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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. 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

  17. 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

  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. 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

  9. 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

  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. 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

  2. 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

  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. 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

  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. 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

  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. 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

  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. 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

  2. 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

  3. 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

  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. 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.

  6. 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

  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. 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

  4. 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

  5. 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

  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. 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

  8. 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.

  9. 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

  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. 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

  13. 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

  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. 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

  20. 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

  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. 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

  3. 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

  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. 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

  12. 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

  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. 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

  16. 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

  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. 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

  1. 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

  2. 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

  3. 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

  4. 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

  5. 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.

  6. 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

  7. 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

  8. 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

  9. 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

  10. 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

  11. 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.

  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. 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

  14. 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

  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. 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

  20. 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

  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. 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

  3. 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

  4. 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

  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. 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

  9. 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

  10. 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

  11. 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

  12. 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

  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. 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

  16. 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

  17. 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

  18. 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

  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. 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

  2. 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

  3. 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

  4. 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

  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. 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

  12. 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

  13. 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

  14. 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

  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. 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

  1. 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

  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. 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

  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

    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

  13. 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

  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. 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

  19. 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

  20. 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

  1. 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

  2. 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

  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. 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

  12. 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

  13. 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

  14. 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

  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. 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

  20. 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

  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. 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

  6. 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

  7. 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

  8. 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

  9. 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

  10. 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

  11. 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

  12. 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

  13. 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

  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. 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

  16. 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

  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. 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

  19. 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

  20. 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

  1. 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

  2. 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

  3. 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

  4. 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.

  5. 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

  6. Riluzole prodrugs for melanoma and ALS: design, synthesis, and in vitro metabolic profiling

    PubMed Central

    McDonnell, Mark E.; Vera, Matthew D.; Blass, Benjamin E.; Pelletier, Jeffrey C.; King, Richard C.; Fernandez-Metzler, Carmen; Smith, Garry R.; Wrobel, Jay; Chen, Suzie; Reitz, Allen B.

    2012-01-01

    Riluzole (1) is an approved therapeutic for the treatment of ALS and has also demonstrated antimelanoma activity in metabotropic glutamate GRM1 positive cell lines, a mouse xenograft assay and human clinical trials. Highly variable drug exposure following oral administration among patients, likely due to variable first pass effects from heterogeneous CYP1A2 expression, hinders its clinical use. In an effort to mitigate effects of this clearance pathway and uniformly administer riluzole at efficacious exposure levels, several classes of prodrugs of riluzole were designed, synthesized, and evaluated in multiple in vitro stability assays to predict in vivo drug levels. The optimal prodrug would possess the following profile: stability while transiting the digestive system, stability towards first pass metabolism, and metabolic lability in the plasma releasing riluzole. (S)-O-Benzyl serine derivative 9 was identified as the most promising therapeutically acceptable prodrug. PMID:22892214

  7. Development of Platinum(iv) Complexes as Anticancer Prodrugs: the Story so Far

    NASA Astrophysics Data System (ADS)

    Wong, Daniel Yuan Qiang; Ang, Wee Han

    2012-06-01

    The serendipitous discovery of the antitumor properties of cisplatin by Barnett Rosenberg some forty years ago brought about a paradigm shift in the field of medicinal chemistry and challenged conventional thinking regarding the role of potentially toxic heavy metals in drugs. Platinum(II)-based anticancer drugs have since become some of the most effective and widely-used drugs in a clinician's arsenal and have saved countless lives. However, they are limited by high toxicity, severe side-effects and the incidence of drug resistance. In recent years, attention has shifted to stable platinum(IV) complexes as anticancer prodrugs. By exploiting the unique chemical and structural attributes of their scaffolds, these platinum(IV) prodrugs offer new strategies of targeting and killing cancer cells. This review summarizes the development of anticancer platinum(IV) prodrugs to date and some of the exciting strategies that utilise the platinum(IV) construct as targeted chemotherapeutic agents against cancer.

  8. 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

  9. 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

  10. 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

  11. Co-delivery of 10-hydroxycamptothecin with doxorubicin conjugated prodrugs for enhanced anticancer efficacy.

    PubMed

    Zhang, Yu; Xiao, Chunsheng; Li, Mingqiang; Chen, Jie; Ding, Jianxun; He, Chaoliang; Zhuang, Xiuli; Chen, Xuesi

    2013-05-01

    Well-defined amphiphilic linear-dendritic prodrugs (MPEG-b-PAMAM-DOX) are synthesized by conjugating doxorubicin (DOX), to MPEG-b-PAMAM through the acid-labile hydrazone bond. The amphiphilic prodrugs form self-assembled nanoparticles in deionized water and encapsulate the hydrophobic anticancer drug 10-hydroxycamptothecin (HCPT) with a high drug loading efficiency. Studies on drug release and cellular uptake of the co-delivery system reveal that both drugs are released in a pH-dependent manner and effectively taken up by MCF-7 cells. In vitro methyl thiazolyl tetrazolium (MTT) assays and drug-induced apoptosis tests demonstrate the HCPT-loaded nanoparticles suppress cancer cell growth more efficiently than the MPEG-b-PAMAM-DOX prodrugs, free HCPT, and physical mixtures of MPEG-b-PAMAM-DOX and HCPT at equivalent DOX or HCPT doses. PMID:23420692

  12. 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

  13. 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

  14. Gastric-sparing nitric oxide-releasable 'true' prodrugs of aspirin and naproxen.

    PubMed

    Gund, Machhindra; Gaikwad, Parikshit; Borhade, Namdev; Burhan, Aslam; Desai, Dattatraya C; Sharma, Ankur; Dhiman, Mini; Patil, Mohan; Sheikh, Javed; Thakre, Gajanan; Tipparam, Santhosh G; Sharma, Somesh; Nemmani, Kumar V S; Satyam, Apparao

    2014-12-15

    Nitric oxide-releasing non-steroidal anti-inflammatory drugs (NO-NSAIDs) are gaining attention as potentially gastric-sparing NSAIDs. Herein, we report a novel class of '1-(nitrooxy)ethyl ester' group-containing NSAIDS as efficient NO releasing 'true' prodrugs of aspirin and naproxen. While an aspirin prodrug exhibited comparable oral bioavailability and antiplatelet activity (i.e., TXB2 inhibition) to those of aspirin, a naproxen prodrug exhibited better bioavailability than naproxen. These promising NO-NSAIDs protected experimental rats from gastric damage. We therefore believe that these promising NO-NSAIDs could represent a new class of potentially 'Safe NSAIDs' for the treatment of arthritic pain, inflammation and cardiovascular disorders in the case of NO-aspirin. PMID:25466180

  15. Lipophilic Prodrugs of SN38: Synthesis and in Vitro Characterization toward Oral Chemotherapy.

    PubMed

    Bala, Vaskor; Rao, Shasha; Li, Peng; Wang, Shudong; Prestidge, Clive A

    2016-01-01

    SN38 (7-ethyl-10-hydroxy camptothecin) is a potent anticancer agent belonging to the camptothecin family; however, its oral delivery is extensively restricted by poor solubility in pharmaceutically acceptable excipients and low transmucosal permeability. Lipid-based carriers are well-known for their ability to improve oral absorption and bioavailability of lipid soluble and highly permeable compounds. Thus, this study has focused on improving solubility in lipid excipients, controlling stability, and enhancing transmucosal permeability of SN38 by specific chemical modification. To achieve these aims, a series of lipophilic prodrugs were designed and synthesized by esterification at the C10 and/or C20 positon(s) of SN38 with dietary fatty acids of diverse hydrocarbon chain lengths. The solubility of these novel prodrugs in long-chain triglycerides was increased up to 444-fold, and cytotoxicity was significantly reduced in comparison to SN38. The prodrugs were stable in simulated gastric fluids but exhibited different rates of hydrolysis (t1/2 < 5 min to t1/2 > 2 h) in simulated intestinal fluids (in the presence of enzymes) depending on the alkyl chain length and the position modified. A predictable reconversion of prodrugs to SN38 in plasma was also confirmed. On the basis of these studies, SN38-undecanoate (C20) was identified as the optimal prodrug. Finally, in vitro permeability and uptake studies in rat intestinal mucosal membrane using an Ussing chamber showed significant improvement in transepithelial drug transport and cellular uptake. Together, these results indicate that well designed lipophilic prodrugs have potential for the efficacious and safe oral delivery of SN38. PMID:26623947

  16. Delivery of a Protease-Activated Cytolytic Peptide Prodrug by Perfluorocarbon Nanoparticles.

    PubMed

    Jallouk, Andrew P; Palekar, Rohun U; Marsh, Jon N; Pan, Hua; Pham, Christine T N; Schlesinger, Paul H; Wickline, Samuel A

    2015-08-19

    Melittin is a cytolytic peptide derived from bee venom that inserts into lipid membranes and oligomerizes to form membrane pores. Although this peptide is an attractive candidate for treatment of cancers and infectious processes, its nonspecific cytotoxicity and hemolytic activity have limited its therapeutic applications. Several groups have reported the development of cytolytic peptide prodrugs that only exhibit cytotoxicity following activation by site-specific proteases. However, systemic administration of these constructs has proven difficult because of their poor pharmacokinetic properties. Here, we present a platform for the design of protease-activated melittin derivatives that may be used in conjunction with a perfluorocarbon nanoparticle delivery system. Although native melittin was substantially hemolytic (HD50: 1.9 μM) and cytotoxic (IC50: 2.4 μM), the prodrug exhibited 2 orders of magnitude less hemolytic activity (HD50: > 100 μM) and cytotoxicity (IC50: > 100 μM). Incubation with matrix metalloproteinase-9 (MMP-9) led to cleavage of the prodrug at the expected site and restoration of hemolytic activity (HD50: 3.4 μM) and cytotoxicity (IC50: 8.1 μM). Incubation of the prodrug with perfluorocarbon nanoparticles led to stable loading of 10,250 peptides per nanoparticle. Nanoparticle-bound prodrug was also cleaved and activated by MMP-9, albeit at a fourfold slower rate. Intravenous administration of prodrug-loaded nanoparticles in a mouse model of melanoma significantly decreased tumor growth rate (p = 0.01). Because MMPs and other proteases play a key role in cancer invasion and metastasis, this platform holds promise for the development of personalized cancer therapies directed toward a patient's individual protease expression profile. PMID:26083278

  17. An overview of prodrug technology and its application for developing abuse-deterrent opioids.

    PubMed

    Gudin, Jeffrey A; Nalamachu, Srinivas R

    2016-01-01

    The Centers for Disease Control and Prevention has classified prescription drug abuse and overdose deaths as an epidemic. Prescription drug overdose is now the leading cause of injury death, with rates that have more than doubled since 1999. This crisis has developed concurrently with the increased prescribing and availability analgesic drugs, especially opioids, resulting from an effort on the part of clinicians to address a critical need for improved pain assessment and treatment. Clinicians have recognized that oftentimes, opioid analgesics are one of the few remaining options for patients who suffer with severe pain. A 2015 fact sheet issued by the Office of National Drug Policy stated: "While we must ensure better access to prescription medications to alleviate suffering, it is also vital that we do all we can to reduce the diversion and abuse of pharmaceuticals." The US Food and Drug Administration has issued guidance that encourages the research and development of abuse-deterrent formulation of opioids which have the potential to curtail abuse. Included among the recommended formulations for development of abuse-deterrent opioids are prodrugs. Prodrugs are chemically modified versions of pharmacological agents that must undergo a biochemical conversion following administration, often by enzymatic cleavage, to free the active drug. Prodrugs may be inherently abuse-deterrent because they are inactive or significantly less active until conversion to the active drug. This requirement for conversion in the GI tract can modify the pharmacokinetic profile and eliminate or reduce the euphoria when abusers change the route of administration. Abusers often attempt to extract the active drug for injection or insufflation. Prodrugs can be designed to be resistant to crushing or dissolving. In this article, we review the concept of prodrugs and introduce and examine the potential of abuse-deterrent opioid prodrugs. PMID:26615852

  18. Combination therapy with bioengineered miR-34a prodrug and doxorubicin synergistically suppresses osteosarcoma growth.

    PubMed

    Zhao, Yong; Tu, Mei-Juan; Yu, Yi-Feng; Wang, Wei-Peng; Chen, Qiu-Xia; Qiu, Jing-Xin; Yu, Ai-Xi; Yu, Ai-Ming

    2015-12-15

    Osteosarcoma (OS) is the most common form of primary malignant bone tumor and prevalent among children and young adults. Recently we have established a novel approach to bioengineering large quantity of microRNA-34a (miR-34a) prodrug for miRNA replacement therapy. This study is to evaluate combination treatment with miR-34a prodrug and doxorubicin, which may synergistically suppress human OS cell growth via RNA interference and DNA intercalation. Synergistic effects were indeed obvious between miR-34a prodrug and doxorubicin for the suppression of OS cell proliferation, as defined by Chou-Talalay method. The strongest antiproliferative synergism was achieved when both agents were administered simultaneously to the cells at early stage, which was associated with much greater degrees of late apoptosis, necrosis, and G2 cell cycle arrest. Alteration of OS cellular processes and invasion capacity was linked to the reduction of protein levels of miR-34a targeted (proto-)oncogenes including SIRT1, c-MET, and CDK6. Moreover, orthotopic OS xenograft tumor growth was repressed to a significantly greater degree in mouse models when miR-34a prodrug and doxorubicin were co-administered intravenously. In addition, multiple doses of miR-34a prodrug and doxorubicin had no or minimal effects on mouse blood chemistry profiles. The results demonstrate that combination of doxorubicin chemotherapy and miR-34a replacement therapy produces synergistic antiproliferative effects and it is more effective than monotherapy in suppressing OS xenograft tumor growth. These findings support the development of mechanism-based combination therapy to combat OS and bioengineered miR-34a prodrug represents a new natural miRNA agent. PMID:26518752

  19. Design, synthesis, and evaluation of new cyclophosphamide-based anticancer prodrugs

    SciTech Connect

    Moon, Ki-Young.

    1993-01-01

    Cyclophosphamide (CP,1) is a prodrug that is activated by hepatic microsomal mixed-function oxidase (MFO) catalyzed C[sub 4]-hydroxylation. The resulting 4-hydroxycyclophosphamide (4-OH-CP) undergoes ring opening to aldophosphamide (Aldo), followed by generation of cytotoxic phosphoramide mustard (PDA,2) and acrolein by [beta]-elimination. The cytotoxic activity of CP is attributed to the aziridinium ion species derived from PDA that cross-links interstrand DNA. The aim of this research is to design, synthesize, and evaluate new cyclophosphamide-based alkylating agents to achieve improved therapeutic efficacy against neoplastic cells. Benzyl phosphoramide mustard (Benzyl PDA,4), 2.4-difluorobenzyl phosphoramide mustard (2,4-Difluorobenzyl PDA,5) and methyl phosphoramide mustard (Methyl PDA,6) were examined as lipophilic, chemically stable prodrugs of PDA (2). These phosphorodiamidic esters were designed to undergo biotransformation by hepatic microsomal enzymes to produce 2 without generation of acrolein and to be active against CP-resistant tumor cells. Several N-methyl-4-(alkylthio)cyclophosphamide derivatives were synthesized and examined as chemically stable, biooxidative prodrugs of 4-OH-CP, the activated species of CP. All of the prodrugs underwent N-demethylation in a time-dependent manner when incubated with rat hepatic microsomes, which resulted in formation of formaldehyde as well as alkylating species. Among the prodrugs, N-methyl-4-(diethyldithiocarbamoyl)cyclophosphamide (N-CH[sub 3]-4-DDTC-CP,15) showed exceptional in vitro cytotoxicity against 3T3 cells as well as against a panel of human tumor cell lines, with a particular sensitivity to leukemia and small cell lung cancer cell lines. Preliminary in vivo antitumor evaluation against L1210 leukemia in mice showed that all of the prodrugs were active.

  20. Design, synthesis and characterization of captopril prodrugs for enhanced percutaneous absorption.

    PubMed

    Moss, Gary P; Gullick, Darren R; Cox, Paul A; Alexander, Cameron; Ingram, Matthew J; Smart, John D; Pugh, W John

    2006-02-01

    Most drugs are designed primarily for oral administration, but the activity and stability profiles desirable for this route often make them unsuitable for transdermal delivery. We were therefore interested in designing analogues of captopril, a model drug with poor percutaneous penetration, for which the sustained steady-state blood plasma level associated with transdermal delivery (and which is unattainable orally) would be particularly beneficial. Quantitative structure-permeability relationships (QSPRs) predicted that ester and thiol prodrug derivatives of captopril would have lower maximal transdermal flux (J(m)) than the parent drug, since the increases in permeability coefficient (k(p)) of prodrugs would be outweighed by the reductions in aqueous solubility. Therefore, the aim of this study was to synthesize a series of prodrugs of captopril and to determine if a QSPR model could be used to design therapeutically viable prodrugs. Molecules with the highest predicted k(p) values were synthesized and characterized, and J(m) measured in Franz diffusion cells from saturated aqueous donor across porcine skin (fresh and frozen). In-vitro metabolism was also measured. Captopril and the prodrugs crossed the skin relatively freely, with J(m) being highest for ethyl to butyl esters. Substantial first-order metabolism of the prodrugs was observed, suggesting that their enhanced percutaneous absorption was complemented by their metabolic performance. The results suggested that QSPR models provided excellent enhancements in drug delivery. This was not seen at higher lipophilicities, suggesting that issues of solubility need to be considered in conjunction with any such use of a QSPR model. PMID:16451744

  1. A new type of prodrug of catecholamines: an opportunity to improve the treatment of Parkinson's disease.

    PubMed

    Venhuis, Bastiaan J; Rodenhuis, Nienke; Wikström, Håkan V; Wustrow, David; Meltzer, Leonard T; Wise, Lawrence D; Johnson, Stephen J; Pugsley, Thomas A; Sundell, Staffan; Dijkstra, Durk

    2002-06-01

    After decades of research around dopamine agonists, we have found a promising compound in S-PD148903 that represents a new type of prodrug, which in the rat is bioactivated to the catecholamine S-5,6-diOH-DPAT, known to display mixed dopamine D(1)/D(2) receptor agonist properties just like apomorphine. This prodrug has an enone structure which by an oxidative bioactivation mechanism is converted to the corresponding catechol and is delivered enantioselectively into the CNS. This novel concept has the potential to revolutionize the treatment of Parkinson's disease by competing with L-DOPA, the current treatment of choice. PMID:12036342

  2. Prodrugs of anthracyclines for chemotherapy via enzyme-monoclonal antibody conjugates.

    PubMed

    Gesson, J P; Jacquesy, J C; Mondon, M; Petit, P; Renoux, B; Andrianomenjanahary, S; Dufat-Trinh Van, H; Koch, M; Michel, S; Tillequin, F

    1994-10-01

    New prodrugs of daunorubicin, 1c, 1e and 2c, including a galactopyranosyl residue linked to the N-3' of the daunosaminyl moiety through substituted o- or p-benzyloxycarbonyl groups were synthesized. Their low cytotoxicity and high stability in plasma fulfil the conditions for antibody-directed enzyme prodrug therapy (ADEPT). Enzymatic hydrolysis using alpha-D-galactosidase gives rise to daunorubicin by subsequent self-elimination of the spacers. However, elimination clearly depends on the aromatic substitution pattern, as demonstrated especially by comparison with non-substituted analogues. PMID:7945725

  3. Acetal-linked polymeric prodrug micelles for enhanced curcumin delivery.

    PubMed

    Li, Man; Gao, Min; Fu, Yunlan; Chen, Chao; Meng, Xuan; Fan, Aiping; Kong, Deling; Wang, Zheng; Zhao, Yanjun

