodrug B1 selectively activated by H2O2, converted rapidly to the active form SN-38 under favorable oxidative conditions in cancer cells with elevated levels of H2O2. The cell survival assay showed that prodrug B1 was equally or more effective in inhibiting the growth of six different cancer cells, as compared to SN-38. Unexpectedly, prodrug B1 displayed even more potent Topo I inhibitory activity than SN-38, suggesting that it was not only a prodrug of SN-38 but also a typical Topo I inhibitor. Prodrug B1 also demonstrated a significant antitumor activity at 2.0 mg/kg in a xenograft model using human brain star glioblastoma cell lines U87MG. PMID:27060760

  2. Discovery of Novel Peptidomimetics as Irreversible CHIKV NsP2 Protease Inhibitors Using Quantum Mechanical-Based Ligand Descriptors.

    PubMed

    El-Labbad, Eman M; Ismail, Mohammed A H; Abou Ei Ella, Dalal A; Ahmed, Marawan; Wang, Feng; Barakat, Khaled H; Abouzid, Khaled A M

    2015-12-01

    Chikungunya virus (CHIKV) is a mosquito-borne alphavirus. Recent outbreaks of CHIKV infections have been reported in Asia, Africa, and Europe. The symptoms of CHIKV infection include fever, headache, nausea, vomiting, myalgia, rash, and chronic persistent arthralgia. To date, no vaccines or selective antiviral drugs against this important emerging virus have been reported. In this study, the design, synthesis, and antiviral activity screening of new topographical peptidomimetics revealed three potential prototype agents 3a, 4b, and 5d showing 93-100% maximum inhibition of CHIKV replication in cell-based assay having EC90 of 8.76-9.57 μg/mL. Intensive molecular modeling studies including covalent docking, lowest unoccupied molecular orbital energies, and the atomic condensed Fukui functions calculations strongly suggested the covalent binding of peptidomimetics 3a, 4b, and 5d to CHIKV nsP2 protease leading to permanent enzyme inactivation via Michael adduct formation between α/β-unsaturated ketone functionality in our designed peptidomimetics and active site catalytic cysteine1013. Furthermore, small molecular weight peptidomimetics 3a and 4b satisfied the Lipinski rule of five for drug-likeness and showed promising intestinal absorption and aqueous solubility via computational admet studies making them promising hits for further optimization. PMID:26212366

  3. Design, synthesis and characterization of peptidomimetic conjugate of BODIPY targeting HER2 protein extracellular domain

    PubMed Central

    Banappagari, Sashikanth; McCall, Alecia; Fontenot, Krystal; Vicente, M. Graca H.; Gujar, Amit; Satyanarayanajois, Seetharama

    2013-01-01

    Among the EGFRs, HER2 is a major heterodimer partner and also has important implications in the formation of particular tumors. Interaction of HER2 protein with other EGFR proteins can be modulated by small molecule ligands and, hence, these protein-protein interactions play a key role in biochemical reactions related to control of cell growth. A peptidomimetic (compound 5-1) that binds to HER2 protein extracellular domain and inhibits protein-protein interactions of EGFRs was conjugated with BODIPY (4,4-difluoro-5,7-dimethyl-4-bora-3a,4a-diaza-s-indacene). Conjugation of BODIPY to the peptidomimetic was investigated by different approaches. The conjugate was characterized for its ability to bind to HER2 overexpressing SKBR-3 and BT-474 cells. Furthermore, cellular uptake of conjugate of BODIPY was studied in the presence of membrane tracker and Lyso tracker using confocal microscopy. Our results suggested that fluorescently labeled compound 5-7 binds to the extracellular domain and stays in the membrane for nearly 24 h. After 24 h there is an indication of internalization of the conjugate. Inhibition of protein-protein interaction and downstream signaling effect of compound 5-1 was also studied by proximity ligation assay and western blot analysis. Results suggested that compound 5-1 inhibits protein-protein interactions of HER2-HER3 and phosphorylation of HER2 in a time-dependent manner. PMID:23688700

  4. An Experimental-Theoretical Analysis of Protein Adsorption on Peptidomimetic Polymer Brushes

    PubMed Central

    Lau, K.H. Aaron; Ren, Chunlai; Park, Sung Hyun; Szleifer, Igal; Messersmith, Phillip B.

    2012-01-01

    Surface-grafted water soluble polymer brushes are being intensely investigated for preventing protein adsorption to improve biomedical device function, prevent marine fouling, and enable applications in biosensing and tissue engineering. In this contribution, we present an experimental-theoretical analysis of a peptidomimetic polymer brush system with regard to the critical brush density required for preventing protein adsorption at varying chain lengths. A mussel adhesive-inspired DOPA-Lys pentapeptide surface grafting motif enabled aqueous deposition of our peptidomimetic polypeptoid brushes over a wide range of chain densities. Critical densities of 0.88 nm−2 for a relatively short polypeptoid 10-mer to 0.42 nm−2 for a 50-mer were identified from measurements of protein adsorption. The experiments were also compared with the protein adsorption isotherms predicted by a molecular theory. Excellent agreements in terms of both the polymer brush structure and the critical chain density were obtained. Furthermore, atomic force microscopy (AFM) imaging is shown to be useful in verifying the critical brush density for preventing protein adsorption. The present co-analysis of experimental and theoretical results demonstrates the significance of characterizing the critical brush density in evaluating the performance of an anti-fouling polymer brush system. The high fidelity of the agreement between the experiments and molecular theory also indicate that the theoretical approach presented can aid in the practical design of antifouling polymer brush systems. PMID:22107438

  5. An Antimicrobial Peptidomimetic Induces Mucorales Cell Death through Mitochondria-Mediated Apoptosis

    PubMed Central

    Barbu, E. Magda; Shirazi, Fazal; McGrath, Danielle M.; Albert, Nathaniel; Sidman, Richard L.; Pasqualini, Renata; Arap, Wadih; Kontoyiannis, Dimitrios P.

    2013-01-01

    The incidence of mucormycosis has dramatically increased in immunocompromised patients. Moreover, the array of cellular targets whose inhibition results in fungal cell death is rather limited. Mitochondria have been mechanistically identified as central regulators of detoxification and virulence in fungi. Our group has previously designed and developed a proteolytically-resistant peptidomimetic motif D(KLAKLAK)2 with pleiotropic action ranging from targeted (i.e., ligand-directed) activity against cancer and obesity to non-targeted activity against antibiotic resistant gram-negative rods. Here we evaluated whether this non-targeted peptidomimetic motif is active against Mucorales. We show that D(KLAKLAK)2 has marked fungicidal action, inhibits germination, and reduces hyphal viability. We have also observed cellular changes characteristic of apoptosis in D(KLAKLAK)2-treated Mucorales cells. Moreover, the fungicidal activity was directly correlated with vacuolar injury, mitochondrial swelling and mitochondrial membrane depolarization, intracellular reactive oxygen species accumulation (ROS), and increased caspase-like enzymatic activity. Finally, these apoptotic features were prevented by the addition of the ROS scavenger N-acetyl-cysteine indicating mechanistic pathway specificity. Together, these findings indicate that D(KLAKLAK)2 makes Mucorales exquisitely susceptible via mitochondrial injury-induced apoptosis. This prototype may serve as a candidate drug for the development of translational applications against mucormycosis and perhaps other fungal infections. PMID:24098573

  6. An antimicrobial peptidomimetic induces Mucorales cell death through mitochondria-mediated apoptosis.

    PubMed

    Barbu, E Magda; Shirazi, Fazal; McGrath, Danielle M; Albert, Nathaniel; Sidman, Richard L; Pasqualini, Renata; Arap, Wadih; Kontoyiannis, Dimitrios P

    2013-01-01

    The incidence of mucormycosis has dramatically increased in immunocompromised patients. Moreover, the array of cellular targets whose inhibition results in fungal cell death is rather limited. Mitochondria have been mechanistically identified as central regulators of detoxification and virulence in fungi. Our group has previously designed and developed a proteolytically-resistant peptidomimetic motif D(KLAKLAK)2 with pleiotropic action ranging from targeted (i.e., ligand-directed) activity against cancer and obesity to non-targeted activity against antibiotic resistant gram-negative rods. Here we evaluated whether this non-targeted peptidomimetic motif is active against Mucorales. We show that D(KLAKLAK)2 has marked fungicidal action, inhibits germination, and reduces hyphal viability. We have also observed cellular changes characteristic of apoptosis in D(KLAKLAK)2-treated Mucorales cells. Moreover, the fungicidal activity was directly correlated with vacuolar injury, mitochondrial swelling and mitochondrial membrane depolarization, intracellular reactive oxygen species accumulation (ROS), and increased caspase-like enzymatic activity. Finally, these apoptotic features were prevented by the addition of the ROS scavenger N-acetyl-cysteine indicating mechanistic pathway specificity. Together, these findings indicate that D(KLAKLAK)2 makes Mucorales exquisitely susceptible via mitochondrial injury-induced apoptosis. This prototype may serve as a candidate drug for the development of translational applications against mucormycosis and perhaps other fungal infections. PMID:24098573

  7. Role of Phosphotyrosine Interaction Domain Containing 1 in Porcine Intramuscular Preadipocyte Proliferation and Differentiation.

    PubMed

    Chen, Xiaoling; Luo, Yanliu; Huang, Zhiqing; Jia, Gang; Liu, Guangmang; Zhao, Hua

    2016-10-01

    Phosphotyrosine interaction domain containing 1 (PID1), a recently identified gene involved in obesity-associated insulin resistance, plays an important role in fat deposition. However, its effect on porcine intramuscular preadipocyte proliferation and differentiation remains poorly understood. In this study, the plasmid pcDNA3.1(+)-pPID1 was transfected into porcine intramuscular preadipocytes with Lipofectamine 3000 reagent to over-express porcine PID1 (pPID1). Over-expression of pPID1 significantly promoted porcine intramuscular preadipocyte proliferation. Expression of pPID1 mRNA was significantly increased upon porcine intramuscular preadipocyte differentiation. Indirect fluorescent immunocytochemistry demonstrated that pPID1 protein was localized predominantly in the nucleus of porcine intramuscular preadipocyte. The mRNA levels of peroxisome proliferators-activated receptor γ, CCAAT/enhancer binding protein α and lipoprotein lipase were significantly increased by pPID1 over-expression. Over-expression of pPID1 also led to an increase in lipid accumulation which was detected by Oil Red O staining, and significantly increased the intramuscular triacylglycerol content. These results indicate that pPID1 may play a role in enhancing porcine intramuscular preadipocyte proliferation and differentiation. PMID:27565873

  8. Phosphotyrosine-dependent targeting of mitogen-activated protein kinase in differentiated contractile vascular cells.

    PubMed

    Khalil, R A; Menice, C B; Wang, C L; Morgan, K G

    1995-06-01

    Tyrosine phosphorylation has been linked to plasmalemmal targeting of src homology-2-containing proteins, activation of mitogen-activated protein (MAP) kinase, nuclear signaling, and proliferation of cultured cells. Significant tyrosine phosphorylation and MAP kinase activities have also been reported in differentiated cells, but the signaling role of tyrosine-phosphorylated MAP kinase in these cells is unclear. The spatial and temporal relation between phosphotyrosine and MAP kinase immunoreactivity was quantified in differentiated contractile vascular smooth muscle cells by using digital imaging microscopy. An initial association of MAP kinase with the plasmalemma required upstream protein kinase C activity but occurred in a tyrosine phosphorylation-independent manner. Subsequent to membrane association, a delayed redistribution of MAP kinase, colocalizing with the actin-binding protein caldesmon, occurred in a tyrosine phosphorylation-dependent manner. The apparent association of MAP kinase with the contractile proteins coincided with contractile activation. Thus, tyrosine phosphorylation appears to target MAP kinase to cytoskeletal proteins in contractile vascular cells. This targeting mechanism may determine the specific destination and thereby the specialized function of MAP kinase in other phenotypes. PMID:7538916

  9. Phosphotyrosine in proteins: chemical and immunological identification and isolation of phosphotyrosyl proteins

    SciTech Connect

    Martensen, T.M.

    1986-05-01

    Proteins which contain phosphotyrosine (Tyr-P) residues can be identified and purified by utilizing the chemical and immunological properties of the Tyr-P moiety. The stability of Tyr-P to basic conditions which cleave most ser/thr phosphoryl bonds enabled the analysis of cellular phosphoproteins after Na dodecyl SO/sub 4/ gel electrophoresis. Analyses were simplified using a nylon electroblot of the gel which is stable to 1 hr incubation in 1 N NaOH at 65/sup 0/C. Autoradiograms of nylon blots before and after base treatment were performed on /sup 32/P-labeled phosphoproteins of normal and retrovirus transformed fibroblasts. Conditions for optimal labeling of cellular proteins were studied by protein tyrosine kinase activity measurements using poly glu/sub 4/tyr. Differences in base resistant phosphoprotein patterns could be seen for each transformed cell line. Confirmation that certain base resistant phosphoproteins contained Tyr-P was achieved by immunostaining with affinity-purified sheep anti-Tyr-P antibodies (Ab). These Ab served as useful reagents to study Tyr-P proteins since they inhibit their dephosphorylation by phosphatases, and precipitate them in the presence of rabbit anti-sheep IgG Ab. Tyr-P protein was immobilized when mixed with anti-Tyr-P Ab and incubated with immobilized protein A sepharose rabbit anti-sheep IgG Ab complex. Selective elution of the Tyr-P protein was achieved with free Tyr-P.

  10. A macromolecular prodrug strategy for combinatorial drug delivery.

    PubMed

    Li, Nan-Nan; Lin, Jiantao; Gao, Di; Zhang, Li-Ming

    2014-03-01

    A novel macromolecular prodrug strategy was developed for the combinatorial delivery of two poorly water-soluble drugs, dexamethasone and doxorubicin. In this work, dexamethasone was firstly conjugated onto a water-soluble modified polysaccharide by an acid-labile hydrazone linkage. The resultant macromolecular prodrug had an amphiphilic character and could self-assemble into spherical polymeric micelles in aqueous system. With these micelles, doxorubicin was then encapsulated into their hydrophobic cores. For the conjugated dexamethasone and encapsulated doxorubicin, they could exhibit independent and acid-sensitive release characteristics. For the doxorubicin-loaded prodrug micelles, they were easily be internalized by living cells and showed obvious antitumor activity. PMID:24407691

  11. N,N'-Dialkylaminoalkylcarbonyl (DAAC) prodrugs and aminoalkylcarbonyl (AAC) prodrugs of 4-hydroxyacetanilide and naltrexone with improved skin permeation properties.

    PubMed

    Devarajan-Ketha, H; Sloan, K B

    2011-07-01

    N,N'-Dialkylaminoalkylcarbonyl (DAAC) and aminoalkylcarbonyl (AAC) prodrugs of phenolic drugs acetaminophen (APAP) and naltrexone (NTX) are reported. The effects of incorporation of a basic amine group into the promoiety of an acyl prodrug of a phenolic drug on its skin permeation properties are also presented. DAAC-APAP prodrugs were synthesized via a three-step procedure starting with haloalkylcarbonyl esters which were reacted with five different amines: dimethylamine, diethylamine, dipropylamine, morpholine, and piperidine. The spacing between the amino group and the carbonyl group of the acyl group was 1-3 CH(2). After the hydrolysis of the ester, the carboxylic acid product was subsequently coupled with the parent drug via a dicyclohexyl carbodiimide (DCC) mediated coupling to yield the DAAC-APAP-HCl prodrugs in excellent yields. The AAC prodrugs were synthesized using commercially available Boc-protected amino acids using DCC or EDCI as coupling agents. The yields of the prodrugs synthesized using these two different methods have been compared. Half-lives (t(1/2)) of a few members of the DAAC and AAC series were measured in buffer (pH 6.0, 20mM). The members evaluated in hydrolysis experiments exhibit a t(1/2) range of 15-113min. Among AAC-APAP prodrugs, the isopropyl group in valinate-APAP-HCl exerted a steric effect that increased the t(1/2) value for this prodrug compared to alaninate-APAP-HCl or prolinate-APAP-HCl. The 2-morpholinylacetate-APAP prodrug was able to achieve twice the flux of APAP in in vitro diffusion cell experiments through hairless mouse skin. PMID:21616664

  12. Platinum(iv) anticancer prodrugs - hypotheses and facts.

    PubMed

    Gibson, Dan

    2016-08-16

    In this manuscript we focus on Pt(iv) anticancer prodrugs. We explore the main working hypotheses for the design of effective Pt(iv) prodrugs and note the exceptions to the common assumptions that are prevalent in the field. Special attention was devoted to the emerging class of "dual action" Pt(iv) prodrugs, where bioactive ligands are conjugated to the axial positions of platinum in order to obtain orthogonal or complementary effects that will increase the efficacy of killing the cancer cells. We discuss the rationale behind the design of the "dual action" prodrugs and the results of the pharmacological studies obtained. Simultaneous release of two bioactive moieties inside the cancer cells often triggers several processes that together determine the fate of the cell. Pt(iv) complexes provide many opportunities for applying new concepts in targeting, synergistic cell killing and exploiting novel nanodelivery systems. PMID:27214873

  13. Bioreductive prodrugs as cancer therapeutics: targeting tumor hypoxia

    PubMed Central

    Guise, Christopher P.; Mowday, Alexandra M.; Ashoorzadeh, Amir; Yuan, Ran; Lin, Wan-Hua; Wu, Dong-Hai; Smaill, Jeff B.; Patterson, Adam V.; Ding, Ke

    2014-01-01

    Hypoxia, a state of low oxygen, is a common feature of solid tumors and is associated with disease progression as well as resistance to radiotherapy and certain chemotherapeutic drugs. Hypoxic regions in tumors, therefore, represent attractive targets for cancer therapy. To date, five distinct classes of bioreactive prodrugs have been developed to target hypoxic cells in solid tumors. These hypoxia-activated prodrugs, including nitro compounds, N-oxides, quinones, and metal complexes, generally share a common mechanism of activation whereby they are reduced by intracellular oxidoreductases in an oxygen-sensitive manner to form cytotoxins. Several examples including PR-104, TH-302, and EO9 are currently undergoing phase II and phase III clinical evaluation. In this review, we discuss the nature of tumor hypoxia as a therapeutic target, focusing on the development of bioreductive prodrugs. We also describe the current knowledge of how each prodrug class is activated and detail the clinical progress of leading examples. PMID:23845143

  14. Azo-reductase activated budesodine prodrugs for colon targeting.

    PubMed

    Marquez Ruiz, Juan F; Kedziora, Kinga; O'Reilly, Mary; Maguire, Jacqueline; Keogh, Brian; Windle, Henry; Kelleher, Dermot P; Gilmer, John F

    2012-12-15

    Budesodine is a synthetic glurocorticoid that undergoes substantial first pass metabolism, limiting systemic exposure. Its use in treatment of inflammatory bowel disease would benefit from a targeting strategy that could lead to a local topical effect, improving safety and increasing anti-inflammatory efficacy. A two-step prodrug strategy involving azoreduction/cyclization that we developed previously for prednisolone is here applied with some variations to budesonide. The budesodine prodrugs were tested using an in vitro azoreductase assay simulating human colonic microflora. The kinetics of amino steroid ester cyclization and its pH dependence was also evaluated. The stability of the prodrugs systems in simulated human duodenal and gastric fluid was evaluated to determine the likelihood of intact intestinal transit. The propionic acid derived prodrug 3 undergoes rapid activation by Clostridium perfingens and its putative reduction product cyclizes with acceptable rapidity when synthesized independently. These properties of 3 suggest that it has potential in management of ulcerative colitis. PMID:23122819

  15. Synthesis of Am80 (tamibarotene) prodrug candidates, congeners and metabolites.

    PubMed

    Muratake, Hideaki; Amano, Yohei; Toda, Takahiro; Sugiyama, Kiyoshi; Shudo, Koichi

    2013-01-01

    Compound 1 (IT-M-07000) was previously reported as a candidate prodrug of Am80 (Tamibarotene; used to treat acute promyelocytic leukemia), and shown to be efficiently metabolized to Am80 via β-oxidation. Here, we describe in detail the synthesis of 1, together with another tetradeuterated candidate prodrug, IT-YA-00616 (2), as well as two congeners, and several metabolic intermediates of 1 previously detected in mouse plasma. PMID:23902867

  16. Ocular Sustained Release Nanoparticles Containing Stereoisomeric Dipeptide Prodrugs of Acyclovir

    PubMed Central

    Jwala, Jwala; Boddu, Sai H.S.; Shah, Sujay; Sirimulla, Suman; Pal, Dhananjay

    2011-01-01

    Abstract Purpose The objective of this study was to develop and characterize polymeric nanoparticles of appropriate stereoisomeric dipeptide prodrugs of acyclovir (L-valine-L-valine-ACV, L-valine-D-valine-ACV, D-valine-L-valine-ACV, and D-valine-D-valine-ACV) for the treatment of ocular herpes keratitis. Methods Stereoisomeric dipeptide prodrugs of acyclovir (ACV) were screened for bioreversion in various ocular tissues, cell proliferation, and uptake across the rabbit primary corneal epithelial cell line. Docking studies were carried out to examine the affinity of prodrugs to the peptide transporter protein. Prodrugs with optimum characteristics were selected for the preparation of nanoparticles using various grades of poly (lactic-co-glycolic acid) (PLGA). Nanoparticles were characterized for the entrapment efficiency, surface morphology, size distribution, and in vitro release. Further, the effect of thermosensitive gels on the release of prodrugs from nanoparticles was also studied. Results L-valine-L-valine-ACV and L-valine-D-valine-ACV were considered to be optimum in terms of enzymatic stability, uptake, and cytotoxicity. Docking results indicated that L-valine in the terminal position increases the affinity of the prodrugs to the peptide transporter protein. Entrapment efficiency values of L-valine-L-valine-ACV and L-valine-D-valine-ACV were found to be optimal with PLGA 75:25 and PLGA 65:35 polymers, respectively. In vitro release of prodrugs from nanoparticles exhibited a biphasic release behavior with initial burst phase followed by sustained release. Dispersion of nanoparticles in thermosensitive gels completely eliminated the burst release phase. Conclusion Novel nanoparticulate systems of dipeptide prodrugs of ACV suspended in thermosensitive gels may provide sustained delivery after topical administration. PMID:21500985

  17. Functional characterization of five different PRXamide receptors of the red flour beetle Tribolium castaneum with peptidomimetics and identification of agonists and antagonists

    PubMed Central

    Jiang, Hongbo; Wei, Zhaojun; Nachman, Ronald J.; Kaczmarek, Krzysztof; Zabrocki, Janusz; Park, Yoonseong

    2014-01-01

    The neuropeptidergic system in insects is an excellent target for pest control strategies. One promising biorational approach is the use of peptidomimetics modified from endogenous ligands to enhance biostability and bioavailability. In this study, we functionally characterized five different G protein-coupled receptors in a phylogenetic cluster, containing receptors for PRXamide in the red flour beetle Tribolium castaneum, by evaluating a series of 70 different peptides and peptidomimetics. Three pyrokinin receptors (TcPKr-A, -B, and –C), cardioacceleratory peptide receptor (TcCAPAr) and ecdysis triggering hormone receptor (TcETHr) were included in the study. Strong agonistic or antagonistic peptidomimetics were identified, and included beta-proline (β3P) modification of the core amino acid residue proline and also a cyclo-peptide. It is common for a ligand to act on multiple receptors. In a number of cases, a ligand acting as an agonist on one receptor was an efficient antagonist on another receptor, suggesting complex outcomes of a peptidomimetic in a biological system. Interestingly, TcPK-A was highly promiscuous with a high number of agonists, while TcPK-C and TcCAPAr had a lower number of agonists, but a higher number of compounds acting as an antagonist. This observation suggests that a target GPCR with more promiscuity will provide better success for peptidomimetic approaches. This study is the first description of peptidomimetics on a CAPA receptor and resulted in the identification of peptidomimetic analogs that demonstrate antagonism of CAPA ligands. The PRXamide receptor assays with peptidomimetics provide useful insights into the biochemical properties of receptors. PMID:25447413

  18. Phenylarsine oxide and vanadate: apparent paradox of inhibition of protein phosphotyrosine phosphatases in rat adipocytes.

    PubMed

    Li, J; Elberg, G; Shechter, Y

    1996-07-24

    Vanadate mimics, whereas phenylarsine oxide (PAO) antagonizes, the effects of insulin in rat adipocytes. Both vanadate and PAO are documented inhibitors of protein-phosphotyrosine phosphatases. The relationship between the inhibition of 'inhibitory' PTPase and 'stimulatory' PTPase has been studied here in primary rat adipocytes. Low concentrations of PAO (IC50 = 0.6-2.0 microM) blocked the stimulating effects of insulin, vanadate and pervanadate on hexose uptake and glucose metabolism. Inhibition of isoproterenol-mediating lipolysis by vanadate and insulin was not blocked by PAO. The activating effects of okadaic acid on hexose uptake and glucose metabolism, which occur at points downstream to tyrosine phosphorylation, were also not blocked by PAO. Subsequent studies suggested that the PAO-sensitive PTPase comprises a minute fraction of the total adipocytic PTPase activity. To identify its location we applied procedures involving fractionations and activation of non-receptor adipocytic protein tyrosine kinase by PAO and vanadate in cell free assays. We found that the 'inhibitory' PTPase is exclusively associated with the membrane fraction whereas the 'stimulatory' PTPases are present in both the cytosolic and plasma membrane compartments. We next searched for markers, possibly associated with PAO-dependent desensitization and found that several proteins became phosphorylated on tyrosine moieties in the supernatant of PAO but not in vanadate pretreated adipocytes. In summary, we propose the presence of a minute, plasma membrane associated PTPase in primary rat adipocytes, inhibition of which arrests the activation of glucose metabolism. In contrast, inhibition of all the other cellular adipose PTPases, ultimately activates rather than inhibits these same bioeffects. PMID:8703991

  19. MiniAp-4: A Venom-Inspired Peptidomimetic for Brain Delivery.

    PubMed

    Oller-Salvia, Benjamí; Sánchez-Navarro, Macarena; Ciudad, Sonia; Guiu, Marc; Arranz-Gibert, Pol; Garcia, Cristina; Gomis, Roger R; Cecchelli, Roméo; García, Jesús; Giralt, Ernest; Teixidó, Meritxell

    2016-01-11

    Drug delivery across the blood-brain barrier (BBB) is a formidable challenge for therapies targeting the central nervous system. Although BBB shuttle peptides enhance transport into the brain non-invasively, their application is partly limited by lability to proteases. The present study proposes the use of cyclic peptides derived from venoms as an affordable way to circumvent this drawback. Apamin, a neurotoxin from bee venom, was minimized by reducing its complexity, toxicity, and immunogenicity, while preserving brain targeting, active transport, and protease resistance. Among the analogues designed, the monocyclic lactam-bridged peptidomimetic MiniAp-4 was the most permeable. This molecule is capable of translocating proteins and nanoparticles in a human-cell-based BBB model. Furthermore, MiniAp-4 can efficiently deliver a cargo across the BBB into the brain parenchyma of mice. PMID:26492861

  20. Macrocyclic peptidomimetics with antimicrobial activity: synthesis, bioassay, and molecular modeling studies.

    PubMed

    Ibrahim, Mohamed A; Panda, Siva S; Oliferenko, Alexander A; Oliferenko, Polina V; Girgis, Adel S; Elagawany, Mohamed; Küçükbay, F Zehra; Panda, Chandramukhi S; Pillai, Girinath G; Samir, Ahmed; Tämm, Kaido; Hall, C Dennis; Katritzky, Alan R

    2015-09-28

    Novel, cyclic peptidomimetics were synthesized by facile acylation reactions using benzotriazole chemistry. Microbiological testing of the synthesized compounds revealed an exceptionally high activity against Candida albicans with a minimum inhibitory concentration (MIC) two orders of magnitude lower than the MIC of the antifungal reference drug amphotericin B. A strikingly high activity was also observed against three Gram-negative bacterial strains (Pseudomonas aeruginosa, Klebsiella pneumoniae and Proteus vulgaris), two of which are known human pathogens. Thus the discovered chemotype is a potential polypharmacological agent. The toxicity against mammalian tumor cells was found to be low, as demonstrated in five different human cell lines (HeLa, cervical; PC-3, prostate; MCF-7, breast; HepG2, liver; and HCT-116, colon). The internal consistency of the experimental data was studied using 3D-pharmacophore and 2D-QSAR. PMID:26256838

  1. Peptidomimetics Based On Dehydroepiandrosterone Scaffold: Synthesis, Antiproliferation Activity, Structure-Activity Relationship, and Mechanisms.

    PubMed

    Wang, Xiaohui; Su, Haihuan; Wang, Wenda; Chen, Changshui; Cao, Xiufang

    2016-01-01

    A series of novel peptidomimetics bearing dehydroepiandrosterone moiety were designed, synthesized, and evaluated for their inhibition activities against cell proliferation. According to the preliminary studies on inhibitory activities, some of the newly prepared compounds indicated significantly inhibition activities against human hepatoma cancer (HepG2), human lung cancer (A549), human melanoma (A875) cell lines compared with the control 5-fluorouracil. Especially, compounds Ii (IC50 < 14 μM) and Ik (IC50 < 13 μM) exhibited obvious inhibition activities against all tested cell lines. The highly potential compound Ik induced apoptosis in HepG2 cells were analyzed by flow cytometry, and the apoptotic effects of compound Ik were further evaluated using Annexin V-FITC/propidium iodide dual staining assay, which revealed these highly potential compounds induced cell death in HepG2 cells at least partly by apoptosis. PMID:27585479

  2. Peptidomimetics Based On Dehydroepiandrosterone Scaffold: Synthesis, Antiproliferation Activity, Structure-Activity Relationship, and Mechanisms

    PubMed Central

    Wang, Xiaohui; Su, Haihuan; Wang, Wenda; Chen, Changshui; Cao, Xiufang

    2016-01-01

    A series of novel peptidomimetics bearing dehydroepiandrosterone moiety were designed, synthesized, and evaluated for their inhibition activities against cell proliferation. According to the preliminary studies on inhibitory activities, some of the newly prepared compounds indicated significantly inhibition activities against human hepatoma cancer (HepG2), human lung cancer (A549), human melanoma (A875) cell lines compared with the control 5-fluorouracil. Especially, compounds Ii (IC50 < 14 μM) and Ik (IC50 < 13 μM) exhibited obvious inhibition activities against all tested cell lines. The highly potential compound Ik induced apoptosis in HepG2 cells were analyzed by flow cytometry, and the apoptotic effects of compound Ik were further evaluated using Annexin V-FITC/propidium iodide dual staining assay, which revealed these highly potential compounds induced cell death in HepG2 cells at least partly by apoptosis. PMID:27585479

  3. MiniAp‐4: A Venom‐Inspired Peptidomimetic for Brain Delivery

    PubMed Central

    Oller‐Salvia, Benjamí; Sánchez‐Navarro, Macarena; Ciudad, Sonia; Guiu, Marc; Arranz‐Gibert, Pol; Garcia, Cristina; Gomis, Roger R.; Cecchelli, Roméo; García, Jesús

    2015-01-01

    Abstract Drug delivery across the blood–brain barrier (BBB) is a formidable challenge for therapies targeting the central nervous system. Although BBB shuttle peptides enhance transport into the brain non‐invasively, their application is partly limited by lability to proteases. The present study proposes the use of cyclic peptides derived from venoms as an affordable way to circumvent this drawback. Apamin, a neurotoxin from bee venom, was minimized by reducing its complexity, toxicity, and immunogenicity, while preserving brain targeting, active transport, and protease resistance. Among the analogues designed, the monocyclic lactam‐bridged peptidomimetic MiniAp‐4 was the most permeable. This molecule is capable of translocating proteins and nanoparticles in a human‐cell‐based BBB model. Furthermore, MiniAp‐4 can efficiently deliver a cargo across the BBB into the brain parenchyma of mice. PMID:26492861

  4. HUMAN SKIN PERMEATION OF 3-O-ALKYL CARBAMATE PRODRUGS OF NALTREXONE

    PubMed Central

    Vaddi, Haranath K.; Banks, Stan L.; Chen, Jianhong; Hammell, Dana C.; Crooks, Peter A.; Stinchcomb, Audra L.

    2009-01-01

    N-Monoalkyl and N,N-dialkyl carbamate prodrugs of naltrexone (NTX), an opioid antagonist, were synthesized and their in vitro permeation across human skin was determined. Relevant physicochemical properties were also determined. Most prodrugs exhibited lower melting points, lower aqueous solubilities, and higher oil solubilities than NTX. The flux values from N-monoalkyl carbamate prodrugs were significantly higher than those from NTX and N,N-dialkyl carbamates. The melting points of N-monoalkyl carbamate prodrugs were quite low compared to the N,N-dialkyl carbamate prodrugs and NTX. Heats of fusion for the N,N-dialkyl carbamate prodrugs were higher than that for NTX. N-Monoalkyl carbamate prodrugs had higher stratum corneum/vehicle partition coefficients than their N,N-dialkyl counterparts. Higher percent prodrug bioconversion to NTX in skin appeared to be related to increased skin flux. N,N-Dialkyl carbamate prodrugs were more stable in buffer and in plasma than N-monoalkyl carbamate prodrugs. In conclusion, N-monoalkyl carbamate prodrugs of NTX improved the systemic delivery of NTX across human skin in vitro. N,N-Dialkyl substitution in the prodrug moiety decreased skin permeation and plasma hydrolysis to the parent drug. The cross-sectional area of the carbamate head group was the major determinant of flux of the N-monoalkyl and N,N-dialkyl carbamate prodrugs of NTX. PMID:18972573

  5. Systemic and Brain Pharmacokinetics of Perforin Inhibitor Prodrugs.

    PubMed

    Gynther, Mikko; Pickering, Darryl S; Spicer, Julie A; Denny, William A; Huttunen, Kristiina M

    2016-07-01

    We have recently reported that by converting a perforin inhibitor into an l-type amino acid transporter 1 (LAT1)-utilizing prodrug its cellular uptake can be greatly increased. The aim of the present study was to determine the in vivo and brain pharmacokinetics of two perforin inhibitors and their LAT1-utilizing prodrugs 1 and 2. In addition, the brain uptake mechanism and entry into primary mouse cortical neurons and astrocytes were evaluated. After 23 μmol/kg i.p. bolus injection, the prodrugs' unbound area under the concentration curve in brain was 0.3 nmol/g × min, whereas the parent drugs could not reach the brain. The unbound brain concentrations of the prodrugs after 100 μM in situ mouse brain perfusion were 521.4 ± 46.9 and 126.9 ± 19.9 pmol/g for prodrugs 1 and 2, respectively. The combination of competing transporter substrates for LAT1, l-tryptophan, and for organic anion transporting polypeptides, probenecid, decreased the brain concentrations to 352.4 ± 44.5 and 70.9 ± 7.0 pmol/g, respectively. In addition, in vitro uptake studies showed that at 100 μM prodrug 1 had 3.4-fold and 4.5-fold higher uptake rate into neurons and astrocytes, respectively, compared to its parent drug. Thus, the prodrugs enhance significantly the therapeutic potential of the parent drugs for the treatment of disorders of central nervous system in which neuroinflammation is involved. PMID:27266990

  6. Targeted anticancer prodrug with mesoporous silica nanoparticles as vehicles

    NASA Astrophysics Data System (ADS)

    Fan, Jianquan; Fang, Gang; Wang, Xiaodan; Zeng, Fang; Xiang, Yufei; Wu, Shuizhu

    2011-11-01

    A targeted anticancer prodrug system was fabricated with 180 nm mesoporous silica nanoparticles (MSNs) as carriers. The anticancer drug doxorubicin (DOX) was conjugated to the particles through an acid-sensitive carboxylic hydrazone linker which is cleavable under acidic conditions. Moreover, folic acid (FA) was covalently conjugated to the particle surface as the targeting ligand for folate receptors (FRs) overexpressed in some cancer cells. The in vitro release profiles of DOX from the MSN-based prodrug systems showed a strong dependence on the environmental pH values. The fluorescent dye FITC was incorporated in the MSNs so as to trace the cellular uptake on a fluorescence microscope. Cellular uptakes by HeLa, A549 and L929 cell lines were tested for FA-conjugated MSNs and plain MSNs respectively, and a much more efficient uptake by FR-positive cancer cells (HeLa) can be achieved by conjugation of folic acid onto the particles because of the folate-receptor-mediated endocytosis. The cytotoxicities for the FA-conjugated MSN prodrug, the plain MSN prodrug and free DOX against three cell lines were determined, and the result indicates that the FA-conjugated MSN prodrug exhibits higher cytotoxicity to FR-positive cells, and reduced cytotoxicity to FR-negative cells. Thus, with 180 nm MSNs as the carriers for the prodrug system, good drug loading, selective targeting and sustained release of drug molecules within targeted cancer cells can be realized. This study may provide useful insights for designing and improving the applicability of MSNs in targeted anticancer prodrug systems.

  7. 1-Arylsulfonyl-2-(pyridylmethylsulfinyl) benzimidazoles as new proton pump inhibitor prodrugs.

    PubMed

    Shin, Jai Moo; Sachs, George; Cho, Young-moon; Garst, Michael

    2009-01-01

    New arylsulfonyl proton pump inhibitor (PPI) prodrug forms were synthesized. These prodrugs provided longer residence time of an effective PPI plasma concentration, resulting in better gastric acid inhibition. PMID:20032890

  8. 1-Arylsulfonyl-2-(Pyridylmethylsulfinyl) Benzimidazoles as New Proton Pump Inhibitor Prodrugs

    PubMed Central

    Shin, Jai Moo; Sachs, George; Cho, Young-moon; Garst, Michael

    2010-01-01

    New arylsulfonyl proton pump inhibitor (PPI) prodrug forms were synthesized. These prodrugs provided longer residence time of an effective PPI plasma concentration, resulting in better gastric acid inhibition. PMID:20032890

  9. αvβ3- or α5β1-Integrin-Selective Peptidomimetics for Surface Coating.

    PubMed

    Mas-Moruno, Carlos; Fraioli, Roberta; Rechenmacher, Florian; Neubauer, Stefanie; Kapp, Tobias G; Kessler, Horst

    2016-06-13

    Engineering biomaterials with integrin-binding activity is a very powerful approach to promote cell adhesion, modulate cell behavior, and induce specific biological responses at the surface level. The aim of this Review is to illustrate the evolution of surface-coating molecules in this field: from peptides and proteins with relatively low integrin-binding activity and receptor selectivity to highly active and selective peptidomimetic ligands. In particular, we will bring into focus the difficult challenge of achieving selectivity between the two closely related integrin subtypes αvβ3 and α5β1. The functionalization of surfaces with such peptidomimetics opens the way for a new generation of highly specific cell-instructive surfaces to dissect the biological role of integrin subtypes and for application in tissue engineering and regenerative medicine. PMID:27258759

  10. High-Affinity, Small-Molecule Peptidomimetic Inhibitors of MLL1/WDR5 Protein-Protein Interaction

    SciTech Connect

    Karatas, Hacer; Townsend, Elizabeth C; Cao, Fang; Chen, Yong; Bernard, Denzil; Liu, Liu; Lei, Ming; Dou, Yali; Wang, Shaomeng

    2013-02-12

    Mixed lineage leukemia 1 (MLL1) is a histone H3 lysine 4 (H3K4) methyltransferase, and targeting the MLL1 enzymatic activity has been proposed as a novel therapeutic strategy for the treatment of acute leukemia harboring MLL1 fusion proteins. The MLL1/WDR5 protein–protein interaction is essential for MLL1 enzymatic activity. In the present study, we designed a large number of peptidomimetics to target the MLL1/WDR5 interaction based upon -CO-ARA-NH–, the minimum binding motif derived from MLL1. Our study led to the design of high-affinity peptidomimetics, which bind to WDR5 with Ki < 1 nM and function as potent antagonists of MLL1 activity in a fully reconstituted in vitro H3K4 methyltransferase assay. Determination of co-crystal structures of two potent peptidomimetics in complex with WDR5 establishes their structural basis for high-affinity binding to WDR5. Evaluation of one such peptidomimetic, MM-102, in bone marrow cells transduced with MLL1-AF9 fusion construct shows that the compound effectively decreases the expression of HoxA9 and Meis-1, two critical MLL1 target genes in MLL1 fusion protein mediated leukemogenesis. MM-102 also specifically inhibits cell growth and induces apoptosis in leukemia cells harboring MLL1 fusion proteins. Our study provides the first proof-of-concept for the design of small-molecule inhibitors of the WDR5/MLL1 protein–protein interaction as a novel therapeutic approach for acute leukemia harboring MLL1 fusion proteins.

  11. Prodrug converting enzyme gene delivery by L. monocytogenes

    PubMed Central

    Stritzker, Jochen; Pilgrim, Sabine; Szalay, Aladar A; Goebel, Werner

    2008-01-01

    Background Listeria monocytogenes is a highly versatile bacterial carrier system for introducing protein, DNA and RNA into mammalian cells. The delivery of tumor antigens with the help of this carrier into tumor-bearing animals has been successfully carried out previously and it was recently reported that L. monocytogenes is able to colonize and replicate within solid tumors after local or even systemic injection. Methods Here we report on the delivery of two prodrug converting enzymes, purine-deoxynucleoside phosphorylase (PNP) and a fusion protein consisting of yeast cytosine deaminase and uracil phosphoribosyl transferase (FCU1) into cancer cells in culture by L. monocytogenes. Transfer of the prodrug converting enzymes was achieved by bacterium mediated transfer of eukaryotic expression plasmids or by secretion of the proteins directly into the host cell cytosol by the infecting bacteria. Results The results indicate that conversion of appropriate prodrugs to toxic drugs in the cancer cells occured after both procedures although L. monocytogenes-mediated bactofection proved to be more efficient than enzyme secretion 4T1, B16 and COS-1 tumor cells. Exchanging the constitutively PCMV-promoter with the melanoma specific P4xTETP-promoter resulted in melanoma cell-specific expression of the prodrug converting enzymes but reduced the efficiencies. Conclusion These experiments open the way for bacterium mediated tumor specific activation of prodrugs in live animals with tumors. PMID:18402662

  12. Recent Trends in Targeted Anticancer Prodrug and Conjugate Design

    PubMed Central

    Singh, Yashveer; Palombo, Matthew; Sinko, Patrick J.

    2009-01-01

    Anticancer drugs are often nonselective antiproliferative agents (cytotoxins) that preferentially kill dividing cells by attacking their DNA at some level. The lack of selectivity results in significant toxicity to noncancerous proliferating cells. These toxicities along with drug resistance exhibited by the solid tumors are major therapy limiting factors that results into poor prognosis for patients. Prodrug and conjugate design involves the synthesis of inactive drug derivatives that are converted to an active form inside the body and preferably at the site of action. Classical prodrug and conjugate design has focused on the development of prodrugs that can overcome physicochemical (e.g., solubility, chemical instability) or biopharmaceutical problems (e.g., bioavailability, toxicity) associated with common anticancer drugs. The recent targeted prodrug and conjugate design, on the other hand, hinges on the selective delivery of anticancer agents to tumor tissues thereby avoiding their cytotoxic effects on noncancerous cells. Targeting strategies have attempted to take advantage of low extracellular pH, elevated enzymes in tumor tissues, the hypoxic environment inside the tumor core, and tumor-specific antigens expressed on tumor cell surfaces. The present review highlights recent trends in prodrug and conjugate rationale and design for cancer treatment. The various approaches that are currently being explored are critically analyzed and a comparative account of the advantages and disadvantages associated with each approach is presented. PMID:18691040

  13. Polymerizable disulfide paclitaxel prodrug for controlled drug delivery.

    PubMed

    Ding, Yi; Chen, Wulian; Hu, Jianhua; Du, Ming; Yang, Dong

    2014-11-01

    A polymerizable disulfide paclitaxel (PTX) prodrug was synthesized by the consequential esterification reactions of 3,3'-dithiodipropionic acid (DTPA), a disulfide compound containing two active carboxyl groups, with 2-hydroxyethyl methacrylate (HEMA) and PTX. The structure of the prodrug was confirmed by (1)H NMR characterization. Then, the polymerizable prodrug was copolymerized with poly(ethylene glycol) methyl ether methacrylate (PEGMEA) to obtain a copolymer with hydrophilic PEG side chains and PTX covalently linked onto the backbone via disulfide bonds. The loading content of PTX was 23%. In aqueous solution, this copolymer prodrug could self-assemble into micelles, with hydrophobic PTX as the cores and hydrophilic PEG-segment as the shells. In vitro cell assay demonstrated that this copolymer prodrug showed more apparent cytotoxicity to cancer cells than to human normal cells. After incubation for 48 h, the cell viability of HEK-293 cells (human embryo kidney cells) at 0.1 μg/mL PTX still remained more than 90%, however, that of HeLa cells (human cervical cancer cells) decreased to 52%. PMID:25280719

  14. PACE4 inhibitors and their peptidomimetic analogs block prostate cancer tumor progression through quiescence induction, increased apoptosis and impaired neovascularisation

    PubMed Central

    Levesque, Christine; Couture, Frédéric; Kwiatkowska, Anna; Desjardins, Roxane; Guérin, Brigitte; Neugebauer, Witold A.; Day, Robert

    2015-01-01

    Prostate cancer is the leading cancer in North American men. Current pharmacological treatments are limited to anti-androgen strategies and the development of new therapeutic approaches remains a challenge. As a fundamentally new approach, we propose the inhibition of PACE4, a member of the proprotein convertases family of enzymes, as a therapeutic target in prostate cancer. We developed an inhibitor named the Multi-Leu peptide, with potent in vitro anti-proliferative effects. However, the Multi-Leu peptide has not been tested under in vivo conditions and its potency under such conditions is most likely limited, due to the labile characteristics of peptides in general. Using a peptidomimetic approach, we modified the initial scaffold, generating the analog Ac-[DLeu]LLLRVK-Amba, which demonstrates increased inhibitory potency and stability. The systemic administration of this peptidomimetic significantly inhibits tumor progression in the LNCaP xenograft model of prostate cancer by inducing tumor cell quiescence, increased apoptosis and neovascularization impairment. Pharmacokinetic and biodistribution profiles of this inhibitor confirm adequate tumor delivery properties of the compound. We conclude that PACE4 peptidomimetic inhibitors could result in stable and potent drugs for a novel therapeutic strategy for prostate cancer. PMID:25682874

  15. Ester Prodrugs of Ketoprofen: Synthesis, Hydrolysis Kinetics and Pharmacological Evaluation.

    PubMed

    Dhokchawle, B V; Tauro, S J; Bhandari, A B

    2016-01-01

    The ester prodrugs of ketoprofen with various naturally available antioxidants; menthol, thymol, eugenol, guiacol, vanillin and sesamol have been synthesized by the dicyclohexyl carbodiimide (DCC) coupling method, purified and characterized by spectral data. Further, their, partition coefficients have been determined as well as, hydrolytic studies performed. The synthesized compounds are more lipophilic compared to the parent moieties and are stable in acidic environment, which is a prerequisite for their oral absorption. Under gastric as well as intestinal pH conditions these prodrugs showed variable susceptibility towards hydrolysis. The title compounds when evaluated for anti-inflammatory, analgesic activities and ulcerogenicity, showed improvement over the parent drug. PMID:25894087

  16. Evaluation of hydroxyimine as cytochrome P450-selective prodrug structure.

    PubMed

    Kumpulainen, Hanna; Mähönen, Niina; Laitinen, Marja-Leena; Jaurakkajärvi, Marja; Raunio, Hannu; Juvonen, Risto O; Vepsäläinen, Jouko; Järvinen, Tomi; Rautio, Jarkko

    2006-02-01

    Hydroxyimine derivatives of ketoprofen (1) and nabumetone (2) were synthesized and evaluated in vitro and in vivo as cytochrome P450-selective intermediate prodrug structures of ketones. 2 released nabumetone in vitro in the presence of isolated rat and human liver microsomes and in different recombinant human CYP isoforms. Bioconversion of 2 to both nabumetone and its active metabolite, 6-methoxy-2-naphthylacetic acid (6-MNA), was further confirmed in rats in vivo. Results indicate that hydroxyimine is a useful intermediate prodrug structure for ketone drugs. PMID:16451086

  17. Evaluating Aziridinyl Nitrobenzamide Compounds as Leishmanicidal Prodrugs

    PubMed Central

    Voak, Andrew A.; Seifert, Karin; Helsby, Nuala A.

    2014-01-01

    Many of the nitroaromatic agents used in medicine function as prodrugs and must undergo activation before exerting their toxic effects. In most cases, this is catalyzed by flavin mononucleotide (FMN)-dependent type I nitroreductases (NTRs), a class of enzyme absent from higher eukaryotes but expressed by bacteria and several eukaryotic microbes, including trypanosomes and Leishmania. Here, we utilize this difference to evaluate whether members of a library of aziridinyl nitrobenzamides have activity against Leishmania major. Biochemical screens using purified L. major NTR (LmNTR) revealed that compounds containing an aziridinyl-2,4-dinitrobenzyl core were effective substrates for the enzyme and showed that the 4-nitro group was important for this activity. To facilitate drug screening against intracellular amastigote parasites, we generated leishmanial cells that expressed the luciferase reporter gene and optimized a mammalian infection model in a 96-well plate format. A subset of aziridinyl-2,4-dinitrobenzyl compounds possessing a 5-amide substituent displayed significant growth-inhibitory properties against the parasite, with the most potent agents generating 50% inhibitory concentrations of <100 nM for the intracellular form. This antimicrobial activity was shown to be LmNTR specific since L. major NTR+/− heterozygote parasites were slightly resistance to most aziridinyl dinitrobenzyl agents tested. When the most potent leishmanicidal agents were screened against the mammalian cells in which the amastigote parasites were propagated, no growth-inhibitory effect was observed at concentrations of up to 100 μM. We conclude that the aziridinyl nitrobenzamides represent a new lead structure that may have the potential to treat leishmanial infections. PMID:24165190

  18. Bisphosphonamidate Clodronate Prodrug Exhibits Selective Cytotoxic Activity Against Melanoma Cell Lines

    PubMed Central

    Webster, Marie R.; Kamat, Chandrashekhar; Connis, Nick; Zhao, Ming; Weeraratna, Ashani T.; Rudek, Michelle A.; Hann, Christine L.; Freel Meyers, Caren L.

    2014-01-01

    Bisphosphonates are used clinically to treat disorders of calcium metabolism and malignant bone disease and are known to inhibit cancer cell growth, adhesion, and invasion. However, clinical use of these agents for the treatment of extraskeletal disease is limited due to low cell permeability. We recently described a bisphosphonamidate prodrug strategy for efficient intracellular release of bisphosphonates, including clodronate (CLO), in NSCLC cells. To evaluate anticancer activity of this prodrug class across many cancer cell types, the bisphosphonamidate clodronate prodrug (CLO prodrug) was screened against the NCI-60 cell line panel, and was found to exhibit selectivity toward melanoma cell lines. Here, we confirm efficient cellular uptake and intracellular activation of this prodrug class in melanoma cells. We further demonstrate inhibition of melanoma cell proliferation, induction of apoptosis, and an anti-tumor effect of CLO prodrug in a xenograft model. These data suggest a novel therapeutic application for the CLO prodrug and potential to selectively target melanoma cells. PMID:24310621

  19. Investigating the Impact of Asp181 Point Mutations on Interactions between PTP1B and Phosphotyrosine Substrate

    NASA Astrophysics Data System (ADS)

    Liu, Mengyuan; Wang, Lushan; Sun, Xun; Zhao, Xian

    2014-05-01

    Protein tyrosine phosphatase 1B (PTP1B) is a key negative regulator of insulin and leptin signaling, which suggests that it is an attractive therapeutic target in type II diabetes and obesity. The aim of this research is to explore residues which interact with phosphotyrosine substrate can be affected by D181 point mutations and lead to increased substrate binding. To achieve this goal, molecular dynamics simulations were performed on wild type (WT) and two mutated PTP1B/substrate complexes. The cross-correlation and principal component analyses show that point mutations can affect the motions of some residues in the active site of PTP1B. Moreover, the hydrogen bond and energy decomposition analyses indicate that apart from residue 181, point mutations have influence on the interactions of substrate with several residues in the active site of PTP1B.

  20. Conformationally Constrained Histidines in the Design of Peptidomimetics: Strategies for the χ-Space Control

    PubMed Central

    Stefanucci, Azzurra; Pinnen, Francesco; Feliciani, Federica; Cacciatore, Ivana; Lucente, Gino; Mollica, Adriano

    2011-01-01

    A successful design of peptidomimetics must come to terms with χ-space control. The incorporation of χ-space constrained amino acids into bioactive peptides renders the χ1 and χ2 torsional angles of pharmacophore amino acids critical for activity and selectivity as with other relevant structural features of the template. This review describes histidine analogues characterized by replacement of native α and/or β-hydrogen atoms with alkyl substituents as well as analogues with α, β-didehydro unsaturation or Cα-Cβ cyclopropane insertion (ACC derivatives). Attention is also dedicated to the relevant field of β-aminoacid chemistry by describing the synthesis of β2- and β3-models (β-hHis). Structural modifications leading to cyclic imino derivatives such as spinacine, aza-histidine and analogues with shortening or elongation of the native side chain (nor-histidine and homo-histidine, respectively) are also described. Examples of the use of the described analogues to replace native histidine in bioactive peptides are also given. PMID:21686155

  1. Guanidino Groups Greatly Enhance the Action of Antimicrobial Peptidomimetics Against Bacterial Cytoplasmic Membranes

    PubMed Central

    Laursen, Jonas S.; Citterio, Linda; Hein-Kristensen, Line; Gram, Lone; Kuzmenko, Ivan; Olsen, Christian A.; Gidalevitz, David

    2014-01-01

    A promising class of potential new antibiotics are the antimicrobial peptides or their synthetic mimics. Herein we assess the effect of the type of cationic side chain (i.e., guanidino vs. amino groups) on the membrane perturbing mechanism of antimicrobial α-peptide–β-peptoid chimeras. Two separate Langmuir monolayers composed of 1,2-dipalmitoyl-sn-glycero-3-phosphatidylglycerol (DPPG) and lipopolysaccharide Kdo2-lipid A were applied to model the outer membranes of Gram-positive and Gram-negative bacteria, respectively. We report the results of the measurements using an array of techniques, including high-resolution synchrotron surface X-ray scattering, epifluorescence microscopy, and in vitro antimicrobial activity to study the molecular mechanisms of peptidomimetic interaction with bacterial membranes. We found guanidino group-containing chimeras to exhibit greater disruptive activity on DPPG monolayers than the amino group-containing analogues. However, this effect was not observed for lipopolysaccharide monolayers where the difference was negligible. Furthermore, the addition of the nitrobenzoxadiazole fluorophore did not reduce the insertion activity of these antimicrobials into both model membrane systems examined, which may be useful for future cellular localization studies. PMID:24878450

  2. Sugar amino acid based scaffolds--novel peptidomimetics and their potential in combinatorial synthesis.

    PubMed

    Chakraborty, Tushar K; Jayaprakash, Sarva; Ghosh, Subhash

    2002-08-01

    To meet the growing demands for the development of new molecular entities for discovering new drugs and materials, organic chemists have started looking for new concepts to supplement traditional approaches. In one such approach, the expertise gained over the years in the area of organic synthesis and the rational drug-design concepts are combined together to create "nature-like" and yet unnatural organic molecules that are expected to provide leads in discovering new molecules. Emulating the basic principles followed by nature to build its vast repertoire of biomolecules, organic chemists are developing many novel multifunctional building blocks. Sugar amino acids constitute an important class of such polyfunctional scaffolds where the carboxyl, amino and hydroxyl groups provide an excellent opportunity for organic chemists to create structural diversities akin to nature's molecular arsenal. Recent advances in the area of combinatorial chemistry give unprecedented technological support for rapid compilations of sugar amino acid-based libraries exploiting the diversities of carbohydrate molecules and well-developed solid-phase peptide synthesis methods. This review chronicles the development of sugar amino acids as a novel class of peptidomimetic building blocks and their applications in generating desired secondary structures in peptides as well as in creating mimics of natural biopolymers. PMID:12180903

  3. Design, Synthesis, and Application of OB2C Combinatorial Peptide and Peptidomimetic Libraries

    PubMed Central

    Liu, Ruiwu; Shih, Tsung-Chieh; Deng, Xiaojun; Anwar, Lara; Ahadi, Sara; Kumaresan, Pappanaicken; Lam, Kit S.

    2015-01-01

    The “one-bead two-compound” (OB2C) combinatorial library is constructed on topologically segregated trifunctional bilayer beads such that each bead has a fixed cell-capturing ligand and a random library compound co-displayed on its surface and a chemical coding tag (bar code) inside the bead. An OB2C library containing thousands to millions of compounds can be synthesized and screened concurrently within a short period of time. When live cells are incubated with such OB2C libraries, every bead will be coated with a monolayer of cells. The cell membranes of the captured cells facing the bead surface are exposed to the library compounds tethered to each bead. A specific biochemical or cellular response can be detected with an appropriate reporter system. The OB2C method enables investigators to rapidly discover synthetic molecules that not only interact with cell-surface receptors but can also stimulate or inhibit downstream cell signaling. To demonstrate this powerful method, one OB2C peptide library and two OB2C peptidomimetic libraries were synthesized and screened against Molt-4 lymphoma cells to discover “death ligands.” Apoptosis of the bead-bound cells was detected with immunocytochemistry using horseradish peroxidase (HRP)-conjugated anti-cleaved caspase-3 antibody and 3,3′-diaminobenzidine as a substrate. Two novel synthetic “death ligands” against Molt-4 cells were discovered using this OB2C library approach. PMID:25616322

  4. Studies toward novel peptidomimetic inhibitors of thioredoxin-thioredoxin reductase system.

    PubMed

    Kłossowski, Szymon; Muchowicz, Angelika; Firczuk, Małgorzata; Swiech, Marta; Redzej, Adam; Golab, Jakub; Ostaszewski, Ryszard

    2012-01-12

    Thioredoxins (Trx) are ubiquitous multifunctional low-molecular weight proteins that together with thioredoxin reductases (TrxR) participate in the maintenance of protein thiol homeostasis in NADPH-dependent reactions. An increasing number of data reveal that the Trx-TrxR system is an attractive target for anticancer therapies. In this work, we have elaborated a new and simple synthetic approach employing Ugi reaction to synthesize several new inhibitors of this system. The influence of various electrophilic fragments of this new class of compounds on the inhibition of the Trx-TrxR system was evaluated. As a result, a new compound 19a (SK053), which inhibits the activity of the Trx-TrxR system and exhibits antitumor activity, was obtained. Biologic analyses revealed that 19a inhibits induction of NF-κB and AP-1 and decreases H(2)O(2) scavenging capacity in tumor cells. Altogether, we show that 19a is a novel potential antitumor peptidomimetic inhibitor that can be used as a starting compound for further optimization. PMID:22128876

  5. Chitosan based nanoparticles functionalized with peptidomimetic derivatives for oral drug delivery.

    PubMed

    Chronopoulou, Laura; Nocca, Giuseppina; Castagnola, Massimo; Paludetti, Gaetano; Ortaggi, Giancarlo; Sciubba, Fabio; Bevilacqua, Melania; Lupi, Alessandro; Gambarini, Gianluca; Palocci, Cleofe

    2016-01-25

    The goal of this study was to develop an optimized drug delivery carrier for oral mucosa applications able to release in situ bioactive molecules by using biopolymeric materials. Among them chitosan and poly(lactic-co-glycolic acid) (PLGA) have gained considerable attention as biocompatible carriers able to improve the delivery of active agents. The formulation of such vehicles in the form of nanoparticles (NPs) could permit to exploit the peculiar properties of nanomaterials in order to enhance the efficacy of active agents. Chitosan (CS) and PLGA chlorexidine dihydrochloride (CHX)-loaded NPs were synthesized by ionotropic gelation and osmosis based methodology respectively. In order to facilitate NPs adhesion on human dental surfaces, two different strategies were employed: PLGA particles with an external shell of CS to produce a positive surface charge allowing CHX loaded PLGA NPs to interact with the negative charged dental surfaces, while CS particles were functionalized with peptidomimetic derivative glutathione (GSH). The morphology was investigated by scanning electron microscopy. A sustained release profile of CHX from CS NPs was achieved. CS-based NPs adhered on human tooth surfaces in a simulated brushing and rinsing process and their in vitro toxicity evaluation on Human Gingival Fibroblasts (HGFs) was between 20 and 60% in all experimental conditions. Thanks to their adhesion properties and low cytotoxicity, the synthesized CS-based formulations may be efficiently exploited for therapy purposes or to enhance in vivo dental care (i.e. preparation of toothpastes or other cosmetics for daily oral care). PMID:26257139

  6. Design, synthesis, and application of OB2C combinatorial peptide and peptidomimetic libraries.

    PubMed

    Liu, Ruiwu; Shih, Tsung-Chieh; Deng, Xiaojun; Anwar, Lara; Ahadi, Sara; Kumaresan, Pappanaicken; Lam, Kit S

    2015-01-01

    The "one-bead two-compound" (OB2C) combinatorial library is constructed on topologically segregated trifunctional bilayer beads such that each bead has a fixed cell-capturing ligand and a random library compound co-displayed on its surface and a chemical coding tag (bar code) inside the bead. An OB2C library containing thousands to millions of compounds can be synthesized and screened concurrently within a short period of time. When live cells are incubated with such OB2C libraries, every bead will be coated with a monolayer of cells. The cell membranes of the captured cells facing the bead surface are exposed to the library compounds tethered to each bead. A specific biochemical or cellular response can be detected with an appropriate reporter system. The OB2C method enables investigators to rapidly discover synthetic molecules that not only interact with cell-surface receptors but can also stimulate or inhibit downstream cell signaling. To demonstrate this powerful method, one OB2C peptide library and two OB2C peptidomimetic libraries were synthesized and screened against Molt-4 lymphoma cells to discover "death ligands." Apoptosis of the bead-bound cells was detected with immunocytochemistry using horseradish peroxidase (HRP)-conjugated anti-cleaved caspase-3 antibody and 3,3'-diaminobenzidine as a substrate. Two novel synthetic "death ligands" against Molt-4 cells were discovered using this OB2C library approach. PMID:25616322

  7. Inhibition of dipeptidyl peptidase IV by fluoroolefin-containing N-peptidyl-O-hydroxylamine peptidomimetics

    PubMed Central

    Lin, Jian; Toscano, Paul J.; Welch, John T.