    2016-04-01

    On-demand curcumin delivery via stimuli-responsive micellar nanocarriers holds promise for addressing its solubility and stability problem. Polymer-curcumin prodrug conjugate micelle is one of such nanosystems. The diversity of linker and conjugation chemistry enabled the generation and optimization of different curcumin micelles with tunable stimuli-responsiveness and delivery efficiency. The aim of the current work was to generate and assess acetal-linked polymeric micelles to enrich the pH-responsive curcumin delivery platforms. Curcumin was slightly modified prior to conjugating to amphiphilic methoxy poly(ethylene glycol)-poly(lactic acid) (mPEG-PLA) copolymer via an acetal bond, whereas an ester bond-linked conjugate was used as the control. The acetal-containing micelles showed a hydrodynamic diameter of 91.1 ± 2.9(nm) and the accompanying core size of 63.5 ± 7.1 (nm) with a zeta potential of -10.9 ± 0.7(mV). Both control and pH-labile micelles displayed similar critical micelle concentration at 1.6 μM. The acetal-containing nanocarriers exhibited a pH-dependent drug release behavior, which was faster at lower pH values. The cytotoxicity study in HepG2 cells revealed a significantly lower IC50 at 51.7 ± 9.0(μM) for acetal-linked micelles in contrast to the control at 103.0 ± 17.8(μM), but the polymer residue showed no cytotoxicity upon drug release. The acetal-linked micellar nanocarrier could be a useful addition to the spectrum of currently available stimuli-responsive curcumin nano-formulations. PMID:26731193

  4. A budesonide prodrug accelerates treatment of colitis in rats.

    PubMed Central

    Cui, N; Friend, D R; Fedorak, R N

    1994-01-01

    Although oral glucocorticoids are the treatment of choice for moderate to severe ulcerative pancolitis, their systemic side effects and adrenal suppression account for considerable morbidity. An oral glucocorticoid-conjugate (prodrug), budesonide-beta-D-glucuronide, which is not absorbed in the small intestine but is hydrolysed by colonic bacterial and mucosal beta-glucuronidase to release free budesonide into the colon was synthesised. The objective of this study was to compare treatment with budesonide-beta-D-glucuronide with treatment with free budesonide by examining: (1) the healing of experimental colitis and (2) the extent of adrenal suppression. Pancolitis was induced with 4% acetic acid. Animals were then randomised to receive oral therapy for 72 hours with (1) budesonide-beta-D-glucuronide, (2) free budesonide, or (3) vehicle. Drug efficacy and colitic healing was determined by measuring gross colonic ulceration, myeloperoxidase activity, and in vivo colonic fluid absorption. Adrenal suppression was determined by measuring plasma adrenocorticotrophic hormone and serum corticosterone. Vehicle-treated colitis animals had gross ulceration, increased myeloperoxidase activity, and net colonic fluid secretion. Treatment with oral budesonide-beta-D-glucuronide accelerated all measures of colitis healing at a fourfold lower dose than did free budesonide. Furthermore, treatment with budesonide-beta-D-glucuronide did not result in adrenal suppression whereas free budesonide treatment did. A newly synthesised orally administered glucocorticoid-conjugate accelerates colitis healing with limited adrenal suppression. Development of an orally administered colon-specific steroid delivery system represents a novel approach to inflammatory bowel disease treatment. PMID:7959202

  5. Synthesis and Characterization of Silicate Ester Prodrugs and Poly(ethylene glycol)-b-poly(lactic-co-glycolic acid) Block Copolymers for Formulation into Prodrug-Loaded Nanoparticles

    NASA Astrophysics Data System (ADS)

    Wohl, Adam Richard

    Fine control of the physical and chemical properties of customized materials is a field that is rapidly advancing. This is especially critical in pursuits to develop and optimize novel nanoparticle drug delivery. Specifically, I aim to apply chemistry concepts to test the hypothesis "Silicate ester prodrugs of paclitaxel, customized to have the proper hydrophobicity and hydrolytic lability, can be formulated with well-defined, biocompatible, amphiphilic block copolymers into nanoparticles that are effective drugs." Chapter 1 briefly describes the context and motivation of the scientific pursuits described in this thesis. In Chapter 2, a family of model silicate esters is synthesized, the hydrolysis rate of each compound is benchmarked, and trends are established based upon the steric bulk and leaving group ability of the silicate substituents. These trends are then applied to the synthesis of labile silicate ester prodrugs in Chapter 3. The bulk of this chapter focuses on the synthesis, hydrolysis, and cytotoxicity of prodrugs based on paclitaxel, a widely used chemotherapeutic agent. In Chapter 4, a new methodology for the synthesis of narrowly dispersed, "random" poly(lactic-co-glycolic acid) polymers by a constant infusion of the glycolide monomer is detailed. Using poly(ethylene glycol) as a macroinitiator, amphiphilic block copolymers were synthesized. Co-formulating a paclitaxel silicate and an amphiphilic block copolymer via flash nanoprecipitation led to highly prodrug-loaded, kinetically trapped nanoparticles. Studies to determine the structure, morphology, behavior, and efficacy of these nanoparticles are described in Chapter 5. Efforts to develop a general strategy for the selective end-functionalization of the polyether block of these amphiphilic block copolymers are discussed in Chapter 6. Examples of this strategy include functionalization of the polyether with an azide or a maleimide. Finally, Chapter 7 provides an outlook for future development of

  6. Structural Characterization and in Vivo Evaluation of β-Hairpin Peptidomimetics as Specific CXCR4 Imaging Agents

    PubMed Central

    Lesniak, Wojciech G.; Sikorska, Emilia; Shallal, Hassan; Azad, Babak Behnam; Lisok, Ala; Pullambhatla, Mrudula; Pomper, Martin G.; Nimmagadda, Sridhar

    2015-01-01

    The CXCR4 chemokine receptor is integral to several biological functions and plays a pivotal role in the pathophysiology of many diseases. As such, CXCR4 is an enticing target for the development of imaging and therapeutic agents. Here we report the evaluation of the POL3026 peptidomimetic template for the development of imaging agents that target CXCR4. Structural and conformational analyses of POL3026 and two of its conjugates, DOTA (POL-D) and PEG12-DOTA (POL-PD), by circular dichroism, two-dimensional NMR spectroscopy and molecular dynamics calculations are reported. In silico observations were experimentally verified with in vitro affinity assays and rationalized using crystal structure-based molecular modeling studies. [111In]-labeled DOTA conjugates were assessed in vivo for target specificity in CXCR4 expressing subcutaneous U87 tumors (U87-stb-CXCR4) through single photon emission computed tomography (SPECT/CT) imaging and biodistribution studies. In silico and in vitro studies show that POL3026 and its conjugates demonstrate similar interactions with different micelles that mimic cellular membrane and that the ε-NH2 of lysine7 is critical to maintain high affinity to CXCR4. Modification of this group with DOTA or PEG12-DOTA led to the decrease of IC50 value from 0.087 nM for POL3026 to 0.47 nM and 1.42 nM for POL-D and POL-PD, respectively. In spite of the decreased affinity toward CXCR4, [111In]POL-D and [111In]POL-PD demonstrated high and significant uptake in U87-stb-CXCR4 tumors compared to the control U87 tumors at 90 min and 24 h post injection. Uptake in U87-stb-CXCR4 tumors could be blocked by unlabeled POL3026, indicating specificity of the agents in vivo. These results suggest POL3026 as a promising template to develop new imaging agents that target CXCR4. PMID:25590535

  7. Carrier-linked mutual prodrugs of biphenylacetic acid as a promising alternative to bioprecursor fenbufen: design, kinetics, and pharmacological studies.

    PubMed

    Dhaneshwar, Suneela; Kusurkar, Manisha; Bodhankar, Subhash; Bihani, Gopal

    2014-08-01

    Novel mutual prodrugs of biphenylacetic acid were designed as a promising gastro-protective alternative to fenbufen. Biphenyacetic acid was covalently linked with two non-essential amino acids (D-phenylalanine and glycine) possessing wound healing, analgesic, and anti-inflammatory properties. The prodrugs exhibited good stability in stomach homogenates while hydrolytic release of biphenylacetic acid was observed in phosphate buffer, small intestinal homogenates, and 80% human plasma. In vivo behavior of prodrugs on oral administration to Wistar rats demonstrated 33-45% release of biphenylacetic acid in blood over a period of 24 h indicating passage of intact prodrugs to colon, colonic release of parent drug followed by its absorption through colonic mucosa into systemic circulation. Prodrugs were extensively evaluated for analgesic, anti-inflammatory, anti-arthritic, and ulcerogenic activities. Biochemical, haemetological, histopathological, and radiological studies were also performed. Conversion of bioprecusor fenbufen into mutual carrier-linked prodrugs proved to be promising alternative in terms of reduced ulcerogenic propensity, longer duration of analgesia, enhanced/prolonged anti-inflammatory activity, and superior anti-arthritic effect. These prodrugs could be developed further for chronotherapy of rheumatoid arthritis. PMID:24178955

  8. An injectable drug-loaded hydrogel based on a supramolecular polymeric prodrug.

    PubMed

    Xiong, Lu; Luo, Qiaojie; Wang, Ying; Li, Xiaodong; Shen, Zhiquan; Zhu, Weipu

    2015-10-01

    We reported a novel injectable doxorubicin-loaded hydrogel based on host-guest interaction and Schiff's base reaction. A supramolecular polymeric prodrug was prepared through the inclusion of adamantane-modified doxorubicin into the β-cyclodextrin cavity on the polyaldehyde dextran chain, which was in situ crosslinked by carboxymethyl chitosan. PMID:26290273

  9. ROS-activated anticancer prodrugs: a new strategy for tumor-specific damage

    PubMed Central

    Peng, Xiaohua; Gandhi, Varsha

    2013-01-01

    Targeting tumor cells is an important strategy to improve the selectivity of cancer therapies. With the advanced studies in cancer biology, we know that cancer cells are usually under increased oxidative stress. The high level of reactive oxygen species in cancer cells has been exploited for developing novel therapeutic strategies to preferentially kill cancer cells. Our group, amongst others, have used boronic acids/esters as triggers for developing ROS-activated anticancer prodrugs that target cancer cells. The selectivity was achieved by combining a specific reaction between boronates and H2O2 with the efficient masking of drug toxicity in the prodrug via boronates. Prodrugs activated via ferrocene-mediated oxidation have also been developed to improve the selectivity of anticancer drugs. We describe how the strategies of ROS-activation can be used for further development of new ROS-targeting prodrugs, eventually leading to novel approaches and/or combined technology for more efficient and selective treatment of cancers. PMID:22900465

  10. Chemotherapeutic Potential of Diazeniumdiolate-based Aspirin Prodrugs in Breast Cancer

    PubMed Central

    Basudhar, Debashree; Cheng, Robert C.; Bharadwaj, Gaurav; Ridnour, Lisa A.; Wink, David A.; Miranda, Katrina M.

    2015-01-01

    Diazeniumdiolate-based aspirin prodrugs have previously been shown to retain the anti-inflammatory properties of aspirin while protecting against the common side effect of stomach ulceration. Initial analysis of two new prodrugs of aspirin that also release either nitroxyl (HNO) or nitric oxide (NO) demonstrated increased cytotoxicity toward human lung carcinoma cells compared to either aspirin or the parent nitrogen oxide donor. In addition, cytotoxicity was significantly lower in endothelial cells, suggesting cancer-specific sensitivity. To assess the chemotherapeutic potential of these new prodrugs in breast cancer, we studied their effect both in cultured cells and in a nude mouse model. Both prodrugs reduced growth of breast adenocarcinoma cells more effectively than the parent compounds while not being appreciably cytotoxic in a related non-tumorigenic cell line (MCF-10A). The HNO donor also was more cytotoxic than the related NO donor. The basis for the observed specificity was investigated in terms of impact on metabolism, DNA damage and repair, apoptosis, angiogenesis and metastasis. The results suggest a significant pharmacological potential for treatment of breast cancer. PMID:25659932

  11. Cytotoxic, Antiangiogenic and Antitelomerase Activity of Glucosyl- and Acyl- Resveratrol Prodrugs and Resveratrol Sulfate Metabolites.

    PubMed

    Falomir, Eva; Lucas, Ricardo; Peñalver, Pablo; Martí-Centelles, Rosa; Dupont, Alexia; Zafra-Gómez, Alberto; Carda, Miguel; Morales, Juan C

    2016-07-15

    Resveratrol (RES) is a natural polyphenol with relevant and varied biological activity. However, its low bioavailability and rapid metabolism to its glucuronate and sulfate conjugates has opened a debate on the mechanisms underlying its bioactivity. RES prodrugs are being developed to overcome these problems. We have synthesized a series of RES prodrugs and RES sulfate metabolites (RES-S) and evaluated their biological activities. RES glucosylated prodrugs (RES-Glc) were more cytotoxic in HT-29 and MCF-7 cells than RES itself whereas RES-S showed similar or higher cytotoxicity than RES. VEGF production was decreased by RES-Glc, and RES-disulfate (RES-diS) diminished it even more than RES. Finally, RES-Glc and RES-diS inhibited hTERT gene expression to a higher extent than RES. In conclusion, resveratrol prodrugs are promising candidates as anticancer drugs. In addition, RES-S showed distinct biological activity, thus indicating they are not simply RES reservoirs. PMID:27147200

  12. "Project ALERT's" Effects on Adolescents' Prodrug Beliefs: A Replication and Extension Study

    ERIC Educational Resources Information Center

    Clark, Heddy Kovach; Ringwalt, Chris L.; Hanley, Sean; Shamblen, Stephen R.

    2010-01-01

    This article represents a replication and extension of previous studies of the effects of "Project ALERT", a school-based substance use prevention program, on the prodrug beliefs of adolescents. Specifically, the authors' research examined "Project ALERT's" effects on adolescents' intentions to use substances in the future, beliefs about substance…

  13. Nucleotide Analog Prodrug, Tenofovir Disoproxil, Enhances Lymphoid Cell Loading Following Oral Administration in Monkeys

    PubMed Central

    Durand-Gasselin, Lucie; Van Rompay, Koen K.A.; Vela, Jennifer E.; Henne, Ilana N.; Lee, William A.; Rhodes, Gerry R.; Ray, Adrian S.

    2009-01-01

    The antiviral drug tenofovir (TFV) is orally administered as the fumarate salt of its disoproxil prodrug (TFV disoproxil fumarate (TDF)). TFV is a di-anion at physiological pH and, as a result, has poor lipid membrane permeability. Administration of the lipophilic and cell permeable prodrug, TFV disoproxil, enhances the oral absorption of TFV. In order to determine if oral administration of TDF also increases distribution to sites of viral infection, the plasma and circulating lymphoid cell pharmacokinetics of TFV and its phosphorylated metabolites were assessed following a single oral TDF or subcutaneous TFV administration at doses yielding equivalent plasma exposures to TFV in macaques. Despite TFV disoproxil’s lack of plasma stability and undetectable levels in the first plasma samples taken, oral administration of TDF resulted in 7.9-fold higher peripheral blood mononuclear cell exposures to the active metabolite, TFV-diphosphate. The apparent plasma terminal half-life (t1/2) of TFV was also longer following oral TDF relative to subcutaneous TFV administration (median t1/2 of 15.3 and 3.9 h, respectively), suggesting broader distribution to cells and tissues outside of the central plasma compartment. In conclusion, the disoproxil pro-moiety not only enhances the oral absorption of TFV but also tissue and lymphoid cell loading. These results illustrate that administration of even a fleeting prodrug can increase target tissue loading and gives valuable insight for future prodrug development. PMID:19545170

  14. Dual-Sensitive Charge-Conversional Polymeric Prodrug for Efficient Codelivery of Demethylcantharidin and Doxorubicin.

    PubMed

    Wu, Yanjuan; Zhou, Dongfang; Zhang, Qingfei; Xie, Zhigang; Chen, Xuesi; Jing, Xiabin; Huang, Yubin

    2016-08-01

    A tumor is a complicated system, and tumor cells are typically heterogeneous in many aspects. Polymeric drug delivery nanocarriers sensitive to a single type of biosignals may not release cargos effectively in all tumor cells, leading to low therapeutic efficacy. To address the challenges, here, we demonstrated a pH/reduction dual-sensitive charge-conversional polymeric prodrug strategy for efficient codelivery. Reduction-sensitive disulfide group and acid-labile anticancer drug (demethylcantharidin, DMC)-conjugated β-carboxylic amide group were repeatedly and regularly introduced into copolymer chain simultaneously via facile CuAAC click polymerization. The obtained multifunctional polymeric prodrug P(DMC), mPEG-b-poly(disulfide-alt-demethylcantharidin)-b-mPEG was further utilized for DOX encapsulation. Under tumor tissue/cell microenvironments (pH 6.5 and 10 mM GSH), the DOX-loaded polymeric prodrug nanoparticles (P(DMC)@DOX NPs) performed surface negative-to-positive charge conversion and accelerated/sufficient release of DMC and DOX. The remarkably enhanced cellular internalization and cytotoxicity in vitro, especially against DOX-resistant SMMC-7721 cells, were demonstrated. P(DMC)@DOX NPs in vivo also exhibited higher tumor accumulation and improved antitumor efficiency compared to P(SA)@DOX NPs with one drug and without charge-conversion ability. The desired multifunctional polymeric prodrug strategy brings a new opportunity for cancer chemotherapy. PMID:27384255

  15. Spiral assembly of amphiphilic cytarabine prodrug assisted by probe sonication: Enhanced therapy index for leukemia.

    PubMed

    Liu, Jing; Ma, Naxin; Zhao, Dujuan; Li, Zhonghao; Luan, Yuxia

    2015-12-01

    In order to overcome the drawbacks of cytarabine (Ara-C), such as low lipophilicity as well as short plasma half-life and rapid inactivation, a new derivative of Ara-C was designed by incorporation into the non-toxic material, oleic acid (OA), obtaining an amphiphilic small molecular weight prodrug (OA-Ara). By a simple amidation reaction, OA-Ara was synthesized successfully with a yield up to 61.32%. It was for the first time to see that the novel prodrug molecules could assemble into the unexpectedly spiral assembly under probe ultrasonication in aqueous solution. The oil/water partition coefficient (Ko/w) and the permeability of cell membrane of the prodrug were significantly increased compared with Ara-C molecules. In addition, OA-Ara molecules were stable in various pH solutions and artificial digestives, which indicated that it could be administrated orally. Cell viability assay showed that the prodrug displayed much higher antiproliferative effect against K562 and HL60 cells due to its improvement of the lipophilicity and penetrability of cell membrane. These findings demonstrate the feasibility of utilizing structural modification to broaden the clinic application of Ara-C and thus provide an effective new therapeutic alternative for leukemia. PMID:26551869

  16. Naloxone pro-drug rescues morphine induced respiratory depression in Sprague-Dawley rats.

    PubMed

    Wallisch, Michael; El Rody, Nehad M; Huang, Baohua; Koop, Dennis R; Baker, James R; Olsen, George D

    2012-01-15

    Respiratory depression is the main obstacle for the safe administration of morphine for acute pain after injury. Due to this complication, new delivery methods are needed to insure that safe and effective doses of opioid analgesics are administered during emergencies. A depot formulation containing a naloxone pro-drug was designed to release the antidote when morphine causes dangerous hypoxic conditions in the blood. The aim of this work was to test the naloxone release in vivo in response to a severe overdose of morphine in the Sprague-Dawley rat model. Non-invasive two-chamber plethysmography was used to monitor and record respiration and to test the capability of the naloxone pro-drug to respond to and rescue morphine-induced respiratory depression in the animal. We show that the pro-drug formulation can both prevent and reverse severe morphine induced respiratory depression. The animal model demonstrates that co-administration of the naloxone pro-drug reliably antagonizes profound respiratory depressive effects of morphine. PMID:22027217

  17. Evolution of an Amino Acid-based Prodrug Approach: Stay Tuned

    PubMed Central

    Krylov, Ivan S.; Kashemirov, Boris A.; Hilfinger, John M.; McKenna, Charles E.