    1998-01-01

    Dipeptidyl peptidase IV (EC 3.4.14.5; DPP IV), also known as the leukocyte differentiation antigen CD26 when found as an extracellular membrane-bound proline specific serine protease, cleaves a dipeptide from the N terminus of a polypeptide chain containing a proline residue in the penultimate position. Here we report that known (Z)-Ala-ψ[CF=C]-Pro dipeptide isosteres 1 and 2, which contain O-acylhydroxylamines, were isolated as diastereomeric pairs u-1, l-1, and l-2. The effect of each diastereomeric pair as an inhibitor of human placental dipeptidyl peptidase DPP IV has been examined. The inhibition of DPP IV by these compounds is rapid and efficient. The diastereomeric pair u-1 exhibits very potent inhibitory activity with a Ki of 188 nM. Fluoroolefin containing N-peptidyl-O-hydroxylamine peptidomimetics, by virtue of their inhibitory potency and stability, are superior to N-peptidyl-O-hydroxylamine inhibitors derived from an Ala-Pro dipeptide. PMID:9826645

  8. The Structural Basis for Peptidomimetic Inhibition of Eukaryotic Ribonucleotide Reductase: A Conformationally Flexible Pharmacophore

    SciTech Connect

    Xu, Hai; Fairman, James W.; Wijerathna, Sanath R.; Kreischer, Nathan R.; LaMacchia, John; Helmbrecht, Elizabeth; Cooperman, Barry S.; Dealwis, Chris

    2008-08-19

    Eukaryotic ribonucleotide reductase (RR) catalyzes nucleoside diphosphate conversion to deoxynucleoside diphosphate. Crucial for rapidly dividing cells, RR is a target for cancer therapy. RR activity requires formation of a complex between subunits R1 and R2 in which the R2 C-terminal peptide binds to R1. Here we report crystal structures of heterocomplexes containing mammalian R2 C-terminal heptapeptide, P7 (Ac-{sup 1}FTLDADF{sup 7}) and its peptidomimetic P6 ({sup 1}Fmoc(Me)PhgLDChaDF{sup 7}) bound to Saccharomyces cerevisiae R1 (ScR1). P7 and P6, both of which inhibit ScRR, each bind at two contiguous sites containing residues that are highly conserved among eukaryotes. Such binding is quite distinct from that reported for prokaryotes. The Fmoc group in P6 peptide makes several hydrophobic interactions that contribute to its enhanced potency in binding to ScR1. Combining all of our results, we observe three distinct conformations for peptide binding to ScR1. These structures provide pharmacophores for designing highly potent nonpeptide class I RR inhibitors.

  9. Guanidino groups greatly enhance the action of antimicrobial peptidomimetics against bacterial cytoplasmic membranes

    SciTech Connect

    Andreev, Konstantin; Bianchi, Christopher; Laursen, Jonas S.; Citterio, Linda; Hein-Kristensen, Line; Gram, Lone; Kuzmenko, Ivan; Olsen, Christian A.; Gidalevitz, David

    2014-01-01

    Antimicrobial peptides or their synthetic mimics are a promising class of potential new antibiotics. Herein we assess the effect of the type of cationic side chain (i.e., guanidino vs. amino groups) on the membrane perturbing mechanismof antimicrobial α-peptide–β-peptoid chimeras. Langmuirmonolayers composed of 1,2-dipalmitoylsn- glycero-3-phosphatidylglycerol (DPPG) were used to model cytoplasmic membranes of both Gram-positive and Gram-negative bacteria,while lipopolysaccharide Kdo2-lipid Amonolayersweremimicking the outer membrane of Gram-negative species.We report the results of themeasurements using an array of techniques, including high-resolution synchrotron surface X-ray scattering, epifluorescence microscopy, and in vitro antimicrobial activity to study the molecularmechanisms of peptidomimetic interaction with bacterialmembranes.We found guanidino group-containing chimeras to exhibit greater disruptive activity on DPPGmonolayers than the amino group-containing analogues. However, this effect was not observed for lipopolysaccharidemonolayerswhere the difference was negligible. Furthermore, the addition of the nitrobenzoxadiazole fluorophore did not reduce the insertion activity of these antimicrobials into both model membrane systems examined, which may be useful for future cellular localization studies.

  10. Mode of action of claudin peptidomimetics in the transient opening of cellular tight junction barriers.

    PubMed

    Staat, Christian; Coisne, Caroline; Dabrowski, Sebastian; Stamatovic, Svetlana M; Andjelkovic, Anuska V; Wolburg, Hartwig; Engelhardt, Britta; Blasig, Ingolf E

    2015-06-01

    In epithelial/endothelial barriers, claudins form tight junctions, seal the paracellular cleft, and limit the uptake of solutes and drugs. The peptidomimetic C1C2 from the C-terminal half of claudin-1's first extracellular loop increases drug delivery through epithelial claudin-1 barriers. However, its molecular and structural mode of action remains unknown. In the present study, >100 μM C1C2 caused paracellular opening of various barriers with different claudin compositions, ranging from epithelial to endothelial cells, preferentially modulating claudin-1 and claudin-5. After 6 h incubation, C1C2 reversibly increased the permeability to molecules of different sizes; this was accompanied by redistribution of claudins and occludin from junctions to cytosol. Internalization of C1C2 in epithelial cells depended on claudin-1 expression and clathrin pathway, whereby most C1C2 was retained in recyclosomes >2 h. In freeze-fracture electron microscopy, C1C2 changed claudin-1 tight junction strands to a more parallel arrangement and claudin-5 strands from E-face to P-face association - drastic and novel effects. In conclusion, C1C2 is largely recycled in the presence of a claudin, which explains the delayed onset of barrier and junction loss, the high peptide concentration required and the long-lasting effect. Epithelial/endothelial barriers are specifically modulated via claudin-1/claudin-5, which can be targeted to improve drug delivery. PMID:25907035

  11. Conformationally Constrained Peptidomimetic Inhibitors of Signal Transducer and Activator of Transcription 3: Evaluation and Molecular Modeling

    PubMed Central

    Mandal, Pijus K.; Limbrick, Donald; Coleman, David R.; Dyer, Garrett A.; Ren, Zhiyong; Birtwistle, J. Sanderson; Xiong, Chiyi; Chen, Xiaomin; Briggs, James M.; McMurray, John S.

    2009-01-01

    Signal transducer and activator of transcription 3 (Stat3) is involved in aberrant growth and survival signals in malignant tumor cells and is a validated target for anti-cancer drug design. We are targeting its SH2 domain to prevent docking to cytokine and growth factor receptors and subsequent signaling. The amino acids of our lead phosphopeptide, Ac-pTyr-Leu-Pro-Gln-Thr-Val-NH2, were replaced with conformationally constrained mimics. Structure-affinity studies led to the peptidomimetic, pCinn-Haic-Gln-NHBn (21) which had an IC50 of 162 nM (fluorescence polarization), as compared to 290 nM for the lead phosphopeptide (pCinn = 4-phosphoryloxycinnamate, Haic = (2S,5S)-5-amino-1,2,4,5,6,7-hexahydro-4-oxo-azepino[3,2,1-hi]indole-2-carboxylic acid). pCinn-Haic-Gln-OH was docked to the SH2 domain (AUTODOCK) and the two highest populated clusters were subjected to molecular dynamics simulations. Both converged to a common peptide conformation. The complex exhibits unique hydrogen bonding between Haic and Gln and Stat3 as well as hydrophobic interactions between the protein and pCinn and Haic. PMID:19334714

  12. Regulation of sarcoma cell migration, invasion and invadopodia formation by AFAP1L1 through a phosphotyrosine-dependent pathway.

    PubMed

    Tie, S R; McCarthy, D J; Kendrick, T S; Louw, A; Le, C; Satiaputra, J; Kucera, N; Phillips, M; Ingley, E

    2016-04-21

    Invasion and metastasis are controlled by the invadopodia, which delivers matrix-degrading enzymes to the invasion interface permitting cancer cell penetration and spread into healthy tissue. We have identified a novel pathway that directs Lyn/Src family tyrosine kinase signals to the invadopodia to regulate sarcoma cell invasion via the molecule AFAP-1-like-1 (AFAP1L1), a new member of the AFAP (actin filament-associated protein) family. We show that AFAP1L1 can transform cells, promote migration and co-expression with active Lyn profoundly influences cell morphology and movement. AFAP1L1 intersects several invadopodia pathway components through its multiple domains and motifs, including the following (i) pleckstrin homology domains that bind phospholipids generated at the plasma membrane by phosphoinositide 3-kinase, (ii) a direct filamentous-actin binding domain and (iii) phospho-tyrosine motifs (pY136 and pY566) that specifically bind Vav2 and Nck2 SH2 domains, respectively. These phosphotyrosine motifs are essential for AFAP1L1-mediated cytoskeleton regulation. Through its interaction with Vav2, AFAP1L1 regulates Rac activity and downstream control of PAK1/2/3 (p21-activated kinases) phosphorylation of myosin light chain (MLC) kinase and MLC2. AFAP1L1 interaction with Nck2 recruits actin-nucleating complexes. Significantly, in osteosarcoma cell lines, knockdown of AFAP1L1 inhibits phosphorylated MLC2 recruitment to filamentous-actin structures, disrupts invadopodia formation, cell attachment, migration and invasion. These data define a novel pathway that directs Lyn/Src family tyrosine kinase signals to sarcoma cell invadopodia through specific recruitment of Vav2 and Nck2 to phosphorylated AFAP1L1, to control cell migration and invasion. PMID:26212012

  13. Prodrugs of anthracycline antibiotics suited for tumor-specific activation.

    PubMed

    Azoulay, M; Florent, J C; Monneret, C; Gesson, J P; Jacquesy, J C; Tillequin, F; Koch, M; Bosslet, K; Czech, J; Hoffman, D

    1995-09-01

    The two novel prodrugs 4 and 11 have been prepared from tetra-O-acetyl-D-galactopyranose and doxorubicin in three and six steps, respectively. Their low cytotoxicity, high stability in plasma and, in the case of 11, efficient hydrolysis in the presence of alpha-galactosidase, fulfill preliminary conditions for their use in combination with monoclonal antibody-enzyme conjugates. PMID:7575986

  14. Simvastatin Prodrug Micelles Target Fracture and Improve Healing

    PubMed Central

    Dusad, Anand; Yuan, Hongjiang; Ren, Ke; Li, Fei; Fehringer, Edward V.; Purdue, P. Edward; Goldring, Steven R.; Daluiski, Aaron; Wang, Dong

    2014-01-01

    Simvastatin (SIM), a widely used anti-lipidaemic drug, has been identified as a bone anabolic agent. Its poor water solubility and the lack of distribution to the skeleton, however, have limited its application in the treatment of bone metabolic diseases. In this study, an amphiphilic macromolecular prodrug of SIM was designed and synthesized to overcome these limitations. The polyethylene glycol (PEG)-based prodrug can spontaneously self-assemble to form micelles. The use of SIM trimer as the prodrug’s hydrophobic segment allows easy encapsulation of additional free SIM. The in vitro studies showed that SIM/SIM-mPEG micelles were internalized by MC3T3 cells via lysosomal trafficking and consistently induced expression of both BMP2 and DKK1 mRNA, suggesting that the prodrug micelle retains the biological functions of SIM. After systemic administration, optical imaging suggests that the micelles would passively target to bone fracture sites associated with hematoma and inflammation. Furthermore, flow cytometry study revealed that SIM/SIM-mPEG micelles had preferred cellular uptake by inflammatory and resident cells within the fracture callus tissue. The treatment study using a mouse osteotomy model validated the micelles’ therapeutic efficacy in promoting bone fracture healing as demonstrated by micro-CT and histological analyses. Collectively, these data suggest that the macromolecular prodrug-based micelle formulation of SIM may have great potential for clinical management of impaired fracture healing. PMID:25542644

  15. Antiangiogenic nanotherapy with lipase-labile Sn-2 fumagillin prodrug

    PubMed Central

    Pan, Dipanjan; Sanyal, Nibedita; Schmieder, Anne H; Senpan, Angana; Kim, Benjamin; Yang, Xiaoxia; Hu, Grace; Allen, John S; Gross, Richard W; Wickline, Samuel A; Lanza, Gregory M

    2012-01-01

    Background The chemical instability of antiangiogenic fumagillin, combined with its poor retention during intravascular transit, requires an innovative solution for clinical translation. We hypothesized that an Sn-2 lipase-labile fumagillin prodrug in combination with a contact-facilitated drug delivery mechanism, could be used to address these problems. Methods αvβ3-targeted and nontargeted nanoparticles with and without fumagillin in the prodrug or native forms were evaluated in vitro and in vivo in the Matrigel™ (BD Biosciences, CA, USA) plug model of angiogenesis in mice. Results In vitro experiments demonstrated that the new fumagillin prodrug decreased viability at least as efficacious as the parent compound, on an equimolar basis. In the Matrigel mouse angiogenesis model, αvβ3-fumagillin prodrug decreased angiogenesis as measured by MRI (3T), while the neovasculature was unaffected with the control nanoparticles. Conclusion The present approach resolved the previously intractable problems of drug instability and premature release in transit to target sites. PMID:22709347

  16. Theranostic reduction-sensitive gemcitabine prodrug micelles for near-infrared imaging and pancreatic cancer therapy

    NASA Astrophysics Data System (ADS)

    Han, Haijie; Wang, Haibo; Chen, Yangjun; Li, Zuhong; Wang, Yin; Jin, Qiao; Ji, Jian

    2015-12-01

    A biodegradable and reduction-cleavable gemcitabine (GEM) polymeric prodrug with in vivo near-infrared (NIR) imaging ability was reported. This theranostic GEM prodrug PEG-b-[PLA-co-PMAC-graft-(IR820-co-GEM)] was synthesized by ring-opening polymerization and ``click'' reaction. The as-prepared reduction-sensitive prodrug could self-assemble into prodrug micelles in aqueous solution confirmed by dynamic light scattering (DLS) and transmission electron microscopy (TEM). In vitro drug release studies showed that these prodrug micelles were able to release GEM in an intracellular-mimicking reductive environment. These prodrug micelles could be effectively internalized by BxPC-3 pancreatic cancer cells, which were observed by confocal laser scanning microscopy (CLSM). Meanwhile, a methyl thiazolyl tetrazolium (MTT) assay demonstrated that this prodrug exhibited high cytotoxicity against BxPC-3 cells. The in vivo whole-animal near-infrared (NIR) imaging results showed that these prodrug micelles could be effectively accumulated in tumor tissue and had a longer blood circulation time than IR820-COOH. The endogenous reduction-sensitive gemcitabine prodrug micelles with the in vivo NIR imaging ability might have great potential in image-guided pancreatic cancer therapy.A biodegradable and reduction-cleavable gemcitabine (GEM) polymeric prodrug with in vivo near-infrared (NIR) imaging ability was reported. This theranostic GEM prodrug PEG-b-[PLA-co-PMAC-graft-(IR820-co-GEM)] was synthesized by ring-opening polymerization and ``click'' reaction. The as-prepared reduction-sensitive prodrug could self-assemble into prodrug micelles in aqueous solution confirmed by dynamic light scattering (DLS) and transmission electron microscopy (TEM). In vitro drug release studies showed that these prodrug micelles were able to release GEM in an intracellular-mimicking reductive environment. These prodrug micelles could be effectively internalized by BxPC-3 pancreatic cancer cells, which

  17. Activation of multiple chemotherapeutic prodrugs by the natural enzymolome of tumour-localised probiotic bacteria.

    PubMed

    Lehouritis, Panos; Stanton, Michael; McCarthy, Florence O; Jeavons, Matthieu; Tangney, Mark

    2016-01-28

    Some chemotherapeutic drugs (prodrugs) require activation by an enzyme for efficacy. We and others have demonstrated the ability of probiotic bacteria to grow specifically within solid tumours following systemic administration, and we hypothesised that the natural enzymatic activity of these tumour-localised bacteria may be suitable for activation of certain such chemotherapeutic drugs. Several wild-type probiotic bacteria; Escherichia coli Nissle, Bifidobacterium breve, Lactococcus lactis and Lactobacillus species, were screened against a panel of popular prodrugs. All strains were capable of activating at least one prodrug. E. coli Nissle 1917 was selected for further studies because of its ability to activate numerous prodrugs and its resistance to prodrug toxicity. HPLC data confirmed biochemical transformation of prodrugs to their toxic counterparts. Further analysis demonstrated that different enzymes can complement prodrug activation, while simultaneous activation of multiple prodrugs (CB1954, 5-FC, AQ4N and Fludarabine phosphate) by E. coli was confirmed, resulting in significant efficacy improvement. Experiments in mice harbouring murine tumours validated in vitro findings, with significant reduction in tumour growth and increase in survival of mice treated with probiotic bacteria and a combination of prodrugs. These findings demonstrate the ability of probiotic bacteria, without the requirement for genetic modification, to enable high-level activation of multiple prodrugs specifically at the site of action. PMID:26655063

  18. Effects of a novel cyclic RGD peptidomimetic on cell proliferation, migration and angiogenic activity in human endothelial cells

    PubMed Central

    2014-01-01

    Background Cyclic RGD peptidomimetics containing a bifunctional diketopiperazine scaffold are a novel class of high-affinity ligands for the integrins αVβ3 and αVβ5. Since integrins are a promising target for the modulation of normal and pathological angiogenesis, the present study aimed at characterizing the ability of the RGD peptidomimetic cyclo[DKP-RGD] 1 proliferation, migration and network formation in human umbilical vein endothelial cells (HUVEC). Methods Cell viability was assessed by flow cytometry and annexin V (ANX)/propidium iodide (PI) staining. Cell proliferation was evaluated by the ELISA measurement of bromodeoxyuridine (BrdU) incorporation. Network formation by HUVEC cultured in Matrigel-coated plates was evaluated by optical microscopy and image analysis. Integrin subunit mRNA expression was assessed by real time-PCR and Akt phosphorylation by western blot analysis. Results Cyclo[DKP-RGD] 1 does not affect cell viability and proliferation either in resting conditions or in the presence of the pro-angiogenic growth factors VEGF, EGF, FGF, and IGF-I. Addition of cyclo[DKP-RGD] 1 however significantly decreased network formation induced by pro-angiogenic growth factors or by IL-8. Cyclo[DKP-RGD] 1 did not affect mRNA levels of αV, β3 or β5 integrin subunits, however it significantly reduced the phosphorylation of Akt. Conclusions Cyclo[DKP-RGD] 1 can be a potential modulator of angiogenesis induced by different growth factors, possibly devoid of the adverse effects of cytotoxic RGD peptidomimetic analogues. PMID:25053992

  19. Supramolecular Crafting of Self-Assembling Camptothecin Prodrugs with Enhanced Efficacy against Primary Cancer Cells.

    PubMed

    Su, Hao; Zhang, Pengcheng; Cheetham, Andrew G; Koo, Jin Mo; Lin, Ran; Masood, Asad; Schiapparelli, Paula; Quiñones-Hinojosa, Alfredo; Cui, Honggang

    2016-01-01

    Chemical modification of small molecule hydrophobic drugs is a clinically proven strategy to devise prodrugs with enhanced treatment efficacy. While this prodrug strategy improves the parent drug's water solubility and pharmacokinetic profile, it typically compromises the drug's potency against cancer cells due to the retarded drug release rate and reduced cellular uptake efficiency. Here we report on the supramolecular design of self-assembling prodrugs (SAPD) with much improved water solubility while maintaining high potency against cancer cells. We found that camptothecin (CPT) prodrugs created by conjugating two CPT molecules onto a hydrophilic segment can associate into filamentous nanostructures in water. Our results suggest that these SAPD exhibit much greater efficacy against primary brain cancer cells relative to that of irinotecan, a clinically used CPT prodrug. We believe these findings open a new avenue for rational design of supramolecular prodrugs for cancer treatment. PMID:27217839

  20. Tunable self-assembly of Irinotecan-fatty acid prodrugs with increased cytotoxicity to cancer cells†

    PubMed Central

    Xue, Xiangdong; Zhang, Tingbin; Jiang, Yonggang

    2016-01-01

    The development of a clinical chemotherapeutic is not an easy task. One challenge is how to deliver the agent to cancer cells. Nano-formulation of prodrugs, which combines the strengths of nanotechnology and prodrugs, possesses many advantages for chemotherapeutic drug delivery, including high drug loading efficiency, improved drug availability and enhanced accumulation in cancer cells. Here, we have constructed a small library of Irinotecan-derived prodrugs, in which the 20-hydroxyl group was derived with fatty-acid moieties through esterification. This conjugation fine-tuned the polarity of the Irinotecan molecule, thus enhancing the lipophilicity of the prodrugs and inducing their self-assembly into nanoparticles with different morphologies. These nano-formulated prodrugs accumulated at higher levels in cancer cells and were much more cytotoxic than free drugs. The rational design of prodrug-based nano-formulations opens a new avenue for the engineering of more efficient drug-delivery systems. PMID:27239311

  1. Supramolecular Crafting of Self-Assembling Camptothecin Prodrugs with Enhanced Efficacy against Primary Cancer Cells

    PubMed Central

    Su, Hao; Zhang, Pengcheng; Cheetham, Andrew G; Koo, Jin Mo; Lin, Ran; Masood, Asad; Schiapparelli, Paula; Quiñones-Hinojosa, Alfredo; Cui, Honggang

    2016-01-01

    Chemical modification of small molecule hydrophobic drugs is a clinically proven strategy to devise prodrugs with enhanced treatment efficacy. While this prodrug strategy improves the parent drug's water solubility and pharmacokinetic profile, it typically compromises the drug's potency against cancer cells due to the retarded drug release rate and reduced cellular uptake efficiency. Here we report on the supramolecular design of self-assembling prodrugs (SAPD) with much improved water solubility while maintaining high potency against cancer cells. We found that camptothecin (CPT) prodrugs created by conjugating two CPT molecules onto a hydrophilic segment can associate into filamentous nanostructures in water. Our results suggest that these SAPD exhibit much greater efficacy against primary brain cancer cells relative to that of irinotecan, a clinically used CPT prodrug. We believe these findings open a new avenue for rational design of supramolecular prodrugs for cancer treatment. PMID:27217839

  2. Preclinical Efficacy of a Carboxylesterase 2-Activated Prodrug of Doxazolidine

    PubMed Central

    Barthel, Benjamin L.; Zhang, Zhiyong; Rudnicki, Daniel L.; Coldren, Christopher D.; Polinkovsky, Margaret; Sun, Hengrui; Koch, Gary G.; Chan, Daniel C.F.; Koch, Tad H.

    2009-01-01

    Doxazolidine (Doxaz) is a functionally distinct formaldehyde conjugate of doxorubicin (Dox) that induces cancer cell death in Dox-sensitive and resistant cells. Pentyl PABC-Doxaz (PPD) is a prodrug of Doxaz that is activated by carboxylesterase 2 (CES2), which is expressed by liver, non-small cell lung, colon, pancreatic, renal, and thyroid cancer cells. Here, we demonstrate that in two murine models, PPD was effective at slowing tumor growth and demonstrated markedly reduced cardiotoxic and nephrotoxic effects, as well as better tolerance, relative to Dox. Hepatotoxicity, consistent with liver expression of the murine CES2 homolog, was induced by PPD. Unlike irinotecan, a clinical CES2-activated prodrug, PPD produced no visible gastrointestinal damage. Finally, we demonstrate that cellular response to PPD may be predicted with good accuracy using CES2 expression and Doxaz sensitivity, suggesting that these metrics may be useful as clinical biomarkers for sensitivity of a specific tumor to PPD treatment. PMID:19634903

  3. Prodrugs of phosphonates and phosphates: crossing the membrane barrier

    PubMed Central

    Wiemer, Andrew J.; Wiemer, David F.

    2016-01-01

    A substantial portion of metabolism involves transformation of phosphate esters, including pathways leading to nucleotides and oligonucleotides, carbohydrates, isoprenoids and steroids, and phosphorylated proteins. Because the natural substrates bear one or more negative charges, drugs that target these enzymes generally must be charged as well but small charged molecules can have difficulty traversing the cell membrane other than by endocytosis. The resulting dichotomy has stimulated abundant effort to develop effective prodrugs, compounds that carry little or no charge to enable them to transit biological membranes but then able to release the parent drug once inside the target cell. This chapter will present recent studies on advances in prodrug forms, along with representative examples of their application to marketed and developmental drugs. PMID:25391982

  4. Formulation and evaluation of suspensions: mefenamic acid prodrugs.

    PubMed

    Shah, Kamal; Shrivastava, Sushant K; Mishra, Pradeep

    2014-07-01

    Gastrosparing novel prodrugs (MAM and MAT) synthesized consisted of mefenamic acid (MA) with menthol (M) and thymol (T). Structural characterizations of synthesized esters were done by Infra red spectroscopy (IR), proton nuclear magnetic resonance (1HNMR), mass spectroscopy. After evaluation of pharmacological i.e. anti-inflammatory, analgesic and ulcerogenic activities, the preformulation studies were undertaken. Based on these a few formulation (suspensions) were designed and prepared. The formulated suspensions were evaluated for content uniformity, sedimentation volume, recovery studies, redispersibility, viscosity, pH, particle size, zeta potential, effect of temperature and in-vitro dissolution rate. All the above parameters were found to be within the limit these indicated that the synthesized esters are good candidate for liquid dosage form. Thus it can be concluded synthesized prodrugs can be formulated in suspension form. PMID:25015460

  5. Synthesis and Characterization of αvβ3-Targeting Peptidomimetic Chelate Conjugates for PET and SPECT Imaging

    PubMed Central

    Kim, Young-Seung; Nwe, Kido; Milenic, Diane E.; Brechbiel, Martin W.; Satz, Stanley; Baidoo, Kwamena E.

    2012-01-01

    There is growing interest in small peptidomimetic αvβ3 integrin antagonists that are readily synthesized and characterized and can be easily handled using physiological conditions. Peptidomimetic 4-[2-(3,4,5,6-tetrahydropyrimidine-2-ylamino)ethyloxy]benzoyl-2-[N-(3-amino-neopenta-1-carbamyl)]-aminoethylsulfonyl-amino-β-alanine (IAC) was successfully conjugated to 1-(1-carboxy-3-carbo-t-butoxypropyl)-4,7-(carbo-tert-butoxymethyl)-1,4,7-triazacyclononane (NODAGA(tBu)3) and 1-(1-carboxy-3-carbotertbutoxymethyl)-1,4,7,10-tetraazacyclododecane (DOTAGA(tBu)4) and radiolabeled with 111In, 67Ga and 203Pb. Results of a radioimmunoassay demonstrated binding to purified αvβ3 integrin when one to four equivalents of integrin were added to the reaction. Based on this promising result, investigations are moving forward to evaluate the NODA-GA-IAC and DOTA-GA-IAC conjugates for the targeting tumor associated angiogenesis and αvβ3 integrin positive tumors to define their PET and SPECT imaging qualities as well as their potential for delivery of therapeutic radionuclides. PMID:22853992

  6. Design, synthesis and anti-HIV-1 evaluation of hydrazide-based peptidomimetics as selective gelatinase inhibitors.

    PubMed

    Yang, Liang; Wang, Ping; Wu, Ji-Feng; Yang, Liu-Meng; Wang, Rui-Rui; Pang, Wei; Li, Yong-Gang; Shen, Yue-Mao; Zheng, Yong-Tang; Li, Xun

    2016-05-01

    As our ongoing work on research of gelatinase inhibitors, an array of hydrazide-containing peptidomimetic derivatives bearing quinoxalinone as well as spiro-heterocyclic backbones were designed, synthesized, and assayed for their in vitro enzymatic inhibitory effects. The results demonstrated that both the quinoxalinone (series I and II) and 1,4-dithia-7-azaspiro[4,4]nonane-based hydrazide peptidomimetics (series III) displayed remarkably selectivity towards gelatinase A as compared to APN, with IC50 values in the micromole range. Structure-activity relationships were herein briefly discussed. Given evidences have validated that gelatinase inhibition may be contributable to the therapy of HIV-1 infection, all the target compounds were also submitted to the preliminary in vitro anti-HIV-1 evaluation. It resulted that gelatinase inhibition really has positive correlation with anti-HIV-1 activity, especially compounds 4m and 7h, which gave enhanced gelatinase inhibition in comparison with the positive control LY52, and also decent anti-HIV-1 potencies. The FlexX docking results provided a straightforward insight into the binding pattern between inhibitors and gelatinase, as well as the selective inhibition towards gelatinase over APN. Collectively, our research encouraged potent gelatinase inhibitors might be used in the development of anti-HIV-1 agents. And else, compounds 4m and 7h might be promising candidates to be considered for further chemical optimization. PMID:27039251

  7. Desirability-based multi-criteria virtual screening of selective antimicrobial cyclic β-hairpin cationic peptidomimetics.

    PubMed

    Cruz-Monteagudo, Maykel; Romero, Yansy; Cordeiro, M Natália D S; Borges, Fernanda

    2013-01-01

    Today, emerging and increasing resistance to antibiotics has become a threat to public health worldwide. Antimicrobial peptides own unique action mechanisms making peptide antibiotics an attractive therapeutic option against resistant bacteria. However, their high haemolytic activity lacks the selectivity required for a human antibiotic. Therefore, additional efforts are needed to develop new antimicrobial peptides that possess greater selectivity for bacterial cells over erythrocytes. In this article, we introduce a chemoinformatics approach to simultaneously deal with these two conflicting properties consisting on a multi-criteria virtual screening strategy based on the use of a desirability-based multi-criteria classifier combined with similarity and chemometrics concepts. Here we propose a new quantitative feature encoding information related to the desirability, the degree of credibility ascribed to this desirability and the similarity of a candidate to a highly desirable query, which can be used as ranking criterion in a virtual screening campaign, the Desirability-Credibility- Similarity (DCS) Score. The enrichment ability of a multi-criteria virtual screening strategy based on the use of the DCS Score it is also assessed and compared to other virtual screening options. The results obtained evidenced that the use of the DCS score seems to be an efficient virtual screening strategy rendering promising overall and initial enrichment performance. Specifically, by using the DCS score it was possible to rank a selective antibacterial peptidomimetic earlier than a biologically inactive or non selective antibacterial peptidomimetic with a probability of ca. 0.9. PMID:23016843

  8. One-Step Way to Form Prodrug Micelles with High Amount Drug Loading.

    PubMed

    Zhang, Jing; Wu, Dan; Feng, Jie

    2016-06-01

    Prodrug micelles with high amount drug loading were obtained via one-step way. Antineoplastic drug doxorubicin (DOX), used as hydrophobic tail, was conjugated to hydrophilic head mPEG via hydrazone bonds, allowing drug release under intracellular condition. Free DOX was loaded into the hydrophobic core of micelles during the conjugation step simultaneously. Total drug content of the prodrug micelles was up to 61.2%. Endocytosis experiments confirmed that the prodrug micelles achieved good cellular-uptake ability. In vitro experiments indicated that the prodrug micelles showed better therapy efficacy than free drug in cancerous cells. PMID:27427600

  9. A Minireview: Usefulness of Transporter-Targeted Prodrugs in Enhancing Membrane Permeability.

    PubMed

    Murakami, Teruo

    2016-09-01

    Orally administered drugs are categorized into 4 classes depending on the solubility and permeability in a Biopharmaceutics Classification System. Prodrug derivatization is one of feasible approaches in modifying the physicochemical properties such as low solubility and low permeability without changing the in vivo pharmacological action of the parent drug. In this article, prodrug-targeted solute carrier (SLC) transporters were searched randomly by PubMed. Collected SLC transporters are amino acid transporter 1, bile acid transporter, carnitine transporter 2, glucose transporter 1, peptide transporter 1, vitamin C transporter 1, and multivitamin transporter. The usefulness of transporter-targeted prodrugs was evaluated in terms of membrane permeability, stability under acidic condition, and conversion to the parent drug. Among prodrugs collected, peptide transporter-targeted prodrugs exhibited the highest number, and some prodrugs such as valaciclovir and valganciclovir are clinically available. ATP-binding cassette efflux transporter, P-glycoprotein (P-gp), reduces the intestinal absorption of lipophilic P-gp substrate drugs, and SLC transporter-targeted prodrugs of P-gp substrate drugs circumvented the P-gp-mediated efflux transport. Thus, SLC transporter-targeted prodrug derivatization seems to be feasible approach to increase the oral bioavailability by overcoming various unwanted physicochemical properties of orally administered drugs, although the effect of food on prodrug absorption should be taken into consideration. PMID:27321236

  10. Stereoisomeric Prodrugs to Improve Corneal Absorption of Prednisolone: Synthesis and In Vitro Evaluation.

    PubMed

    Sheng, Ye; Yang, Xiaoyan; Wang, Zhiying; Mitra, Ashim K

    2016-06-01

    A series of stereoisomeric prodrugs have been designed to examine efficacy in generating higher corneal absorption relative to prednisolone. Prodrugs have been studied and identified with LC/MS/MS and NMR analyses. Prodrugs have been characterized for aqueous solubility, buffer stability, and cytotoxicity. Cellular uptake and permeability studies have been conducted across MDCK-MDR1 cells to determine prodrug affinity towards P-glycoprotein (P-gp) and peptide transporters. Enzyme-mediated degradation of prodrugs has been determined using Statens Seruminstitut rabbit cornea (SIRC) cell homogenates. Prodrugs exhibited higher aqueous solubility relative to prednisolone. Prodrugs circumvented P-gp-mediated cellular efflux and were recognized by peptide transporters. Prodrugs (DP, DDP) produced with D-isomers (D-valine) were significantly stable against both chemical and enzymatic hydrolyses. The order of degradation rate constants observed in chemical and enzymatic hydrolyses were in the same order, i.e., L-valine-L-valine-prednisolone (LLP) > L-valine-D-valine-prednisolone (LDP) > D-valine-L-valine-prednisolone (DLP) > D-valine-D-valine-prednisolone (DDP). Results obtained from this study clearly suggest that stereoisomeric prodrug approach is an effective strategy to overcome P-gp-mediated efflux and improve transcorneal permeability of prednisolone following topical administration. PMID:26335418

  11. Amino Acid Prodrugs: An Approach to Improve the Absorption of HIV-1 Protease Inhibitor, Lopinavir

    PubMed Central

    Patel, Mitesh; Mandava, Nanda; Gokulgandhi, Mitan; Pal, Dhananjay; Mitra, Ashim K.

    2014-01-01

    Poor systemic concentrations of lopinavir (LPV) following oral administration occur due to high cellular efflux by P-glycoprotein (P-gp) and multidrug resistance-associated proteins (MRPs) and extensive metabolism by CYP3A4 enzymes. In this study, amino acid prodrugs of LPV were designed and investigated for their potential to circumvent efflux processes and first pass effects. Three amino acid prodrugs were synthesized by conjugating isoleucine, tryptophan and methionine to LPV. Prodrug formation was confirmed by the LCMS/MS and NMR technique. Interaction of LPV prodrugs with efflux proteins were carried out in P-gp (MDCK-MDR1) and MRP2 (MDCK-MRP2) transfected cells. Aqueous solubility studies demonstrated that prodrugs generate higher solubility relative to LPV. Prodrugs displayed higher stability under acidic conditions and degraded significantly with rise in pH. Uptake and transport data suggested that prodrugs carry significantly lower affinity towards P-gp and MRP2 relative to LPV. Moreover, prodrugs exhibited higher liver microsomal stability relative to LPV. Hence, amino acid prodrug modification might be a viable approach for enhancing LPV absorption across intestinal epithelial and brain endothelial cells which expresses high levels of P-gp and MRP2. PMID:24727459

  12. Liposomal formulation of a glycerolipidic prodrug for lymphatic delivery of didanosine via oral route.

    PubMed

    Lalanne, M; Andrieux, K; Paci, A; Besnard, M; Ré, M; Bourgaux, C; Ollivon, M; Desmaele, D; Couvreur, P

    2007-11-01

    Didanosine is a polar drug with poor membrane absorption and high hepatic first pass metabolism. This study aimed at developing a lipidic formulation of a glycerolipidic prodrug of didanosine in order to improve its bioavailability. In the course of a preformulation study, the glycerolipidic prodrug of didanosine was characterized by microscopy, DSC and XRDT. In anhydrous conditions, the prodrug displayed a polymorphic behaviour similar to that of triglycerides. Then, we evaluated three types of lipidic formulations (emulsions, mixed micelles and liposomes) in order to encapsulate the prodrug. Solubilities in water - even in the presence of taurocholate micelles - but also in some oils were very low (max 244 microg/mL) as the prodrug was found to be amphiphilic (log P=2). On the contrary, the prodrug was found to be perfectly incorporated in dipalmitoylphosphatidylcholine (DPPC) multilamellar liposomes up to a ratio of 1:5 (mol:mol) prodrug:DPPC as suggested by HPLC-UV and DSC experiments. Moreover, these liposomes could be freeze-dried whereas the chemical integrity of the prodrug was preserved. Then, the freeze-dried liposomal preparation could be formulated as gastro-resistant capsules to prevent didanosine from acidic degradation. Further experiments are on the way to evaluate in vitro the absorption of prodrug incorporated in liposomes by enterocytes. PMID:17616448

  13. Follicle-stimulating Hormone Activates Extracellular Signal-regulated Kinase but Not Extracellular Signal-regulated Kinase Kinase through a 100-kDa Phosphotyrosine Phosphatase*

    PubMed Central

    Cottom, Joshua; Salvador, Lisa M.; Maizels, Evelyn T.; Reierstad, Scott; Park, Youngkyu; Carr, Daniel W.; Davare, Monika A.; Hell, Johannes W.; Palmer, Stephen S.; Dent, Paul; Kawakatsu, Hisaaki; Ogata, Masato; Hunzicker-Dunn, Mary

    2006-01-01

    In this report we sought to elucidate the mechanism by which the follicle-stimulating hormone (FSH) receptor signals to promote activation of the p42/p44 extracellular signal-regulated protein kinases (ERKs) in granulosa cells. Results show that the ERK kinase MEK and upstream intermediates Raf-1, Ras, Src, and L-type Ca2+ channels are already partially activated in vehicle-treated cells and that FSH does not further activate them. This tonic stimulatory pathway appears to be restrained at the level of ERK by a 100-kDa phosphotyrosine phosphatase that associates with ERK in vehicle-treated cells and promotes dephosphorylation of its regulatory Tyr residue, resulting in ERK inactivation. FSH promotes the phosphorylation of this phosphotyrosine phosphatase and its dissociation from ERK, relieving ERK from inhibition and resulting in its activation by the tonic stimulatory pathway and consequent translocation to the nucleus. Consistent with this premise, FSH-stimulated ERK activation is inhibited by the cell-permeable protein kinase A-specific inhibitor peptide Myr-PKI as well as by inhibitors of MEK, Src, a Ca2+ channel blocker, and chelation of extracellular Ca2+. These results suggest that FSH stimulates ERK activity in immature granulosa cells by relieving an inhibition imposed by a 100-kDa phosphotyrosine phosphatase. PMID:12493768

  14. A Screen for and Validation of Prodrug Antimicrobials

    PubMed Central

    Fleck, Laura E.; North, E. Jeffrey; Lee, Richard E.; Mulcahy, Lawrence R.; Casadei, Gabriele

    2014-01-01

    The rise of resistant pathogens and chronic infections tolerant to antibiotics presents an unmet need for novel antimicrobial compounds. Identifying broad-spectrum leads is challenging due to the effective penetration barrier of Gram-negative bacteria, formed by an outer membrane restricting amphipathic compounds, and multidrug resistance (MDR) pumps. In chronic infections, pathogens are shielded from the immune system by biofilms or host cells, and dormant persisters tolerant to antibiotics are responsible for recalcitrance to chemotherapy with conventional antibiotics. We reasoned that the dual need for broad-spectrum and sterilizing compounds could be met by developing prodrugs that are activated by bacterium-specific enzymes and that these generally reactive compounds could kill persisters and accumulate over time due to irreversible binding to targets. We report the development of a screen for prodrugs, based on identifying compounds that nonspecifically inhibit reduction of the viability dye alamarBlue, and then eliminate generally toxic compounds by testing for cytotoxicity. A large pilot of 55,000 compounds against Escherichia coli produced 20 hits, 3 of which were further examined. One compound, ADC111, is an analog of a known nitrofuran prodrug nitrofurantoin, and its activity depends on the presence of activating enzymes nitroreductases. ADC112 is an analog of another known antimicrobial tilbroquinol with unknown mechanism of action, and ADC113 does not belong to an approved class. All three compounds had a good spectrum and showed good to excellent activity against persister cells in biofilm and stationary cultures. These results suggest that screening for overlooked prodrugs may present a viable platform for antimicrobial discovery. PMID:24342644

  15. Cell-permeable succinate prodrugs bypass mitochondrial complex I deficiency.

    PubMed

    Ehinger, Johannes K; Piel, Sarah; Ford, Rhonan; Karlsson, Michael; Sjövall, Fredrik; Frostner, Eleonor Åsander; Morota, Saori; Taylor, Robert W; Turnbull, Doug M; Cornell, Clive; Moss, Steven J; Metzsch, Carsten; Hansson, Magnus J; Fliri, Hans; Elmér, Eskil

    2016-01-01

    Mitochondrial complex I (CI) deficiency is the most prevalent defect in the respiratory chain in paediatric mitochondrial disease. This heterogeneous group of diseases includes serious or fatal neurological presentations such as Leigh syndrome and there are very limited evidence-based treatment options available. Here we describe that cell membrane-permeable prodrugs of the complex II substrate succinate increase ATP-linked mitochondrial respiration in CI-deficient human blood cells, fibroblasts and heart fibres. Lactate accumulation in platelets due to rotenone-induced CI inhibition is reversed and rotenone-induced increase in lactate:pyruvate ratio in white blood cells is alleviated. Metabolomic analyses demonstrate delivery and metabolism of [(13)C]succinate. In Leigh syndrome patient fibroblasts, with a recessive NDUFS2 mutation, respiration and spare respiratory capacity are increased by prodrug administration. We conclude that prodrug-delivered succinate bypasses CI and supports electron transport, membrane potential and ATP production. This strategy offers a potential future therapy for metabolic decompensation due to mitochondrial CI dysfunction. PMID:27502960

  16. Macromolecular Pt(IV) Prodrugs from Poly(organo)phosphazenes.

    PubMed

    Henke, Helena; Kryeziu, Kushtrim; Banfić, Jelena; Theiner, Sarah; Körner, Wilfried; Brüggemann, Oliver; Berger, Walter; Keppler, Bernhard K; Heffeter, Petra; Teasdale, Ian

    2016-08-01

    The preparation of novel macromolecular prodrugs via the conjugation of two platinum(IV) complexes to suitably functionalized poly(organo)phosphazenes is presented. The inorganic/organic polymers provide carriers with controlled dimensions due to the use of living cationic polymerization and allow the preparation of conjugates with excellent aqueous solubility but long-term hydrolytic degradability. The macromolecular Pt(IV) prodrugs are designed to undergo intracellular reduction and simultaneous release from the macromolecular carrier to present the active Pt(II) drug derivatives. In vitro investigations show a significantly enhanced intracellular uptake of Pt for the macromolecular prodrugs when compared to small molecule Pt complexes, which is also reflected in an increase in cytotoxicity. Interestingly, drug-resistant sublines also show a significantly smaller resistance against the conjugates compared to clinically established platinum drugs, indicating that an alternative uptake route of the Pt(IV) conjugates might also be able to overcome acquired resistance against Pt(II) drugs. In vivo studies of a selected conjugate show improved tumor shrinkage compared to the respective Pt(IV) complex. PMID:27169668

  17. Disposition of a new tamibarotene prodrug in mice.

    PubMed

    Sugitani, Megumi; Abe, Rieko; Ikarashi, Nobutomo; Ito, Kiyomi; Muratake, Hideaki; Shudo, Koichi; Sugiyama, Kiyoshi

    2009-12-01

    Recently, a new compound IT-M-07000 was designed as a prodrug of tamibarotene, one of the therapeutic agents for acute promyelocytic leukemia. In the present study, IT-M-07000 was administered to mice to investigate whether it is actually metabolized to tamibarotene. Its metabolic pathway and the utility as a tamibarotene prodrug were also evaluated. After oral administration of IT-M-07000, IT-M-07000, tamibarotene and two compounds that were supposed to be metabolic intermediates in a beta-oxidation pathway of IT-M-07000 to tamibarotene were detected in mouse plasma. It was thus shown that IT-M-07000 is probably beta-oxidized to tamibarotene in mice. Comparison of tamibarotene concentration profiles after oral administration of IT-M-07000 or tamibarotene showed that the plasma tamibarotene concentration increased slower and was retained stable, and the area under the plasma concentration-time curve (AUC) of tamibarotene was larger in mice administered IT-M-07000 than tamibarotene. These results indicate that IT-M-07000 is possibly useful as a prodrug of tamibarotene. PMID:19952418

  18. Macromolecular Pt(IV) Prodrugs from Poly(organo)phosphazenes

    PubMed Central

    Banfić, Jelena; Theiner, Sarah; Körner, Wilfried; Brüggemann, Oliver; Berger, Walter; Keppler, Bernhard K.; Heffeter, Petra; Teasdale, Ian

    2016-01-01

    The preparation of novel macromolecular prodrugs via the conjugation of two platinum(IV) complexes to suitably functionalized poly(organo)phosphazenes is presented. The inorganic/organic polymers provide carriers with controlled dimensions due to the use of living cationic polymerization and allow the preparation of conjugates with excellent aqueous solubility but long-term hydrolytic degradability. The macromolecular Pt(IV) prodrugs are designed to undergo intracellular reduction and simultaneous release from the macromolecular carrier to present the active Pt(II) drug derivatives. In vitro investigations show a significantly enhanced intracellular uptake of Pt for the macromolecular prodrugs when compared to small molecule Pt complexes, which is also reflected in an increase in cytotoxicity. Interestingly, drug-resistant sublines also show a significantly smaller resistance against the conjugates compared to clinically established platinum drugs, indicating that an alternative uptake route of the Pt(IV) conjugates might also be able to overcome acquired resistance against Pt(II) drugs. In vivo studies of a selected conjugate show improved tumor shrinkage compared to the respective Pt(IV) complex. PMID:27169668

  19. Cell-permeable succinate prodrugs bypass mitochondrial complex I deficiency

    PubMed Central

    Ehinger, Johannes K.; Piel, Sarah; Ford, Rhonan; Karlsson, Michael; Sjövall, Fredrik; Frostner, Eleonor Åsander; Morota, Saori; Taylor, Robert W.; Turnbull, Doug M.; Cornell, Clive; Moss, Steven J.; Metzsch, Carsten; Hansson, Magnus J.; Fliri, Hans; Elmér, Eskil

    2016-01-01

    Mitochondrial complex I (CI) deficiency is the most prevalent defect in the respiratory chain in paediatric mitochondrial disease. This heterogeneous group of diseases includes serious or fatal neurological presentations such as Leigh syndrome and there are very limited evidence-based treatment options available. Here we describe that cell membrane-permeable prodrugs of the complex II substrate succinate increase ATP-linked mitochondrial respiration in CI-deficient human blood cells, fibroblasts and heart fibres. Lactate accumulation in platelets due to rotenone-induced CI inhibition is reversed and rotenone-induced increase in lactate:pyruvate ratio in white blood cells is alleviated. Metabolomic analyses demonstrate delivery and metabolism of [13C]succinate. In Leigh syndrome patient fibroblasts, with a recessive NDUFS2 mutation, respiration and spare respiratory capacity are increased by prodrug administration. We conclude that prodrug-delivered succinate bypasses CI and supports electron transport, membrane potential and ATP production. This strategy offers a potential future therapy for metabolic decompensation due to mitochondrial CI dysfunction. PMID:27502960

  20. Targeting the cyclin-binding groove site to inhibit the catalytic activity of CDK2/cyclin A complex using p27(KIP1)-derived peptidomimetic inhibitors.

    PubMed

    Karthiga, Arumugasamy; Tripathi, Sunil Kumar; Shanmugam, Ramasamy; Suryanarayanan, Venkatesan; Singh, Sanjeev Kumar

    2015-01-01

    Functionally activated cyclin-dependent kinase 2 (CDK2)/cyclin A complex has been validated as an interesting therapeutic target to develop the efficient antineoplastic drug based on the cell cycle arrest. Cyclin A binds to CDK2 and activates the kinases as well as recruits the substrate and inhibitors using a hydrophobic cyclin-binding groove (CBG). Blocking the cyclin substrate recruitment on CBG is an alternative approach to override the specificity hurdle of the currently available ATP site targeting CDK2 inhibitors. Greater understanding of the interaction of CDK2/cyclin A complex with p27 (negative regulator) reveals that the Leu-Phe-Gly (LFG) motif region of p27 binds with the CBG site of cyclin A to arrest the malignant cell proliferation that induces apoptosis. In the present study, Replacement with Partial Ligand Alternatives through Computational Enrichment (REPLACE) drug design strategies have been applied to acquire LFG peptide-derived peptidomimetics library. The peptidomimetics function is equivalent with respect to substrate p27 protein fashion but does not act as an ATP antagonist. The combined approach of molecular docking, molecular dynamics (MD), and molecular electrostatic potential and ADME/T prediction were carried out to evaluate the peptidomimetics. Resultant interaction and electrostatic potential maps suggested that smaller substituent is desirable at the position of phenyl ring to interact with Trp217, Arg250, and Gln254 residues in the active site. The best docked poses were refined by the MD simulations which resulted in conformational changes. After equilibration, the structure of the peptidomimetic and receptor complex was stable. The results revealed that the various substrate protein-derived peptidomimetics could serve as perfect leads against CDK2 protein. PMID:25584078

  1. Phosphotyrosine-mediated LAT assembly on membranes drives kinetic bifurcation in recruitment dynamics of the Ras activator SOS.

    PubMed

    Huang, William Y C; Yan, Qingrong; Lin, Wan-Chen; Chung, Jean K; Hansen, Scott D; Christensen, Sune M; Tu, Hsiung-Lin; Kuriyan, John; Groves, Jay T

    2016-07-19

    The assembly of cell surface receptors with downstream signaling molecules is a commonly occurring theme in multiple signaling systems. However, little is known about how these assemblies modulate reaction kinetics and the ultimate propagation of signals. Here, we reconstitute phosphotyrosine-mediated assembly of extended linker for the activation of T cells (LAT):growth factor receptor-bound protein 2 (Grb2):Son of Sevenless (SOS) networks, derived from the T-cell receptor signaling system, on supported membranes. Single-molecule dwell time distributions reveal two, well-differentiated kinetic species for both Grb2 and SOS on the LAT assemblies. The majority fraction of membrane-recruited Grb2 and SOS both exhibit fast kinetics and single exponential dwell time distributions, with average dwell times of hundreds of milliseconds. The minor fraction exhibits much slower kinetics, extending the dwell times to tens of seconds. Considering this result in the context of the multistep process by which the Ras GEF (guanine nucleotide exchange factor) activity of SOS is activated indicates that kinetic stabilization from the LAT assembly may be important. This kinetic proofreading effect would additionally serve as a stochastic noise filter by reducing the relative probability of spontaneous SOS activation in the absence of receptor triggering. The generality of receptor-mediated assembly suggests that such effects may play a role in multiple receptor proximal signaling processes. PMID:27370798

  2. Increased expression of receptor phosphotyrosine phosphatase-β/ζ is associated with molecular, cellular, behavioral and cognitive schizophrenia phenotypes

    PubMed Central

    Takahashi, N; Sakurai, T; Bozdagi-Gunal, O; Dorr, N P; Moy, J; Krug, L; Gama-Sosa, M; Elder, G A; Koch, R J; Walker, R H; Hof, P R; Davis, K L; Buxbaum, J D

    2011-01-01

    Schizophrenia is a serious and chronic mental disorder, in which both genetic and environmental factors have a role in the development of the disease. Neuregulin-1 (NRG1) is one of the most established genetic risk factors for schizophrenia, and disruption of NRG1 signaling has been reported in this disorder. We reported previously that NRG1/ErbB4 signaling is inhibited by receptor phosphotyrosine phosphatase-β/ζ (RPTP β/ζ) and that the gene encoding RPTPβ/ζ (PTPRZ1) is genetically associated with schizophrenia. In this study, we examined the expression of RPTPβ/ζ in the brains of patients with schizophrenia and observed increased expression of this gene. We developed mice overexpressing RPTPβ/ζ (PTPRZ1-transgenic mice), which showed reduced NRG1 signaling, and molecular and cellular changes implicated in the pathogenesis of schizophrenia, including altered glutamatergic, GABAergic and dopaminergic activity, as well as delayed oligodendrocyte development. Behavioral analyses also demonstrated schizophrenia-like changes in the PTPRZ1-transgenic mice, including reduced sensory motor gating, hyperactivity and working memory deficits. Our results indicate that enhanced RPTPβ/ζ signaling can contribute to schizophrenia phenotypes, and support both construct and face validity for PTPRZ1-transgenic mice as a model for multiple schizophrenia phenotypes. Furthermore, our results implicate RPTPβ/ζ as a therapeutic target in schizophrenia. PMID:22832403

  3. Global phosphotyrosine survey in triple-negative breast cancer reveals activation of multiple tyrosine kinase signaling pathways

    PubMed Central

    Ma, Binyun; Liu, Ren; Renuse, Santosh; Sahasrabuddhe, Nandini A.; Chen, Lily; Chaerkady, Raghothama; Kim, Min-Sik; Zhong, Jun; Jelinek, Christine; Barbhuiya, Mustafa A.; Leal-Rojas, Pamela; Yang, Yi; Kashyap, Manoj Kumar; Marimuthu, Arivusudar; Ling, Min; Fackler, Mary Jo; Merino, Vanessa; Zhang, Zhen; Zahnow, Cynthia A.; Gabrielson, Edward; Stearns, Vered; Roa, Juan Carlos; Sukumar, Saraswati; Gill, Parkash S.; Pandey, Akhilesh

    2015-01-01

    Breast cancer is the most prevalent cancer in women worldwide. About 15–20% of all breast cancers are triple negative breast cancer (TNBC) and are often highly aggressive when compared to other subtypes of breast cancers. To better characterize the biology that underlies the TNBC phenotype, we profiled the phosphotyrosine proteome of a panel of twenty-six TNBC cell lines using quantitative high resolution Fourier transform mass spectrometry. A heterogeneous pattern of tyrosine kinase activation was observed based on 1,789 tyrosine-phosphorylated peptides identified from 969 proteins. One of the tyrosine kinases, AXL, was found to be activated in a majority of aggressive TNBC cell lines and was accompanied by a higher level of AXL expression. High levels of AXL expression are correlated with a significant decrease in patient survival. Treatment of cells bearing activated AXL with a humanized AXL antibody inhibited cell proliferation and migration in vitro, and tumor growth in mice. Overall, our global phosphoproteomic analysis provided new insights into the heterogeneity in the activation status of tyrosine kinase pathways in TNBCs. Our approach presents an effective means of identifying important novel biomarkers and targets for therapy such as AXL in TNBC. PMID:26356563

  4. Light-switched inhibitors of protein tyrosine phosphatase PTP1B based on phosphonocarbonyl phenylalanine as photoactive phosphotyrosine mimetic.

    PubMed

    Wagner, Stefan; Schütz, Anja; Rademann, Jörg

    2015-06-15

    Phosphopeptide mimetics containing the 4-phosphonocarbonyl phenylalanine (pcF) as a photo-active phosphotyrosine isoster are developed as potent, light-switchable inhibitors of the protein tyrosine phosphatase PTP1B. The photo-active inhibitors 6-10 are derived from phosphopeptide substrates and are prepared from the suitably protected pcF building block 12 by Fmoc-based solid phase peptide synthesis. All pcF-containing peptides are moderate inhibitors of PTP1B with KI values between 10 and 50μM. Irradiation of the inhibitors at 365nm in the presence of the protein PTP1B amplify the inhibitory activity of pcF-peptides up to 120-fold, switching the KI values of the best inhibitors to the sub-micromolar range. Photo-activation of the inhibitors results in the formation of triplet intermediates of the benzoylphosphonate moiety, which deactivate PTP1B following an oxidative radical mechanism. Deactivation of PTP1B proceeds without covalent crosslinking of the protein target with the photo-switched inhibitors and can be reverted by subsequent addition of reducing agent dithiothreitol (DTT). PMID:25907367

  5. Jointly handling potency and toxicity of antimicrobial peptidomimetics by simple rules from desirability theory and chemoinformatics.

    PubMed

    Cruz-Monteagudo, Maykel; Borges, Fernanda; Cordeiro, M Natália D S

    2011-12-27

    Today, emerging and increasing resistance to antibiotics has become a threat to public health worldwide. Antimicrobial peptides have unique action mechanisms making them an attractive therapeutic prospect to be applied against resistant bacteria. However, the major drawback is related with their high hemolytic activity which cancels out the safety requirements for a human antibiotic. Therefore, additional efforts are needed to develop new antimicrobial peptides that possess a greater potency for bacterial cells and less or no toxicity over erythrocytes. In this paper, we introduce a practical approach to simultaneously deal with these two conflicting properties. The convergence of machine learning techniques and desirability theory allowed us to derive a simple, predictive, and interpretable multicriteria classification rule for simultaneously handling the antibacterial and hemolytic properties of a set of cyclic β-hairpin cationic peptidomimetics (Cβ-HCPs). The multicriteria classification rule exhibited a prediction accuracy of about 80% on training and external validation sets. Results from an additional concordance test have shown an excellent agreement between the multicriteria classification rule predictions and the predictions from independent classifiers for complementary antibacterial and hemolytic activities, respectively, evidencing the reliability of the multicriteria classification rule. The rule was also consistent with the general mode of action of cationic peptides pointing out its biophysical relevance. We also propose a multicriteria virtual screening strategy based on the joint use of the multicriteria classification rule, desirability, similarity, and chemometrics concepts. The ability of such a virtual screening strategy to prioritize selective (nonhemolytic) antibacterial Cβ-HCPs was assessed and challenged for their predictivity regarding the training, validation, and overall data. In doing so, we were able to rank a selective

  6. Delivery of siRNA Complexed with Palmitoylated α-Peptide/β-Peptoid Cell-Penetrating Peptidomimetics: Membrane Interaction and Structural Characterization of a Lipid-Based Nanocarrier System.

    PubMed

    Jing, Xiaona; Foged, Camilla; Martin-Bertelsen, Birte; Yaghmur, Anan; Knapp, Kolja M; Malmsten, Martin; Franzyk, Henrik; Nielsen, Hanne M

    2016-06-01

    Proteolytically stable α-peptide/β-peptoid peptidomimetics constitute promising cell-penetrating carrier candidates exhibiting superior cellular uptake as compared to commonly used cell-penetrating peptides (CPPs). The aim of the present study was to explore the potential of these peptidomimetics for delivery of small interfering RNA (siRNA) to the cytosol by incorporation of a palmitoylated peptidomimetic construct into a cationic lipid-based nanocarrier system. The optimal construct was selected on the basis of the effect of palmitoylation and the influence of the length of the peptidomimetic on the interaction with model membranes and the cellular uptake. Palmitoylation enhanced the peptidomimetic adsorption to supported lipid bilayers as studied by ellipsometry. However, both palmitoylation and increased peptidomimetic chain length were found to be beneficial in the cellular uptake studies using fluorophore-labeled analogues. Thus, the longer palmitoylated peptidomimetic was chosen for further formulation of siRNA in a dioleoylphosphatidylethanolamine/cholesteryl hemisuccinate (DOPE/CHEMS) nanocarrier system, and the resulting nanoparticles were found to mediate efficient gene silencing in vitro. Cryo-transmission electron microscopy (cryo-TEM) revealed multilamellar, onion-like spherical vesicles, and small-angle X-ray scattering (SAXS) analysis confirmed that the majority of the lipids in the nanocarriers were organized in lamellar structures, yet coexisted with a hexagonal phase, which is important for efficient nanocarrier-mediated endosomal escape of siRNA ensuring cytosolic delivery. The present work is a proof-of-concept for the use of α-peptides/β-peptoid peptidomimetics in an efficient delivery system that may be more generally exploited for the intracellular delivery of biomacromolecular drugs. PMID:26654841

  7. A novel anticancer theranostic pro-prodrug based on hypoxia and photo sequential control.

    PubMed

    Feng, Weipei; Gao, Chunyue; Liu, Wei; Ren, Huihui; Wang, Chao; Ge, Kun; Li, Shenghui; Zhou, Guoqiang; Li, Hongyan; Wang, Shuxiang; Jia, Guang; Li, Zhenhua; Zhang, Jinchao

    2016-08-01

    A novel anticancer pro-prodrug (GMC-CAE-NO2) with diagnosis and therapy functions based on hypoxia and photo sequential control was designed. It provides a platform for constructing theranostic pro-prodrugs to release active drugs controlled by hypoxic status and UV illumination. PMID:27379361

  8. In vivo evidence for a prodrug activation mechanism during colibactin maturation.

    PubMed

    Bian, Xiaoying; Fu, Jun; Plaza, Alberto; Herrmann, Jennifer; Pistorius, Dominik; Stewart, A Francis; Zhang, Youming; Müller, Rolf

    2013-07-01

    Releasing the cytopath: We have identified an N-myristoyl-D-asparagine (1) as the free N-terminal prodrug scaffold in cytopathogenic Escherichia coli strains expressing the colibactin gene cluster. Colibactin is released in vivo upon cleavage of precolibactin. We provide for the first time in vivo evidence of the prodrug-like release mechanism of colibactin. PMID:23744512

  9. The Dipeptide Monoester Prodrugs of Floxuridine and Gemcitabine—Feasibility of Orally Administrable Nucleoside Analogs

    PubMed Central

    Tsume, Yasuhiro; Bermejo, Blanca Borras; Amidon, Gordon L.