    2013-01-01

    Certain acyclic nucleoside phosphonates (ANPs) such as (S)-HPMPC (cidofovir, Vistide®) and (S)-HPMPA have been shown to be active against a broad spectrum of DNA and retroviruses. However, their poor absorption as well as their toxicity limit the utilization of these therapeutics in the clinic. Nucleoside phosphonates are poorly absorbed primarily due to the presence of the phosphonic acid group, which ionizes at physiological pH. When dosed intravenously they display dose-limiting nephrotoxicity due to their accumulation in the kidney. To overcome these limitations, nucleoside phosphonate prodrug strategies have taken center stage in the development pathway and a number of different approaches are at various stages of development. Our efforts have focused on the development of ANP prodrugs in which a benign amino acid promoiety masks a phosphonate P-OH via a hydroxyl side chain. The design of these prodrugs incorporates multiple chemical groups (the P−X−C linkage, the amino acid stereochemistry, the C-terminal and N-terminal functional groups) that can be been tuned to modify absorption, pharmacokinetic and efficacy properties with the goal of improving overall prodrug performance. PMID:23339402

  18. Combined strategies of apomorphine diester prodrugs and nanostructured lipid carriers for efficient brain targeting

    NASA Astrophysics Data System (ADS)

    Liu, Kuo-Sheng; Wen, Chih-Jen; Yen, Tzu-Chen; Sung, K. C.; Ku, Ming-Chuan; Wang, Jhi-Joung; Fang, Jia-You

    2012-03-01

    Our aim is to develop nanostructured lipid carriers (NLCs) for loading the apomorphine diester prodrugs, diacetyl apomorphine (DAA) and diisobutyryl apomorphine (DIA), into the brain. NLCs were prepared using sesame oil/cetyl palmitate as the lipid matrices. Experiments were performed with the objective of evaluating the physicochemical characteristics, drug release, safety and brain-targeting efficacy of the NLCs. The size of regular NLCs (N-NLCs) was 214 nm. The addition of Forestall (FE) and polyethylene glycol (PEG) to the NLCs (P-NLCs) increased the particle diameter to 250 nm. The zeta potentials of N-NLCs and P-NLCs were respectively shown to be - 21 and 48 mV. Diester prodrugs were more lipophilic and more chemically stable than the parent apomorphine. The hydrolysis study indicated that the prodrugs underwent bioconversion in plasma and brain extract, with DAA exhibiting faster degradation than DIA. Sustained release was achieved through the synergistic effect of integrating strategies of prodrugs and NLCs, with the longer carbon chain showing the slower release (DIA < DAA). None of the NLCs tested here exhibited a toxicity problem according to the examination of neutrophil lactate dehydrogenase (LDH) release and hemolysis. Results of a bioimaging study in mice showed that P-NLCs largely accumulated in the brain. The distribution duration of the fluorescent dye in the brain region was also prolonged by the nanocarriers.

  19. Synthesis, Screening and Pharmacokinetic Evaluation of Potential Prodrugs of Bupropion. Part One: In Vitro Development

    PubMed Central

    O’Byrne, Paul Matthew; Williams, Robert; Walsh, John J.; Gilmer, John F.

    2014-01-01

    In general, prodrugs are developed to circumvent deficiencies associated with the absorption, distribution, metabolism, excretion or toxicological (ADMET) profile associated with the active drug. In our study, we select bupropion, a drug with broad pharmacology incorporating dopaminergic, noradrenergic, nicotinic and cytokine modulation properties, but which is rapidly metabolized in vivo. We exploited its carbonyl and secondary amine functionality to facilitate the synthesis of bioprecursor prodrug forms with the sole objective of identifying analogues with enhanced properties over bupropion. A range of analogues were synthesized, ranging from N-methyl, N-benzyl, oximes, enol acetate and ether forms to examples where both functional groups were utilized to form oxadiazine, oxadiazinone, oxazolone and acetylated derivatives. We then developed an in vitro metabolic screen to simulate the human oral delivery route for these analogues. The selection of media in the screens contained a variety of pH, enzymatic and co-factor systems which mimic metabolic in vivo environments that drugs encounter when delivered orally. By coupling our in vitro screening tool to a selective hyphenated technique such as LC-MS, we were able to quickly select potential prodrugs for further in vitro and in vivo development. From the data generated, the N-alkylated bupropion analogues were shown to have the highest potential to act as bioprecursor prodrugs of bupropion. PMID:24830986

  20. Dual pH-responsive 5-aminolevulinic acid pseudopolyrotaxane prodrug micelles for enhanced photodynamic therapy.

    PubMed

    Tong, Hongxin; Wang, Yin; Li, Huan; Jin, Qiao; Ji, Jian

    2016-03-11

    Novel 5-aminolevulinic acid (ALA) pseudopolyrotaxane prodrug micelles with dual pH-responsive properties were prepared by the host-guest interaction of α-cyclodextrin (α-CD) and poly(ethylene glycol) (PEG). The micelles exhibited pH dependent cellular uptake and pH-sensitive ALA release, enabling enhanced photodynamic therapy. PMID:26882232

  1. Targeting and Treatment of Tumor Hypoxia by Newly Designed Prodrug Possessing High Permeability in Solid Tumors.

    PubMed

    Ikeda, Yutaka; Hisano, Hikaru; Nishikawa, Yuji; Nagasaki, Yukio

    2016-07-01

    Tumor hypoxia, which is associated with poor prognosis in cancer, is known to lead to resistance to radiotherapy and anticancer chemotherapy. Impaired drug penetration in hypoxic regions has been recognized as an essential barrier to drug development in solid tumors. Here, we propose novel hypoxia-activated prodrugs, which drastically improved the penetration property of commonly used anticancer drugs in the hypoxic region. In this design, conventional anticancer drugs were modified with 2-nitroimidazole derivatives. The most important point of this study was that the prodrug designed formed a 6-membered cyclic structure to allow liberation of the active drug in the hypoxic region. This design markedly increased the selectivity of the hypoxia-targeted prodrug, resulting in significant reduction of adverse effects in the normoxic region. In vitro studies confirmed the selective activation under hypoxic conditions. In vivo studies showed drastic reduction of adverse effects associated with conventional anticancer drugs and improvement of the survival rate of mice. Immunofluorescence analyses confirmed that the designed prodrug had a tendency to localize at the hypoxic region, in contrast to conventional anticancer drugs, which localize only at the normoxic region. PMID:27187083

  2. Preparation, characterization, cytotoxicity and pharmacokinetics of liposomes containing water-soluble prodrugs of paclitaxel.

    PubMed

    Ceruti, M; Crosasso, P; Brusa, P; Arpicco, S; Dosio, F; Cattel, L

    2000-01-01

    Paclitaxel (Taxol) is a diterpenoid isolated from Taxus brevifolia, used clinically for the treatment of ovarian and breast cancer. Due to its aqueous insolubility it is administered dissolved in ethanol and Cremophor EL (polyethoxylated castor oil), which has serious side effects. In order to eliminate this vehicle, in previous work we entrapped paclitaxel in conventional and in polyethylene glycol coated liposomes. However, in neither formulation did we obtain satisfactory entrapment efficiency. In this study we increased the paclitaxel concentration entrapped in liposomes by incorporating different water-soluble prodrugs, such as the 2'-succinyl, 2'-methylpyridinium acetate and 2'-mPEG ester paclitaxel derivatives, in the lipid vesicles. Liposomes containing 2'-mPEG (5000)-paclitaxel showed the best performance in terms of stability, entrapment efficiency and drug concentration (6.5 mgml(-1)). The in vitro cytotoxic activity of this liposomal prodrug was similar to that of the parent drug. The pharmacokinetic parameters for the free and for the liposomal prodrugs fitted a bi-exponential plasma disposition. The most important change in pharmacokinetic values of the prodrug vs. the free drug liposomal formulations was t(1/2)beta, plasma lifetime, which was longer in liposomes containing 2'-mPEG (5000)-paclitaxel. PMID:10640588

  3. Ester prodrugs of acyclic nucleoside thiophosphonates compared to phosphonates: synthesis, antiviral activity and decomposition study.

    PubMed

    Roux, Loïc; Priet, Stéphane; Payrot, Nadine; Weck, Clément; Fournier, Maëlenn; Zoulim, Fabien; Balzarini, Jan; Canard, Bruno; Alvarez, Karine

    2013-05-01

    9-[2-(Thiophosphonomethoxy)ethyl]adenine [S-PMEA, 8] and (R)-9-[2-(Thiophosphonomethoxy)propyl]adenine [S-PMPA, 9] are acyclic nucleoside thiophosphonates we described recently that display the same antiviral spectrum (DNA viruses) as approved and potent phosphonates PMEA and (R)-PMPA. Here, we describe the synthesis, antiviral activities in infected cell cultures and decomposition study of bis(pivaloyloxymethoxy)-S-PMEA [Bis-POM-S-PMEA, 13] and bis(isopropyloxymethylcarbonyl)-S-PMPA [Bis-POC-S-PMPA, 14] as orally bioavailable prodrugs of the S-PMEA 8 and S-PMPA 9, in comparison to the equivalent "non-thio" derivatives [Bis-POM-PMEA, 11] and [Bis-POC-PMPA, 12]. Compounds 11, 12, 13 and 14 were evaluated for their in vitro antiviral activity against HIV-1-, HIV-2-, HBV- and a broad panel of DNA viruses, and found to exhibit moderate to potent antiviral activity. In order to determine the decomposition pathway of the prodrugs 11, 12, 13 and 14 into parent compounds PMEA, PMPA, 8 and 9, kinetic data and decomposition pathways in several media are presented. As expected, bis-POM-S-PMEA 13 and bis-POC-S-PMPA 14 behaved as prodrugs of S-PMEA 8 and S-PMPA 9. However, thiophosphonates 8 and 9 were released very smoothly in cell extracts, in contrast to the release of PMEA and PMPA from "non-thio" prodrugs 11 and 12. PMID:23603046

  4. Nasal chitosan microparticles target a zidovudine prodrug to brain HIV sanctuaries.

    PubMed

    Dalpiaz, Alessandro; Fogagnolo, Marco; Ferraro, Luca; Capuzzo, Antonio; Pavan, Barbara; Rassu, Giovanna; Salis, Andrea; Giunchedi, Paolo; Gavini, Elisabetta

    2015-11-01

    Zidovudine (AZT) is an antiretroviral drug that is a substrate of active efflux transporters (AETs) that extrude the drug from the central nervous system (CNS) and macrophages, which are considered to be sanctuaries of HIV. The conjugation of AZT to ursodeoxycholic acid is known to produce a prodrug (UDCA-AZT) that is able to elude the AET systems, indicating the potential ability of this prodrug to act as a carrier of AZT in the CNS and in macrophages. Here, we demonstrate that UDCA-AZT is able to permeate and remain in murine macrophages with an efficiency twenty times higher than that of AZT. Moreover, we propose the nasal administration of this prodrug in order to induce its uptake into the CNS. Chitosan chloride-based microparticles (CP) were prepared by spray-drying and were characterized with respect to size, morphology, density, water uptake and the dissolution profile of UDCA-AZT. The CP sample was then nasally administered to rats. All in vitro and in vivo measurements were also performed for a CP parent physical mixture. The CP sample was able to increase the dissolution rate of UDCA-AZT and to reduce water uptake with respect to its parent physical mixture, inducing better uptake of UDCA-AZT into the cerebrospinal fluid of rats, where the prodrug can act as an AZT carrier in macrophages. PMID:26427553

  5. NMR analysis of interactions of a phosphatidylinositol 3'-kinase SH2 domain with phosphotyrosine peptides reveals interdependence of major binding sites.

    PubMed

    Günther, U L; Liu, Y; Sanford, D; Bachovchin, W W; Schaffhausen, B

    1996-12-01

    The interactions of the N-terminal src homology (SH2) domain (N-SH2) of the 85 kDa subunit of phosphatidylinositol 3'-kinase (PI-3K) with phosphotyrosine (ptyr) and a series of ptyr-containing peptides have been examined by NMR spectroscopy. HSQC (heteronuclear single-quantum coherence) NMR spectra of 15N-labeled SH2 were used to evaluate its interactions with ptyr-containing ligands. The ability of ligands to cause chemical shift changes was compared to their potency as competitors in in vitro binding experiments using polyoma virus middle T antigen (MT). The results suggest the interdependence of SH2 binding elements. Chemical shifts of residues involved in the ptyr binding were altered by variations of the sequence of the bound peptide, suggesting that the ptyr fit can be adjusted by the peptide sequence. Perturbations of chemical shifts of residues coordinating the methionine three residues C-terminal to the ptyr (the +3 residue) were affected by substitution in the binding peptide at +1 and vice versa. Such results show synergistic interplay between regions of the SH2 binding residues C-terminal to the ptyr. PMID:8952511

  6. Identification of Phosphotyrosine Binding Domain-Containing Proteins as Novel Downstream Targets of the EphA8 Signaling Function▿ †

    PubMed Central

    Shin, Jongdae; Gu, Changkyu; Park, Eunjeong; Park, Soochul

    2007-01-01

    Eph receptors and ephrins have been implicated in a variety of cellular processes, including morphology and motility, because of their ability to modulate intricate signaling networks. Here we show that the phosphotyrosine binding (PTB) domain-containing proteins AIDA-1b and Odin are tightly associated with the EphA8 receptor in response to ligand stimulation. Both AIDA-1b and Odin belong to the ankyrin repeat and sterile alpha motif domain-containing (Anks) protein family. The PTB domain of Anks family proteins is crucial for their association with the juxtamembrane domain of EphA8, whereas EphA8 tyrosine kinase activity is not required for this protein-protein interaction. In addition, we found that Odin is a more physiologically relevant partner of EphA8 in mammalian cells. Interestingly, overexpression of the Odin PTB domain alone attenuated EphA8-mediated inhibition of cell migration in HEK293 cells, suggesting that it acts as a dominant-negative mutant of the endogenous Odin protein. More importantly, small interfering RNA-mediated Odin silencing significantly diminished ephrinA5-induced EphA8 signaling effects, which inhibit cell migration in HEK293 cells and retract growing neurites of Neuro2a cells. Taken together, our findings support a possible function for Anks family proteins as scaffolding proteins of the EphA8 signaling pathway. PMID:17875921

  7. Chemical Reactivity Window Determines Prodrug Efficiency toward Glutathione Transferase Overexpressing Cancer Cells.

    PubMed

    van Gisbergen, Marike W; Cebula, Marcus; Zhang, Jie; Ottosson-Wadlund, Astrid; Dubois, Ludwig; Lambin, Philippe; Tew, Kenneth D; Townsend, Danyelle M; Haenen, Guido R M M; Drittij-Reijnders, Marie-José; Saneyoshi, Hisao; Araki, Mika; Shishido, Yuko; Ito, Yoshihiro; Arnér, Elias S J; Abe, Hiroshi; Morgenstern, Ralf; Johansson, Katarina

    2016-06-01

    Glutathione transferases (GSTs) are often overexpressed in tumors and frequently correlated to bad prognosis and resistance against a number of different anticancer drugs. To selectively target these cells and to overcome this resistance we previously have developed prodrugs that are derivatives of existing anticancer drugs (e.g., doxorubicin) incorporating a sulfonamide moiety. When cleaved by GSTs, the prodrug releases the cytostatic moiety predominantly in GST overexpressing cells, thus sparing normal cells with moderate enzyme levels. By modifying the sulfonamide it is possible to control the rate of drug release and specifically target different GSTs. Here we show that the newly synthesized compounds, 4-acetyl-2-nitro-benzenesulfonyl etoposide (ANS-etoposide) and 4-acetyl-2-nitro-benzenesulfonyl doxorubicin (ANS-DOX), function as prodrugs for GSTA1 and MGST1 overexpressing cell lines. ANS-DOX, in particular, showed a desirable cytotoxic profile by inducing toxicity and DNA damage in a GST-dependent manner compared to control cells. Its moderate conversion of 500 nmol/min/mg, as catalyzed by GSTA1, seems hereby essential since the more reactive 2,4-dinitrobenzenesulfonyl doxorubicin (DNS-DOX) (14000 nmol/min/mg) did not display a preference for GSTA1 overexpressing cells. DNS-DOX, however, effectively killed GSTP1 (20 nmol/min/mg) and MGST1 (450 nmol/min/mg) overexpressing cells as did the less reactive 4-mononitrobenzenesulfonyl doxorubicin (MNS-DOX) in a MGST1-dependent manner (1.5 nmol/min/mg) as shown previously. Furthermore, we show that the mechanism of these prodrugs involves a reduction in GSH levels as well as inhibition of the redox regulatory enzyme thioredoxin reductase 1 (TrxR1) by virtue of their electrophilic sulfonamide moiety. TrxR1 is upregulated in many tumors and associated with resistance to chemotherapy and poor patient prognosis. Additionally, the prodrugs potentially acted as a general shuttle system for DOX, by overcoming resistance

  8. Zosuquidar and an albumin-binding prodrug of zosuquidar reverse multidrug resistance in breast cancer cells of doxorubicin and an albumin-binding prodrug of doxorubicin.

    PubMed

    Abu Ajaj, Khalid; Graeser, Ralph; Kratz, Felix

    2012-07-01

    The P-glycoprotein (P-gp) is a 170-kDa protein that acts as an energy dependent, transmembrane efflux pump and is encoded by the MDR1 gene. It has been shown to be responsible for multidrug resistance (MDR) in a defined subpopulation of breast cancer patients and thus represents a molecular target for circumventing MDR in this tumor indication. MDR modulators have been developed and demonstrated high selectivity for P-gp with inhibitory activities in the low nanomolar range. Although some objective responses were achieved in clinical trials, combination therapy with these MDR modulators, such as Ca2+ antagonists caused unacceptable toxicity. Targeting P-gp inhibitors to the tumor site is a mean to increase their therapeutic index, and in this context binding of tailor-made prodrugs to circulating albumin is an established technology to reduce the toxicity and enhance the efficacy of anticancer drugs. In this study, we consequently developed an acid-sensitive albumin-binding prodrug of the P-gp inhibitor zosuquidar (LY335979) in a two-step synthesis using a maleimide hydrazone linker system established in our laboratory that first introduces acetylbenzoic acid at the HO-group of zosuquidar followed by derivatization with 6-maleimidocaproyl hydrazide to form the acid-sensitive hydrazone bond. The maleimide group enables the prodrug to bind rapidly and selectively to the cysteine-34 position of endogenous albumin after intravenous administration. HPLC analysis demonstrated rapid albumin binding of the zosuquidar prodrug as well as the quantitative release of the acetylbenzoic ester derivative of zosuquidar at pH 5.0. Subsequently, its ability to circumvent MDR was tested in two doxorubicin-resistant breast carcinoma cell lines (MCF-7/ADR and MT-3/ADR). The MDR status of these cell lines can be reversed by zosuquidar which was confirmed in a rhodamine 123 assay using fluorescence microscopy and FACS analysis. Furthermore, zosuquidar as well its acid-sensitive albumin

  9. Synthesis and in vitro stability of amino acid prodrugs of 6-β-naltrexol for microneedle-enhanced transdermal delivery.

    PubMed

    Eldridge, Joshua A; Milewski, Mikolaj; Stinchcomb, Audra L; Crooks, Peter A

    2014-11-15

    A small library of amino acid ester prodrugs of 6-β-naltrexol (NTXOL, 1) was prepared in order to investigate the candidacy of these prodrugs for microneedle-enhanced transdermal delivery. Six amino acid ester prodrugs were synthesized (6a-f). 6b, 6d, and 6 e were stable enough at skin pH (pH 5.0) to move forward to studies in 50% human plasma. The lead compound (6 e) exhibited the most rapid bioconversion to NTXOL in human plasma (t1/2 = 2.2 ± 0.1h). PMID:25442314

  10. Synthesis and in vitro stability of amino acid prodrugs of 6-β-naltrexol for microneedle-enhanced transdermal delivery

    PubMed Central

    Eldridge, Joshua A.; Milewski, Mikolaj; Stinchcomb, Audra L.; Crooks, Peter A.

    2014-01-01

    A small library of amino acid ester prodrugs of 6-β-naltrexol (NTXOL, 1) was prepared in order to investigate the candidacy of these prodrugs for microneedle-enhanced transdermal delivery. Six amino acid ester prodrugs were synthesized (6a-f). 6b, 6d, and 6e were stable enough at skin pH (pH 5.0) to move forward to studies in 50% human plasma. The lead compound (6e) exhibited the most rapid bioconversion to NTXOL in human plasma (t½ = 2.2 ± 0.1 h). PMID:25442314

  11. Design of Optimized Hypoxia-Activated Prodrugs Using Pharmacokinetic/Pharmacodynamic Modeling

    PubMed Central

    Foehrenbacher, Annika; Secomb, Timothy W.; Wilson, William R.; Hicks, Kevin O.

    2013-01-01

    Hypoxia contributes to resistance of tumors to some cytotoxic drugs and to radiotherapy, but can in principle be exploited with hypoxia-activated prodrugs (HAP). HAP in clinical development fall into two broad groups. Class I HAP (like the benzotriazine N-oxides tirapazamine and SN30000), are activated under relatively mild hypoxia. In contrast, Class II HAP (such as the nitro compounds PR-104A or TH-302) are maximally activated only under extreme hypoxia, but their active metabolites (effectors) diffuse to cells at intermediate O2 and thus also eliminate moderately hypoxic cells. Here, we use a spatially resolved pharmacokinetic/pharmacodynamic (SR-PK/PD) model to compare these two strategies and to identify the features required in an optimal Class II HAP. The model uses a Green’s function approach to calculate spatial and longitudinal gradients of O2, prodrug, and effector concentrations, and resulting killing in a digitized 3D tumor microregion to estimate activity as monotherapy and in combination with radiotherapy. An analogous model for a normal tissue with mild hypoxia and short intervessel distances (based on a cremaster muscle microvessel network) was used to estimate tumor selectivity of cell killing. This showed that Class II HAP offer advantages over Class I including higher tumor selectivity and greater freedom to vary prodrug diffusibility and rate of metabolic activation. The model suggests that the largest gains in class II HAP antitumor activity could be realized by optimizing effector stability and prodrug activation rates. We also use the model to show that diffusion of effector into blood vessels is unlikely to materially increase systemic exposure for realistic tumor burdens and effector clearances. However, we show that the tumor selectivity achievable by hypoxia-dependent prodrug activation alone is limited if dose-limiting normal tissues are even mildly hypoxic. PMID:24409417

  12. The self-assembling camptothecin-tocopherol prodrug: An effective approach for formulating camptothecin.

    PubMed

    Lu, Jianqin; Liu, Chuang; Wang, Pengcheng; Ghazwani, Mohammed; Xu, Jieni; Huang, Yixian; Ma, Xiaochao; Zhang, Peijun; Li, Song

    2015-09-01

    Camptothecin (CPT) is a potent antitumor agent and functions via inhibiting the activity of topoisomerase I during DNA replication. However, the clinical application of CPT has been greatly hindered by its extremely poor solubility, the instability of its active lactone ring in blood stream, as well as the non-specific toxicity to normal tissues. In addition, most of the formulations developed so far are not applicable for formulating CPT. In this study, two novel CPT prodrugs were developed by conjugating CPT to α-tocopherol via a carbonate ester bond (CPT-VE) or disulfide linkage (CPT-S-S-VE). Both CPT prodrugs were able to self-assemble into nanofibers with the facilitation of a PEG5K-Fmoc-VE2-based nanomicellar carrier. Both prodrug nanoassemblies exhibited excellent stability. Fluorescence quenching, UV absorbance, and FT-IR studies demonstrated strong interactions between carrier and prodrugs, including hydrophobic interaction, π-π stacking, as well as hydrogen bonding. NMR studies suggested that prodrugs were successfully incorporated into PEG5K-Fmoc-VE2 during self-assembly process. In vitro, PEG5K-Fmoc-VE2/CPT-S-S-VE presented significantly higher level of cytotoxicity on tumor cells compared to PEG5K-Fmoc-VE2/CPT-VE. Biodistribution study showed that CPT-S-S-VE formulated in PEG5K-Fmoc-VE2 micelles was effectively converted to parent CPT following delivery to tumor tissues. Finally, PEG5K-Fmoc-VE2/CPT-S-S-VE nanofibers showed superior tumor growth inhibition in an aggressive murine breast cancer model (4T1.2). PMID:26057133

  13. Comparative efficacies of 2 cysteine prodrugs and a glutathione delivery agent in a colitis model.

    PubMed

    Oz, Helieh S; Chen, Theresa S; Nagasawa, Herbert

    2007-08-01

    Oxidant-mediated injury plays an important role in the pathophysiology of inflammatory bowel disease (IBD). Recently, antioxidants were shown to modulate colitis in mice. In this study, the protective effects of L-cysteine and glutathione (GSH) prodrugs are further evaluated against progression of colitis in a murine model. ICR mice were fed compounds incorporated into chow as follows: Group (A) received chow supplemented with vehicle. Group (B) was provided 2-(RS)-n-propylthiazolidine-4(R)-carboxylic-acid (PTCA), a cysteine prodrug. Group (C) received D-ribose-L-cysteine (RibCys), another cysteine prodrug that releases L-cysteine. Group (D) was fed L-cysteine-glutathione mixed sulfide (CySSG), a ubiquitous GSH derivative present in mammalian cells. After 3 days, the animals were further provided with normal drinking water or water supplemented with dextran sodium sulfate (DSS). Mice administered DSS developed severe colitis and suffered weight loss. Colonic lesions significantly improved in animals treated with PTCA and RibCys and, to a lesser extent, with CySSG therapy. Hepatic GSH levels were depleted in colitis animals (control vs DSS, P < 0.001), and normalized with prodrug therapies (control vs treatments, P > 0.05). Protein expressions of serum amyloid A and inflammatory cytokines [interleukin (IL)-6, IL-12, tumor necrosis factor-alpha (TNF-alpha), osteopontin (OPN)] were significantly increased in colitis animals and improved with therapies. Immunohistochemistry and Western blot analyses showed significant upregulation of the macrophage-specific markers, COX-2 and CD68, which suggests macrophage activation and infiltration in the colonic lamina propria in colitis animals. These abnormalities were attenuated in prodrug-treated mice. In conclusion, these data strongly support the novel action of the PTCA against colitis, which further supports a possible therapeutic application for IBD patients. PMID:17656332

  14. Prodrugs of fumarate esters for the treatment of psoriasis and multiple sclerosis--a computational approach.

    PubMed

    Karaman, Rafik; Dokmak, Ghadeer; Bader, Maryam; Hallak, Hussein; Khamis, Mustafa; Scrano, Laura; Bufo, Sabino Aurelio

    2013-01-01

    Density functional theory (DFT) calculations at B3LYP/6-31 G (d,p) and B3LYP/6-311+G(d,p) levels for the substituted pyridine-catalyzed isomerization of monomethyl maleate revealed that isomerization proceeds via four steps, with the rate-limiting step being proton transfer from the substituted pyridinium ion to the C=C double bond in INT1. In addition, it was found that the isomerization rate (maleate to fumarate) is solvent dependent. Polar solvents, such as water, tend to accelerate the isomerization rate, whereas apolar solvents, such as chloroform, act to slow down the reaction. A linear correlation was obtained between the isomerization activation energy and the dielectric constant of the solvent. Furthermore, linearity was achieved when the activation energy was plotted against the pKa value of the catalyst. Substituted-pyridine derivatives with high pKa values were able to catalyze isomerization more efficiently than those with low pKa values. The calculated relative rates for prodrugs 1-6 were: 1 (406.7), 2 (7.6×10(6)), 3 (1.0), 4 (20.7), 5 (13.5) and 6 (2.2×10(3)). This result indicates that isomerizations of prodrugs 1 and 3-5 are expected to be slow and that of prodrugs 2 and 6 are expected to be relatively fast. Hence, prodrugs 2 and 3-5 have the potential to be utilized as prodrugs for the slow release of monomethylfumarate in the treatment of psoriasis and multiple sclerosis. PMID:22940872

  15. Comparative plasma disposition kinetics of albendazole and its new benzimidazol prodrug in dog.

    PubMed

    Khalil, Z; El Karbane, M; Faouzi, M E A; Ansar, M; Azougagh, M; El Harti, J; Taoufik, J

    2016-01-01

    The comparative pharmacokinetic behavior of albendazole (ABZ) and its new benzimidazol prodrug [1-tert-butyloxycarbonyl-5-propylthio-1-H-benzimidazol-2ylcarbamate of methyl] (ABZBoc), following their oral administration (10mg/kg) to healthy dogs was explored. Blood samples were obtained serially over a 24h period after treatment, then the plasma was analyzed by high-performance liquid chromatography (HPLC) to search the albendazole metabolites (ABZSO and ABZSO2). However, the albendazole parent drug was not detectable at any time after both treatments (ABZ and ABZBoc). By albendazole metabolites (ABZSO and ABZSO2) were the analytes recovered in the plasma after oral administration of ABZ and ABZBoc. Furthermore, some amounts of ABZBoc were also available in the plasma samples treated with this new produg. The plasma profile of each analyte followed a similar pattern after both treatments, the active metabolite (ABZSO) was the major analyte recovered in plasma (between 1 and 24h post-treatment). The pharmacokinetic parameters of both groups were calculated (Cmax, Tmax, t1/2, AUC0-›∞), and analyzed using the Student's t-test, P<0.05. Thus,the pharmacokinetic analysis indicated four statistically significant changes in the pharmacokinetic parameters defined above of the albendazole metabolites (ABZSO, ABZSO2) between the group treated with albendazole (group A) and that treated with ABZBoc prodrug (group B). Hence, the levels of the various pharmacokinetics parameters were low in the group treated with prodrug, as well they did not reach equivalent concentrations to that of albendazole. These differences between albendazole and its new prodrug may be explained by the fact that ABZBoc prodrug was not effectively reduced in the intestine of dogs. PMID:26530448

  16. Phospholipase A(2)-susceptible liposomes of anticancer double lipid-prodrugs.

    PubMed

    Arouri, Ahmad; Mouritsen, Ole G

    2012-03-12

    A novel approach to anticancer drug delivery is presented based on lipid-like liposome-forming anticancer prodrugs that are susceptible to secretory phospholipase A(2) (sPLA(2)) that is overexpressed in several cancer types. The approach provides a selective unloading of anticancer drugs at the target tissues, as well as circumvents the necessity for "conventional" drug loading. In our attempts to improve the performance of the liposomes in vivo, several PEGylated and non-PEGylated liposomal formulations composed of a retinoid prodrug premixed with the sPLA(2)-hydrolyzable DPPC (1,2-dipalmitoyl-sn-glycero-3-phosphocholine) were prepared. Besides favorably modifying the physicochemical properties of the liposomes, the incorporation of DPPC and PEG-lipids in the liposomes should substantially enhance the enzymatic activity, as concluded from literature. In addition, one can reap benefits from the presumed permeability enhancing effect of the liberated fatty acids and lysolipids. The size distribution of the prepared liposomes as well as their phase behavior, enzymatic hydrolysis, and cytotoxicity, in the presence and absence of sPLA(2), were determined. The liposomes were around 100nm in diameter and in the gel/fluid coexistence region at 37°C. The enzymatic hydrolysis of the prodrug was pronouncedly accelerated upon the premixing with DPPC, and the hydrolysis was further enhanced by PEGylation. Interestingly, the faster hydrolysis of the prodrug and the released fatty acids and lysolipids from DPPC did not improve the cytotoxicity of the mixture; the effect of combining the prodrug with DPPC was additive and not synergistic. The data presented here question the significance of the permeability enhancing effects claimed for fatty acids and lysolipids at the target cell membrane, and whether these effects can be achieved using physiologically achievable concentrations of fatty acids and lysolipids. PMID:21946258

  17. Folate Receptor-Mediated Enhanced and Specific Delivery of Far-Red Light-Activatable Prodrugs of Combretastatin A-4 to FR-Positive Tumor

    PubMed Central

    2015-01-01

    We examined the concept of a novel prodrug strategy in which anticancer drug can be locally released by visible/near IR light, taking advantage of the photodynamic process and photo-unclick chemistry. Our most recently formulated prodrug of combretastatin A-4, Pc-(L-CA4)2, showed multifunctionality for fluorescence imaging, light-activated drug release, and the combined effects of PDT and local chemotherapy. In this formulation, L is a singlet oxygen cleavable linker. Here, we advanced this multifunctional prodrug by adding a tumor-targeting group, folic acid (FA). We designed and prepared four FA-conjugated prodrugs 1–4 (CA4-L-Pc-PEGn-FA: n = 0, 2, 18, ∼45) and one non-FA-conjugated prodrug 5 (CA4-L-Pc-PEG18-boc). Prodrugs 3 and 4 had a longer PEG spacer and showed higher hydrophilicity, enhanced uptake to colon 26 cells via FR-mediated mechanisms, and more specific localization to SC colon 26 tumors in Balb/c mice than prodrugs 1 and 2. Prodrug 4 also showed higher and more specific uptake to tumors, resulting in selective tumor damage and more effective antitumor efficacy than non-FA-conjugated prodrug 5. FR-mediated targeting seemed to be an effective strategy to spare normal tissues surrounding tumors in the illuminated area during treatment with this prodrug. PMID:25351441

  18. Synthesis and Biological Evaluation of Phosphate Prodrugs of 4-Phospho-d -erythronohydroxamic Acid, an Inhibitor of 6-Phosphogluconate Dehydrogenase

    PubMed Central

    Ruda, Gian Filippo; Alibu, Vincent P; Mitsos, Christos; Bidet, Olivier; Kaiser, Marcel; Brun, Reto; Barrett, Michael P; Gilbert, Ian H

    2007-01-01

    We have previously reported the discovery of potent and selective inhibitors of 6-phosphogluconate dehydrogenase, the third enzyme of the phosphate pentose pathway, from Trypanosoma brucei, the causative organism of human African trypanosomiasis. These inhibitors were charged phosphate derivatives with restricted capacity to enter cells. Herein, we report the synthesis of five different classes of prodrugs: phosphoramidate; bis-S-acyl thioethyl esters (bis-SATE); bis-pivaloxymethyl (bis-POM); CycloSaligenyl; and phenyl, S-acyl thioethyl mixed phosphate esters (mix-SATE). Prodrugs were studied for stability and activity against the intact parasites. Most prodrugs caused inhibition of the growth of the parasites. The activity of the prodrugs against the parasites appeared to be related to their stability in aqueous buffer. PMID:17615587

  19. Design and synthesis of imidazole N-H substituted amide prodrugs as inhibitors of hepatitis C virus replication.

    PubMed

    Zong, Xi; Cai, Jin; Chen, Junqing; Wang, Peng; Zhou, Gaoxin; Chen, Bo; Li, Wei; Ji, Min

    2015-08-15

    Twenty-five novel imidazole N-H substituted Daclatasvir (BMS-790052, DCV) analogues (8a-8y) were designed and synthesized as potential prodrugs. Structure modifications were performed in order to improve potency and pharmacokinetic (PK) properties. All target compounds were evaluated in a hepatitis C virus (HCV) genotype 1b replicon, and the 2-oxoethyl acetate substituted compound 8t showed similar anti-HCV activity (EC50 = 0.08 nM) to that of the lead compound Daclatasvir. Moreover, the utility of prodrug 8t was demonstrated through similar exposure of the parent compound when the prodrugs were dosed in vivo. PK studies showed that prodrug 8t was an ideal candidate for a slower and sustained release form of Daclatasvir. PMID:26077493

  20. Synthesis and characterization of brain penetrant prodrug of neuroprotective D-264: Potential therapeutic application in the treatment of Parkinson's disease.

    PubMed

    Dholkawala, Fahd; Voshavar, Chandrashekhar; Dutta, Aloke K

    2016-06-01

    Parkinson's disease (PD) is one of the major debilitating neurodegenerative disorders affecting millions of people worldwide. Progressive loss of dopamine neurons resulting in development of motor dysfunction and other related non-motor symptoms is the hallmark of PD. Previously, we have reported on the neuroprotective property of a potent D3 preferring agonist D-264. In our goal to increase the bioavailability of D-264 in the brain, we have synthesized a modified cysteine based prodrug of D-264 and evaluated its potential in crossing the blood-brain barrier. Herein, we report the synthesis of a novel modified cysteine conjugated prodrug of potent neuroprotective D3 preferring agonist D-264 and systematic evaluation of the hydrolysis pattern of the prodrug to yield D-264 at different time intervals in rat plasma and brain homogenates using HPLC analysis. Furthermore, we have also performed in vivo experiments with the prodrug to evaluate its enhanced brain penetration ability. PMID:26994936

  1. Nanostructured nanoparticles of self-assembled lipid pro-drugs as a route to improved chemotherapeutic agents

    NASA Astrophysics Data System (ADS)

    Sagnella, Sharon M.; Gong, Xiaojuan; Moghaddam, Minoo J.; Conn, Charlotte E.; Kimpton, Kathleen; Waddington, Lynne J.; Krodkiewska, Irena; Drummond, Calum J.

    2011-03-01

    We demonstrate that oral delivery of self-assembled nanostructured nanoparticles consisting of 5-fluorouracil (5-FU) lipid prodrugs results in a highly effective, target-activated, chemotherapeutic agent, and offers significantly enhanced efficacy over a commercially available alternative that does not self-assemble. The lipid prodrug nanoparticles have been found to significantly slow the growth of a highly aggressive mouse 4T1 breast tumour, and essentially halt the growth of a human MDA-MB-231 breast tumour in mouse xenografts. Systemic toxicity is avoided as prodrug activation requires a three-step, enzymatic conversion to 5-FU, with the third step occurring preferentially at the tumour site. Additionally, differences in the lipid prodrug chemical structure and internal nanostructure of the nanoparticle dictate the enzymatic conversion rate and can be used to control sustained release profiles. Thus, we have developed novel oral nanomedicines that combine sustained release properties with target-selective activation.

  2. Nanostructured nanoparticles of self-assembled lipid pro-drugs as a route to improved chemotherapeutic agents

    SciTech Connect

    Sagnella, Sharon M.; Gong, Xiaojuan; Moghaddam, Minoo J.; Conn, Charlotte E.; Kimpton, Kathleen; Waddington, Lynne J.; Krodkiewska, Irena; Drummond, Calum J.

    2014-09-24

    We demonstrate that oral delivery of self-assembled nanostructured nanoparticles consisting of 5-fluorouracil (5-FU) lipid prodrugs results in a highly effective, target-activated, chemotherapeutic agent, and offers significantly enhanced efficacy over a commercially available alternative that does not self-assemble. The lipid prodrug nanoparticles have been found to significantly slow the growth of a highly aggressive mouse 4T1 breast tumour, and essentially halt the growth of a human MDA-MB-231 breast tumour in mouse xenografts. Systemic toxicity is avoided as prodrug activation requires a three-step, enzymatic conversion to 5-FU, with the third step occurring preferentially at the tumour site. Additionally, differences in the lipid prodrug chemical structure and internal nanostructure of the nanoparticle dictate the enzymatic conversion rate and can be used to control sustained release profiles. Thus, we have developed novel oral nanomedicines that combine sustained release properties with target-selective activation.

  3. Topical iontophoretic delivery of ionizable, biolabile aciclovir prodrugs: A rational approach to improve cutaneous bioavailability.