    2014-01-01

    Dipeptide monoester prodrugs of floxuridine and gemcitabine were synthesized. Their chemical stability in buffers, enzymatic stability in cell homogenates, permeability in mouse intestinal membrane along with drug concentration in mouse plasma, and anti-proliferative activity in cancer cells were determined and compared to their parent drugs. Floxuridine prodrug was more enzymatically stable than floxuridine and the degradation from prodrug to parent drug works as the rate-limiting step. On the other hand, gemcitabine prodrug was less enzymatically stable than gemcitabine. Those dipeptide monoester prodrugs exhibited 2.4- to 48.7-fold higher uptake than their parent drugs in Caco-2, Panc-1, and AsPC-1 cells. Floxuridine and gemcitabine prodrugs showed superior permeability in mouse jejunum to their parent drugs and exhibited the higher drug concentration in plasma after in situ mouse perfusion. Cell proliferation assays in ductal pancreatic cancer cells, AsPC-1 and Panc-1, indicated that dipeptide prodrugs of floxuridine and gemcitabine were more potent than their parent drugs. The enhanced potency of nucleoside analogs was attributed to their improved membrane permeability. The prodrug forms of 5′-l-phenylalanyl-l-tyrosyl-floxuridine and 5′-l-phenylalanyl-l-tyrosyl-gemcitabine appeared in mouse plasma after the permeation of intestinal membrane and the first-pass effect, suggesting their potential for the development of oral dosage form for anti-cancer agents. PMID:24473270

  10. Acid-responsive PEGylated doxorubicin prodrug nanoparticles for neuropilin-1 receptor-mediated targeted drug delivery.

    PubMed

    Song, Huijuan; Zhang, Ju; Wang, Weiwei; Huang, Pingsheng; Zhang, Yumin; Liu, Jianfeng; Li, Chen; Kong, Deling

    2015-12-01

    Self-assembled prodrug nanoparticles have demonstrated great promise in cancer chemotherapy. In the present study, we developed a new kind of prodrug nanoparticles for targeted drug delivery. PEGylated doxorubicin conjugate with an acid-cleavable cis-aconityl spacer was prepared. Then it was functionalized with a tumor-penetrating peptide, Cys-Arg-Gly-Asp-Lys (CRGDK), providing the prodrug nanoparticles with the specific binding ability to neurophilin-1 receptor. In acid mediums, doxorubicin could be released from the prodrug nanoparticles with an accumulative release around 60% through the acid-triggered hydrolysis of cis-aconityl bond and nanoparticle disassembly. Whereas, drug release was slow under a neutral pH and the accumulative drug release was less than 16%. In the cell culture tests, our prodrug nanoparticles showed enhanced endocytosis and cytotoxicity in cancer cells including HepG2, MCF-7 and MDA-MB-231 cells, but lower cytotoxicity in human cardiomyocyte H2C9. In the animal experiments, the prodrug nanoparticles were intravenously injected into Balb/c nude mice bearing MDA-MB-231 tumors. Enhanced drug penetration and accumulation in tumors, accompanying with a rapid early tumor-binding behavior, was observed after intravenous injection of the peptide modified prodrug nanoparticles. These data suggests that the acid-sensitive and tumor-targeting PEGylated doxorubicin prodrug nanoparticle may be an efficient drug delivery system for cancer chemotherapy. PMID:26433349

  11. Efficacious Cyclic N-Acyl O-Amino Phenol Duocarmycin Prodrugs

    PubMed Central

    Wolfe, Amanda L.; Duncan, Katharine K.; Parelkar, Nikhil K.; Brown, Douglas; Vielhauer, George A.; Boger, Dale L.

    2013-01-01

    Two novel cyclic N-acyl O-amino phenol prodrugs are reported as new members of a unique class of reductively cleaved prodrugs of the duocarmycin family of natural products. These prodrugs were explored with the expectation that they may be cleaved selectively within hypoxic tumor environments that have intrinsically higher concentrations of reducing nucleophiles and were designed to liberate the free drug without the release of an extraneous group. In vivo evaluation of the prodrug 6 showed that it exhibits extraordinary efficacy (T/C > 1500, L1210; 6/10 one year survivors) substantially exceeding that of the free drug, that its therapeutic window of activity is much larger permitting a dosing ≥ 40-fold higher than the free drug, and yet that it displays a potency in vivo that approaches the free drug (within 3-fold). Clearly, the prodrug 6 benefits from either its controlled slow release of the free drug or its preferential intracellular reductive cleavage. PMID:23627265

  12. Erythrocytes as carriers of new anti-opioid prodrugs: in vitro studies.

    PubMed

    Noël-Hocquet, S; Jabbouri, S; Lazar, S; Maunier, J C; Guillaumet, G; Ropars, C

    1992-01-01

    In using a reversed hypoosmotic lysis and resealing process, internalization of Naloxone or Naltrexone prodrugs as new opioid antagonists into red blood cells (RBC) can be obtained. We describe the entrapment method and haematological parameters of such RBC carriers. Percentages of internalization range between 35% and 70%, depending on the prodrug. The stability of the different prodrugs within RBC, after in vitro internalization have been compared at 4 degrees C and 37 degrees C. These studies were performed in order to screen and choose the most interesting prodrugs for in vivo studies. Some prodrugs tested appeared to be stable after 24 hours of incubation, but others were not. The intracellular ATP and 2,3 BPG concentrations of RBC carriers were similar to the control. PMID:1295310

  13. A Novel, Unusually Efficacious Duocarmycin Carbamate Prodrug That Releases No Residual Byproduct

    PubMed Central

    Wolfe, Amanda L.; Duncan, Katharine K.; Parelkar, Nikhil K.; Weir, Scott J.; Vielhauer, George A.; Boger, Dale L.

    2012-01-01

    A unique heterocyclic carbamate prodrug of seco-CBI-indole2 that releases no residual byproduct is reported as a new member of a class of hydrolyzable prodrugs of the duocarmycin and CC-1065 family of natural products. The prodrug was designed to be activated by hydrolysis of a carbamate releasing the free drug without the cleavage release of a traceable extraneous group. Unlike prior carbamate prodrugs examined that are rapidly cleaved in vivo, the cyclic carbamate was found to be exceptionally stable to hydrolysis under both chemical and biological conditions providing a slow, sustained release of the exceptionally potent free drug. An in vivo evaluation of the prodrug found that its efficacy exceeded that of the parent drug, that its therapeutic window of efficacy versus toxicity is much larger than the parent drug, and that its slow free drug release permitted the safe and efficacious use of doses 150-fold higher than the parent compound. PMID:22650244

  14. Pharmacological Evaluation and Preliminary Pharmacokinetics Studies of a New Diclofenac Prodrug without Gastric Ulceration Effect

    PubMed Central

    dos Santos, Jean Leandro; Moreira, Vanessa; Campos, Michel Leandro; Chelucci, Rafael Consolin; Barbieri, Karina Pereira; de Castro Souto, Pollyana Cristina Maggio; Matsubara, Márcio Hideki; Teixeira, Catarina; Bosquesi, Priscila Longhin; Peccinini, Rosângela Gonçalves; Chin, Chung Man

    2012-01-01

    Long-term nonsteroidal anti-inflammatory drugs (NSAIDs) therapy has been associated with several adverse effects such as gastric ulceration and cardiovascular events. Among the molecular modifications strategies, the prodrug approach is a useful tool to discover new safe NSAIDs. The 1-(2,6-dichlorophenyl)indolin-2-one is a diclofenac prodrug which demonstrated relevant anti-inflammatory properties without gastro ulceration effect. In addition, the prodrug decreases PGE2 levels, COX-2 expression and cellular influx into peritoneal cavity induced by carrageenan treatment. Preliminary pharmacokinetic studies have shown in vivo bioconversion of prodrug to diclofenac. This prodrug is a new nonulcerogenic NSAID useful to treat inflammatory events by long-term therapy. PMID:23203127

  15. A Potent Peptidomimetic Inhibitor of Botulinum Neurotoxin Serotype a Has a Very Different Conformation Than SNAP-25 Substrate

    SciTech Connect

    Zuniga, J.E.; Schmidt, J.J.; Fenn, T.; Burnett, J.C.; Arac, D.; Gussio, R.; Stafford, R.G.; Badie, S.S.; Bavari, S.; Brunger, A.T.

    2009-05-28

    Botulinum neurotoxin serotype A is the most lethal of all known toxins. Here, we report the crystal structure, along with SAR data, of the zinc metalloprotease domain of BoNT/A bound to a potent peptidomimetic inhibitor (Ki = 41 nM) that resembles the local sequence of the SNAP-25 substrate. Surprisingly, the inhibitor adopts a helical conformation around the cleavage site, in contrast to the extended conformation of the native substrate. The backbone of the inhibitor's P1 residue displaces the putative catalytic water molecule and concomitantly interacts with the 'proton shuttle' E224. This mechanism of inhibition is aided by residue contacts in the conserved S1' pocket of the substrate binding cleft, and the induction of new hydrophobic pockets, which are not present in the apo form, especially for the P2' residue of the inhibitor. Our inhibitor is specific for BoNT/A as it does not inhibit other BoNT serotypes or thermolysin.

  16. Substrate-competitive activity-based profiling of ester prodrug activating enzymes

    PubMed Central

    Xu, Hao; Majmudar, Jaimeen D.; Davda, Dahvid; Ghanakota, Phani; Kim, Ki H.; Carlson, Heather A.; Showalter, Hollis D.; Martin, Brent R.; Amidon, Gordon L.

    2015-01-01

    Understanding the mechanistic basis of prodrug delivery and activation is critical for establishing species-specific prodrug sensitivities necessary for evaluating pre-clinical animal models and potential drug-drug interactions. Despite significant adoption of prodrug methodologies for enhanced pharmacokinetics, functional annotation of prodrug activating enzymes is laborious and often unaddressed. Activity-based protein profiling (ABPP) describes an emerging chemoproteomic approach to assay active site occupancy within a mechanistically similar enzyme class in native proteomes. The serine hydrolase enzyme family is broadly reactive with reporter-linked fluorophosphonates, which have shown to provide a mechanism-based covalent labeling strategy to assay the activation state and active site occupancy of cellular serine amidases, esterases, and thioesterases. Here we describe a modified ABPP approach using direct substrate competition to identify activating enzymes for an ethyl ester prodrug, the influenza neuraminidase inhibitor oseltamivir. Substrate-competitive ABPP analysis identified carboxylesterase 1 (CES1) as an oseltamivir-activating enzyme in intestinal cell homogenates. Saturating concentrations of oseltamivir lead to a 4-fold reduction in the observed rate constant for CES1 inactivation by fluorophosphonates. WWL50, a reported carbamate inhibitor of mouse CES1, blocked oseltamivir hydrolysis activity in human cell homogenates, confirming CES1 is the primary prodrug activating enzyme for oseltamivir in human liver and intestinal cell lines. The related carbamate inhibitor WWL79 inhibited mouse, but not human CES1, providing a series of probes for analyzing prodrug activation mechanisms in different preclinical models. Overall, we present a substrate-competitive activity-based profiling approach for broadly surveying candidate prodrug hydrolyzing enzymes and outline the kinetic parameters for activating enzyme discovery, ester prodrug design and preclinical

  17. A Food Effect Study of an Oral Thrombin Inhibitor and Prodrug Approach To Mitigate It.

    PubMed

    Lee, Jihye; Kim, Bongchan; Kim, Tae Hun; Lee, Sun Hwa; Park, Hee Dong; Chung, Kyungha; Lee, Sung-Hack; Paek, Seungyup; Kim, Eunice EunKyeong; Yoon, SukKyoon; Kim, Aeri

    2016-04-01

    LB30870, a new direct thrombin inhibitor, showed 80% reduction in oral bioavailability in fed state. The present study aims to propose trypsin binding as a mechanism for such negative food effect and demonstrate a prodrug approach to mitigate food effect. Effect of food composition on fed state oral bioavailability of LB30870 was studied in dogs. Various prodrugs were synthesized, and their solubility, permeability, and trypsin binding affinity were measured. LB30870 and prodrugs were subject to cocrystallization with trypsin, and the X-ray structures of cocrystals were determined. Food effect was studied in dogs for selected prodrugs. Protein or lipid meal appeared to affect oral bioavailability of LB30870 in dogs more than carbohydrate meal. Blocking both carboxyl and amidine groups of LB30870 resulted in trypsin Ki values orders of magnitude higher than that of LB30870. Prodrugs belonged to either Biopharmaceutical Classification System I, II, or III. X-ray crystallography revealed that prodrugs did not bind to trypsin, but instead their hydrolysis product at the amidine blocking group formed cocrystal with trypsin. A prodrug with significantly less food effect than LB30870 was identified. Binding of prodrugs to food components such as dietary fiber appeared to counteract the positive effect brought with the prodrug approach. Further formulation research is warranted to enhance the oral bioavailability of prodrugs. In conclusion, this study is the first to demonstrate that the negative food effect of LB30870 can be attributed to trypsin binding. Trypsin binding study is proposed as a screening tool during lead optimization to minimize food effect. PMID:26886576

  18. Rational Design of Antirheumatic Prodrugs Specific for Sites of Inflammation

    PubMed Central

    Onuoha, Shimobi C.; Ferrari, Mathieu; Sblattero, Daniele

    2015-01-01

    Objective Biologic drugs, such as the anti–tumor necrosis factor (anti‐TNF) antibody adalimumab, have represented a breakthrough in the treatment of rheumatoid arthritis. Yet, concerns remain over their lack of efficacy in a sizable proportion of patients and their potential for systemic side effects such as infection. Improved biologic prodrugs specifically targeted to the site of inflammation have the potential to alleviate current concerns surrounding biologic anticytokine therapies. The purpose of this study was to design, construct, and evaluate in vitro and ex vivo the targeting and antiinflammatory capacity of activatable bispecific antibodies. Methods Activatable dual variable domain (aDVD) antibodies were designed and constructed to target intercellular adhesion molecule 1 (ICAM‐1), which is up‐regulated at sites of inflammation, and anti‐TNF antibodies (adalimumab and infliximab). These bispecific molecules included an external arm that targets ICAM‐1 and an internal arm that comprises the therapeutic domain of an anti‐TNF antibody. Both arms were linked to matrix metalloproteinase (MMP)–cleavable linkers. The constructs were tested for their ability to bind and neutralize both in vitro and ex vivo targets. Results Intact aDVD constructs demonstrated significantly reduced binding and anti‐TNF activity in the prodrug formulation as compared to the parent antibodies. Human synovial fluid and physiologic concentrations of MMP enzyme were capable of cleaving the external domain of the antibody, revealing a fully active molecule. Activated antibodies retained the same binding and anti‐TNF inhibitory capacities as the parent molecules. Conclusion The design of a biologic prodrug with enhanced specificity for sites of inflammation (synovium) and reduced specificity for off‐target TNF is described. This construct has the potential to form a platform technology that is capable of enhancing the therapeutic index of drugs for the treatment of

  19. Phosphotyrosine signaling analysis in human tumors is confounded by systemic ischemia-driven artifacts and intra-specimen heterogeneity.

    PubMed

    Gajadhar, Aaron S; Johnson, Hannah; Slebos, Robbert J C; Shaddox, Kent; Wiles, Kerry; Washington, Mary Kay; Herline, Alan J; Levine, Douglas A; Liebler, Daniel C; White, Forest M

    2015-04-01

    Tumor protein phosphorylation analysis may provide insight into intracellular signaling networks underlying tumor behavior, revealing diagnostic, prognostic or therapeutic information. Human tumors collected by The Cancer Genome Atlas program potentially offer the opportunity to characterize activated networks driving tumor progression, in parallel with the genetic and transcriptional landscape already documented for these tumors. However, a critical question is whether cellular signaling networks can be reliably analyzed in surgical specimens, where freezing delays and spatial sampling disparities may potentially obscure physiologic signaling. To quantify the extent of these effects, we analyzed the stability of phosphotyrosine (pTyr) sites in ovarian and colon tumors collected under conditions of controlled ischemia and in the context of defined intratumoral sampling. Cold-ischemia produced a rapid, unpredictable, and widespread impact on tumor pTyr networks within 5 minutes of resection, altering up to 50% of pTyr sites by more than 2-fold. Effects on adhesion and migration, inflammatory response, proliferation, and stress response pathways were recapitulated in both ovarian and colon tumors. In addition, sampling of spatially distinct colon tumor biopsies revealed pTyr differences as dramatic as those associated with ischemic times, despite uniform protein expression profiles. Moreover, intratumoral spatial heterogeneity and pTyr dynamic response to ischemia varied dramatically between tumors collected from different patients. Overall, these findings reveal unforeseen phosphorylation complexity, thereby increasing the difficulty of extracting physiologically relevant pTyr signaling networks from archived tissue specimens. In light of this data, prospective tumor pTyr analysis will require appropriate sampling and collection protocols to preserve in vivo signaling features. PMID:25670172

  20. Neighboring group catalysis in the design of nucleotide prodrugs.

    PubMed

    Khamnei, S; Torrence, P F

    1996-09-27

    An approach is described for potential application to the delivery of polar nucleosides and nucleotides across lipophilic membranes, namely, nucleotide prodrugs based on salicyl phosphate. 3'-Azido-3'-deoxythymidine (AZT) and 3'-deoxythymidine (ddT) were chosen as models. For the synthesis of prototype compounds 1 and 2, the approach was first to react either methyl salicylate (for 1) or phenyl salicylate (for 2) with phosphorus oxychloride in dry methylene chloride at 0 degree C with the addition of triethylamine as acid scavenger. The resulting intermediate phosphorodichloridate was reacted immediately with excess nucleoside under the same conditions. The control model compound 3 was prepared by reaction of phenyl phosphorodichloridate and excess nucleoside in pyridine/methylene chloride at 0 degree C to give 3 in 82% yield. The synthesis of triester 7 involved reaction of alpha-(chloroacetyl)salicyl chloride with 2,3,4,6-tetra-O-benzyl-D-glucopyranose to give [[(2,3,4,6-tetra-O-benzyl-D-glucopyranosyl)-oxy]carbonyl]-2- (1-chloroacetoxy)benzene (4) which was dechloroacetylated to 5,2,3,4,6-tetra-O-benzyl-D-glucopyranosyl salicylate. Phosphorylation of 5 with phosphorus oxychloride provided the phosphorodichloridate which was directly converted to 6 by reaction with dideoxythymidine. Removal of benzyl groups by catalytic hydrogenation gave compound 7, bis(2',3'-dideoxythymidin-5'-yl) D-glucopyranosyl phosphate. The AZT prodrug triesters, 1 and 2, underwent much more rapid hydrolysis than the triester 3, most probably due to the formation of an acyl phosphate complex from the attack on phosphorus of the salicylate carboxylate. The hydrolysis of the less lipophilic 7 was significantly slower than that of 1 or 2. Both pig liver esterase and rat brain cytosol were able to effect the cleavage to dinucleotide or mononucleotide of prodrug forms 2 and 7, much more rapidly than either 3 or 1, suggesting that the esterase-like enzymatic activity of rat brain was similar to

  1. Supramolecular curcumin-barium prodrugs for formulating with ceramic particles.

    PubMed

    Kamalasanan, Kaladhar; Anupriya; Deepa, M K; Sharma, Chandra P

    2014-10-01

    A simple and stable curcumin-ceramic combined formulation was developed with an aim to improve curcumin stability and release profile in the presence of reactive ceramic particles for potential dental and orthopedic applications. For that, curcumin was complexed with barium (Ba(2+)) to prepare curcumin-barium (BaCur) complex. Upon removal of the unbound curcumin and Ba(2+) by dialysis, a water-soluble BaCur complex was obtained. The complex was showing [M+1](+) peak at 10,000-20,000 with multiple fractionation peaks of MALDI-TOF-MS studies, showed that the complex was a supramolecular multimer. The (1)H NMR and FTIR studies revealed that, divalent Ba(2+) interacted predominantly through di-phenolic groups of curcumin to form an end-to-end complex resulted in supramolecular multimer. The overall crystallinity of the BaCur was lower than curcumin as per XRD analysis. The complexation of Ba(2+) to curcumin did not degrade curcumin as per HPLC studies. The fluorescence spectrum was blue shifted upon Ba(2+) complexation with curcumin. Monodisperse nanoparticles with size less than 200dnm was formed, out of the supramolecular complex upon dialysis, as per DLS, and upon loading into pluronic micelles the size was remaining in similar order of magnitude as per DLS and AFM studies. Stability of the curcumin was improved greater than 50% after complexation with Ba(2+) as per UV/Vis spectroscopy. Loading of the supramloecular nanoparticles into pluronic micelles had further improved the stability of curcumin to approx. 70% in water. These BaCur supramolecule nanoparticles can be considered as a new class of prodrugs with improved solubility and stability. Subsequently, ceramic nanoparticles with varying chemical composition were prepared for changing the material surface reactivity in terms of the increase in, degradability, surface pH and protein adsorption. Further, these ceramic particles were combined with curcumin prodrug formulations and optimized the curcumin release

  2. Discovery and Characterization of a Water-Soluble Prodrug of a Dual Inhibitor of Bacterial DNA Gyrase and Topoisomerase IV

    PubMed Central

    2015-01-01

    Benzimidazole 1 is the lead compound resulting from an antibacterial program targeting dual inhibitors of bacterial DNA gyrase and topoisomerase IV. With the goal of improving key drug-like properties, namely, the solubility and the formulability of 1, an effort to identify prodrugs was undertaken. This has led to the discovery of a phosphate ester prodrug 2. This prodrug is rapidly cleaved to the parent drug molecule upon both oral and intravenous administration. The prodrug achieved equivalent exposure of 1 compared to dosing the parent in multiple species. The prodrug 2 has improved aqueous solubility, simplifying both intravenous and oral formulation. PMID:26191374

  3. Novel Biotinylated Lipid Prodrugs of Acyclovir for the Treatment of Herpetic Keratitis (HK): Transporter Recognition, Tissue Stability and Antiviral Activity

    PubMed Central

    Vadlapudi, Aswani Dutt; Vadlapatla, Ramya Krishna; Earla, Ravinder; Sirimulla, Suman; Bailey, Jake Brain; Pal, Dhananjay; Mitra, Ashim K.

    2013-01-01

    Purpose Biotinylated lipid prodrugs of acyclovir (ACV) were designed to target the sodium dependent multivitamin transporter (SMVT) on the cornea to facilitate enhanced cellular absorption of ACV. Methods All the prodrugs were screened for in vitro cellular uptake, interaction with SMVT, docking analysis, cytotoxicity, enzymatic stability and antiviral activity. Results Uptake of biotinylated lipid prodrugs of ACV (B-R-ACV and B-12HS-ACV) was significantly higher than biotinylated prodrug (B-ACV), lipid prodrugs (R-ACV and 12HS-ACV) and ACV in corneal cells. Transepithelial transport across rabbit corneas indicated the recognition of the prodrugs by SMVT. Average Vina scores obtained from docking studies further confirmed that biotinylated lipid prodrugs possess enhanced affinity towards SMVT. All the prodrugs studied did not cause any cytotoxicity and were found to be safe and non-toxic. B-R-ACV and B-12HS-ACV were found to be relatively more stable in ocular tissue homogenates and exhibited excellent antiviral activity. Conclusions Biotinylated lipid prodrugs demonstrated synergistic improvement in cellular uptake due to recognition of the prodrugs by SMVT on the cornea and lipid mediated transcellular diffusion. These biotinylated lipid prodrugs appear to be promising drug candidates for the treatment of herpetic keratitis (HK) and may lower ACV resistance in patients with poor clinical response. PMID:23657675

  4. Opioid Peptidomimetics: Leads for the Design of Bioavailable Mixed Efficacy Mu Opioid Receptor (MOR) Agonist/Delta Opioid Receptor (DOR) Antagonist Ligands

    PubMed Central

    Mosberg, Henry I.; Yeomans, Larisa; Harland, Aubrie A.; Bender, Aaron M.; Sobczyk-Kojiro, Katarzyna; Anand, Jessica P.; Clark, Mary J.; Jutkiewicz, Emily M.; Traynor, John R.

    2013-01-01

    We have previously described opioid peptidomimetic, 1, employing a tetrahydroquinoline scaffold and modeled on a series of cyclic tetrapeptide opioid agonists. We have recently described modifications to these peptides that confer a mu opioid receptor (MOR) agonist, delta opioid receptor (DOR) antagonist profile, which has been shown to reduce the development of tolerance to the analgesic actions of MOR agonists. Several such bifunctional ligands have been reported, but none has been demonstrated to cross the blood brain barrier. Here we describe the transfer of structural features that evoked MOR agonist/DOR antagonist behavior in the cyclic peptides to the tetrahydroquinoline scaffold and show that the resulting peptidomimetics maintain the desired pharmacological profile. Further, the 4R diastereomer of 1 was fully efficacious and approximately equipotent to morphine in the mouse warm water tail withdrawal assay following intraperitoneal administration and thus a promising lead for the development of opioid analgesics with reduced tolerance. PMID:23419026

  5. ALA-Butyrate prodrugs for Photo-Dynamic Therapy

    NASA Astrophysics Data System (ADS)

    Berkovitch, G.; Nudelman, A.; Ehenberg, B.; Rephaeli, A.; Malik, Z.

    2010-05-01

    The use of 5-aminolevulinic acid (ALA) administration has led to many applications of photodynamic therapy (PDT) in cancer. However, the hydrophilic nature of ALA limits its ability to penetrate the cells and tissues, and therefore the need for ALA derivatives became an urgent research target. In this study we investigated the activity of novel multifunctional acyloxyalkyl ester prodrugs of ALA that upon metabolic hydrolysis release active components such as, formaldehyde, and the histone deacetylase inhibitory moiety, butyric acid. Evaluation of these prodrugs under photo-irradiation conditions showed that butyryloxyethyl 5-amino-4-oxopentanoate (ALA-BAC) generated the most efficient photodynamic destruction compared to ALA. ALA-BAC stimulated a rapid biosynthesis of protoporphyrin IX (PpIX) in human glioblastoma U-251 cells which resulted in generation of intracellular ROS, reduction of mitochondrial activity, leading to apoptotic and necrotic death of the cells. The apoptotic cell death induced by ALA / ALA-BAC followed by PDT equally activate intrinsic and extrinsic apoptotic signals and both pathways may occur simultaneously. The main advantage of ALA-BAC over ALA stems from its ability to induce photo-damage at a significantly lower dose than ALA.

  6. Macrophage mediated PCI enhanced gene-directed enzyme prodrug therapy

    NASA Astrophysics Data System (ADS)

    Christie, Catherine E.; Zamora, Genesis; Kwon, Young J.; Berg, Kristian; Madsen, Steen J.; Hirschberg, Henry

    2015-03-01

    Photochemical internalization (PCI) is a photodynamic therapy-based approach for improving the delivery of macromolecules and genes into the cell cytosol. Prodrug activating gene therapy (suicide gene therapy) employing the transduction of the E. coli cytosine deaminase (CD) gene into tumor cells, is a promising method. Expression of this gene within the target cell produces an enzyme that converts the nontoxic prodrug, 5-FC, to the toxic metabolite, 5-fluorouracil (5-FU). 5-FC may be particularly suitable for brain tumors, because it can readily cross the bloodbrain barrier (BBB). In addition the bystander effect, where activated drug is exported from the transfected cancer cells into the tumor microenvironment, plays an important role by inhibiting growth of adjacent tumor cells. Tumor-associated macrophages (TAMs) are frequently found in and around glioblastomas. Monocytes or macrophages (Ma) loaded with drugs, nanoparticles or photosensitizers could therefore be used to target tumors by local synthesis of chemo attractive factors. The basic concept is to combine PCI, to enhance the ex vivo transfection of a suicide gene into Ma, employing specially designed core/shell NP as gene carrier.

  7. Preparation of alginate beads containing a prodrug of diethylenetriaminepentaacetic acid

    PubMed Central

    Yang, Yu-Tsai; Di Pasqua, Anthony J.; He, Weiling; Tsai, Tsuimin; Sueda, Katsuhiko; Zhang, Yong; Jay, Michael

    2012-01-01

    A penta-ethyl ester prodrug of the radionuclide decorporation agent diethylenetriaminepentaacetic acid (DTPA), which exists as an oily liquid, was encapsulated in alginate beads by the ionotropic gelation method. An optimal formulation was found by varying initial concentrations of DTPA pentaethyl ester, alginate polymer, Tween 80 surfactant and calcium chloride. All prepared alginate beads were ~1.6 mm in diameter, and the optimal formulation had loading and encapsulation efficiencies of 91.0 ± 1.1 and 72.6 ± 2.2%, respectively, and only 3.2 ± 0.8% water absorption after storage at room temperature in ~80% relative humidity. Moreover, Fourier transform infrared spectroscopy showed that DTPA penta-ethyl ester did not react with excipients during formation of the DTPA penta-ethyl ester-containing alginate beads. Release of prodrug from alginate beads was via anomalous transport, and its stability enhanced by encapsulation. Collectively, these data suggest that this solid dosage form may be suitable for oral administration after radionuclide contamination. PMID:23399237

  8. Kinetic Studies and Bioactivity of Potential Manzamine Prodrugs

    PubMed Central

    Shilabin, Abbas Gholipour; Kasanah, Noer; Tekwani, Babu L.; Hamann, Mark T.

    2016-01-01

    The manzamines represent a class of marine natural products that show considerable promise in the control of malaria but generate GI distress in rodents when administered orally in high doses. In an effort to generate manzamine prodrugs with improved antimalarial activity and reduced GI toxicity, we prepared acetylated 8-hydroxymanzamine A analogues including 8-acetoxymanzamine A (3) and 8,12-diacetoxymanzamine A (4), and 8-methoxymanzamine A (5) beginning with 8-hydroxymanzamine A (2). The semisynthetic analogues were assayed for antimalarial and antimicrobial activities, cytotoxicity, and biological and chemical stability. Due to gradual hydrolysis of the ester group, application of monoacetate 3 as an antimalarial prodrug was investigated. The in vitro and in vivo bioassays show that acetylated analogues exhibit significant antimalarial activity (IC50(3) 9.6–30 ng/mL), which are comparable to the parent molecule; however the monoaceate 3 was shown to actually produce higher toxicity at 30 mg/kg when administered orally. PMID:18598080

  9. Crystal Structure of Human E-Cadherin-EC1EC2 in Complex with a Peptidomimetic Competitive Inhibitor of Cadherin Homophilic Interaction.

    PubMed

    Nardone, Valentina; Lucarelli, Anna Paola; Dalle Vedove, Andrea; Fanelli, Roberto; Tomassetti, Antonella; Belvisi, Laura; Civera, Monica; Parisini, Emilio

    2016-05-26

    Cadherins are transmembrane cell adhesion proteins whose aberrant expression often correlates with cancer development and proliferation. We report the crystal structure of an E-cadherin extracellular fragment in complex with a peptidomimetic compound that was previously shown to partially inhibit cadherin homophilic adhesion. The structure reveals an unexpected binding mode and allows the identification of a druggable cadherin interface, thus paving the way to a future structure-guided design of cell adhesion inhibitors against cadherin-expressing solid tumors. PMID:27120112

  10. IN VITRO PERMEATION OF A PEGYLATED NALTREXONE PRODRUG ACROSS MICRONEEDLE-TREATED SKIN

    PubMed Central

    Milewski, Mikolaj; Yerramreddy, Thirupathi Reddy; Ghosh, Priyanka; Crooks, Peter A.; Stinchcomb, Audra L.

    2010-01-01

    Microneedles (MN) are a useful tool for increasing skin permeability to xenobiotics. Previous research showed marked improvement in the percutaneous flux of naltrexone (NTX) hydrochloride by the use of MN skin pretreatment alone; however, for better therapeutic effect, further enhancement is desired. The goal of this in vitro study was to combine microneedle skin pretreatment with the use of a highly water-soluble PEGylated naltrexone prodrug (polyethyleneglycol-NTX, PEG-NTX) to investigate its transdermal transport at varying concentrations. Solubility and stability of the prodrug were investigated. In vitro diffusion experiments employing MN-treated minipig skin were used to evaluate the performance of the PEGylated prodrug. The results revealed substantial deviation from ideal behavior, with the flux through MN-treated skin having a nonlinear relationship to the prodrug concentration in the donor solution. While in the lower concentration range tested the prodrug flux increase was proportional to the concentration increase, at high concentrations it showed no such dependence. Accounting for the decrease in the effective prodrug diffusivity accompanying the increase in viscosity, as predicted by the Stokes-Einstein equation, provided a rationale for the observed flux values. Increasing the viscosity of the donor solution is hypothesized to afford a curvilinear permeation profile for the PEGylated NTX prodrug. PMID:20678989

  11. In vitro permeation of a pegylated naltrexone prodrug across microneedle-treated skin.

    PubMed

    Milewski, Mikolaj; Yerramreddy, Thirupathi Reddy; Ghosh, Priyanka; Crooks, Peter A; Stinchcomb, Audra L

    2010-08-17

    Microneedles (MN) are a useful tool for increasing skin permeability to xenobiotics. Previous research showed marked improvement in the percutaneous flux of naltrexone (NTX) hydrochloride by the use of MN skin pretreatment alone; however, for better therapeutic effect, further enhancement is desired. The goal of this in vitro study was to combine microneedle skin pretreatment with the use of a highly water-soluble PEGylated naltrexone prodrug (polyethyleneglycol-NTX, PEG-NTX) to investigate its transdermal transport at varying concentrations. Solubility and stability of the prodrug were investigated. In vitro diffusion experiments employing MN-treated minipig skin were used to evaluate the performance of the PEGylated prodrug. The results revealed substantial deviation from ideal behavior, with the flux through MN-treated skin having a nonlinear relationship to the prodrug concentration in the donor solution. While in the lower concentration range tested the prodrug flux increase was proportional to the concentration increase, at high concentrations it showed no such dependence. Accounting for the decrease in the effective prodrug diffusivity accompanying the increase in viscosity, as predicted by the Stokes-Einstein equation, provided a rationale for the observed flux values. Increasing the viscosity of the donor solution is hypothesized to afford a curvilinear permeation profile for the PEGylated NTX prodrug. PMID:20678989

  12. Mutual Amide Prodrug of Etodolac-glucosamine: Synthesis, Characterization and Pharmacological Screening

    PubMed Central

    Pandey, Preeti; Pandey, S.; Dubey, Shaifali

    2013-01-01

    Etodolac, a nonsteroidal antiinflammatory drug, widely used in arthritis is associated with gastric ulceration and irritation due to presence of free carboxylic group. The current investigation reports synthesis of mutual amide prodrug of etodolac by masking free carboxylic group with glucosamine, a nutritional supplement for treatment of arthritis. Confirmation and characterization of the structure of the synthesized prodrug done by elemental and spectroscopy analysis, melting point, determination of migration parameters (Rf, RM, and Rt) by using thin layer chromatography and high performance liquid chromatography, respectively. Partition coefficient and solubility study confirms its lipophilic character so can be suitable candidate for controlled release delivery. In vitro hydrolytic studies of prodrug confirms good rate of hydrolysis in blood plasma, fecal matter, and simulated intestinal fluid while stable in gastric simulated fluid. In vivo pharmacological screening performed on animals. Prodrug with respect to etodolac shows good analgesic, antiinflammatory, and antiarthritic activity. The prodrug was assessed for their probable damaging effects by ulcerogeniticity and histopathological analysis. The histopathological studies showed less ulceration in the gastric region when treated with prodrug, thereby proving the prodrug to be better in action as compared to etodolac and are advantageous in having less gastrointestinal side effects. PMID:24302794

  13. Mutual Amide Prodrug of Etodolac-glucosamine: Synthesis, Characterization and Pharmacological Screening.

    PubMed

    Pandey, Preeti; Pandey, S; Dubey, Shaifali

    2013-07-01

    Etodolac, a nonsteroidal antiinflammatory drug, widely used in arthritis is associated with gastric ulceration and irritation due to presence of free carboxylic group. The current investigation reports synthesis of mutual amide prodrug of etodolac by masking free carboxylic group with glucosamine, a nutritional supplement for treatment of arthritis. Confirmation and characterization of the structure of the synthesized prodrug done by elemental and spectroscopy analysis, melting point, determination of migration parameters (Rf, RM, and Rt) by using thin layer chromatography and high performance liquid chromatography, respectively. Partition coefficient and solubility study confirms its lipophilic character so can be suitable candidate for controlled release delivery. In vitro hydrolytic studies of prodrug confirms good rate of hydrolysis in blood plasma, fecal matter, and simulated intestinal fluid while stable in gastric simulated fluid. In vivo pharmacological screening performed on animals. Prodrug with respect to etodolac shows good analgesic, antiinflammatory, and antiarthritic activity. The prodrug was assessed for their probable damaging effects by ulcerogeniticity and histopathological analysis. The histopathological studies showed less ulceration in the gastric region when treated with prodrug, thereby proving the prodrug to be better in action as compared to etodolac and are advantageous in having less gastrointestinal side effects. PMID:24302794

  14. Identification of novel nitroreductases from Bacillus cereus and their interaction with the CB1954 prodrug.

    PubMed

    Gwenin, Vanessa V; Poornima, Paramasivan; Halliwell, Jennifer; Ball, Patrick; Robinson, George; Gwenin, Chris D

    2015-12-01

    Directed enzyme prodrug therapy is a form of cancer chemotherapy in which bacterial prodrug-activating enzymes, or their encoding genes, are directed to the tumour before administration of a prodrug. The prodrug can then be activated into a toxic drug at the tumour site, reducing off-target effects. The bacterial nitroreductases are a class of enzymes used in this therapeutic approach and although very promising, the low turnover rate of prodrug by the most studied nitroreductase enzyme, NfnB from Escherichia coli (NfnB_Ec), is a major limit to this technology. There is a continual search for enzymes with greater efficiency, and as part of the search for more efficient bacterial nitroreductase enzymes, two novel enzymes from Bacillus cereus (strain ATCC 14579) have been identified and shown to reduce the CB1954 (5-(aziridin-1-yl)-2,4-dinitrobenzamide) prodrug to its respective 2'-and 4'-hydroxylamine products. Both enzymes shared features characteristic of the nitro-FMN-reductase superfamily including non-covalently associated FMN, requirement for the NAD(P)H cofactor, homodimeric, could be inhibited by Dicoumarol (3,3'-methylenebis(4-hydroxy-2H-chromen-2-one)), and displayed ping pong bi bi kinetics. Based on the biochemical characteristics and nucleotide alignment with other nitroreductase enzymes, one enzyme was named YdgI_Bc and the other YfkO_Bc. Both B. cereus enzymes had greater turnover for the CB1954 prodrug compared with NfnB_Ec, and in the presence of added NADPH cofactor, YfkO_Bc had superior cell killing ability, and produced mainly the 4'-hydroxylamine product at low prodrug concentration. The YfkO_Bc was identified as a promising candidate for future enzyme prodrug therapy. PMID:26415543

  15. Synthesis and pharmacological evaluation of nucleoside prodrugs designed to target siderophore biosynthesis in Mycobacterium tuberculosis.

    PubMed

    Dawadi, Surendra; Kawamura, Shuhei; Rubenstein, Anja; Remmel, Rory; Aldrich, Courtney C

    2016-03-15

    The nucleoside antibiotic, 5'-O-[N-(salicyl)sulfamoyl]adenosine (1), possesses potent whole-cell activity against Mycobacterium tuberculosis (Mtb), the etiological agent of tuberculosis (TB). This compound is also active in vivo, but suffers from poor drug disposition properties that result in poor bioavailability and rapid clearance. The synthesis and evaluation of a systematic series of lipophilic ester prodrugs containing linear and α-branched alkanoyl groups from two to twelve carbons at the 3'-position of a 2'-fluorinated analog of 1 is reported with the goal to improve oral bioavailability. The prodrugs were stable in simulated gastric fluid (pH 1.2) and under physiological conditions (pH 7.4). The prodrugs were also remarkably stable in mouse, rat, and human serum (relative serum stability: human∼rat≫mouse) displaying a parabolic trend in the SAR with hydrolysis rates increasing with chain length up to eight carbons (t1/2=1.6 h for octanoyl prodrug 7 in mouse serum) and then decreasing again with higher chain lengths. The permeability of the prodrugs was also assessed in a Caco-2 cell transwell model. All of the prodrugs were found to have reduced permeation in the apical-to-basolateral direction and enhanced permeation in the basolateral-to-apical direction relative to the parent compound 2, resulting in efflux ratios 5-28 times greater than 2. Additionally, Caco-2 cells were found to hydrolyze the prodrugs with SAR mirroring the serum stability results and a preference for hydrolysis on the apical side. Taken together, these results suggest that the described prodrug strategy will lead to lower than expected oral bioavailability of 2 and highlight the contribution of intestinal esterases for prodrug hydrolysis. PMID:26875934

  16. An In Silico Insight into Novel Therapeutic Interaction of LTNF Peptide-LT10 and Design of Structure Based Peptidomimetics for Putative Anti-Diabetic Activity

    PubMed Central

    Chavan, Sonali Gopichand; Deobagkar, Deepti Dileep

    2015-01-01

    Lethal Toxin Neutralizing Factor (LTNF) obtained from Opossum serum (Didephis virginiana) is known to exhibit toxin-neutralizing activity for envenomation caused by animals, plants and bacteria. Small synthetic peptide- LT10 (10mer) derived from N-terminal fraction of LTNF exhibit similar anti-lethal and anti-allergic property. In our in silico study, we identified Insulin Degrading Enzyme (IDE) as a potential target of LT10 peptide followed by molecular docking and molecular dynamic (MD) simulation studies which revealed relatively stable interaction of LT10 peptide with IDE. Moreover, their detailed interaction analyses dictate IDE-inhibitory interactions of LT10 peptide. This prediction ofLT10 peptide as a novel putative IDE-inhibitor suggests its possible role in anti-diabetic treatment since IDE- inhibitors are known to assist treatment of Diabetes mellitus by enhancing insulin signalling. Furthermore, series of structure based peptidomimetics were designed from LT10 peptide and screened for their inhibitory interactions which ultimately led to a small set of peptidomimetic inhibitors of IDE. These peptidomimetic thus might provide a new class of IDE-inhibitors, those derived from LT10 peptide. PMID:25816209

  17. In situ activation of a doxorubicin prodrug using imaging-capable nanoparticles.

    PubMed

    Khan, Irfan; Agris, Paul F; Yigit, Mehmet V; Royzen, Maksim

    2016-05-01

    A general strategy for image-guided prodrug activation using fluorescently-labeled magnetic iron oxide nanoparticles is described. It is based on a recently developed concept in bio-orthogonal inverse-electron demand Diels-Alder chemistry, which is termed 'click-to-release'. To illustrate a potential new biomedical application of the chemistry, the nanoparticles were modified with tetrazine, as well as near infrared fluorescent (NIRF) cy5.5 dye, while doxorubicin was converted into a prodrug. The nanoparticles taken up by the MDA-MB-231 breast cancer cells efficiently converted the prodrug of doxorubicin into the biologically active chemotherapeutic doxorubicin form. PMID:27076271

  18. Increasing oral absorption of polar neuraminidase inhibitors: a prodrug transporter approach applied to oseltamivir analogue.

    PubMed

    Gupta, Deepak; Varghese Gupta, Sheeba; Dahan, Arik; Tsume, Yasuhiro; Hilfinger, John; Lee, Kyung-Dall; Amidon, Gordon L

    2013-02-01

    Poor oral absorption is one of the limiting factors in utilizing the full potential of polar antiviral agents. The neuraminidase target site requires a polar chemical structure for high affinity binding, thus limiting oral efficacy of many high affinity ligands. The aim of this study was to overcome this poor oral absorption barrier, utilizing prodrug to target the apical brush border peptide transporter 1 (PEPT1). Guanidine oseltamivir carboxylate (GOCarb) is a highly active polar antiviral agent with insufficient oral bioavailability (4%) to be an effective therapeutic agent. In this report we utilize a carrier-mediated targeted prodrug approach to improve the oral absorption of GOCarb. Acyloxy(alkyl) ester based amino acid linked prodrugs were synthesized and evaluated as potential substrates of mucosal transporters, e.g., PEPT1. Prodrugs were also evaluated for their chemical and enzymatic stability. PEPT1 transport studies included [(3)H]Gly-Sar uptake inhibition in Caco-2 cells and cellular uptake experiments using HeLa cells overexpressing PEPT1. The intestinal membrane permeabilities of the selected prodrugs and the parent drug were then evaluated for epithelial cell transport across Caco-2 monolayers, and in the in situ rat intestinal jejunal perfusion model. Prodrugs exhibited a pH dependent stability with higher stability at acidic pHs. Significant inhibition of uptake (IC(50) <1 mM) was observed for l-valyl and l-isoleucyl amino acid prodrugs in competition experiments with [(3)H]Gly-Sar, indicating a 3-6 times higher affinity for PEPT1 compared to valacyclovir, a well-known PEPT1 substrate and >30-fold increase in affinity compared to GOCarb. The l-valyl prodrug exhibited significant enhancement of uptake in PEPT1/HeLa cells and compared favorably with the well-absorbed valacyclovir. Transepithelial permeability across Caco-2 monolayers showed that these amino acid prodrugs have a 2-5-fold increase in permeability as compared to the parent drug and

  19. Hydrogen Peroxide Inducible DNA Cross-Linking Agents: Targeted Anticancer Prodrugs

    PubMed Central

    Kuang, Yunyan; Balakrishnan, Kumudha; Gandhi, Varsha; Peng, Xiaohua

    2011-01-01

    The major concern for anticancer chemotherapeutic agents is the host toxicity. The development of anti-cancer prodrugs targeting the unique biochemical alterations in cancer cells is an attractive approach to achieve therapeutic activity and selectivity. We designed and synthesized a new type of nitrogen mustard prodrug that can be activated by high level of reactive oxygen species (ROS) found in cancer cells to release the active chemotherapy agent. The activation mechanism was determined by NMR analysis. The activity and selectivity of these prodrugs towards ROS was determined by measuring DNA interstrand crosslinks and/or DNA alkylations. These compounds showed 60–90% inhibition toward various cancer cells, while normal lymphocytes were not affected. To the best of our knowledge, this is the first example of H2O2-activated anticancer prodrugs. PMID:22035519

  20. Synthesis and evaluation of mutual prodrugs of ibuprofen with menthol, thymol and eugenol.

    PubMed

    Redasani, Vivekkumar K; Bari, Sanjay B

    2012-10-01

    The present works deals with simple and efficient method of improving therapeutic efficacy of racemic ibuprofen by retarding gastrointestinal side effects through masking of carboxylic group chemically. This is achieved by synthesis and evaluation of ester derivatives of ibuprofen as mutual prodrugs with naturally occurring phenolic and alcoholic compounds. Promoieties like menthol; thymol and eugenol were selected with the aim of getting synergistic effect as these are natural analgesic having traditional medicinal values. Prodrugs are found to be highly lipophilic as compared to parent drug. All the prodrugs are found to be highly stable at acidic pH while undergoes hydrolysis at neutral and alkaline pH as indicated by their t(1/2) values. Synthesized prodrugs derivatives show increased anti-inflammatory activity that might be attributed to synergistic effect as ibuprofen conjugates to natural analgesics. Ulcer index shows much reduction in gastric ulceration compared to ibuprofen concluding the successful masking of acidic group. PMID:22982120

  1. Development and characterization of nanoparticulate formulation of a water soluble prodrug of dexamethasone by HIP complexation

    PubMed Central

    Gaudana, Ripal; Parenky, Ashwin; Vaishya, Ravi; Samanta, Swapan K.; Mitra, Ashim K.

    2015-01-01

    The objective of this study was to develop and characterize a nanoparticulate-based sustained release formulation of a water soluble dipeptide prodrug of dexamethasone, valine–valine-dexamethasone (VVD). Being hydrophilic in nature, it readily leaches out in the external aqueous medium and hence partitions poorly into the polymeric matrix resulting in minimal entrapment in nanoparticles. Hence, hydrophobic ion pairing (HIP) complexation of the prodrug was employed with dextran sulphate as a complexing polymer. A novel, solid in oil in water emulsion method was employed to encapsulate the prodrug in HIP complex form in poly(lactic-co-glycolic acid) matrix. Nanoparticles were characterized with respect to size, zeta potential, crystallinity of entrapped drug and surface morphology. A significant enhancement in the entrapment of the prodrug in nanoparticles was achieved. Finally, a simple yet novel method was developed which can also be applicable to encapsulate other charged hydrophilic molecules, such as peptides and proteins. PMID:20939702

  2. Design of Prodrugs to Enhance Colonic Absorption by Increasing Lipophilicity and Blocking Ionization

    PubMed Central

    Nofsinger, Rebecca; Clas, Sophie-Dorothee; Sanchez, Rosa I.; Walji, Abbas; Manser, Kimberly; Nissley, Becky; Balsells, Jaume; Nair, Amrithraj; Dang, Qun; Bennett, David Jonathan; Hafey, Michael; Wang, Junying; Higgins, John; Templeton, Allen; Coleman, Paul; Grobler, Jay; Smith, Ronald; Wu, Yunhui

    2014-01-01

    Prodrugs are chemistry-enabled drug delivery modifications of active molecules designed to enhance their pharmacokinetic, pharmacodynamic and/or biopharmaceutical properties. Ideally, prodrugs are efficiently converted in vivo, through chemical or enzymatic transformations, to the active parent molecule. The goal of this work is to enhance the colonic absorption of a drug molecule with a short half-life via a prodrug approach to deliver sustained plasma exposure and enable once daily (QD) dosing. The compound has poor absorption in the colon and by the addition of a promoiety to block the ionization of the molecule as well as increase lipophilicity, the relative colonic absorption increased from 9% to 40% in the retrograde dog colonic model. A combination of acceptable solubility and stability in the gastrointestinal tract (GI) as well as permeability was used to select suitable prodrugs to optimize colonic absorption. PMID:24566521

  3. Membrane-permeable Triphosphate Prodrugs of Nucleoside Analogues.

    PubMed

    Gollnest, Tristan; Dinis de Oliveira, Thiago; Rath, Anna; Hauber, Ilona; Schols, Dominique; Balzarini, Jan; Meier, Chris

    2016-04-18

    The metabolic conversion of nucleoside analogues into their triphosphates often proceeds insufficiently. Rate-limitations can be at the mono-, but also at the di- and triphosphorylation steps. We developed a nucleoside triphosphate (NTP) delivery system (TriPPPro-approach). In this approach, NTPs are masked by two bioreversible units at the γ-phosphate. Using a procedure involving H-phosphonate chemistry, a series of derivatives bearing approved, as well as potentially antivirally active, nucleoside analogues was synthesized. The enzyme-triggered delivery of NTPs was demonstrated by pig liver esterase, in human T-lymphocyte cell extracts and by a polymerase chain reaction using a prodrug of thymidine triphosphate. The TriPPPro-compounds of some HIV-inactive nucleoside analogues showed marked anti-HIV activity. For cellular uptake studies, a fluorescent TriPPPro-compound was prepared that delivered the triphosphorylated metabolite to intact CEM cells. PMID:27008042

  4. Pivalase catalytic antibodies: towards abzymatic activation of prodrugs.

    PubMed

    Bensel, N; Reymond, M T; Reymond, J L

    2001-11-01

    Screening of monoclonal-antibody libraries generated against the tert-butyl phosphonate hapten 2 and the chloromethyl phosphonate hapten 3 with pivaloyloxymethyl-umbelliferone 1 as a fluorogenic substrate led to the isolation of eleven catalytic antibodies with rate accelerations around kcat/ kuncat = 10(3). The antibodies are not inhibited by the product and accept different acyloxymethyl derivatives of acidic phenols as substrates. The highest activity was found for the bulky, chemically less-reactive pivaloyloxymethyl group: there is no activity with acetoxymethyl or acetyl esters. This difference might reflect the preference of the immune system for hydrophobic interactions in binding and catalysis. Pivalase catalytic antibodies might be useful for activating orally available pivaloyloxymethyl prodrugs. PMID:11757652

  5. Hydrophobic ion pairing of isoniazid using a prodrug approach.

    PubMed

    Zhou, Huiyu; Lengsfeld, Corinne; Claffey, David J; Ruth, James A; Hybertson, Brooks; Randolph, Theodore W; Ng, Ka-Yun; Manning, Mark C

    2002-06-01

    Inhalation therapy for infectious lung diseases, such as tuberculosis, is currently being explored, with microspheres being used to target alveolar macrophages. One method of drug encapsulation into polymeric microspheres to form hydrophobic ion-paired (HIP) complexes, and then coprecipitate the complex and polymer using supercritical fluid methodology. For the potent antituberculosis drug, isoniazid (isonicotinic acid hydrazide, INH), to be used in this fashion, it was modified into an ionizable form suitable for HIP. The charged prodrug, sodium isoniazid methanesulfonate (Na-INHMS), was then ion paired with hydrophobic cations, such as alkyltrimethylammonium or tetraalkylammonium. The logarithms of the apparent partition coefficients (log P') of various HIP complexes of INHMS display a roughly linear relationship with the numbers of carbon atoms in the organic counterions. The water solubility of the tetraheptylammonium-INHMS complex is about 220-fold lower than that of Na-INHMS, while the solubility in dichloromethane exceeds 10 mg/mL, which is sufficient for microencapsulation of the drug into poly(lactide) microspheres. The actual logarithm of the dichloromethane/water partition coefficient (log P) for tetraheptylammonium-INHMS is 1.55, compared to a value of - 1.8 for the sodium salt of INHMS. The dissolution kinetics of the tetraheptylammonium-INHMS complex in 0.9% aqueous solutions of NaCl was also investigated. Dissolution of tetraheptylammonium-INHMS exhibited a first-order time constant of about 0.28 min(-1), followed by a slower reverse ion exchange process to form Na-INHMS. The half-life of this HIP complex is on the order of 30 min, making the enhanced transport of the drug across biological barriers possible. This work represents the first use of a prodrug approach to introduce functionality that would allow HIP complex formation for a neutral molecule. PMID:12115849

  6. Studies on a novel series of acyl ester prodrugs of prostaglandin F2 alpha.

    PubMed Central

    Cheng-Bennett, A; Chan, M F; Chen, G; Gac, T; Garst, M E; Gluchowski, C; Kaplan, L J; Protzman, C E; Roof, M B; Sachs, G

    1994-01-01

    A novel series of prostaglandin F2 alpha (PGF2 alpha) prodrugs, with acyl ester groups at the 9, 11, and 15 positions, was prepared in order to design clinically acceptable prostaglandins for treating glaucoma. Studies involving isolated esterases and ocular tissue homogenates indicated that 9-acyl esters cannot provide a prodrug since PGF2 alpha would not be formed as a product. In contrast, 11-mono, 15-mono, and 11, 15-diesters were converted to PGF2 alpha in ocular tissues and could, therefore, be considered as prodrugs of PGF2 alpha. Carboxylesterase (CE) appeared critically important for the hydrolytic conversion of those PGF2 alpha prodrugs where the 11 or 15-OH group was esterified and such prodrugs were not substrates for acetylcholinesterase (ACHE) or butyrylcholinesterase (BuCHE). The enzymatic hydrolysis of PGF2 alpha-1-isopropyl ester was also investigated for comparative purposes. This PGF2 alpha prodrug was a good substrate for CE, but was also hydrolysed by BuCHE, albeit at a much slower rate. The most striking feature of the enzymatic hydrolysis of PGF2 alpha-1-isopropyl ester in ocular tissue homogenates was that it was much faster than for prodrugs esterified at the 11 and/or 15 positions. In terms of ocular hypotensive activity, all prodrugs which showed detectable conversion to nascent PGF2 alpha were potent ocular hypotensives. Although no separation of ocular hypotensive and ocular surface hyperaemic effects was apparent for PGF2 alpha-1-isopropyl ester, a temporal separation of these effects was apparent for the novel PGF2 alpha ester series. This difference may reflect an unfavourably rapid conversion of PGF2 alpha-1-isopropyl ester in ocular surface tissues compared with anterior segment tissues. PMID:7918269

  7. Diaryl ester prodrugs of FR900098 with improved in vivo antimalarial activity.

    PubMed

    Reichenberg, A; Wiesner, J; Weidemeyer, C; Dreiseidler, E; Sanderbrand, S; Altincicek, B; Beck, E; Schlitzer, M; Jomaa, H

    2001-03-26

    The fosmidomycin derivative FR900098 represents an inhibitor of the 1-deoxy-D-xylulose 5-phosphate (DOXP) reductoisomerase with potent antimalarial activity. Prodrugs of FR900098 with increased activity after oral administration were obtained by chemical modification of the phosphonate moiety to yield phosphodiaryl esters. One diaryl ester prodrug demonstrated efficacy in mice infected with the rodent malaria parasite Plasmodium vinckei comparable to i.p. drug administration. PMID:11277531

  8. Hypoxia-activated prodrugs: paths forward in the era of personalised medicine

    PubMed Central

    Hunter, Francis W; Wouters, Bradly G; Wilson, William R

    2016-01-01

    Tumour hypoxia has been pursued as a cancer drug target for over 30 years, most notably using bioreductive (hypoxia-activated) prodrugs that target antineoplastic agents to low-oxygen tumour compartments. Despite compelling evidence linking hypoxia with treatment resistance and adverse prognosis, a number of such prodrugs have recently failed to demonstrate efficacy in pivotal clinical trials; an outcome that demands reflection on the discovery and development of these compounds. In this review, we discuss a clear disconnect between the pathobiology of tumour hypoxia, the pharmacology of hypoxia-activated prodrugs and the manner in which they have been taken into clinical development. Hypoxia-activated prodrugs have been evaluated in the manner of broad-spectrum cytotoxic agents, yet a growing body of evidence suggests that their activity is likely to be dependent on the coincidence of tumour hypoxia, expression of specific prodrug-activating reductases and intrinsic sensitivity of malignant clones to the cytotoxic effector. Hypoxia itself is highly variable between and within individual tumours and is not treatment-limiting in all cancer subtypes. Defining predictive biomarkers for hypoxia-activated prodrugs and overcoming the technical challenges of assaying them in clinical settings will be essential to deploying these agents in the era of personalised cancer medicine. PMID:27070712

  9. Albumin-binding caspase-cleavable prodrug that is selectively activated in radiation exposed local tumor.

    PubMed

    Chung, Seung Woo; Choi, Jeong Uk; Lee, Beom Seok; Byun, Julia; Jeon, Ok-Cheol; Kim, Seong Who; Kim, In-San; Kim, Sang Yoon; Byun, Youngro

    2016-07-01

    Existence of the genomically and epigenomically diverse subclones in a tumor severely limits the therapeutic efficacy of targeted agents. To overcome such a limitation, we prepared a novel targeted prodrug, EMC-DEVD-S-DOX, which comprises two important features: radiation-induced apoptosis targeting and albumin-binding properties. In particular, the prodrug binds circulating albumin after intravenous administration and then activated by caspase-3, which is upregulated from apoptotic cells that responded to radiotherapy. The prodrug was designed to bind circulating albumin to extend half-life and facilitate tumor accumulation in order to increase the possibility of contacting caspase-3, which is only transiently upregulated during apoptosis. Our results showed that EMC-DEVD-S-DOX had a prolonged half-life with enhanced tumor accumulation, which clearly benefited the therapeutic effect of the prodrug. Also, agreeing with the in vitro studies that showed ignorable cytotoxic effect in the absence of caspase-3, the prodrug was effective only when combined with radiotherapy without any noticeable systemic toxicity in vivo. Due to the highly selective action of EMC-DEVD-S-DOX independent to the complex genomic profiles of tumor, the prodrug would overcome the limitation of current targeted therapy and potentiate radiotherapy in the clinical oncology. PMID:27085176

  10. Development of Novel Polymeric Prodrugs Synthesized by Mechanochemical Solid-State Copolymerization of Hydroxyethylcellulose and Vinyl Monomers.