    PubMed

    Chen, Yong; Alberti, Ingo; Kalia, Yogeshvar N

    2016-02-01

    The objective was to investigate the topical iontophoretic delivery of a series of amino acid ester prodrugs of aciclovir (ACV-X, where ACV=aciclovir and X=Arg, Gly, Ile, Phe, Trp and Val) as a means to enhance cutaneous delivery of ACV. The newly synthesized prodrugs were characterized by (1)H NMR and high resolution mass spectrometry. Analytical methods using HPLC-UV were developed for their quantification and each method was validated. Investigation of solution stability as a function of pH showed that all ACV-X prodrugs were relatively stable in acid conditions at pH 2.0 and pH 5.5 for up to 8h but susceptible to extensive hydrolysis at pH 7.4 and under alkaline conditions (pH 10). No ACV-X hydrolysis was observed after contact for 2h with the external surface of porcine stratum corneum. However, there was significant hydrolysis following contact with the dermal surface of dermatomed porcine skin, in particular, for ACV-Arg. Passive transport of ACV and ACV-X prodrugs from aqueous solution after 2h was below the limit of detection. Iontophoresis of ACV at 0.5 mA/cm(2) for 2h led to modest ACV skin deposition (QDEP,ACV) of 4.6 ± 0.3 nmol/cm(2). In contrast, iontophoresis of ACV-X prodrugs under the same conditions produced order of magnitude increases in cutaneous deposition of ACV species, that is, QDEP,TOTAL=QDEP,ACV+QDEP,ACV-X. QDEP,TOTAL for ACV-Gly, ACV-Val, ACV-Ile, ACV-Phe, ACV-Trp and ACV-Arg was 412.8 ± 44.0, 358.8 ± 66.8, 434.1 ± 68.2, 249.8 ± 81.4, 156.1 ± 76.3, 785.9 ± 78.1 nmol/cm(2), respectively. The extent of bioconversion of ACV-X to ACV in the skin was high and the proportion of ACV present ranged from 81% to 100%. The skin retention ratio, a measure of the selectivity of ACV species for deposition over permeation after iontophoretic delivery of ACV-X prodrugs, was dependent on both the rate of transport and the susceptibility to hydrolysis of the prodrugs. Skin deposition of ACV and its six prodrugs were investigated further as a

  4. Enhanced Antitumor Activity of Monophosphate Ester Prodrugs of Gemcitabine: In Vitro and In Vivo Evaluation.

    PubMed

    Qi, Huixin; Lu, Jia; Li, Jiajun; Wang, Meiyu; Xu, Yunting; Wang, Yedong; Zhang, Hongjian

    2016-09-01

    The prodrug strategy has been explored frequently for a number of marked drugs to obtain better pharmaceutical properties and efficacy and safety profiles. For gemcitabine, a nucleoside analog that has been used widely as a chemotherapeutic agent for the treatment of a variety of cancers, the protection of the amino group from extensive deamination and increase of permeability have been used for oral prodrug development. In the present study, several novel and proprietary monophosphate ester prodrugs of gemcitabine representing different "tail" structures were evaluated for their antiproliferation activities in various tumor cell lines. As compared to LY2334737, a prototype oral prodrug of gemcitabine, the monophosphate ester prodrugs exhibited superior in vitro antiproliferation activity. Among those, compound-3 emerged as a promising prodrug candidate. Data revealed that cellular concentrations of compound-3 were correlated well with its antiproliferation activity and its cellular uptake did not involve human equilibrative nucleoside transporter, suggesting a potential to treat gemcitabine resistant tumors. Compound-3 demonstrated equal or better antitumor efficacy after oral administration as compared to intraperitoneally injected gemcitabine. Taken together, compound-3 has the potential for further development as an orally active antitumor agent. PMID:26994559

  5. A non-canonical ESCRT pathway, including histidine domain phosphotyrosine phosphatase (HD-PTP), is used for down-regulation of virally ubiquitinated MHC class I

    PubMed Central

    Parkinson, Michael D.J.; Piper, Siân C.; Bright, Nicholas A.; Evans, Jennifer L.; Boname, Jessica M.; Bowers, Katherine; Lehner, Paul J.; Luzio, J. Paul

    2015-01-01

    The Kaposi's sarcoma-associated herpes virus (KSHV) K3 viral gene product effectively down-regulates cell surface MHC class I. K3 is an E3 ubiquitin ligase that promotes Lys63-linked polyubiquitination of MHC class I, providing the signal for clathrin-mediated endocytosis. Endocytosis is followed by sorting into the intralumenal vesicles (ILVs) of multivesicular bodies (MVBs) and eventual delivery to lysosomes. The sorting of MHC class I into MVBs requires many individual proteins of the four endosomal sorting complexes required for transport (ESCRTs). In HeLa cells expressing the KSHV K3 ubiquitin ligase, the effect of RNAi-mediated depletion of individual proteins of the ESCRT-0 and ESCRT-I complexes and three ESCRT-III proteins showed that these are required to down-regulate MHC class I. However, depletion of proteins of the ESCRT-II complex or of the ESCRT-III protein, VPS20 (vacuolar protein sorting 20)/CHMP6 (charged MVB protein 6), failed to prevent the loss of MHC class I from the cell surface. Depletion of histidine domain phosphotyrosine phosphatase (HD-PTP) resulted in an increase in the cell surface concentration of MHC class I in HeLa cells expressing the KSHV K3 ubiquitin ligase. Rescue experiments with wild–type (WT) and mutant HD-PTP supported the conclusion that HD-PTP acts as an alternative to ESCRT-II and VPS20/CHMP6 as a link between the ESCRT-I and those ESCRT-III protein(s) necessary for ILV formation. Thus, the down-regulation of cell surface MHC class I, polyubiquitinated by the KSHV K3 ubiquitin ligase, does not employ the canonical ESCRT pathway, but instead utilizes an alternative pathway in which HD-PTP replaces ESCRT-II and VPS20/CHMP6. PMID:26221024

  6. A non-canonical ESCRT pathway, including histidine domain phosphotyrosine phosphatase (HD-PTP), is used for down-regulation of virally ubiquitinated MHC class I.

    PubMed

    Parkinson, Michael D J; Piper, Siân C; Bright, Nicholas A; Evans, Jennifer L; Boname, Jessica M; Bowers, Katherine; Lehner, Paul J; Luzio, J Paul

    2015-10-01

    The Kaposi's sarcoma-associated herpes virus (KSHV) K3 viral gene product effectively down-regulates cell surface MHC class I. K3 is an E3 ubiquitin ligase that promotes Lys(63)-linked polyubiquitination of MHC class I, providing the signal for clathrin-mediated endocytosis. Endocytosis is followed by sorting into the intralumenal vesicles (ILVs) of multivesicular bodies (MVBs) and eventual delivery to lysosomes. The sorting of MHC class I into MVBs requires many individual proteins of the four endosomal sorting complexes required for transport (ESCRTs). In HeLa cells expressing the KSHV K3 ubiquitin ligase, the effect of RNAi-mediated depletion of individual proteins of the ESCRT-0 and ESCRT-I complexes and three ESCRT-III proteins showed that these are required to down-regulate MHC class I. However, depletion of proteins of the ESCRT-II complex or of the ESCRT-III protein, VPS20 (vacuolar protein sorting 20)/CHMP6 (charged MVB protein 6), failed to prevent the loss of MHC class I from the cell surface. Depletion of histidine domain phosphotyrosine phosphatase (HD-PTP) resulted in an increase in the cell surface concentration of MHC class I in HeLa cells expressing the KSHV K3 ubiquitin ligase. Rescue experiments with wild-type (WT) and mutant HD-PTP supported the conclusion that HD-PTP acts as an alternative to ESCRT-II and VPS20/CHMP6 as a link between the ESCRT-I and those ESCRT-III protein(s) necessary for ILV formation. Thus, the down-regulation of cell surface MHC class I, polyubiquitinated by the KSHV K3 ubiquitin ligase, does not employ the canonical ESCRT pathway, but instead utilizes an alternative pathway in which HD-PTP replaces ESCRT-II and VPS20/CHMP6. PMID:26221024

  7. Functional analysis of the promoter region of the human phosphotyrosine phosphatase activator gene: Yin Yang 1 is essential for core promoter activity.

    PubMed Central

    Janssens, V; Van Hoof, C; De Baere, I; Merlevede, W; Goris, J

    1999-01-01

    The phosphotyrosine phosphatase activator (PTPA) has been isolated as an in vitro regulator of protein phosphatase 2A. Human PTPA is encoded by a single gene, the structure and chromosomal localization of which have been determined in our previous work. Here we describe the further isolation, sequencing and functional characterization of the PTPA promoter region. In agreement with its ubiquitous expression, the PTPA promoter displays several characteristics of housekeeping genes: it lacks both a TATA-box and a CAAT-box, it is very GC-rich and it contains an unmethylated CpG island surrounding the transcription initiation site. Transient transfection experiments in different cell types with several truncated chimaeric luciferase reporter gene plasmids revealed the importance of the region between positions -67 and -39 for basal promoter activity. This region coincides remarkably well with the determined CpG island. Further analysis of this region demonstrated the presence of a Yin Yang 1 (YY1) binding motif at positions -52 to -44. Binding of YY1 to this sequence is demonstrated in bandshift and DNase I footprinting experiments. Another YY1 binding motif is found in the 5' untranslated region, at positions +27 to +35. Mutations in either of these sites, abolishing YY1 binding in vitro, have differential effects on promoter activity. Point mutations in both sites completely abolish promoter activity. Moreover, induction of promoter activity by co-transfection with a YY1 expression plasmid is fully dependent upon the presence of both intact YY1 binding sites. Thus YY1 apparently mediates basal transcription of the human PTPA gene through two binding sites within its proximal promoter. PMID:10585862

  8. Novel 3-O-pegylated carboxylate and 3-O-pegylated carbamate prodrugs of naltrexone for microneedle-enhanced transdermal delivery.

    PubMed

    Yerramreddy, Thirupathi Reddy; Milewski, Mikolaj; Penthala, Narsimha Reddy; Stinchcomb, Audra L; Crooks, Peter A

    2010-06-01

    A small library of novel 3-O-pegylated carboxylate prodrugs (4a-4b) and 3-O-pegylated carbamate prodrugs (9a-9b) of naltrexone were synthesized. The goal behind the design of these prodrugs was to investigate their potential for microneedle-enhanced transdermal delivery. All the synthesized 3-O-pegylated carboxylate prodrugs (4a-4b) and 3-O-pegylated carbamate prodrugs (9a-9b) of naltrexone were found to have adequate stability in a transdermal formulation and improved apparent solubility compared to naltrexone. Viscosity effects were postulated to be responsible for the observed non-linearity in the flux-concentration profile of these prodrugs. PMID:20451376

  9. A study on liposomal encapsulation of a lipophilic prodrug of LHRH.

    PubMed

    Pignatello, Rosario; Musumeci, Teresa; Graziano, Adriana C E; Lo Furno, Debora; Varamini, Pegah; Mansfeld, Friederike M; Cardile, Venera; Toth, Istvan

    2016-09-01

    This study aimed at evaluating whether derivatization of luteinizing hormone-releasing hormone (LHRH) peptide with an amphiphilic lipoamino acid moiety could allow, along with other technological and/or pharmacokinetic advantages, to improve its encapsulation in liposomes, potentially driving its further body distribution and cellular uptake. Experimental data confirmed that a lipophilic derivative of LHRH was efficiently incorporated in various liposomal systems, differing in lipid composition and surface charge, and obtained using different methods of production. Incubation of liposomes, loaded with a fluorescent derivative of the LHRH prodrug, with NCTC keratinocytes or Caco-2 cell cultures showed that the carriers can be rapidly internalized. Conversely, the internalization of the free prodrug occurred only at very high concentrations. PMID:25946073

  10. Serine Peptide Phosphoester Prodrugs of Cyclic Cidofovir: Synthesis, Transport, and Antiviral Activity

    PubMed Central

    Eriksson, Ulrika; Peterson, Larryn W.; Kashemirov, Boris A.; Hilfinger, John M.; Drach, John C.; Borysko, Katherine Z.; Breitenbach, Julie M.; Kim, Jae Seung; Mitchell, Stefanie; Kijek, Paul; McKenna, Charles E.

    2008-01-01

    Cidofovir (HPMPC, 1), a broad-spectrum antiviral agent, is currently used to treat AIDS-related human cytomegalovirus (HCMV) retinitis and has recognized therapeutic potential for orthopox virus infections, but is limited by its low oral bioavailability. Cyclic cidofovir (2) displays decreased nephrotoxicity compared to 1, while also exhibiting potent antiviral activity. Here we describe in detail the synthesis and evaluation as prodrugs of four cHPMPC dipeptide conjugates in which the free POH of 2 is esterified by the Ser side chain alcohol group of an X-l-Ser(OMe) dipeptide: 3 (X = l-Ala), 4 (X = l-Val), 5 (X = l-Leu), and 6 (X = l-Phe). Perfusion studies in the rat establish that the mesenteric permeability to 4 is more than 30-fold greater than to 1, and the bioavailability of 4 is increased 8-fold relative to 1 in an in vivo murine model. In gastrointestinal and liver homogenates, the cHPMPC prodrugs are rapidly hydrolyzed to 2. Prodrugs 3, 4, and 5 are nontoxic at 100 μM in HFF and KB cells and in cell-based plaque reduction assays had IC50 values of 0.1–0.5 μM for HCMV and 10 μM for two orthopox viruses (vaccinia and cowpox). The enhanced transport properties of 3–6, conferred by incorporation of a toxicologically benign dipeptide moiety, and the facile cleavage of the Ser–O–P linkage suggest that these prodrugs represent a promising new approach to enhancing the bioavailability of 2. PMID:18481868

  11. Enhanced cellular uptake and intracellular drug controlled release of VESylated gemcitabine prodrug nanocapsules.

    PubMed

    Fang, Yanfen; Du, Fang; Xu, Yanyun; Meng, Haijing; Huang, Jin; Zhang, Xiongwen; Lu, Wei; Liu, Shiyuan; Yu, Jiahui

    2015-04-01

    Gemcitabine, 2',2'-difluoro-2'-deoxycytidine (dFdC), is the first-line antitumor agent in the treatment of pancreatic tumors. However, it possesses certain drawbacks, such as poor biological half-life resulted from rapid metabolism and the induction of resistance, leading to its restricted therapeutic potential. With the purpose of overcoming the above drawbacks, we developed a novel VESylated gemcitabine (VES-dFdC) prodrug by coupling the N4-amino group of the pyrimidine ring of dFdC to the carboxylic group of vitamin E succinate (VES). The resulting amphiphilic compound could protect the N4-amino group of the pyrimidine ring of dFdC from being degraded by cytidine deaminase. What is more, the prodrug was able to form nanocapsules in aqueous media (similar to the structure of cytomembrane), confirmed by transmission electron microscope (TEM). Their average particle size is about 107 nm with zeta potential of -33.4 mV measured by dynamic light scattering (DLS). VES-dFdC nanocapsules showed accelerated accumulative drug release profile in simulated lysosome environment (sodium acetate buffer pH 5+cathepsin B, an enzyme in lysosome), due to the easily hydrolyzed property of amide bond by cathepsin B, while rather stable in PBS (pH 7.4) or sodium acetate buffer (pH 5.0) without cathepsin B, indicating their enhanced intracellular drug controlled release manner. Besides, VES-dFdC prodrug nanocapsules showed enhanced cellular uptake ability, and the amount of cellular uptake of the nanocapsules by the pancreatic cancer cell line BxPC-3 is seventy times higher than that of native gemcitabine in the first 1.5 h. Compared with free gemcitabine, VES-dFdC nanocapsules showed essentially increased growth inhibition activity against BxPC-3 cells, indicating its great potential as prodrug for pancreatic tumor therapy with improved antitumor activity. PMID:25746328

  12. Synthesis, crystallization, and biological evaluation of an orally active prodrug of gemcitabine.

    PubMed

    Bender, David M; Bao, Jingqi; Dantzig, Anne H; Diseroad, William D; Law, Kevin L; Magnus, Nicholas A; Peterson, Jeffrey A; Perkins, Everett J; Pu, Yangwei J; Reutzel-Edens, Susan M; Remick, David M; Starling, James J; Stephenson, Gregory A; Vaid, Radhe K; Zhang, Deyi; McCarthy, James R

    2009-11-26

    The design, synthesis, and biological characterization of an orally active prodrug (3) of gemcitabine are described. Additionally, the identification of a novel co-crystal solid form of the compound is presented. Valproate amide 3 is orally bioavailable and releases gemcitabine into the systemic circulation after passing through the intestinal mucosa. The compound has entered clinical trials and is being evaluated as a potential new anticancer agent. PMID:19860433

  13. Bispecific Antibodies for Targeted Delivery of Dendritic Polyglycerol (dPG) Prodrug Conjugates.

    PubMed

    Mehrabadi, Fatemeh Sheikhi; Adelmann, Juliane; Gupta, Shilpi; Wedepohl, Stefanie; Calderón, Marcelo; Brinkmann, Ulrich; Haag, Rainer

    2016-01-01

    One approach to further improve the therapeutic efficacy of nanoparticles is employment of active targeting strategies. Bispecific antibodies (bsAbs) that bind to both tumor specific antigens on the cell surface and to haptens such as digoxigenin (Dig) can direct digoxigeninylated payloads to tumor cells. In this study, we investigate the potential of dendritic polyglycerol (dPG) conjugates, which consist of a doxorubicin (DOX) prodrug, Dig moiety, and a poly (ethylene glycol) (PEG) shell, in combination with bsAb, as a novel approach for targeted prodrug delivery. We could show successful binding of the bsAbs to dPGDigMal- DOX-PEG conjugates, as well as binding of these complexes to the cell surface of Lewis Y (LeY) expressing MCF-7 cells. Using flow cytometry, we could show the preferential binding of the targeting complex over the complex of control conjugate lacking Dig moieties. At concentrations that are usually applied for drug delivery, antibodycomplexed nanoparticles (independent of antibody specificity) released cytotoxic compounds into cells to the same degree as unmodified nanoparticles. This indicates that antibody-attachment does not interfere with the inherent cell binding and drug delivery properties of nanoparticles. At low doxorubicin concentrations and short incubation times, however, we were able to see a slightly increased target specific cytotoxicity in vitro which is mediated by complexation of the digoxigeninylated NP with the Dig-binding moiety of a bsAb that in turns direct the complexed bsAb to target cells. This study demonstrates the potential of digoxigeninylated dPG prodrug conjugates in combination with bsAbs as a new platform for targeted prodrug delivery into cancerous tissues. However the nanoparticle design needs to be further optimized for significant targeted delivery. PMID:26853135

  14. Strategy for Imidazotetrazine Prodrugs with Anticancer Activity Independent of MGMT and MMR

    PubMed Central

    2012-01-01

    The imidazotetrazine ring is an acid-stable precursor and prodrug of highly reactive alkyl diazonium ions. We have shown that this reactivity can be managed productively in an aqueous system for the generation of aziridinium ions with 96% efficiency. The new compounds are potent DNA alkylators and have antitumor activity independent of the O6-methylguanine-DNA methyltransferase and DNA mismatch repair constraints that limit the use of Temozolomide. PMID:24900418

  15. Pharmacokinetic and pharmacodynamic assessment of bioavailability for two prodrugs of methylprednisolone

    PubMed Central

    Daley-Yates, P. T.; Gregory, A. J.; Brooks, C. D.