    PubMed

    Doi, Naoki; Sasai, Yasushi; Yamauchi, Yukinori; Adachi, Tetsuo; Kuzuya, Masayuki; Kondo, Shin-ichi

    2015-01-01

    Novel polymeric prodrugs were synthesized by mechanochemical solid-state copolymerization of hydroxyethylcellulose and the methacryloyloxy derivative of 5-fluorouracil (5-FU). Copolymerization was about 94% complete after 4 h, and the polymeric prodrug was quantitatively obtained after 14 h of reaction. The number average molecular weight (Mn) and polydispersity (H) of the polymeric prodrug were 39000 g/mol and 6.20, respectively. Mechanical fracturing of the polymer in a stainless steel twin-shell blender improved these properties (Mn=16000 g/mol and H=1.94). 5-FU was sustainably released from the polymeric prodrugs, and the rate was not affected by the molecular weight or molecular weight distribution of the prodrug under the experimental conditions used. These results suggest that novel polymeric prodrugs composed of a polysaccharide and a synthetic polymer can be fabricated by mechanochemical solid-state copolymerization under anaerobic conditions. PMID:26423117

  11. Gold(III)-Dithiocarbamato Peptidomimetics in the Forefront of the Targeted Anticancer Therapy: Preclinical Studies against Human Breast Neoplasia

    PubMed Central

    Nardon, Chiara; Schmitt, Sara M.; Yang, Huanjie; Zuo, Jian

    2014-01-01

    Since the serendipitous discovery of cisplatin, platinum-based drugs have become well-established antitumor agents, despite the fact that their clinical use is limited by many severe side-effects. In order to both improve the chemotherapeutic index and broaden the therapeutic spectrum of current drugs, our most recent anti-neoplastic agents, Au(III) complexes, were designed as carrier-mediated delivery systems exploiting peptide transporters, which are up-regulated in some cancers. Among all, we focused on two compounds and tested them on human MDA-MB-231 (resistant to cisplatin) breast cancer cell cultures and xenografts, discovering the proteasome as a major target both in vitro and in vivo. 53% inhibition of breast tumor growth in mice was observed after 27 days of treatment at 1.0 mg kg−1 d−1, compared to control. Remarkably, if only the most responsive mice are taken into account, 85% growth inhibition, with some animals showing tumor shrinkage, was observed after 13 days. These results led us to file an international patent, recognizing this class of gold(III) peptidomimetics as suitable candidates for entering phase I clinical trials. PMID:24392119

  12. Peptide vaccines and peptidomimetics targeting HER and VEGF proteins may offer a potentially new paradigm in cancer immunotherapy

    PubMed Central

    Kaumaya, Pravin TP; Foy, Kevin Chu

    2013-01-01

    The ErbB family (HER-1, HER-2, HER-3 and HER-4) of receptor tyrosine kinases has been the focus of cancer immunotherapeutic strategies while antiangiogenic therapies have focused on VEGF and its receptors VEGFR-1 and VEGFR-2. Agents targeting receptor tyrosine kinases in oncology include therapeutic antibodies to receptor tyrosine kinase ligands or the receptors themselves, and small-molecule inhibitors. Many of the US FDA-approved therapies targeting HER-2 and VEGF exhibit unacceptable toxicities, and show problems of efficacy, development of resistance and unacceptable safety profiles that continue to hamper their clinical progress. The combination of dif ferent peptide vaccines and peptidomimetics targeting specific molecular pathways that are dysregulated in tumors may potentiate anticancer immune responses, bypass immune tolerance and circumvent resistance mechanisms. The focus of this review is to discuss efforts in our laboratory spanning two decades of rationally developing peptide vaccines and therapeutics for breast cancer. This review highlights the prospective benefit of a new, untapped category of therapies biologically targeted to EGF receptor (HER-1), HER-2 and VEGF with potential peptide ‘blockbusters‘ that could lay the foundation of a new paradigm in cancer immunotherapy by creating clinical breakthroughs for safe and efficacious cancer cures. PMID:22894670

  13. Rapid Synthesis of Boc-2',6'-dimethyl-l-tyrosine and Derivatives and Incorporation into Opioid Peptidomimetics.

    PubMed

    Bender, Aaron M; Griggs, Nicholas W; Gao, Chao; Trask, Tyler J; Traynor, John R; Mosberg, Henry I

    2015-12-10

    The unnatural amino acid 2',6'-dimethyl-l-tyrosine has found widespread use in the development of synthetic opioid ligands. Opioids featuring this residue at the N-terminus often display superior potency at one or more of the opioid receptor types, but the availability of the compound is hampered by its cost and difficult synthesis. We report here a short, three-step synthesis of Boc-2',6'-dimethyl-l-tyrosine (3a) utilizing a microwave-assisted Negishi coupling for the key carbon-carbon bond forming step, and employ this chemistry for the expedient synthesis of other unnatural tyrosine derivatives. Three of these derivatives (3c, 3d, 3f) have not previously been examined as Tyr(1) replacements in opioid ligands. We describe the incorporation of these tyrosine derivatives in a series of opioid peptidomimetics employing our previously reported tetrahydroquinoline (THQ) scaffold, and the binding and relative efficacy of each of the analogues at the three opioid receptor subtypes: mu (MOR), delta (DOR), and kappa (KOR). PMID:26713104

  14. Eco-Friendly Insecticide Discovery via Peptidomimetics: Design, Synthesis, and Aphicidal Activity of Novel Insect Kinin Analogues.

    PubMed

    Zhang, Chuanliang; Qu, Yanyan; Wu, Xiaoqing; Song, Dunlun; Ling, Yun; Yang, Xinling

    2015-05-13

    Insect kinin neuropeptides are pleiotropic peptides that are involved in the regulation of hindgut contraction, diuresis, and digestive enzyme release. They share a common C-terminal pentapeptide sequence of Phe(1)-Xaa(2)-Yaa(3)-Trp(4)-Gly(5)-NH2 (where Xaa(2) = His, Asn, Phe, Ser, or Tyr; Yaa(3) = Pro, Ser, or Ala). Recently, the aphicidal activity of insect kinin analogues has attracted the attention of researchers. Our previous work demonstrated that the sequence-simplified insect kinin pentapeptide analogue Phe-Phe-[Aib]-Trp-Gly-NH2 could retain good aphicidal activity and be the lead compound for the further discovery of eco-friendly insecticides which encompassed a broad array of biochemicals derived from micro-organisms and other natural sources. Using the peptidomimetics strategy, we chose Phe-Phe-[Aib]-Trp-Gly-NH2 as the lead compound, and we designed and synthesized three series, including 31 novel insect kinin analogues. The aphicidal activity of the new analogues against soybean aphid was determined. The results showed that all of the analogues exhibited aphicidal activity. Of particular interest was the analogue II-1, which exhibited improved aphicidal activity with an LC50 of 0.019 mmol/L compared with the lead compound (LC50 = 0.045 mmol/L) or the commercial insecticide pymetrozine (LC50 = 0.034 mmol/L). This suggests that the analogue II-1 could be used as a new lead for the discovery of potential eco-friendly insecticides. PMID:25912216

  15. The phosphotyrosine interaction domains of X11 and FE65 bind to distinct sites on the YENPTY motif of amyloid precursor protein.

    PubMed Central

    Borg, J P; Ooi, J; Levy, E; Margolis, B

    1996-01-01

    The phosphotyrosine interaction (PI) domains (also known as the PTB, or phosphotyrosine binding, domains) of Shc and IRS-1 are recently described domains that bind peptides phosphorylated on tyrosine residues. The PI/PTB domains differ from Src homology 2 (SH2) domains in that their binding specificity is determined by residues that lie amino terminal and not carboxy terminal to the phosphotyrosine. Recently, it has been appreciated that other cytoplasmic proteins also contain PI domains. We now show that the PI domain of X11 and one of the PI domains of FE65, two neuronal proteins, bind to the cytoplasmic domain of the amyloid precursor protein ((beta)APP). (beta)APP is an integral transmembrane glycoprotein whose cellular function is unknown. One of the processing pathways of (beta)APP leads to the secretion of A(beta), the major constituent of the amyloid deposited in the brain parenchyma and vessel walls of Alzheimer's disease patients. We have found that the X11 PI domain binds a YENPTY motif in the intracellular domain of (beta)APP that is strikingly similar to the NPXY motifs that bind the Shc and IRS-1 PI/PTB domains. However, unlike the case for binding of the Shc PI/PTB domain, tyrosine phosphorylation of the YENPTY motif is not required for the binding of (beta)APP to X11 or FE65. The binding site of the FE65 PI domain appears to be different from that of X11, as mutations within the YENPTY motif differentially affect the binding of X11 and FE65. Using site-directed mutagenesis, we have identified a crucial residue within the PI domain involved in X11 and FE65 binding to (beta)APP. The binding of X11 or FE65 PI domains to residues of the YENPTY motif of (beta)APP identifies PI domains as general protein interaction domains and may have important implications for the processing of (beta)APP. PMID:8887653

  16. Water soluble cyclosporine monomethoxy poly(ethyleneglycol) conjugates as potential prodrugs.

    PubMed

    Cho, Hoon; Chung, Yongseog

    2004-06-01

    The highly water-soluble monomethoxypoly(ethyleneglycol) (mPEG) prodrugs of cyclosporin A (CsA) were synthesized. These prodrugs were prepared by initially preparing intermediate in the form of carbonate at the 3'-positions of CsA with chloromethyl chloroformate, in the presence of a base to provide a 3'-carbonated CsA intermediate. Reaction of the CsA intermediate with mPEG derivative in the presence of a base provides the desired water-soluble prodrugs. As a model, we chose molecular weight 5 kDa mPEG in the reaction with CsA to give water soluble prodrugs. To prove that the prodrug is decomposed in the body to produce CsA, the enzymatic hydrolysis test was conducted using human liver homogenate at 37 degrees C. The prodrug was decomposed in human liver homogenate to produce the active material, CsA, and the hydrolysis half-life (t(1/2)) of the prodrug, KI-306 was 2.2 minutes at 37 degrees C. However, a demonstration of non-enzymatic conversion in pH 7.4 phosphate buffer was provided by the fact that the half-life (t(1/2)) is 21 hours at 37 degrees C. The hydrolysis test in rat whole blood was also conducted. The hydrolysis was seen with half-life (t(1/2)) of about 9.9, 65.0, 14.2, 3.4, 2.1 9.5, and 1.6 minutes for KI-306, 309, 312, 313, 315, 316, and 317, respectively. This is the ideal for CsA prodrug. The pharmacokinetic study of the prodrug, KI-306, in comparison to the commercial product (Sandimmune Neoral Solution) was also carried out after single oral dose. Each rat received 7 mg/kg of CsA equivalent dose. Especially, the prodrug KI-306 exhibits higher AUC and Cmax than the conventional Neoral. The AUC and Cmax were increased nearly 1.5 fold. The kinetic value was also seen with Tmax of about 1.43 and 2.44 hours for KI-306 and Neoral, respectively. PMID:15283470

  17. Crystal Structures of HLA-A*0201 Complexed with Melan-A/MART-1[subscript 26(27L)-35] Peptidomimetics Reveal Conformational Heterogeneity and Highlight Degeneracy of T Cell Recognition

    SciTech Connect

    Douat-Casassus, Celine; Borbulevych, Oleg; Tarbe, Marion; Gervois, Nadine; Jotereau, Francine; Baker, Brian M.; Quideau, Stphane

    2010-10-07

    There is growing interest in using tumor associated antigens presented by class I major histocompatibility complex (MHC-I) proteins as cancer vaccines. As native peptides are poorly stable in biological fluids, researchers have sought to engineer synthetic peptidomimetics with greater biostability. Here, we demonstrate that antigenic peptidomimetics of the Melan-A/MART-1{sub 26(27L)-35} melanoma antigen adopt strikingly different conformations when bound to MHC-I, highlighting the degeneracy of T cell recognition and revealing the challenges associated with mimicking native peptide conformation.

  18. Surface coating of siRNA-peptidomimetic nano-self-assemblies with anionic lipid bilayers: enhanced gene silencing and reduced adverse effects in vitro

    NASA Astrophysics Data System (ADS)

    Zeng, Xianghui; de Groot, Anne Marit; Sijts, Alice J. A. M.; Broere, Femke; Oude Blenke, Erik; Colombo, Stefano; van Eden, Willem; Franzyk, Henrik; Nielsen, Hanne Mørck; Foged, Camilla

    2015-11-01

    Cationic vectors have demonstrated the potential to facilitate intracellular delivery of therapeutic oligonucleotides. However, enhanced transfection efficiency is usually associated with adverse effects, which also proves to be a challenge for vectors based on cationic peptides. In this study a series of proteolytically stable palmitoylated α-peptide/β-peptoid peptidomimetics with a systematically varied number of repeating lysine and homoarginine residues was shown to self-assemble with small interfering RNA (siRNA). The resulting well-defined nanocomplexes were coated with anionic lipids giving rise to net anionic liposomes. These complexes and the corresponding liposomes were optimized towards efficient gene silencing and low adverse effects. The optimal anionic liposomes mediated a high silencing effect, which was comparable to that of the control (cationic Lipofectamine 2000), and did not display any noticeable cytotoxicity and immunogenicity in vitro. In contrast, the corresponding nanocomplexes mediated a reduced silencing effect with a more narrow safety window. The surface coating with anionic lipid bilayers led to partial decomplexation of the siRNA-peptidomimetic nanocomplex core of the liposomes, which facilitated siRNA release. Additionally, the optimal anionic liposomes showed efficient intracellular uptake and endosomal escape. Therefore, these findings suggest that a more efficacious and safe formulation can be achieved by surface coating of the siRNA-peptidomimetic nano-self-assemblies with anionic lipid bilayers.Cationic vectors have demonstrated the potential to facilitate intracellular delivery of therapeutic oligonucleotides. However, enhanced transfection efficiency is usually associated with adverse effects, which also proves to be a challenge for vectors based on cationic peptides. In this study a series of proteolytically stable palmitoylated α-peptide/β-peptoid peptidomimetics with a systematically varied number of repeating lysine

  19. Transport of levovirin prodrugs in the human intestinal Caco-2 cell line.

    PubMed

    Li, Fujun; Hong, Lei; Mau, Cheng-I; Chan, Rebecca; Hendricks, Than; Dvorak, Chuck; Yee, Calvin; Harris, Jason; Alfredson, Tom

    2006-06-01

    The transport of 10 amino acid ester prodrugs of levovirin (LVV) was investigated in the human intestinal Caco-2 cell line in order to overcome the poor oral bioavailability of LVV, an investigational drug for the treatment of hepatitis C infection. The prodrugs were designed to improve the permeability of LVV across the intestinal epithelium by targeting the di/tri-peptide carrier, PepT1. Caco-2 cell monolayers were employed to study the transport and hydrolysis properties of the prodrugs. Among all mono amino acid ester prodrugs studied, the LVV-5'-(L)-valine prodrug (R1518) exhibited the maximum increase (48-fold) in permeability with nearly complete conversion to LVV within 1 h. Di-amino acid esters did not offer significant enhancement in permeability comparing with mono amino acid esters and exhibited slower conversion to LVV in Caco2 cell monolayers. Pharmacokinetic screening studies of the prodrugs in rats yielded the highest fold increase (6.9-fold) of AUC with R1518 and in general displayed a similar trend to that observed in increases of permeability in Caco-2 cells. Mechanisms involved in the Caco-2 cell transport of R1518 were also investigated. Results of bi-directional transport studies support the involvement of carrier-mediated transport mechanisms for R1518, but not for the LVV-5'-(D)-valine prodrug or LVV. Moreover, the permeability of R1518 was found to be proton dependent. PepT1-mediated transport of R1518 was supported by results of competitive transport studies of R1518 with the PepT1 substrates enalapril, Gly-Sar, valganciclovir, and cephalexin. R1518 was also found to inhibit the permeability of valganciclovir and cephalexin. These results suggest that R1518 is a PepT1 substrate as well as an inhibitor. PMID:16634069

  20. Adrenocorticotrophic hormone stimulates phosphotyrosine phosphatase SHP2 in bovine adrenocortical cells: phosphorylation and activation by cAMP-dependent protein kinase.

    PubMed Central

    Rocchi, S; Gaillard, I; van Obberghen, E; Chambaz, E M; Vilgrain, I

    2000-01-01

    During activation of adrenocortical cells by adrenocorticotrophic hormone (ACTH), tyrosine dephosphorylation of paxillin is stimulated and this correlates with protrusion of filopodial structures and a decreased number of focal adhesions. These effects are inhibited by Na(3)VO(4), a phosphotyrosine phosphatase inhibitor [Vilgrain, Chinn, Gaillard, Chambaz and Feige (1998) Biochem. J. 332, 533-540]. However, the tyrosine phosphatases involved in these processes remain to be identified. In this study, we provide evidence that the Src homology domain (SH)2-containing phosphotyrosine phosphatase (SHP)2, but not SHP1, is expressed in adrenocortical cells and is phosphorylated upon ACTH challenge. ACTH (10(-8) M) treatment of (32)P-labelled adrenocortical cells resulted in an increase in phosphorylated SHP2. By probing SHP2-containing immunoprecipitates with an antibody to phosphoserine we found that SHP2 was phosphorylated on serine in ACTH-treated cells in a dose- and time-dependent manner. Furthermore, using an in vitro kinase assay, we showed that SHP2 was a target for cAMP-dependent protein kinase (PKA). Serine was identified as the only target amino acid phosphorylated in SHP2. Phosphorylation of SHP2 by PKA resulted in a dramatic stimulation of phosphatase activity measured either with insulin receptor substrate-1 or with the synthetic peptide [(32)P]poly(Glu/Tyr) as substrate. In an in-gel assay of SHP2-containing immunoprecipitates, phosphorylated in vitro by PKA or isolated from adrenocortical cells treated with 10 nM ACTH, a pronounced activation of SHP2 activity was shown. These observations clearly support the idea that a PKA-mediated signal transduction pathway contributes to SHP2 regulation in adrenocortical cells and point to SHP2 as a possible mediator of the effects of ACTH. PMID:11085942

  1. Flurbiprofen-antioxidant mutual prodrugs as safer nonsteroidal anti-inflammatory drugs: synthesis, pharmacological investigation, and computational molecular modeling.

    PubMed

    Ashraf, Zaman; Alamgeer; Kanwal, Munazza; Hassan, Mubashir; Abdullah, Sahar; Waheed, Mamuna; Ahsan, Haseeb; Kim, Song Ja

    2016-01-01

    Flurbiprofen-antioxidant mutual prodrugs were synthesized to reduce the gastrointestinal (GI) effects associated with flurbiprofen. For reducing the GI toxicity, the free carboxylic group (-COOH) was temporarily masked by esterification with phenolic -OH of natural antioxidants vanillin, thymol, umbelliferone, and sesamol. The in vitro hydrolysis of synthesized prodrugs showed that they were stable in buffer solution at pH 1.2, indicating their stability in the stomach. The synthesized prodrugs undergo significant hydrolysis in 80% human plasma and thus release free flurbiprofen. The minimum reversion was observed at pH 1.2, suggesting that prodrugs are less irritating to the stomach than flurbiprofen. The anti-inflammatory, analgesic, antipyretic, and ulcerogenic activities of prodrugs were evaluated. All the synthesized prodrugs significantly (P<0.001) reduced the inflammation against carrageenan and egg albumin-induced paw edema at 4 hours of study. The reduction in the size of the inflamed paw showed that most of the compounds inhibited the later phase of inflammation. The prodrug 2-oxo-2H-chromen-7-yl-2-(2-fluorobiphenyl-4-yl)propanoate (4b) showed significant reduction in paw licking with percentage inhibition of 58%. It also exhibited higher analgesic activity, reducing the number of writhes with a percentage of 75%, whereas flurbiprofen showed 69% inhibition. Antipyretic activity was investigated using brewer's yeast-induced pyrexia model, and significant (P<0.001) reduction in rectal temperature was shown by all prodrugs at all times of assessment. The results of ulcerogenic activity showed that all prodrugs produced less GI irritation than flurbiprofen. Molecular docking and simulation studies were carried out with cyclooxygenase (COX-1 and COX-2) proteins, and it was observed that our prodrugs have more potential to selectively bind to COX-2 than to COX-1. It is concluded that the synthesized prodrugs have promising pharmacological activities with

  2. Flurbiprofen–antioxidant mutual prodrugs as safer nonsteroidal anti-inflammatory drugs: synthesis, pharmacological investigation, and computational molecular modeling

    PubMed Central

    Ashraf, Zaman; Alamgeer; Kanwal, Munazza; Hassan, Mubashir; Abdullah, Sahar; Waheed, Mamuna; Ahsan, Haseeb; Kim, Song Ja

    2016-01-01

    Flurbiprofen–antioxidant mutual prodrugs were synthesized to reduce the gastrointestinal (GI) effects associated with flurbiprofen. For reducing the GI toxicity, the free carboxylic group (–COOH) was temporarily masked by esterification with phenolic –OH of natural antioxidants vanillin, thymol, umbelliferone, and sesamol. The in vitro hydrolysis of synthesized prodrugs showed that they were stable in buffer solution at pH 1.2, indicating their stability in the stomach. The synthesized prodrugs undergo significant hydrolysis in 80% human plasma and thus release free flurbiprofen. The minimum reversion was observed at pH 1.2, suggesting that prodrugs are less irritating to the stomach than flurbiprofen. The anti-inflammatory, analgesic, antipyretic, and ulcerogenic activities of prodrugs were evaluated. All the synthesized prodrugs significantly (P<0.001) reduced the inflammation against carrageenan and egg albumin-induced paw edema at 4 hours of study. The reduction in the size of the inflamed paw showed that most of the compounds inhibited the later phase of inflammation. The prodrug 2-oxo-2H-chromen-7-yl-2-(2-fluorobiphenyl-4-yl)propanoate (4b) showed significant reduction in paw licking with percentage inhibition of 58%. It also exhibited higher analgesic activity, reducing the number of writhes with a percentage of 75%, whereas flurbiprofen showed 69% inhibition. Antipyretic activity was investigated using brewer’s yeast-induced pyrexia model, and significant (P<0.001) reduction in rectal temperature was shown by all prodrugs at all times of assessment. The results of ulcerogenic activity showed that all prodrugs produced less GI irritation than flurbiprofen. Molecular docking and simulation studies were carried out with cyclooxygenase (COX-1 and COX-2) proteins, and it was observed that our prodrugs have more potential to selectively bind to COX-2 than to COX-1. It is concluded that the synthesized prodrugs have promising pharmacological activities

  3. The development of orally administrable gemcitabine prodrugs with D-enantiomer amino acids: Enhanced membrane permeability and enzymatic stability

    PubMed Central

    Tsume, Yasuhiro; Incecayir, Tuba; Song, Xueqin; Hilfinger, John M.; Amidon, Gordon L.

    2014-01-01

    Gemcitabine prodrugs with D- and L-configuration amino acids were synthesized and their chemical stability in buffers, resistance to glycosidic bond metabolism, enzymatic activation, permeability in Caco-2 cells and mouse intestinal membrane, anti-proliferation activity in cancer cell were determined and compared to that of parent drug, gemcitabine. Prodrugs containing D-configuration amino acids were enzymatically more stable than ones with L-configuration amino acids. The activation of all gemcitabine prodrugs was 1.3–17.6-fold faster in cancer cell homogenate than their hydrolysis in buffer, suggesting enzymatic action. The enzymatic activation of amino acid monoester prodrugs containing D-configuration amino acids in cell homogenates was 2.2–10.9-fold slower than one of amino acid monoester prodrugs with L-configuration amino acids. All prodrugs exhibited enhanced resistance to glycosidic bond metabolism by thymidine phosphorylase compared to parent gemcitabine. Gemcitabine prodrugs showed superior the effective permeability in mouse jejunum to gemcitabine. More importantly, the high plasma concentration of D-amino acid gemcitabine prodrugs was observed more than one of L-amino acid gem-citabine prodrugs. In general, the 5′-mono-amino acid monoester gemcitabine prodrugs exhibited higher permeability and uptake than their parent drug, gemcitabine. Cell proliferation assays in AsPC-1 pancreatic ductal cell line indicated that gemcitabine prodrugs were more potent than their parent drug, gemcitabine. The transport and enzymatic profiles of 5′-D-valyl-gemcitabine and 5′-D-phenylalanyl-gem-citabine suggest their potential for increased oral uptake and delayed enzymatic bioconversion as well as enhanced uptake and cytotoxic activity in cancer cells, would facilitate the development of oral dosage form for anti-cancer agents and, hence, improve the quality of life for the cancer patients. PMID:24361461

  4. A diversity-oriented synthesis strategy enabling the combinatorial-type variation of macrocyclic peptidomimetic scaffolds† †Electronic supplementary information (ESI) available: Experimental procedures, characterization data and details of the computational analyses. See DOI: 10.1039/c5ob00371g Click here for additional data file.

    PubMed Central

    Isidro-Llobet, Albert; Hadje Georgiou, Kathy; Galloway, Warren R. J. D.; Giacomini, Elisa; Hansen, Mette R.; Méndez-Abt, Gabriela; Tan, Yaw Sing; Carro, Laura; Sore, Hannah F.

    2015-01-01

    Macrocyclic peptidomimetics are associated with a broad range of biological activities. However, despite such potentially valuable properties, the macrocyclic peptidomimetic structural class is generally considered as being poorly explored within drug discovery. This has been attributed to the lack of general methods for producing collections of macrocyclic peptidomimetics with high levels of structural, and thus shape, diversity. In particular, there is a lack of scaffold diversity in current macrocyclic peptidomimetic libraries; indeed, the efficient construction of diverse molecular scaffolds presents a formidable general challenge to the synthetic chemist. Herein we describe a new, advanced strategy for the diversity-oriented synthesis (DOS) of macrocyclic peptidomimetics that enables the combinatorial variation of molecular scaffolds (core macrocyclic ring architectures). The generality and robustness of this DOS strategy is demonstrated by the step-efficient synthesis of a structurally diverse library of over 200 macrocyclic peptidomimetic compounds, each based around a distinct molecular scaffold and isolated in milligram quantities, from readily available building-blocks. To the best of our knowledge this represents an unprecedented level of scaffold diversity in a synthetically derived library of macrocyclic peptidomimetics. Cheminformatic analysis indicated that the library compounds access regions of chemical space that are distinct from those addressed by top-selling brand-name drugs and macrocyclic natural products, illustrating the value of our DOS approach to sample regions of chemical space underexploited in current drug discovery efforts. An analysis of three-dimensional molecular shapes illustrated that the DOS library has a relatively high level of shape diversity. PMID:25778821

  5. Lisdexamfetamine Dimesylate: Prodrug Delivery, Amphetamine Exposure and Duration of Efficacy.

    PubMed

    Ermer, James C; Pennick, Michael; Frick, Glen

    2016-05-01

    Lisdexamfetamine dimesylate (LDX) is a long-acting d-amphetamine prodrug used to treat attention-deficit/hyperactivity disorder (ADHD) in children, adolescents and adults. LDX is hydrolysed in the blood to yield d-amphetamine, and the pharmacokinetic profile of d-amphetamine following oral administration of LDX has a lower maximum plasma concentration (Cmax), extended time to Cmax (Tmax) and lower inter- and intra-individual variability in exposure compared with the pharmacokinetic profile of an equivalent dose of immediate-release (IR) d-amphetamine. The therapeutic action of LDX extends to at least 13 h post-dose in children and 14 h post-dose in adults, longer than that reported for any other long-acting formulation. Drug-liking scores for LDX are lower than for an equivalent dose of IR d-amphetamine, which may result from the reduced euphorigenic potential associated with its pharmacokinetic profile. These pharmacokinetic and pharmacodynamic characteristics of LDX may be beneficial in the management of symptoms in children, adolescents and adults with ADHD. PMID:27021968

  6. Click synthesis of a polyamidoamine dendrimer-based camptothecin prodrug

    PubMed Central

    Zolotarskaya, Olga Yu.; Xu, Leyuan; Valerie, Kristoffer; Yang, Hu

    2015-01-01

    In the present work we report on the click synthesis of a new camptothecin (CPT) prodrug based on anionic polyamidoamine (PAMAM) dendrimer intended for cancer therapy. We applied ‘click’ chemistry to improve polymer-drug coupling reaction efficiency. Specifically, CPT was functionalized with a spacer, 1-azido-3,6,9,12,15-pentaoxaoctadecan-18-oic acid (APO), via EDC/DMAP coupling reaction. In parallel, propargylamine (PPA) and methoxypoly(ethylene glycol) amine were conjugated to PAMAM dendrimer G4.5 in sequence using an effective coupling agent 4-(4,6-dimethoxy-(1,3,5)triazin-2-yl)-4-methyl-morpholinium chloride (DMTMM). CPT-APO was then coupled to PEGylated PAMAM dendrimer G4.5-PPA via a click reaction using copper bromide/2,2’-bipyridine/ dimethyl sulfoxide (catalyst/ligand/solvent). Human glioma cells were exposed to the CPT-conjugate to determine toxicity and cell cycle effects using WST-1 assay and flow cytometry. The CPT-conjugate displayed a dose-dependent toxicity with an IC50 of 5 μM, a 185-fold increase relative to free CPT, presumably as a result of slow release. As expected, conjugated CPT resulted in G2/M arrest and cell death while the dendrimer itself had little to no toxicity. Altogether, highly efficient click chemistry allows for the synthesis of multifunctional dendrimers for sustained drug delivery. PMID:26640689

  7. Preparation and Characterization of Lipophilic Doxorubicin Pro-drug Micelles.

    PubMed

    Li, Feng; Snow-Davis, Candace; Du, Chengan; Bondarev, Mikhail L; Saulsbury, Marilyn D; Heyliger, Simone O

    2016-01-01

    Micelles have been successfully used for the delivery of anticancer drugs. Amphiphilic polymers form core-shell structured micelles in an aqueous environment through self-assembly. The hydrophobic core of micelles functions as a drug reservoir and encapsulates hydrophobic drugs. The hydrophilic shell prevents the aggregation of micelles and also prolongs their systemic circulation in vivo. In this protocol, we describe a method to synthesize a doxorubicin lipophilic pro-drug, doxorubicin-palmitic acid (DOX-PA), which will enhance drug loading into micelles. A pH-sensitive hydrazone linker was used to conjugate doxorubicin with the lipid, which facilitates the release of free doxorubicin inside cancer cells. Synthesized DOX-PA was purified with a silica gel column using dichloromethane/methanol as the eluent. Purified DOX-PA was analyzed with thin layer chromatography (TLC) and (1)H-Nuclear Magnetic Resonance Spectroscopy ((1)H-NMR). A film dispersion method was used to prepare DOX-PA loaded DSPE-PEG micelles. In addition, several methods for characterizing micelle formulations are described, including determination of DOX-PA concentration and encapsulation efficiency, measurement of particle size and distribution, and assessment of in vitro anticancer activities. This protocol provides useful information regarding the preparation and characterization of drug-loaded micelles and thus will facilitate the research and development of novel micelle-based cancer nanomedicines. PMID:27584689

  8. Integrase Inhibitor Prodrugs: Approaches to Enhancing the Anti-HIV Activity of β-Diketo Acids.

    PubMed

    Nair, Vasu; Okello, Maurice

    2015-01-01

    HIV integrase, encoded at the 3'-end of the HIV pol gene, is essential for HIV replication. This enzyme catalyzes the incorporation of HIV DNA into human DNA, which represents the point of "no-return" in HIV infection. Integrase is a significant target in anti-HIV drug discovery. This review article focuses largely on the design of integrase inhibitors that are β-diketo acids constructed on pyridinone scaffolds. Methodologies for synthesis of these compounds are discussed. Integrase inhibition data for the strand transfer (ST) step are compared with in vitro anti-HIV data. The review also examines the issue of the lack of correlation between the ST enzymology data and anti-HIV assay results. Because this disconnect appeared to be a problem associated with permeability, prodrugs of these inhibitors were designed and synthesized. Prodrugs dramatically improved the anti-HIV activity data. For example, for compound, 96, the anti-HIV activity (EC50) improved from 500 nM for this diketo acid to 9 nM for its prodrug 116. In addition, there was excellent correlation between the IC50 and IC90 ST enzymology data for 96 (6 nM and 97 nM, respectively) and the EC50 and EC90 anti-HIV data for its prodrug 116 (9 nM and 94 nM, respectively). Finally, it was confirmed that the prodrug 116 was rapidly hydrolyzed in cells to the active compound 96. PMID:26184144

  9. Design, Synthesis, and In Vitro Kinetics Study of Atenolol Prodrugs for the Use in Aqueous Formulations

    PubMed Central

    Qtait, Alaa; Dajani, Khulod Khayyat; Abu Lafi, Saleh

    2014-01-01

    Based on DFT, MP2, and the density functional from Truhlar group (hybrid GGA: MPW1k) calculations for an acid-catalyzed hydrolysis of nine Kirby's N-alkylmaleamic acids and two atenolol prodrugs were designed. The calculations demonstrated that the amide bond cleavage is due to intramolecular nucleophilic catalysis by the adjacent carboxylic acid group and the rate-limiting step is determined based on the nature of the amine leaving group. In addition, a linear correlation of the calculated and experimental rate values has drawn credible basis for designing atenolol prodrugs that are bitterless, are stable in neutral aqueous solutions, and have the potential to release the parent drug in a sustained release manner. For example, based on the calculated B3LYP/6-31 G (d,p) rates, the predicted t1/2 (a time needed for 50% of the prodrug to be converted into drug) values for atenolol prodrugs ProD 1-ProD 2 at pH 2 were 65.3 hours (6.3 hours as calculated by GGA: MPW1K) and 11.8 minutes, respectively. In vitro kinetic study of atenolol prodrug ProD 1 demonstrated that the t1/2 was largely affected by the pH of the medium. The determined t1/2 values in 1N HCl, buffer pH 2, and buffer pH 5 were 2.53, 3.82, and 133 hours, respectively. PMID:24526887

  10. Biological conversion of aripiprazole lauroxil - An N-acyloxymethyl aripiprazole prodrug

    PubMed Central

    Rohde, Morten; M?rk, Niels; Håkansson, Anders E.; Jensen, Klaus G.; Pedersen, Henrik; Dige, Tina; J?rgensen, Erling B.; Holm, René

    2014-01-01

    N-acyloxyalkylation of NH-acidic compounds can be a prodrug approach for e.g. tertiary or some N-heterocyclic amines and secondary amides and have the potential to modify the properties of the parent drug for specific uses, for example its physicochemical, pharmacokinetic or biopharmaceutical properties. Aripiprazole lauroxil was prepared as a model compound for such prodrugs and its bioconversion was investigated both in vitro and in vivo. Theoretically, N-acyloxyalkyl derivates of NH-acid compounds undergo a two-step bioconversion into the parent NH-acidic drug through an N-hydroxyalkyl intermediate. However, to our knowledge no published studies have investigated the formation of an intermediate in vivo. In the present study, it was demonstrated that the assumed N-hydroxymethyl intermediate was readily observed both in vitro and in vivo. In vivo, the observed plasma concentration of the intermediate was at the same level as the drug (aripiprazole). When prodrug intermediates are formed, it is important to make a proper pharmacological, pharmacokinetic and toxicological evaluation of the intermediates to ensure patient safety; however, several challenges were identified when testing an N-acyloxyalkyl prodrug. These included the development of a suitable bioanalytical method, the accurate prediction of prodrug bioconversion and thereby the related pharmacokinetics in humans and the toxicological potential of the intermediate. PMID:25756003

  11. Mechanism of brain targeting by dexibuprofen prodrugs modified with ethanolamine-related structures.

    PubMed

    Li, Yanping; Zhou, Yangyang; Jiang, Jiayu; Wang, Xinyi; Fu, Yao; Gong, Tao; Sun, Xun; Zhang, Zhirong

    2015-12-01

    The first molecular insights into how prodrugs modified with ethanolamine-related structures target the brain were generated using an in vitro BBB model and in situ perfusion technique. Prodrugs were delivered safely and efficiently to the brain through tight interaction with the anionic membrane of brain capillary endothelial cells, observed as a shift in zeta potential, followed by uptake into the cells. Prodrugs III and IV carrying primary and secondary amine modifications appeared to enter the brain via energy-independent passive diffusion. In contrast, besides the passive diffusion, prodrugs I and II carrying tertiary amine modifications also appeared to enter via an active process that was energy and pH dependent but was independent of sodium or membrane potential. This active process involved, at least in part, the pyrilamine-sensitive H(+)/OC antiporter, for which the N,N-diethyl-based compound II showed a much lower affinity than the N,N-dimethyl-based compound I, likely due to steric hindrance. These new insights into brain-targeting mechanisms may help guide efforts to design new prodrugs. PMID:26154870

  12. Nitroreductase gene-directed enzyme prodrug therapy: insights and advances toward clinical utility.

    PubMed

    Williams, Elsie M; Little, Rory F; Mowday, Alexandra M; Rich, Michelle H; Chan-Hyams, Jasmine V E; Copp, Janine N; Smaill, Jeff B; Patterson, Adam V; Ackerley, David F

    2015-10-15

    This review examines the vast catalytic and therapeutic potential offered by type I (i.e. oxygen-insensitive) nitroreductase enzymes in partnership with nitroaromatic prodrugs, with particular focus on gene-directed enzyme prodrug therapy (GDEPT; a form of cancer gene therapy). Important first indications of this potential were demonstrated over 20 years ago, for the enzyme-prodrug pairing of Escherichia coli NfsB and CB1954 [5-(aziridin-1-yl)-2,4-dinitrobenzamide]. However, it has become apparent that both the enzyme and the prodrug in this prototypical pairing have limitations that have impeded their clinical progression. Recently, substantial advances have been made in the biodiscovery and engineering of superior nitroreductase variants, in particular development of elegant high-throughput screening capabilities to enable optimization of desirable activities via directed evolution. These advances in enzymology have been paralleled by advances in medicinal chemistry, leading to the development of second- and third-generation nitroaromatic prodrugs that offer substantial advantages over CB1954 for nitroreductase GDEPT, including greater dose-potency and enhanced ability of the activated metabolite(s) to exhibit a local bystander effect. In addition to forging substantial progress towards future clinical trials, this research is supporting other fields, most notably the development and improvement of targeted cellular ablation capabilities in small animal models, such as zebrafish, to enable cell-specific physiology or regeneration studies. PMID:26431849

  13. Synthesis and Evaluation as Prodrugs of Hydrophilic Carbamate Ester Analogues of Resveratrol.

    PubMed

    Azzolini, Michele; Mattarei, Andrea; La Spina, Martina; Marotta, Ester; Zoratti, Mario; Paradisi, Cristina; Biasutto, Lucia

    2015-09-01

    Resveratrol (3,5,4'-trihydroxy-trans-stilbene) is an unfulfilled promise for health care: its exploitation is hindered by rapid conjugative metabolism in enterocytes and hepatocytes; low water solubility is a serious practical problem. To advantageously modify the physicochemical properties of the compound we have developed prodrugs in which all or part of the hydroxyl groups are linked via an N-monosubstituted carbamate ester bond to promoieties derived from glycerol or galactose, conferring higher water solubility. Kinetic studies of hydrolysis in aqueous solutions and in blood indicated that regeneration of resveratrol takes place in an appropriate time frame for delivery via oral administration. Despite their hydrophilicity some of the synthesized compounds were absorbed in the gastrointestinal tract of rats. In these cases the species found in blood after administration of a bolus consisted mainly of partially deprotected resveratrol derivatives and of the products of their glucuronidation, thus providing proof-of-principle evidence of behavior as prodrugs. The soluble compounds largely reached the lower intestinal tract. Upon administration of resveratrol, the major species found in this region was dihydroresveratrol, produced by enzymes of the intestinal flora. In experiments with a fully protected (trisubstituted) deoxygalactose containing prodrug, the major species were the prodrug itself and partially deprotected derivatives, along with small amounts of dihydroresveratrol. We conclude that the N-monosubstituted carbamate moiety is suitable for use in prodrugs of polyphenols. PMID:26252229

  14. Dipeptidyl peptidase IV dependent water-soluble prodrugs of highly lipophilic bicyclic nucleoside analogues.

    PubMed

    Diez-Torrubia, Alberto; Balzarini, Jan; Andrei, Graciela; Snoeck, Robert; De Meester, Ingrid; Camarasa, María-José; Velázquez, Sonsoles

    2011-03-24

    We present the first report of the application of the dipeptidyl peptidase IV (DPPIV/CD26) based prodrug approach to hydroxy-containing drug derivatives. In particular, we applied this strategy to the highly lipophilic antiviral drug family of bicyclic furanopyrimidine nucleoside analogues (BCNA) in order to improve their physicochemical and pharmacokinetic properties. Our stability data demonstrated that the prodrugs efficiently release the parent BCNA drug upon selective conversion by purified DPPIV/CD26 and by soluble DPPIV/CD26 present in bovine, murine, and human serum. Vildagliptin, a specific inhibitor of DPPIV/CD26, was able to completely block the hydrolysis of the prodrugs in the presence of purified DPPIV/CD26 human, murine, and bovine serum. Several novel prodrugs showed remarkable increases in water solubility (up to more than 3 orders of magnitude) compared to the poorly soluble parent drug. We also demonstrated a markedly enhanced oral bioavailability of the prodrugs versus the parent drug in mice. PMID:21332170

  15. Amorphous Solid Dispersions or Prodrugs: Complementary Strategies to Increase Drug Absorption.

    PubMed

    Rumondor, Alfred C F; Dhareshwar, Sundeep S; Kesisoglou, Filippos

    2016-09-01

    Maximizing oral bioavailability of drug candidates represents a challenge in the pharmaceutical industry. In recent years, there has been an increase in the use of amorphous solid dispersions (ASDs) to address this issue, where a growing number of solid dispersion formulations have been introduced to the market. However, an increase in solubility or dissolution rate through ASD does not always result in sufficient improvement of oral absorption because solubility limitations may still exist at high doses. Chemical modification in the form of a prodrug may offer an alternative approach for these cases. Although prodrugs have been primarily used to improve membrane permeability, examples are available in which prodrugs have been used to increase drug solubility beyond what can be achieved via formulation approaches. In this mini review, the role of ASDs and prodrugs as 2 complementary approaches in improving oral bioavailability of drug candidates is discussed. We discuss the fundamental principles of absorption and bioavailability, and review available literature on both solid dispersions and prodrugs, providing a summary of their use and examples of successful applications, and cover some of the biopharmaceutics evaluation aspects for these approaches. PMID:26886316

  16. Synthesis of Mevalonate- and Fluorinated Mevalonate Prodrugs and Their in vitro Human Plasma Stability

    PubMed Central

    Kang, Soosung; Watanabe, Mizuki; Jacobs, JC; Yamaguchi, Masaya; Dahesh, Samira; Nizet, Victor; Leyh, Thomas S.; Silverman, Richard B.

    2014-01-01

    The mevalonate pathway is essential for the production of many important molecules in lipid biosynthesis. Inhibition of this pathway is the mechanism of statin cholesterol-lowering drugs, as well as the target of drugs to treat osteoporosis, to combat parasites, and to inhibit tumor cell growth. Unlike the human mevalonate pathway, the bacterial pathway appears to be regulated by diphosphomevalonate (DPM). Enzymes in the mevalonate pathway act to produce isopentenyl diphosphate, the product of the DPM decarboxylase reaction, utilize phosphorylated (charged) intermediates, which are poorly bioavailable. It has been shown that fluorinated DPMs (6-fluoro- and 6,6,6-trifluoro-5-diphosphomevalonate) are excellent inhibitors of the bacterial pathway; however, highly charged DPM and analogues are not bioavailable. To increase cellular permeability of mevalonate analogues, we have synthesized various prodrugs of mevalonate and 6-fluoro- and 6,6,6-trifluoromevalonate that can be enzymatically transformed to the corresponding DPM or fluorinated DPM analogues by esterases or amidases. To probe the required stabilities as potentially bioavailable prodrugs, we measured the half-lives of esters, amides, carbonates, acetals, and ketal promoieties of mevalonate and the fluorinated mevalonate analogues in human blood plasma. Stability studies showed that the prodrugs are converted to the mevalonates in human plasma with a wide range of half-lives. These studies provide stability data for a variety of prodrug options having varying stabilities and should be very useful in the design of appropriate prodrugs of mevalonate and fluorinated mevalonates. PMID:25461893

  17. Synthesis of a highly water-soluble acacetin prodrug for treating experimental atrial fibrillation in beagle dogs

    PubMed Central

    Liu, Hui; Wang, Ya-Jing; Yang, Lei; Zhou, Mei; Jin, Man-Wen; Xiao, Guo-Sheng; Wang, Yan; Sun, Hai-Ying; Li, Gui-Rong

    2016-01-01

    We previously reported that duodenal administration of the natural flavone acacetin can effectively prevent the induction of experimental atrial fibrillation (AF) in canines; however, it may not be used intravenously to terminate AF due to its poor water-solubility. The present study was to design a water-soluble prodrug of acacetin and investigate its anti-AF effect in beagle dogs. Acacetin prodrug was synthesized by a three-step procedure. Aqueous solubility, bioconversion and anti-AF efficacy of acacetin prodrug were determined with different methodologies. Our results demonstrated that the synthesized phosphate sodium salt of acacetin prodrug had a remarkable increase of aqueous solubility in H2O and clinically acceptable solution (5% glucose or 0.9% NaCl). The acacetin prodrug was effectively converted into acacetin in ex vivo rat plasma and liver microsome, and in vivo beagle dogs. Intravenous infusion of acacetin prodrug (3, 6 and 12 mg/kg) terminated experimental AF without increasing ECG QTc interval in beagle dogs. The intravenous LD50 of acacetin prodrug was 721 mg/kg in mice. Our preclinical study indicates that the synthesized acacetin prodrug is highly water-soluble and safe; it effectively terminates experimental AF in beagle dogs and therefore may be a promising drug candidate for clinical trial to treat patients with acute AF. PMID:27160397

  18. Modulating lipophilicity of rohitukine via prodrug approach: Preparation, characterization, and in vitro enzymatic hydrolysis in biorelevant media.

    PubMed

    Kumar, Vikas; Bharate, Sonali S; Vishwakarma, Ram A

    2016-09-20

    Rohitukine is a medicinally important natural product which has inspired the discovery of two anticancer clinical candidates. Rohitukine is highly hydrophilic in nature which hampers its oral bioavailability. Thus, herein our objective was to improve the drug-like properties of rohitukine via prodrug-strategy. Various ester prodrugs were synthesized and studied for solubility, lipophilicity, chemical stability and enzymatic hydrolysis in plasma/esterase. All prodrugs displayed lower aqueous solubility and improved lipophilicity compared with rohitukine, which was in accordance with the criteria of compounds in drug-discovery. The stability of synthesized prodrugs was evaluated in buffers at different pH, SGF, SIF, rat plasma and in esterase enzyme. The rate of hydrolysis in all incubation media was dependent primarily on the acyl promoieties. Hexanoyl ester prodrug of rohitukine, 3d, was stable under chemical conditions; however it was completely hydrolyzed to rohitukine, in plasma and in esterase in 4h. Hexanoate ester 3d appeared to be the most promising prodrug as it remained intact at gastric/intestinal pH and was completely transformed to the parent compound in plasma as desired for an ideal prodrug. The data presented herein, will help in designing prodrugs with desired physicochemical properties in future in structurally similar chemotypes. PMID:27422078

  19. Synthesis of a highly water-soluble acacetin prodrug for treating experimental atrial fibrillation in beagle dogs.

    PubMed

    Liu, Hui; Wang, Ya-Jing; Yang, Lei; Zhou, Mei; Jin, Man-Wen; Xiao, Guo-Sheng; Wang, Yan; Sun, Hai-Ying; Li, Gui-Rong

    2016-01-01

    We previously reported that duodenal administration of the natural flavone acacetin can effectively prevent the induction of experimental atrial fibrillation (AF) in canines; however, it may not be used intravenously to terminate AF due to its poor water-solubility. The present study was to design a water-soluble prodrug of acacetin and investigate its anti-AF effect in beagle dogs. Acacetin prodrug was synthesized by a three-step procedure. Aqueous solubility, bioconversion and anti-AF efficacy of acacetin prodrug were determined with different methodologies. Our results demonstrated that the synthesized phosphate sodium salt of acacetin prodrug had a remarkable increase of aqueous solubility in H2O and clinically acceptable solution (5% glucose or 0.9% NaCl). The acacetin prodrug was effectively converted into acacetin in ex vivo rat plasma and liver microsome, and in vivo beagle dogs. Intravenous infusion of acacetin prodrug (3, 6 and 12 mg/kg) terminated experimental AF without increasing ECG QTc interval in beagle dogs. The intravenous LD50 of acacetin prodrug was 721 mg/kg in mice. Our preclinical study indicates that the synthesized acacetin prodrug is highly water-soluble and safe; it effectively terminates experimental AF in beagle dogs and therefore may be a promising drug candidate for clinical trial to treat patients with acute AF. PMID:27160397

  20. Glutathione- and pH-responsive nonporous silica prodrug nanoparticles for controlled release and cancer therapy

    NASA Astrophysics Data System (ADS)

    Xu, Zhigang; Liu, Shiying; Kang, Yuejun; Wang, Mingfeng

    2015-03-01

    A myriad of drug delivery systems such as liposomes, micelles, polymers and inorganic nanoparticles (NPs) have been developed for cancer therapy. Very few of them, however, have the ability to integrate multiple functionalities such as specific delivery, high circulation stability, controllable release and good biocompatibility and biodegradability in a single system to improve the therapeutic efficacy. Herein, we report two types of stimuli-responsive nonporous silica prodrug NPs towards this goal for controlled release of anticancer drugs and efficient combinatorial cancer therapy. As a proof of concept, anticancer drugs camptothecin (CPT) and doxorubicin (DOX) were covalently encapsulated into silica matrices through glutathione (GSH)-responsive disulfide and pH-responsive hydrazone bonds, respectively, resulting in NPs with sizes tunable in the range of 50-200 nm. Both silica prodrug NPs showed stimuli-responsive controlled release upon exposure to a GSH-rich or acidic environment, resulting in improved anticancer efficacy. Notably, two prodrug NPs simultaneously taken up by HeLa cells showed a remarkable combinatorial efficacy compared to free drug pairs. These results suggest that the stimuli-responsive silica prodrug NPs are promising anticancer drug carriers for efficient cancer therapy.A myriad of drug delivery systems such as liposomes, micelles, polymers and inorganic nanoparticles (NPs) have been developed for cancer therapy. Very few of them, however, have the ability to integrate multiple functionalities such as specific delivery, high circulation stability, controllable release and good biocompatibility and biodegradability in a single system to improve the therapeutic efficacy. Herein, we report two types of stimuli-responsive nonporous silica prodrug NPs towards this goal for controlled release of anticancer drugs and efficient combinatorial cancer therapy. As a proof of concept, anticancer drugs camptothecin (CPT) and doxorubicin (DOX) were

  1. The application of prodrug-based nano-drug delivery strategy in cancer combination therapy.

    PubMed

    Ge, Yanxiu; Ma, Yakun; Li, Lingbing

    2016-10-01

    Single drug therapy that leads to the multidrug resistance of cancer cells and severe side-effect is a thing of the past. Combination therapies that affect multiple signaling pathways have been the focus of recent active research. Due to the successful development of prodrug-based nano-drug delivery systems (P-N-DDSs), their use has been extended to combination therapy as drug delivery platforms. In this review, we focus specifically on the P-N-DDSs in the field of combination therapy including the combinations of prodrugs with different chemotherapeutic agents, other therapeutic agents, nucleic acid or the combination of different types of therapy (e.g. chemotherapy and phototherapy). The relevant examples of prodrug-based nanoparticulate drug delivery strategy in combination cancer therapy from the recent literature are discussed to demonstrate the feasibilities of relevant technology. PMID:27400243

  2. Dual-Mode HDAC Prodrug for Covalent Modification and Subsequent Inhibitor Release

    PubMed Central

    2016-01-01

    Histone deacetylase inhibitors (HDACi) target abnormal epigenetic states associated with a variety of pathologies, including cancer. Here, the development of a prodrug of the canonical broad-spectrum HDACi suberoylanilide hydroxamic acid (SAHA) is described. Although hydroxamic acids are utilized universally in the development of metalloenzyme inhibitors, they are considered to be poor pharmacophores with reduced activity in vivo. We developed a prodrug of SAHA by appending a promoiety, sensitive to thiols, to the hydroxamic acid warhead (termed SAHA-TAP). After incubation of SAHA-TAP with an HDAC, the thiol of a conserved HDAC cysteine residue becomes covalently tagged with the promoiety, initiating a cascade reaction that leads to the release of SAHA. Mass spectrometry and enzyme kinetics experiments validate that the cysteine residue is covalently appended with the TAP promoiety. SAHA-TAP demonstrates cytotoxicity activity against various cancer cell lines. This strategy represents an original prodrug design with a dual mode of action for HDAC inhibition. PMID:25974739

  3. Horseradish peroxidase-encapsulated chitosan nanoparticles for enzyme-prodrug cancer therapy.

    PubMed

    Cao, Xiaodan; Chen, Chao; Yu, Haijun; Wang, Ping

    2015-01-01

    Among various enzyme-based therapies, enzyme-prodrug therapy (EPT) promises minimized side effects in that it activates non-toxic prodrugs locally where the enzymes are placed. The success of such an approach requires high enzyme stability against both structural denaturation and potential immunogenicity. This work examines the efficiency of nanoparticles for enzyme protection in EPT applications. Specifically, horseradish peroxidase (HRP)-encapsulated chitosan nanoparticles (HRP-CSNP) were constructed and examined with respect to stability enhancement. HRP-CSNP retained enzyme activity and had improved stability at 37 °C in the presence of a denaturant, urea. The nanoparticles effectively bound to the surface of human breast cancer cell Bcap37 and led to over 80 % cell death when applied with a prodrug indole-3-acetic acid. PMID:25257586

  4. Poly(ethylene glycol)-Prodrug Conjugates: Concept, Design, and Applications

    PubMed Central

    Banerjee, Shashwat S.; Aher, Naval; Patil, Rajesh; Khandare, Jayant

    2012-01-01

    Poly(ethylene glycol) (PEG) is the most widely used polymer in delivering anticancer drugs clinically. PEGylation (i.e., the covalent attachment of PEG) of peptides proteins, drugs, and bioactives is known to enhance the aqueous solubility of hydrophobic drugs, prolong circulation time, minimize nonspecific uptake, and achieve specific tumor targetability through the enhanced permeability and retention effect. Numerous PEG-based therapeutics have been developed, and several have received market approval. A vast amount of clinical experience has been gained which has helped to design PEG prodrug conjugates with improved therapeutic efficacy and reduced systemic toxicity. However, more efforts in designing PEG-based prodrug conjugates are anticipated. In light of this, the current paper highlights the synthetic advances in PEG prodrug conjugation methodologies with varied bioactive components of clinical relevance. In addition, this paper discusses FDA-approved PEGylated delivery systems, their intended clinical applications, and formulations under clinical trials. PMID:22645686

  5. Novel Resveratrol-Based Aspirin Prodrugs: Synthesis, Metabolism, and Anticancer Activity.

    PubMed

    Zhu, Yingdong; Fu, Junsheng; Shurlknight, Kelly L; Soroka, Dominique N; Hu, Yuhui; Chen, Xiaoxin; Sang, Shengmin

    2015-08-27

    Regular aspirin use has been convincingly shown to reduce the risk of colorectal cancer. However, long-term use of aspirin leads to gastrotoxicity. Herein, we designed and synthesized a novel class of resveratrol-based aspirin prodrugs to simultaneously release aspirin and resveratrol to attenuate the side effects caused by aspirin. Prodrug RAH exerted enhanced anticancer activities which are better than a physical mixture of aspirin and resveratrol as well as each individually. Metabolism of RAH in mice showed that the majority of RAH is decomposed to release resveratrol and aspirin or salicylic acid either in the intestine or after absorption. Mechanistic studies demonstrate RAH inhibits cell cycle arrest through downregulation of cyclins and induces apoptosis by activation of caspase-3 in cancer cells. These findings highlighted the improved anticancer properties of resveratrol-based aspirin prodrugs. RAH may represent novel and safe alternatives of aspirin for the purpose of daily use in the future. PMID:26204233

  6. Modulation of Activity Profiles for Largazole-Based HDAC Inhibitors through Alteration of Prodrug Properties

    PubMed Central

    2014-01-01

    Largazole is a potent and class I-selective histone deacetylase (HDAC) inhibitor purified from marine cyanobacteria and was demonstrated to possess antitumor activity. Largazole employs a unique prodrug strategy, via a thioester moiety, to liberate the bioactive species largazole thiol. Here we report alternate prodrug strategies to modulate the pharmacokinetic and pharmacodynamics profiles of new largazole-based compounds. The in vitro effects of largazole analogues on cancer cell proliferation and enzymatic activities of purified HDACs were comparable to the natural product. However, in vitro and in vivo histone hyperacetylation in HCT116 cells and implanted tumors, respectively, showed differences, particularly in the onset of action and oral bioavailability. These results indicate that, by employing a different approach to disguise the “warhead” moiety, the functional consequence of these prodrugs can be significantly modulated. Our data corroborate the role of the pharmacokinetic properties of this class of compounds to elicit the desired and timely functional response. PMID:25147612

  7. Drug-Initiated Synthesis of Polymer Prodrugs: Combining Simplicity and Efficacy in Drug Delivery†

    PubMed Central

    2016-01-01

    In the field of nanomedicine, the global trend over the past few years has been toward the design of highly sophisticated drug delivery systems with active targeting and/or imaging capabilities, as well as responsiveness to various stimuli to increase their therapeutic efficacy. However, providing sophistication generally increases complexity that could be detrimental in regards to potential pharmaceutical development. An emerging concept to design efficient yet simple drug delivery systems, termed the “drug-initiated” method, consists of growing short polymer chains from drugs in a controlled fashion to yield well-defined drug–polymer prodrugs. These materials are obtained in a reduced amount of synthetic steps and can be self-assembled into polymer prodrug nanoparticles, be incorporated into lipid nanocarriers or be used as water-soluble polymer prodrugs. This Perspective article will capture the recent achievements from the “drug-initiated” method and highlight the great biomedical potential of these materials. PMID:27041820

  8. Synthesis and Characterization of a New Peptide Prodrug of Glucosamine with Enhanced Gut Permeability

    PubMed Central

    Gilzad Kohan, Hamed; Kaur, Kamaljit; Jamali, Fakhreddin

    2015-01-01

    The aim of this study was to synthesize a peptide prodrug of glucosamine (GlcN) with increased gut permeability through the gut peptide transporter 1 (PepT1). Glycine-Valine ester derivative of GlcN (GVG) was synthesised using solid phase synthesis followed by characterization and evaluation of its physicochemical and intestinal stability. In addition, GVG was evaluated for its ability to be biotransformed to GlcN in the liver homogenate. In vitro absorption of the new prodrug through everted rat gut was also assessed. GVG demonstrated significant and meaningful increased gut permeability as compared with GlcN. It showed favorable stability in the gut and a quick cleavage to GlcN after exposure to the liver homogenate. In conclusion, a novel prodrug of glucosamine with superior gut permeability compared to GlcN was developed and successfully tested in vitro. PMID:25978315

  9. Novel multifunctional acyloxyalkyl ester prodrugs of 5-aminolevulinic acid display improved anticancer activity dependent on photoactivation

    NASA Astrophysics Data System (ADS)

    Berkovitch, G.; Nudelman, A.; Ehenberg, B.; Rephaeli, A.; Malik, Z.

    2009-06-01

    New approaches to PDT using multifunctional 5-aminolevulinic acid (ALA) based prodrugs activating mutual routes of toxicity are described. We investigated the mutual anti-cancer activity of ALA prodrugs which upon metabolic hydrolysis by unspecific esterases release ALA, formaldehyde or acetaldehye and the histone deacetylase inhibitor (HDACI) butyric acid. The most potent prodrug in this study was butyryloxyethyl 5-amino-4-oxopentanoate (AN-233) that stimulated a rapid biosynthesis of protoporphyrin IX (PpIX) in human glioblastoma U-251 cells and generated an efficient photodynamic destruction. AN-233 induced a considerable high level of intracellular ROS in the cells following light irradiation, reduction of mitochondrial activity, dissipation of the mitochondrial membrane potential resulting in necrotic and apoptotic cell death. The main advantage of AN-233 over ALA stems from its ability to induce photodamage at a significantly lower dose than ALA.

  10. Antimalarial activity of prodrugs of N-branched acyclic nucleoside phosphonate inhibitors of 6-oxopurine phosphoribosyltransferases.

    PubMed

    Hocková, Dana; Janeba, Zlatko; Naesens, Lieve; Edstein, Michael D; Chavchich, Marina; Keough, Dianne T; Guddat, Luke W

    2015-09-01

    Acyclic nucleoside phosphonates (ANPs) that contain a 6-oxopurine base are good inhibitors of the human and Plasmodium falciparum 6-oxopurine phosphoribosyltransferases (PRTs), key enzymes of the purine salvage pathway. Chemical modifications, based on the crystal structures of several inhibitors in complex with the human PRTase, led to the design of a new class of inhibitors--the aza-ANPs. Because of the negative charges of the phosphonic acid moiety, their ability to cross cell membranes is, however, limited. Thus, phosphoramidate prodrugs of the aza-ANPs were prepared to improve permeability. These prodrugs arrest parasitemia with IC50 values in the micromolar range against Plasmodium falciparum-infected erythrocyte cultures (both chloroquine-sensitive and chloroquine-resistant Pf strains). The prodrugs exhibit low cytotoxicity in several human cell lines. Thus, they fulfill two essential criteria to qualify them as promising antimalarial drug leads. PMID:26275679