    1997-01-01

    Aims The aim of this study was to establish whether pharmacokinetic differences between two pro-drugs of methylprednisolone (MP) are likely to be of clinical significance. Methods This study was a single-blind, randomized, crossover design comparing the bioequivalence of MP released from the pro-drugs Promedrol (MP suleptanate) and Solu-Medrol (MP succinate) after a single 250 mg (MP equivalent) intramuscular injection to 20 healthy male volunteers. Bioequivalence was assessed by conventional pharmacokinetic analysis, by measuring pharmacodynamic responses plus a novel approach using pharmacokinetic/pharmacodynamic modeling. The main measure of pharmacodynamic response was whole blood histamine (WBH), a measure of basophil numbers. Results The MP Cmax was less for MP suleptanate due to a longer absorption half-life of the prodrug from the intramuscular injection site. The bioavailability of MP was equivalent when based on AUC with a MP suleptanate median 108% of the MP succinate value (90% CI: 102–114%). For Cmax the MP suleptanate median was 81% of the MP succinate value (90% CI: 75–88%). The tmax for MP from MP suleptanate was delayed relative to MP succinate. The median difference was 200% (90% non-parametric CI: 141–283%). The area under the WBH effect-time curve (AUEC) and the maximum response (Emax ) were found to be equivalent (90% CI: 98–113% and 93–109% respectively). The maximum changes in other white blood cell counts, blood glucose concentration and the parameters of the pharmacodynamic sigmoid Emax model (EC50, Emax and γ) were also not significantly different between prodrugs. Conclusions MP suleptanate is an acceptable pharmaceutical alternative to MP succinate. The use of both pharmacokinetic and pharmacodynamic response data together gives greater confidence in the conclusions compared with those based only on conventional pharmacokinetic bioequivalence analysis. PMID:9205819

  16. Evaluation of Antimalarial Activity and Toxicity of a New Primaquine Prodrug

    PubMed Central

    Davanço, Marcelo Gomes; Aguiar, Anna Caroline Campos; dos Santos, Leandro Alves; Padilha, Elias Carvalho; Campos, Michel Leandro; de Andrade, Cleverton Roberto; da Fonseca, Luiz Marcos; dos Santos, Jean Leandro; Chin, Chung Man; Krettli, Antoniana Ursine; Peccinini, Rosangela Gonçalves

    2014-01-01

    Plasmodium vivax is the most prevalent of the five species causing malaria in humans. The current available treatment for P. vivax malaria is limited and unsatisfactory due to at least two drawbacks: the undesirable side effects of primaquine (PQ) and drug resistance to chloroquine. Phenylalanine-alanine-PQ (Phe-Ala-PQ) is a PQ prodrug with a more favorable pharmacokinetic profile compared to PQ. The toxicity of this prodrug was evaluated in in vitro assays using a human hepatoma cell line (HepG2), a monkey kidney cell line (BGM), and human red blood cells deficient in the enzyme glucose-6-phosphate-dehydrogenase (G6PD). In addition, in vivo toxicity assays were performed with rats that received multiple doses of Phe-Ala-PQ to evaluate biochemical, hematological, and histopathological parameters. The activity was assessed by the inhibition of the sporogonic cycle using a chicken malaria parasite. Phe-Ala-PQ blocked malaria transmission in Aedes mosquitoes. When compared with PQ, it was less cytotoxic to BGM and HepG2 cells and caused less hemolysis of G6PD-deficient red blood cells at similar concentrations. The prodrug caused less alteration in the biochemical parameters than did PQ. Histopathological analysis of the liver and kidney did show differences between the control and Phe-Ala-PQ-treated groups, but they were not statistically significant. Taken together, the results highlight the prodrug as a novel lead compound candidate for the treatment of P. vivax malaria and as a blocker of malaria transmission. PMID:25133630

  17. Dipeptidyl Peptidase IV as a Potential Target for Selective Prodrug Activation and Chemotherapeutic Action in Cancers

    PubMed Central

    2015-01-01

    The efficacy of chemotherapeutic drugs is often offset by severe side effects attributable to poor selectivity and toxicity to normal cells. Recently, the enzyme dipeptidyl peptidase IV (DPPIV) was considered as a potential target for the delivery of chemotherapeutic drugs. The purpose of this study was to investigate the feasibility of targeting chemotherapeutic drugs to DPPIV as a strategy to enhance their specificity. The expression profile of DPPIV was obtained for seven cancer cell lines using DNA microarray data from the DTP database, and was validated by RT-PCR. A prodrug was then synthesized by linking the cytotoxic drug melphalan to a proline-glycine dipeptide moiety, followed by hydrolysis studies in the seven cell lines with a standard substrate, as well as the glycyl-prolyl-melphalan (GP-Mel). Lastly, cell proliferation studies were carried out to demonstrate enzyme-dependent activation of the candidate prodrug. The relative RT-PCR expression levels of DPPIV in the cancer cell lines exhibited linear correlation with U95Av2 Affymetrix data (r2 = 0.94), and with specific activity of a standard substrate, glycine-proline-p-nitroanilide (r2 = 0.96). The significantly higher antiproliferative activity of GP-Mel in Caco-2 cells (GI50 = 261 μM) compared to that in SK-MEL-5 cells (GI50 = 807 μM) was consistent with the 9-fold higher specific activity of the prodrug in Caco-2 cells (5.14 pmol/min/μg protein) compared to SK-MEL-5 cells (0.68 pmol/min/μg protein) and with DPPIV expression levels in these cells. Our results demonstrate the great potential to exploit DPPIV as a prodrug activating enzyme for efficient chemotherapeutic drug targeting. PMID:25365774

  18. Developing Precisely Defined Drug-Loaded Nanoparticles by Ring-Opening Polymerization of a Paclitaxel Prodrug.

    PubMed

    Liu, Jinyao; Pang, Yan; Bhattacharyya, Jayanta; Liu, Wenge; Weitzhandler, Isaac; Li, Xinghai; Chilkoti, Ashutosh

    2016-08-01

    Nanoparticles with high paclitaxel (PTX) loading and low systemic toxicity are prepared in scalable and versatile manner via one-step ring-opening polymerization of a prodrug monomer consisting of PTX that is appended to a cyclic carbonate through a hydrolysable ester linker. Initiating this monomer from a hydrophilic macroinitiator results in an amphiphilic diblock copolymer that spontaneously self-assembles into well-defined nanoparticles with tunable size. PMID:27111757

  19. Mitigation of nitrogen mustard mediated skin injury by a novel indomethacin bifunctional prodrug.

    PubMed

    Composto, Gabriella M; Laskin, Jeffrey D; Laskin, Debra L; Gerecke, Donald R; Casillas, Robert P; Heindel, Ned D; Joseph, Laurie B; Heck, Diane E

    2016-06-01

    Nitrogen mustard (NM) is a bifunctional alkylating agent that is highly reactive in the skin causing extensive tissue damage and blistering. In the present studies, a modified cutaneous murine patch model was developed to characterize NM-induced injury and to evaluate the efficacy of an indomethacin pro-drug in mitigating toxicity. NM (20μmol) or vehicle control was applied onto 6mm glass microfiber filters affixed to the shaved dorsal skin of CD-1 mice for 6min. This resulted in absorption of approximately 4μmol of NM. NM caused localized skin damage within 1 d, progressing to an eschar within 2-3 d, followed by wound healing after 4-5 d. NM-induced injury was associated with increases in skin thickness, inflammatory cell infiltration, reduced numbers of sebocytes, basal keratinocyte double stranded DNA breaks, as measured by phospho-histone 2A.X expression, mast cell degranulation and increases in inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2). Wound healing was characterized by epidermal hyperplasia and marked increases in basal cells expressing proliferating cell nuclear antigen. A novel indomethacin-anticholinergic prodrug (4338) designed to target cyclooxygenases and acetylcholinesterase (AChE), was found to markedly suppress NM toxicity, decreasing wound thickness and eschar formation. The prodrug also inhibited mast cell degranulation, suppressed keratinocyte expression of iNOS and COX-2, as well as markers of epidermal proliferation. These findings indicate that a novel bifunctional pro-drug is effective in limiting NM mediated dermal injury. Moreover, our newly developed cutaneous patch model is a sensitive and reproducible method to assess the mechanism of action of countermeasures. PMID:27189522

  20. Metabolism of ethyl 2-carbamoyloxybenzoate (4003/2), a prodrug of salicylic acid, carsalam and salicylamide.

    PubMed

    Kamal, A

    1990-10-01

    The metabolites of 4003/2 observed in vivo have been produced by incubation in vitro of 4003/2 with liver post-mitochondrial supernatants from rat, rabbit and dog. The metabolites were characterized by UV, i.r., NMR, MS and HPLC. All the metabolites detected have been or are in use as drugs for the relief of pain and inflammation. Hence, the new drug 4003/2 is a pro-drug of salicylic acid, carsalam and salicylamide. PMID:2222521

  1. Alkoxycarbonyloxyethyl ester prodrugs of FR900098 with improved in vivo antimalarial activity.

    PubMed

    Ortmann, Regina; Wiesner, Jochen; Reichenberg, Armin; Henschker, Dajana; Beck, Ewald; Jomaa, Hassan; Schlitzer, Martin

    2005-07-01

    FR900098 represents a derivative of the new antimalarial drug fosmidomycin with enhanced activity. The mechanism of action is the inhibition of the 1-desoxy-D-xylulose 5-phosphate (DOXP) reductoisomerase, an essential enzyme of the mevalonate independent pathway of isoprenoid biosynthesis. Prodrugs with increased oral activity in mice infected with the rodent malaria parasite Plasmodium vinckei were obtained by masking the phosphonate moiety of FR900098 as alkoxycarbonyloxyethyl esters. PMID:15996004

  2. Acyloxyalkyl ester prodrugs of FR900098 with improved in vivo anti-malarial activity.

    PubMed

    Ortmann, Regina; Wiesner, Jochen; Reichenberg, Armin; Henschker, Dajana; Beck, Ewald; Jomaa, Hassan; Schlitzer, Martin

    2003-07-01

    FR900098 represents an improved derivative of the new antimalarial drug fosmidomycin and acts through inhibition of the 1-deoxy-D-xylulose 5-phosphate (DOXP) reductoisomerase, an essential enzyme of the mevalonate independent pathway of isoprenoid biosynthesis. Prodrugs with increased activity after oral administration were obtained by chemical modification of the phosphonate moiety to yield acyloxyalkyl esters. The most successful compound demonstrated 2-fold increased activity in mice infected with the rodent malaria parasite Plasmodium vinckei. PMID:12798327

  3. Metabolism and Pharmacokinetics of the Anti-Hepatitis C Virus Nucleotide Prodrug GS-6620

    PubMed Central

    Wang, Ting; Babusis, Darius; Lepist, Eve-Irene; Sauer, Dorothea; Park, Yeojin; Vela, Jennifer E.; Shih, Robert; Birkus, Gabriel; Stefanidis, Dimitrios; Kim, Choung U.; Cho, Aesop

    2014-01-01

    The anti-hepatitis C virus nucleotide prodrug GS-6620 employs a double-prodrug approach, with l-alanine-isopropyl ester and phenol moieties attached to the 5′-phosphate that release the nucleoside monophosphate in hepatocytes and a 3′-isobutyryl ester added to improve permeability and oral bioavailability. Consistent with the stability found in intestinal homogenates, following oral administration, intact prodrug levels in blood plasma were the highest in dogs, followed by monkeys, and then were the lowest in hamsters. In contrast, liver levels of the triphosphate metabolite at the equivalent surface area-adjusted doses were highest in hamsters, followed by in dogs and monkeys. Studies in isolated primary hepatocytes suggest that relatively poor oral absorption in hamsters and monkeys was compensated for by relatively efficient hepatocyte activation. As intestinal absorption was found to be critical to the effectiveness of GS-6620 in nonclinical species, stomach pH, formulation, and food effect studies were completed in dogs. Consistent with in vitro absorption studies in Caco-2 cells, the absorption of GS-6620 was found to be complex and highly dependent on concentration. Higher rates of metabolism were observed at lower concentrations that were unable to saturate intestinal efflux transporters. In first-in-human clinical trials, the oral administration of GS-6620 resulted in poor plasma exposure relative to that observed in dogs and in large pharmacokinetic and pharmacodynamic variabilities. While a double-prodrug approach, including a 3′-isobutyryl ester, provided higher intrinsic intestinal permeability, this substitution appeared to be a metabolic liability, resulting in extensive intestinal metabolism and relatively poor oral absorption in humans. PMID:24419340

  4. Targeting Carcinoma-Associated Fibroblasts Within the Tumor Stroma With a Fibroblast Activation Protein-Activated Prodrug

    PubMed Central

    2012-01-01

    Background Fibroblasts undergo a morphological transformation to a reactive phenotype in the tumor microenvironment characterized by the expression of proteins such as fibroblast activation protein (FAP), a post-prolyl endopeptidase with expression largely restricted to carcinoma-associated fibroblasts. Thapsigargin (TG) is a highly toxic natural plant product that triggers a rise in intracellular calcium levels and apoptosis. FAP is therefore a provocative target for the activation of prodrugs consisting of a FAP-specific peptide coupled to a potent cytotoxic analog of TG. Methods The efficacy of FAP-activated peptidyl-TG prodrugs was tested in vitro in cell proliferation assays and effects on intracellular calcium in human cancer cell lines. The effects of FAP-activated prodrugs on tumor growth and host toxicity were tested in Balb-C nude MCF-7 and LNCaP xenograft mice (n = 9–11 per group). P values were calculated using permutation tests based on 50 000 permutations. Mixed effects models were used to account for correlations among replicate measures. All statistical tests were two-sided. Results FAP-activated prodrugs killed human cancer cells at low nanomolar concentrations (MCF-7 cells: IC50 = 3.5nM). Amino acid-12ADT analogs from FAP-cleaved prodrugs, but not uncleaved prodrugs, produced a rapid rise in intracellular calcium within minutes of exposure. Immunohistochemical analysis of xenografts exposed to FAP-prodrugs documented stromal-selective cell death of fibroblasts, pericytes, and endothelial cells of sufficient magnitude to inhibit growth of MCF-7 and LNCaP xenografts with minimal systemic toxicity, whereas non-FAP cleavable prodrugs were inactive. MCF-7 and LNCaP xenografts treated with a FAP-activated prodrug had maximal treated-to-control tumor volume ratios of 0.36 (treated: mean = 0.206mm3, 95% CI = 0.068 to 0.344mm3; control: mean = 0.580mm3, 95% CI = 0.267 to 0.893mm3) and 0.24 (treated: mean = 0.131mm3, 95% CI = 0.09 to 0.180mm3; control

  5. Synthesis of an enzyme-dependent prodrug and evaluation of its potential for colon targeting

    PubMed Central

    Pang, Yi-Nuo; Zhang, Yan; Zhang, Zhi-Rong

    2002-01-01

    AIM: To synthesize dexamethasone-succinate-dextran (DSD) conjugate and to evaluate the potentiality of DSD for the treatment of inflammatory bowel diseases. METHODS: Dexamethasone was attached to dextran (average molecular weight = 70400 Dalton) using succinate anhydride in an anhydrous environment catalyzed by 4-dimethylaminopyridine and 1,1’-carbonyldiimidazole. The chemical structure of DSD was identified by UV, IR and NMR, and the in vivo drug release behavior of this prodrug was investigated after oral administration of DSD suspension. RESULTS: The DSD conjugate was obtained in two steps and the content of dexamethasone in DSD was 11.28%. The dextran prodrug was stable in rat stomach and small intestine and negligibly absorbed from these tracts. Four to nine hours after the oral administration, most of the prodrug (> 95%) had moved to the cecum and colon, and was easily hydrolyzed by an endodextranase. Recover of dexamethasone from colon and cecum after administration of DSD conjugate was 6-12 folds higher than the recovery after administration of unmodified dexamethasone (t = 2.74, P < 0.05). The preferential release of free dexamethasone in cecum and colon over that in the small intestine was statistically significant (t = 2.27, P < 0.05). CONCLUSION: The results of this study indicate that dextran conjugates may be useful in selectively delivering glucocorticoids to the colon. PMID:12378641

  6. A novel prodrug strategy to improve the oral absorption of O-desmethylvenlafaxine

    PubMed Central

    Liu, Mingyuan; Sun, Yantong; Zhao, Sen; Li, Youxin; Piao, Riyang; Yang, Yan; Gu, Jingkai

    2016-01-01

    O-Desmethylvenlafaxine (desvenlafaxine, ODV) is the active metabolite of venlafaxine, with similar activity and less risk for pharmacokinetic drug interactions compared to its parent compound venlafaxine. The purpose of this study was to design a series of esters of ODV and assess their potential as ODV prodrugs with improved bioavailability and brain uptake. Seven esters were synthesized and pharmacokinetic screening was performed in rats. The monoester formed on the phenolic hydroxyl of ODV (ODVP-1, ODVP-2, ODVP-3 and ODVP-5) could be degraded to ODV in rat plasma. These four compounds confirmed as possible prodrugs were then studied to evaluated the relative bioavailability of ODV they produced in beagle dogs. ODVP-1, ODVP-2 and ODVP-3 demonstrated higher relative bioavailability of ODV. Finally, ODVP-1, ODVP-2 and ODVP-3 were studied to evaluate their brain uptake in rats. The concentration of ODV in the rat plasma, brain and hypothalamus after administration of ODVP-1, ODVP-2 or ODVP-3 was higher compared with that of ODV. The higher bioavailability, improved pharmacokineics properties and more rapid penetration and translation of ODV suggest that ODVP-1, ODVP-2 or ODVP-3 may warrant further development and application as ODV prodrugs. PMID:27588083

  7. Biocompatible polymeric nanocomplexes as an intracellular stimuli-sensitive prodrug for type-2 diabetes combination therapy.

    PubMed

    Wang, Feng-Zhen; Xie, Zhi-Shen; Xing, Lei; Zhang, Bing-Feng; Zhang, Jia-Liang; Cui, Peng-Fei; Qiao, Jian-Bin; Shi, Kun; Cho, Chong-Su; Cho, Myung-Haing; Xu, Xiaojun; Li, Ping; Jiang, Hu-Lin

    2015-12-01

    Combination therapy is usually considered as a promising strategy owing to its advantages such as reduced doses, minimized side effects and improved therapeutic efficiency in a variety of diseases including diabetes. Here we synthesized a new highly intracellular stimuli-sensitive chitosan-graft-metformin (CS-MET) prodrug by imine reaction between oxidative chitosan and metformin for type 2 diabetes (T2D) therapy. Hypothetically, CS-MET functions dually as an anti-diabetes prodrug as well as a gene delivery vector without superfluous materials. CS-MET formed nanocomplexes with therapeutic gene through electrostatic interactions and entered cells by Organic Cation Transporter (OCT)-independent endocytosis. The incorporation of metformin into chitosan has been found to increase endosomal escape via the proton sponge effect. When vector carrying a short-hairpin RNA (shRNA) silencing sterol regulatory element-binding protein (SREBP), a major transcription factor involved in de novo lipogenisis, it reduced the SREBP mRNA and proteins efficiently. Furthermore, by intraperitoneal injection, CS-MET/shSREBP nanocomplexes effectively knocked down SREBP in livers of western-type diet (WD)-induced obese C57BL/6J mice, markedly reversed insulin resistance and alleviated the fatty liver phenotype without obvious toxic effects. Thus we were able to show that the intracellular stimuli-sensitive CS-MET prodrug renders a potential platform to increase the anti-diabetes activity with synergistic enhancement of gene therapy. PMID:26409000

  8. N-Monosubstituted Methoxy-oligo(ethylene glycol) Carbamate Ester Prodrugs of Resveratrol.

    PubMed

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

    2015-01-01

    Resveratrol is a natural polyphenol with many interesting biological activities. Its pharmacological exploitation in vivo is, however, hindered by its rapid elimination via phase II conjugative metabolism at the intestinal and, most importantly, hepatic levels. One approach to bypass this problem relies on prodrugs. We report here the synthesis, characterization, hydrolysis, and in vivo pharmacokinetic behavior of resveratrol prodrugs in which the OH groups are engaged in an N-monosubstituted carbamate ester linkage. As promoiety, methoxy-oligo(ethylene glycol) groups (m-OEG) (CH₃-[OCH₂CH₂]n-) of defined chain length (n = 3, 4, 6) were used. These are expected to modulate the chemico-physical properties of the resulting derivatives, much like longer poly(ethylene glycol) (PEG) chains, while retaining a relatively low MW and, thus, a favorable drug loading capacity. Intragastric administration to rats resulted in the appearance in the bloodstream of the prodrug and of the products of its partial hydrolysis, confirming protection from first-pass metabolism during absorption. PMID:26404221

  9. Stoichiometric molecularly imprinted polymers for the recognition of anti-cancer pro-drug tegafur.

    PubMed

    Mattos Dos Santos, Paula; Hall, Andrew J; Manesiotis, Panagiotis

    2016-05-15

    Molecularly imprinted polymers (MIPs) targeting tegafur, an anti-cancer 5-fluorouracil pro-drug, have been prepared by stoichiometric imprinting using 2,6-bis(acrylamido)pyridine (BAAPy) as the functional monomer. Solution association between tegafur and BAAPy was studied by (1)H NMR titration, which confirmed the formation of 1:1 complexes with an affinity constant of 574±15M(-1) in CDCl3. Evaluation of the synthesised materials by HPLC and equilibrium rebinding experiments revealed high selectivity of the imprinted polymer for the pro-drug vs. 5-fluorouracil and other competing analytes, with maximum imprinting factors of 25.3 and a binding capacity of 45.1μmolg(-1). The synthesised imprinted polymer was employed in solid-phase extraction of the pro-drug using an optimised protocol that included a simple wash with the porogen used in the preparation of the material. Tegafur recoveries of up to 96% were achieved from aqueous samples and 92% from urine samples spiked with the template and three competing analytes. The results demonstrate the potential of the prepared polymers in the pre-concentration of tegafur from biological samples, which could be an invaluable tool in the monitoring of patient compliance and drug uptake and excretion. PMID:26711233

  10. Studies on synthesis, stability, release and pharmacodynamic profile of a novel diacerein-thymol prodrug.

    PubMed

    Dhaneshwar, Suneela; Patel, Vriha; Patil, Dipmala; Meena, Gourav

    2013-01-01

    Involvement of oxidative stress, leading to chondrocyte senescence and cartilage ageing has been implicated in the pathogenesis of osteoarthritis (OA). New efforts to prevent the development and progression of OA include strategies and interventions aimed at reducing oxidative damage in articular cartilage using antioxidants as adjuncts to conservative therapy. Diacerein is an anthraquinone derivative with a marked disease modifying effect on OA owing to IL-1 β inhibition. In the present work an attempt was made at design and development of a co-drug of diacerein with antioxidant thymol. Structural elucidation was carried out by spectral analysis. When release kinetics of prodrug was studied in phosphate buffer (pH 7.4) and small intestinal homogenates of rats, 91% and 94% diacerein was available respectively at the end of 4.5 h. Chemical linkage of thymol with diacerein improved its lipophilicity and hence bioavailability. Screening of prodrug in Freud's adjuvant-induced arthritis and ulcerogenic potential by Rainsford's cold stress model exhibited significant reduction in paw volume, joint diameter and ulcer index with superior anti-inflammatory/anti-arthritic activities than the standards. Results of histopathology of tibio-tarsal joint indicated that animals treated with diacerein exhibited moderate synovitis while thymol and physical mixture-treated animals showed mild synovitis. Interestingly in prodrug-treated animals synovitis was not observed. The results of this study underline the promising potential of co-drug of diacerein and thymol in the management of OA. PMID:23218603

  11. Dipeptide Prodrug Approach to Evade Efflux Pumps and CYP3A4 Metabolism of Lopinavir

    PubMed Central

    Patel, Mitesh; Sheng, Ye; Mandava, Nanda K.; Pal, Dhananjay; Mitra, Ashim K.

    2014-01-01

    Oral absorption of lopinavir (LPV) is limited due to P-glycoprotein (P-gp) and multidrug resistance-associated protein2 (MRP2) mediated efflux by intestinal epithelial cells. Moreover, LPV is extensively metabolized by CYP3A4 enzymes. In the present study, dipeptide prodrug approach was employed to circumvent efflux pumps (P-gp and MRP2) and CYP3A4 mediated metabolism of LPV. Valine-isoleucine-LPV (Val-Ile-LPV) was synthesized and identified by LCMS and NMR techniques. The extent of LPV and Val-Ile-LPV interactions with P-gp and MRP2 was studied by uptake and transport studies across MDCK-MDR1 and MDCK-MRP2 cells. To determine the metabolic stability, time and concentration dependent degradation study was performed in liver microsomes. Val-Ile-LPV exhibited significantly higher aqueous solubility relative to LPV. This prodrug generated higher stability under acidic pH. Val-Ile-LPV demonstrated significantly lower affinity towards P-gp and MRP2 relative to LPV. Transepithelial transport of Val-Ile-LPV was significantly higher in the absorptive direction (apical to basolateral) relative to LPV. Importantly, Val-Ile-LPV was recognized as an excellent substrate by peptide transporter. Moreover, Val-Ile-LPV displayed significantly higher metabolic stability relative to LPV. Results obtained from this study suggested that dipeptide prodrug approach is a viable option to elevate systemic levels of LPV following oral administration PMID:25261710

  12. A prodrug-doped cellular Trojan Horse for the potential treatment of prostate cancer.

    PubMed

    Levy, Oren; Brennen, W Nathaniel; Han, Edward; Rosen, David Marc; Musabeyezu, Juliet; Safaee, Helia; Ranganath, Sudhir; Ngai, Jessica; Heinelt, Martina; Milton, Yuka; Wang, Hao; Bhagchandani, Sachin H; Joshi, Nitin; Bhowmick, Neil; Denmeade, Samuel R; Isaacs, John T; Karp, Jeffrey M

    2016-06-01

    Despite considerable advances in prostate cancer research, there is a major need for a systemic delivery platform that efficiently targets anti-cancer drugs to sites of disseminated prostate cancer while minimizing host toxicity. In this proof-of-principle study, human mesenchymal stem cells (MSCs) were loaded with poly(lactic-co-glycolic acid) (PLGA) microparticles (MPs) that encapsulate the macromolecule G114, a thapsigargin-based prostate specific antigen (PSA)-activated prodrug. G114-particles (∼950 nm in size) were internalized by MSCs, followed by the release of G114 as an intact prodrug from loaded cells. Moreover, G114 released from G114 MP-loaded MSCs selectively induced death of the PSA-secreting PCa cell line, LNCaP. Finally, G114 MP-loaded MSCs inhibited tumor growth when used in proof-of-concept co-inoculation studies with CWR22 PCa xenografts, suggesting that cell-based delivery of G114 did not compromise the potency of this pro-drug in-vitro or in-vivo. This study demonstrates a potentially promising approach to assemble a cell-based drug delivery platform, which inhibits cancer growth in-vivo without the need of genetic engineering. We envision that upon achieving efficient homing of systemically infused MSCs to cancer sites, this MSC-based platform may be developed into an effective, systemic 'Trojan Horse' therapy for targeted delivery of therapeutic agents to sites of metastatic PCa. PMID:27019026

  13. Design, synthesis and biological evaluation of phosphorodiamidate prodrugs of antiviral and anticancer nucleosides

    PubMed Central

    McGuigan, Christopher; Bourdin, Claire; Derudas, Marco; Hamon, Nadège; Hinsinger, Karen; Kandil, Sahar; Madela, Karolina; Meneghesso, Silvia; Pertusati, Fabrizio; Serpi, Michaela; Slusarczyk, Magdalena; Chamberlain, Stanley; Kolykhalov, Alexander; Vernachio, John; Vanpouille, Christophe; Introini, Andrea; Margolis, Leonid; Balzarini, Jan

    2014-01-01

    We herein report the application of the phosphorodiamidate phosphate prodrug approach to a series of thirteen nucleoside analogs with antiviral or anticancer activity. Twenty-five symmetrical phosphorodiamidates were synthesized, bearing esterified l-Alanine (and in one case d-alanine) in the prodrug moiety, each as single stereoisomer. The presence of an achiral phosphorus represents a potential advantage over the phosphoramidate ProTide approach, where diastereoisomeric mixtures are routinely obtained, and different biological profiles may be expected from the diastereoisomers. Optimization of the synthetic pathway allowed us to identify two general methods depending on the particular nucleoside analogs. All the compounds were biologically evaluated in antiviral and anticancer assays and several showed improvement of activity compared to their parent nucleosides, as in the case of ddA, d4T, abacavir and acyclovir against HIV-1 and/or HIV-2. The biological results were supported by metabolism studies with carboxypeptidase Y monitored by 31P NMR to investigate their bioactivation. This work further validates the phosphorodiamidate approach as a monophosphate prodrug motif with broad application in the antiviral and anticancer fields. PMID:24177359

  14. Aminocarbonyloxymethyl ester prodrugs of flufenamic acid and diclofenac: suppressing the rearrangement pathway in aqueous media.

    PubMed

    Ribeiro, Lina; Silva, Nuno; Iley, Jim; Rautio, Jarkko; Järvinen, Tomi; Mota-Filipe, Hélder; Moreira, Rui; Mendes, Eduarda

    2007-01-01

    Aminocarbonyloxymethyl ester prodrugs are known to undergo rearrangement in aqueous solutions to form the corresponding N-acylamine side product via an O-->N intramolecular acyl transfer from the carbamate conjugate base. Novel aminocarbonyloxymethyl esters of diclofenac and flufenamic acid containing amino acid amide carriers were synthesized and evaluated as potential prodrugs displaying less ability to undergo rearrangement. These compounds were prepared in reasonable yield by a four-step synthetic method that uses the appropriate N-Boc-protected amino acid N-hydroxysuccinimide ester and secondary amine and chloromethyl chloroformate as key reactants. Their reactivity in pH 7.4 buffer and 80% human plasma at 37 degrees C was assessed by RP-HPLC. The aminocarbonyloxymethyl esters containing a secondary carbamate group derived from amino acids such as glycine or phenylalanine were hydrolyzed quantitatively to the parent drug both in non-enzymatic and enzymatic conditions, with no rearrangement product being detected. The oral bioavailability in rats was determined for selected diclofenac derivatives. These derivatives displayed a bioavailability of 25 to 68% relative to that of diclofenac, probably due to their poor aqueous solubility and lipophilicity. These results suggest that further optimization of aminocarbonyloxymethyl esters as potential prodrugs for non-steroidal anti-inflammatory drugs require the use of amino acid carriers with ionizable groups to improve aqueous solubility. PMID:17206608

  15. Direct chemical grafted curcumin on halloysite nanotubes as dual-responsive prodrug for pharmacological applications.

    PubMed

    Massaro, M; Amorati, R; Cavallaro, G; Guernelli, S; Lazzara, G; Milioto, S; Noto, R; Poma, P; Riela, S

    2016-04-01

    Covalently functionalized halloysite nanotubes (HNTs) were successfully employed as dual-responsive nanocarriers for curcumin (Cur). Particularly, we synthesized HNT-Cur prodrug with a controlled curcumin release on dependence of both intracellular glutathione (GSH) and pH conditions. In order to obtain HNT-Cur produgs, halloysite was firstly functionalized with cysteamine through disulphide linkage. Afterwards, curcumin molecules were chemically conjugated to the amino end groups of halloysite via Schiff's base formation. The successful functionalization of halloysite was proved by thermogravimetric analysis, FT-IR spectroscopy, dynamic light scattering and scanning electron microscopy. Experimental data confirmed the presence of curcumin on HNT external surface. Moreover, we investigated the kinetics of curcumin release by UV-vis spectroscopy, which highlighted that HNT-Cur prodrug possesses dual stimuli-responsive ability upon exposure to GSH-rich or acidic environment. In vitro antiproliferative and antioxidant properties of HNT-Cur prodrug were studied with the aim to explore their potential applications in pharmaceutics. This work puts forward an efficient strategy to prepare halloysite based nanocarriers with controlled drug delivery capacity through direct chemical grafting with stimuli-responsive linkage. PMID:26812638

  16. Synthesis and Antitumor Properties of BQC-Glucuronide, a Camptothecin Prodrug for Selective Tumor Activation.

    PubMed

    Prijovich, Zeljko M; Burnouf, Pierre-Alain; Chou, Hua-Cheng; Huang, Ping-Ting; Chen, Kai-Chuan; Cheng, Tian-Lu; Leu, Yu-Lin; Roffler, Steve R

    2016-04-01

    Major limitations of camptothecin anticancer drugs (toxicity, nonselectivity, water insolubility, inactivation by human serum albumin) may be improved by creating glucuronide prodrugs that rely on beta-glucuronidase for their activation. We found that the camptothecin derivative 5,6-dihydro-4H-benzo[de]quinoline-camptothecin (BQC) displays greater cytotoxicity against cancer cells than the clinically used camptothecin derivatives SN-38 and topotecan even in the presence of human serum albumin. We synthesized the prodrug BQC-glucuronide (BQC-G), which was 4000 times more water soluble and 20-40 times less cytotoxic than BQC. Importantly, even in the presence of human serum albumin, BQC-G was efficiently hydrolyzed by beta-glucuronidase and produced greater cytotoxicity (IC50 = 13 nM) than camptothecin, 9-aminocamptothecin, SN-38, or topotecan (IC50 > 3000, 1370, 48, and 28 nM, respectively). BQC-G treatment of mice bearing human colon cancer xenografts with naturally or artificially elevated beta-glucuronidase activity produced significant antitumor activity, showing that BQC-G is a potent prodrug suitable for selective intratumoral drug activation. PMID:26824303

  17. 4-Azidobenzyl ferrocenylcarbamate as an anticancer prodrug activated under reductive conditions.

    PubMed

    Kinski, Elisa; Marzenell, Paul; Hofer, Walter; Hagen, Helen; Raskatov, Jevgenij A; Knaup, Karl X; Zolnhofer, Eva M; Meyer, Karsten; Mokhir, Andriy

    2016-07-01

    Aminoferrocene-based prodrugs are activated in the presence of cancer-specific amounts of reactive oxygen species, e.g. H2O2, with the formation of products of two types: Fe-containing complexes, which catalyze generation of HO and O2(-), and quinone methides, which alkylate glutathione and inhibit the antioxidative system of the cell. Both processes act synergistically by increasing the oxidative stress in cancer cells thereby leading to their death. However, in the activation step including the cleavage of a B-C bond one molecule of H2O2 is consumed that counteracts the desired effect of the products released from aminoferrocenes. We replaced an H2O2-sensitive trigger in original prodrugs with an azide group. This trigger is slowly reduced in the presence of glutathione with the formation of an unstable arylamine intermediate, which decomposes with the release of iron ions and iminoquinone methides. These products induce strong oxidative stress in cells as we confirmed using 2',7'-dichlorodihydrofluorescin diacetate reagent in combination with flow cytometry. In this case the activation process does not consume H2O2. Correspondingly, we observed that the azide-containing prodrug is substantially more toxic towards human promyelocytic leukemia cell line HL-60 (IC50=27±4μM) than its H2O2-responsive analogue (IC50>50μM). PMID:26970945

  18. A Prodrug Approach to the Use of Coumarins as Potential Therapeutics for Superficial Mycoses

    PubMed Central

    Mercer, Derry K.; Robertson, Jennifer; Wright, Kristine; Miller, Lorna; Smith, Shane; Stewart, Colin S.; O′Neil, Deborah A.

    2013-01-01

    Superficial mycoses are fungal infections of the outer layers of the skin, hair and nails that affect 20–25% of the world's population, with increasing incidence. Treatment of superficial mycoses, predominantly caused by dermatophytes, is by topical and/or oral regimens. New therapeutic options with improved efficacy and/or safety profiles are desirable. There is renewed interest in natural product-based antimicrobials as alternatives to conventional treatments, including the treatment of superficial mycoses. We investigated the potential of coumarins as dermatophyte-specific antifungal agents and describe for the first time their potential utility as topical antifungals for superficial mycoses using a prodrug approach. Here we demonstrate that an inactive coumarin glycone, esculin, is hydrolysed to the antifungal coumarin aglycone, esculetin by dermatophytes. Esculin is hydrolysed to esculetin β-glucosidases. We demonstrate that β-glucosidases are produced by dermatophytes as well as members of the dermal microbiota, and that this activity is sufficient to hydrolyse esculin to esculetin with concomitant antifungal activity. A β-glucosidase inhibitor (conduritol B epoxide), inhibited antifungal activity by preventing esculin hydrolysis. Esculin demonstrates good aqueous solubility (<6 g/l) and could be readily formulated and delivered topically as an inactive prodrug in a water-based gel or cream. This work demonstrates proof-of-principle for a therapeutic application of glycosylated coumarins as inactive prodrugs that could be converted to an active antifungal in situ. It is anticipated that this approach will be applicable to other coumarin glycones. PMID:24260474

  19. Dose selection as it pertains to testing a prodrug in carcinogenicity bioassays.

    PubMed

    Szczech, G M; Tucker, W E

    1983-01-01

    Toxicologists need more information than is usually available in the early stages of development of a drug in order to choose proper dose levels for testing in the bioassays. The approach most likely to result in successful bioassays involves an early multidisciplinary effort in which there is pharmacokinetic characterization of the test material in both rats and mice. Preliminary 3 month studies are desirable. Periodic sampling of plasma is essential to detect possible non-linear kinetics (as in the example we report herein) reflected as accumulation of the test material or metabolites. This is true regardless of the test substance. However, if one tests prodrugs it may be particularly helpful to know if chemical or enzymatic conversion of the prodrug is linear and if there is reversion to prodrug or other abberant metabolism. Failure to rule out these possibilities could result in subsequent clinically irrelevant organ damage or could compromise longevity or the interpretation of results in lifetime studies. Pharmacokinetic considerations are as valid as the more traditional biologic or morphologic end points used to estimate maximum tolerated or no-effect dose levels. PMID:6681397

  20. Diabetes mellitus: novel insights, analysis and interpretation of pathophysiology and complications management with imidazole-containing peptidomimetic antioxidants.

    PubMed

    Babizhayev, Mark A; Lankin, Vadim Z; Savel'Yeva, Ekaterina L; Deyev, Anatoliy I; Yegorov, Yegor E

    2013-12-01

    peroxidase type of activity and protection of antioxidant enzymes from inactivation (such as in a case of superoxide dismutase). Carnosine biological mimetics react with methylglyoxal and they are described in this study as a glyoxalase mimetics. The imidazole-containing carnosine biological mimetics can react with a number of deleterious aldehydic products of lipid peroxidation and thereby suppress their toxicity. Carnosine and carcinine can also react with glycated proteins and inhibit advanced glycation end product formation. These studies indicate a therapeutic role for imidazole-containing antioxidants (non-hydrolized carnosine, carcinine, D-carnosine, ophthalmic prodrug N-acetylcarnosine, leucyl-histidylhidrazide and patented formulations thereof) in therapeutic management strategies for Type 2 Diabetes. PMID:24236935

  1. Prediction of Nanoparticle Prodrug Metabolism by Pharmacokinetic Modeling of Biliary Excretion

    PubMed Central

    Stern, Stephan T.; Zou, Peng; Skoczen, Sarah; Xie, Sherwin; Liboiron, Barry; Harasym, Troy; Tardi, Paul; Mayer, LawrenceD.; McNeil, Scott E.

    2013-01-01

    Pharmacokinetic modeling and simulation is a powerful tool for the prediction of drug concentrations in the absence of analytical techniques that allow for direct quantification. The present study applied this modeling approach to determine active drug release from a nanoparticle prodrug formulation. A comparative pharmacokinetic study of a nanoscale micellar docetaxel (DTX) prodrug, Procet 8, and commercial DTX formulation, Taxotere, was conducted in bile duct cannulated rats. The nanoscale (~40 nm) size of the Procet 8 formulation resulted in confinement within the plasma space and high prodrug plasma concentrations. Ex vivo prodrug hydrolysis during plasma sample preparation resulted in unacceptable error that precluded direct measurement of DTX concentrations. Pharmacokinetic modeling of Taxotere and Procet 8 plasma concentrations, and their associated biliary metabolites, allowed for prediction of the DTX concentration profile and DTX bioavailability, and thereby evaluation of Procet 8 metabolism. Procet 8 plasma decay and in vitro plasma hydrolytic rates were identical, suggesting systemic clearance of the prodrug was primarily metabolic. The Procet 8 and Taxotere plasma profiles, and associated docetaxel hydroxy-tert-butyl carbamate (HDTX) metabolite biliary excretion, were best fit by a two compartment model, with both linear and non-linear DTX clearance, and first order Procet 8 hydrolysis. The model estimated HDTX clearance rate agreed with in vitro literature values, supporting the predictability of the proposed model. Model simulation at the 10 mg DTX equivalent/kg dose level predicted DTX formation rate-limited kinetics and a peak plasma DTX concentration of 39 ng/mL at 4h for Procet 8, in comparison to 2826 ng/mL for Taxotere. As a result of nonlinear DTX clearance, the DTX AUCinf for the Procet 8 formulation was predicted to be 2.6 times lower than Taxotere (775 vs. 2017 h x ng/mL, respectively), resulting in an absolute bioavailability estimate of

  2. Platinum(iv) prodrug conjugated Pd@Au nanoplates for chemotherapy and photothermal therapy

    NASA Astrophysics Data System (ADS)

    Shi, Saige; Chen, Xiaolan; Wei, Jingping; Huang, Yizhuan; Weng, Jian; Zheng, Nanfeng

    2016-03-01

    Owing to the excellent near infrared (NIR) light absorption and efficient passive targeting toward tumor tissue, two-dimensional (2D) core-shell PEGylated Pd@Au nanoplates have great potential in both photothermal therapy and drug delivery systems. In this work, we successfully conjugate Pd@Au nanoplates with a platinum(iv) prodrug c,c,t-[Pt(NH3)2Cl2(O2CCH2CH2CO2H)2] to obtain a nanocomposite (Pd@Au-PEG-Pt) for combined photothermal-chemotherapy. The prepared Pd@Au-PEG-Pt nanocomposite showed excellent stability in physiological solutions and efficient Pt(iv) prodrug loading. Once injected into biological tissue, the Pt(iv) prodrug was easily reduced by physiological reductants (e.g. ascorbic acid or glutathione) into its cytotoxic and hydrophilic Pt(ii) form and released from the original nanocomposite, and the NIR laser irradiation could accelerate the release of Pt(ii) species. More importantly, Pd@Au-PEG-Pt has high tumor accumulation (29%ID per g), which makes excellent therapeutic efficiency at relatively low power density possible. The in vivo results suggested that, compared with single therapy the combined thermo-chemotherapy treatment with Pd@Au-PEG-Pt resulted in complete destruction of the tumor tissue without recurrence, while chemotherapy using Pd@Au-PEG-Pt without irradiation or photothermal treatment using Pd@Au-PEG alone did not. Our work highlights the prospects of a feasible drug delivery strategy of the Pt prodrug by using 2D Pd@Au nanoplates as drug delivery carriers for multimode cancer treatment.Owing to the excellent near infrared (NIR) light absorption and efficient passive targeting toward tumor tissue, two-dimensional (2D) core-shell PEGylated Pd@Au nanoplates have great potential in both photothermal therapy and drug delivery systems. In this work, we successfully conjugate Pd@Au nanoplates with a platinum(iv) prodrug c,c,t-[Pt(NH3)2Cl2(O2CCH2CH2CO2H)2] to obtain a nanocomposite (Pd@Au-PEG-Pt) for combined photothermal-chemotherapy. The

  3. Phosphotyrosine binding domain-dependent upregulation of the platelet-derived growth factor receptor alpha signaling cascade by transforming mutants of Cbl: implications for Cbl's function and oncogenicity.

    PubMed Central

    Bonita, D P; Miyake, S; Lupher, M L; Langdon, W Y; Band, H

    1997-01-01

    Recent studies have demonstrated that Cbl, the 120-kDa protein product of the c-cbl proto-oncogene, serves as a substrate of a number of receptor-coupled tyrosine kinases and forms complexes with SH3 and SH2 domain-containing proteins, pointing to its role in signal transduction. Based on genetic evidence that the Caenorhabditis elegans Cbl homolog, SLI-1, functions as a negative regulator of the LET-23 receptor tyrosine kinase and our demonstration that Cbl's evolutionarily conserved N-terminal transforming region (Cbl-N; residues 1 to 357) harbors a phosphotyrosine binding (PTB) domain that binds to activated ZAP-70 tyrosine kinase, we examined the possibility that oncogenic Cbl mutants may activate mitogenic signaling by deregulating cellular tyrosine kinase machinery. Here, we show that expression of Cbl-N and two other transforming Cbl mutants (CblY368 delta and Cbl366-382 delta or Cb170Z), but not wild-type Cbl, in NIH 3T3 fibroblasts leads to enhancement of endogenous tyrosine kinase signaling. We identified platelet-derived growth factor receptor alpha (PDGFR alpha) as one target of mutant Cbl-induced deregulation. In mutant Cbl transfectants, PDGFR alpha was hyperphosphorylated and constitutively complexed with a number of SH2 domain-containing proteins. PDGFR alpha hyperphosphorylation and enhanced proliferation of mutant Cbl-transfected NIH 3T3 cells were drastically reduced upon serum starvation, and PDGF-AA substituted for the maintenance of these traits. PDGF-AA stimulation of serum-starved Cbl transfectants induced the in vivo association of transfected Cbl proteins with PDGFR alpha. In vitro, Cbl-N directly bound to PDGFR alpha derived from PDGF-AA-stimulated cells but not to that from unstimulated cells, and this binding was abrogated by a point mutation (G306E) corresponding to a loss-of-function mutation in SLI-1. The Cbl-N/G306E mutant protein, which failed to induce enhanced growth and transformation of NIH 3T3 cells, also failed to induce

  4. Targeted delivery of a model immunomodulator to the lymphatic system: comparison of alkyl ester versus triglyceride mimetic lipid prodrug strategies.

    PubMed

    Han, Sifei; Quach, Tim; Hu, Luojuan; Wahab, Anisa; Charman, William N; Stella, Valentino J; Trevaskis, Natalie L; Simpson, Jamie S; Porter, Christopher J H

    2014-03-10

    A lipophilic prodrug approach has been used to promote the delivery of a model immunomodulator, mycophenolic acid (MPA), to the lymphatic system after oral administration. Lymphatic transport was employed to facilitate enhanced drug uptake into lymphocytes, as recent studies demonstrate that targeted drug delivery to lymph resident lymphocytes may enhance immunomodulatory effects. Two classes of lymph-directing prodrugs were synthesised. Alkyl chain derivatives (octyl mycophenolate, MPA-C8E; octadecyl mycophenolate, MPA-C18E; and octadecyl mycophenolamide, MPA-C18AM), to promote passive partitioning into lipids in lymphatic transport pathways, and a triglyceride mimetic prodrug (1,3-dipalmitoyl-2-mycophenoloyl glycerol, 2-MPA-TG) to facilitate metabolic integration into triglyceride deacylation-reacylation pathways. Lymphatic transport, lymphocyte uptake and plasma pharmacokinetics were assessed in mesenteric lymph and carotid artery cannulated rats following intraduodenal infusion of lipid-based formulations containing MPA or MPA prodrugs. Patterns of prodrug hydrolysis in rat digestive fluid, and cellular re-esterification in vivo, were evaluated to examine the mechanisms responsible for lymphatic transport. Poor enzyme stability and low absorption appeared to limit lymphatic transport of the alkyl derivatives, although two of the three alkyl chain prodrugs - MPA-C18AM (6-fold) and MPA-C18E (13-fold) still increased lymphatic drug transport when compared to MPA. In contrast, 2-MPA-TG markedly increased lymphatic drug transport (80-fold) and drug concentrations in lymphocytes (103-fold), and this was achieved via biochemical incorporation into triglyceride deacylation-reacylation pathways. The prodrug was hydrolysed rapidly to 2-mycophenoloyl glycerol (2-MPA-MG) in the presence of rat digestive fluid, and 2-MPA-MG was subsequently re-esterified in the enterocyte with oleic acid (most likely originating from the co-administered formulation) prior to accessing the

  5. Design, Synthesis and Biological Evaluation of Brain-Targeted Thiamine Disulfide Prodrugs of Ampakine Compound LCX001.

    PubMed

    Xiao, Dian; Meng, Fan-Hua; Dai, Wei; Yong, Zheng; Liu, Jin-Qiu; Zhou, Xin-Bo; Li, Song

    2016-01-01

    Ampakine compounds have been shown to reverse opiate-induced respiratory depression by activation of amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) glutamate receptors. However, their pharmacological exploitations are hindered by low blood-brain barrier (BBB) permeability and limited brain distribution. Here, we explored whether thiamine disulfide prodrugs with the ability of "lock-in" can be used to solve these problems. A series of thiamine disulfide prodrugs 7a-7f of ampakine compound LCX001 was synthesized and evaluated. The trials in vitro showed that prodrugs 7e, 7d, 7f possessed a certain stability in plasma and quickly decomposed in brain homogenate by the disulfide reductase. In vivo, prodrug 7e decreased the peripheral distribution of LCX001 and significantly increased brain distribution of LCX001 after i.v. administration. This compound showed 2.23- and 3.29-fold greater increases in the AUC0-t and MRT0-t of LCX001 in brain, respectively, than did LCX001 itself. A preliminary pharmacodynamic study indicated that the required molar dose of prodrug 7e was only one eighth that of LCX001 required to achieve the same effect in mice. These findings provide an important reference to evaluate the clinical outlook of ampakine compounds. PMID:27089316

  6. Pharmacokinetic studies and LC-MS/MS method development of ganciclovir and dipeptide monoester prodrugs in Sprague Dawley rats.

    PubMed

    Gunda, Sriram; Earla, Ravinder; Cholkar, Kishore; Mitra, Ashim K

    2015-09-01

    Ganciclovir (GCV) is utilized as an anti-herpetic agent. Reports from our laboratory have suggested that dipeptide ester prodrugs of GCV exhibit high affinity towards the oligopeptide transporter hPEPT1 and therefore seem to be promising candidates for the treatment of oral herpes virus infections. In this study, we have examined the bio-availability of a dipeptide prodrug of GCV after oral administration in jugular cannulated Sprague-Dawley rats. A new bio-analytical method was developed with Q-TRAP liquid chromatography tandem mass spectroscopy (LC-MS/MS) for simultaneous analysis of GCV, Valine-GCV (VGCV) and Tyrosine-Valine-GCV (YVGCV). Acyclovir (ACV) was used as an internal standard in the analysis. Area under plasma-concentration time curves for total concentration of GCV after oral administration of YVGCV was found to be approximately 200 % more than that of GCV following intestinal absorption. A complete conversion of the dipeptide prodrug (YVGCV) to parent compound, GCV, by hepatic first-pass metabolism was evident due to the absence of intermediate metabolite VGCV and administered prodrug YVGCV. The dipeptide prodrugs of GCV exhibit higher systemic availability of regenerated GCV upon oral administration and thus seem to be promising drug candidate in the treatment of systemic herpes infections. PMID:24943988

  7. Synthesis of novel pregnane-diosgenin prodrugs via Ring A and Ring A connection: A combined experimental and theoretical studies

    NASA Astrophysics Data System (ADS)

    Sethi, Arun; Singh, Ranvijay Pratap; Shukla, Dolly; Singh, Praveer

    2016-12-01

    Novel pregnane-diosgenin prodrugs have been synthesized. The route involved preparation of 3β-25R-spirost-5ene 3yl-benzoate-2-carboxylic acid (2) by the esterification of diosgenin (1) with phthalic anhydride. The pregnane-diosgenin prodrugs 5 &6 were synthesized by treating 3β-25R-spirost-5ene 3yl-benzoate-2-carboxylic acid (2) with 3β-hydroxy16α-methoxy pregn-5-ene-20-one (3) and 3β-hydroxypregn-5, 16-diene-20-one (4) respectively. The synthesized compounds have been characterized with the help of spectroscopic techniques like 1H, 13C NMR, FT-IR, UV-visible spectroscopy and mass spectrometry. Density functional theory (DFT) with B3LYP functional and 6-31G (d, p) basis set has been used for the Quantum chemical calculations. UV-Vis spectra of the synthesized compounds were also recorded and electronic properties such as frontier orbitals and band gap energies were calculated by TD-DFT approach. Intramolecular interactions have been identified by AIM approach and vibrational wavenumbers have been calculated using DFT method. The reactivity and reactive site within the synthesized prodrugs were examined with reactivity descriptors (global and local). Dipole moment, polarizability and first static hyperpolarizability were calculated to get a better insight of the properties of synthesized prodrugs. The probable reaction paths of prodrugs were calculated with molecular electrostatic potential (MEP) surface analysis.

  8. Chirally Pure Prodrugs and Their Converting Enzymes Lead to High Supersaturation and Rapid Transcellular Permeation of Benzodiazepines.

    PubMed

    Kapoor, Mamta; Cheryala, Narsihmulu; Rautiola, Davin; Georg, Gunda I; Cloyd, James C; Siegel, Ronald A

    2016-08-01

    Water-soluble prodrugs can be rapidly converted by enzymes to hydrophobic drugs, whose aqueous thermodynamic solubilities are low, but are maintained in aqueous solution at supersaturated concentrations due to slow precipitation kinetics. Recently, we investigated avizafone (AVF) in combination with Aspergillus oryzae protease as a prodrug/enzyme system intended to produce supersaturated diazepam (DZP). Several fold enhancement of permeation of supersaturated DZP across Madin-Darby canine kidney II-wild type (MDCKII-wt) monolayers was observed, compared to saturated DZP solutions. However, prodrug conversion was incomplete, putatively due to partial racemization of AVF and stereoselectivity of A oryzae protease. Here we report synthesis of chirally pure AVF, and demonstrate complete conversion to supersaturated DZP followed by complete DZP permeation at enhanced rates across MDCKII-wt cell monolayers. We also synthesized, for the first time, a chirally pure prodrug of midazolam (MDZ-pro) and carried out the same sequence of studies. A oryzae protease was identified as a benign and efficient activating enzyme for MDZ-pro. The MDZ-pro/A oryzae protease system showed greater than 25-fold increase in absorption rate of MDZ across MDCKII-wt monolayers, compared to saturated MDZ. Such chirally pure prodrug/enzyme systems are promising candidates for efficient intranasal delivery of benzodiazepine drugs used in the treatment of seizure emergencies. PMID:27342435

  9. The peptidomimetic Vasotide targets two retinal VEGF receptors and reduces pathological angiogenesis in murine and nonhuman primate models of retinal disease

    PubMed Central

    Sidman, Richard L.; Li, Jianxue; Lawrence, Matthew; Hu, Wenzheng; Musso, Gary F.; Giordano, Ricardo J.; Cardó-Vila, Marina; Pasqualini, Renata; Arap, Wadih

    2016-01-01

    Blood vessel growth from preexisting vessels (angiogenesis) underlies many severe diseases including major blinding retinal diseases such as retinopathy of prematurity (ROP) and aged macular degeneration (AMD). This observation has driven development of antibody inhibitors that block a central factor in AMD, named vascular endothelial growth factor (VEGF), from binding to its receptors VEGFR-1 and VEGFR-2. However, some patients are insensitive to current anti-VEGF drugs or develop resistance, and the required repeated intravitreal injection of these large molecules is costly and clinically problematic. Here, we have evaluated a small cyclic retro-inverted peptidomimetic, D(Cys-Leu-Pro-Arg-Cys), abbreviated as D(CLPRC), and hereafter named Vasotide, that inhibits retinal angiogenesis by binding selectively to the VEGF receptors, VEGFR-1 and Neuropilin-1 (NRP-1). Delivery of Vasotide in eye drops or via intraperitoneal injection in a laser-induced monkey model of human wet AMD, a mouse genetic knockout model of the AMD subtype called retinal angiomatous proliferation (RAP), and a mouse oxygen-induced model of retinopathy of prematurity (ROP) markedly decreased retinal angiogenesis in all three animal models. This prototype drug candidate is a promising new dual receptor inhibitor of the VEGF ligand with potential for translation into safer, less invasive applications to combat pathological angiogenesis in retinal disorders. PMID:26468327

  10. Structural basis for recognition of Emi2 by Polo-like kinase 1 and development of peptidomimetics blocking oocyte maturation and fertilization

    PubMed Central

    Jia, Jia-Lin; Han, Young-Hyun; Kim, Hak-Cheol; Ahn, Mija; Kwon, Jeong-Woo; Luo, Yibo; Gunasekaran, Pethaiah; Lee, Soo-Jae; Lee, Kyung S.; Kyu Bang, Jeong; Kim, Nam-Hyung; Namgoong, Suk

    2015-01-01

    In a mammalian oocyte, completion of meiosis is suspended until fertilization by a sperm, and the cell cycle is arrested by a biochemical activity called cytostatic factor (CSF). Emi2 is one of the CSFs, and it maintains the protein level of maturation promoting factor (MPF) by inhibiting ubiquitin ligase anaphase promoting complex/cyclosome (APC/C). Degradation of Emi2 via ubiquitin-mediated proteolysis after fertilization requires phosphorylation by Polo-like kinase 1 (Plk1). Therefore, recognition and phosphorylation of Emi2 by Plk1 are crucial steps for cell cycle resumption, but the binding mode of Emi2 and Plk1 is poorly understood. Using biochemical assays and X-ray crystallography, we found that two phosphorylated threonines (Thr152 and Thr176) in Emi2 are each responsible for the recruitment of one Plk1 molecule by binding to its C-terminal polo box domain (PBD). We also found that meiotic maturation and meiosis resumption via parthenogenetic activation were impaired when Emi2 interaction with Plk1-PBD was blocked by a peptidomimetic called 103-8. Because of the inherent promiscuity of kinase inhibitors, our results suggest that targeting PBD of Plk1 may be an effective strategy for the development of novel and specific contraceptive agents that block oocyte maturation and/or fertilization. PMID:26459104

  11. Palladium-Mediated Dealkylation of N-Propargyl-Floxuridine as a Bioorthogonal Oxygen-Independent Prodrug Strategy

    NASA Astrophysics Data System (ADS)

    Weiss, Jason T.; Carragher, Neil O.; Unciti-Broceta, Asier

    2015-03-01

    Herein we report the development and biological screening of a bioorthogonal palladium-labile prodrug of the nucleoside analogue floxuridine, a potent antineoplastic drug used in the clinic to treat advanced cancers. N-propargylation of the N3 position of its uracil ring resulted in a vast reduction of its biological activity (~6,250-fold). Cytotoxic properties were bioorthogonally rescued in cancer cell culture by heterogeneous palladium chemistry both in normoxia and hypoxia. Within the same environment, the reported chemo-reversible prodrug exhibited up to 1,450-fold difference of cytotoxicity whether it was in the absence or presence of the extracellular palladium source, underlining the precise modulation of bioactivity enabled by this bioorthogonally-activated prodrug strategy.

  12. Modular adeno-associated virus (rAAV) vectors used for cellular virus-directed enzyme prodrug therapy.

    PubMed

    Hagen, Sven; Baumann, Tobias; Wagner, Hanna J; Morath, Volker; Kaufmann, Beate; Fischer, Adrian; Bergmann, Stefan; Schindler, Patrick; Arndt, Katja M; Müller, Kristian M

    2014-01-01

    The pre-clinical and clinical development of viral vehicles for gene transfer increased in recent years, and a recombinant adeno-associated virus (rAAV) drug took center stage upon approval in the European Union. However, lack of standardization, inefficient purification methods and complicated retargeting limit general usability. We address these obstacles by fusing rAAV-2 capsids with two modular targeting molecules (DARPin or Affibody) specific for a cancer cell-surface marker (EGFR) while simultaneously including an affinity tag (His-tag) in a surface-exposed loop. Equipping these particles with genes coding for prodrug converting enzymes (thymidine kinase or cytosine deaminase) we demonstrate tumor marker specific transduction and prodrug-dependent apoptosis of cancer cells. Coding terminal and loop modifications in one gene enabled specific and scalable purification. Our genetic parts for viral production adhere to a standardized cloning strategy facilitating rapid prototyping of virus directed enzyme prodrug therapy (VDEPT). PMID:24457557

  13. Discovery of BC-01, a novel mutual prodrug (hybrid drug) of ubenimex and fluorouracil as anticancer agent.

    PubMed

    Jiang, Yuqi; Li, Xiaoyang; Hou, Jinning; Huang, Yongxue; Jia, Yuping; Zou, Mingming; Zhang, Jian; Wang, Xuejian; Xu, Wenfang; Zhang, Yingjie

    2016-10-01

    We designed and synthesized a novel mutual prodrug, named BC-01 (3), by integrating ubenimex and Fluorouracil (5-FU) into one molecule based on prior research results that showed that a combination of the aminopeptidase N (CD13) inhibitor, ubenimex, and the cytotoxic antitumor agent, 5-FU, exhibited improved in vitro and in vivo antitumor efficiency. 3 showed potent inhibitory activity against CD13 enzymatic activity. Compared with ubenimex, 3 exhibited more potent anti-angiogenesis effects, and compared with the approved 5-FU prodrug, capecitabine, 3 exhibited more potent tumor growth inhibitory and anti-metastasis effects. Additionally, compared with 5-FU or 5-FU plus ubenimex, 3 also exhibited a superior antitumor efficiency even in our 5-FU-resistant mice model. Other antitumor agents could be conjugated with ubenimex using this strategy to obtain novel mutual prodrugs with promising antitumor potency. PMID:27322756

  14. Design, Synthesis, and Biological Evaluation of Potential Prodrugs Related to the Experimental Anticancer Agent Indotecan (LMP400).

    PubMed

    Lv, Peng-Cheng; Elsayed, Mohamed S A; Agama, Keli; Marchand, Christophe; Pommier, Yves; Cushman, Mark

    2016-05-26

    Indenoisoquinoline topoisomerase I (Top1) inhibitors are a novel class of anticancer agents with two compounds in clinical trials. Recent metabolism studies of indotecan (LMP400) led to the discovery of the biologically active 2-hydroxylated analogue and 3-hydroxylated metabolite, thus providing strategically placed functional groups for the preparation of a variety of potential ester prodrugs of these two compounds. The current study details the design and synthesis of two series of indenoisoquinoline prodrugs, and it also reveals how substituents on the O-2 and O-3 positions of the A ring, which are next to the cleaved DNA strand in the drug-DNA-Top1 ternary cleavage complex, affect Top1 inhibitory activity and cytotoxicity. Many of the indenoisoquinoline prodrugs were very potent antiproliferative agents with GI50 values below 10 nM in a variety of human cancer cell lines. PMID:27097152

  15. The Role of Bystander Effects in the Antitumor Activity of the Hypoxia-Activated Prodrug PR-104

    PubMed Central

    Foehrenbacher, Annika; Patel, Kashyap; Abbattista, Maria R.; Guise, Chris P.; Secomb, Timothy W.; Wilson, William R.; Hicks, Kevin O.

    2013-01-01

    Activation of prodrugs in tumors (e.g., by bioreduction in hypoxic zones) has the potential to generate active metabolites that can diffuse within the tumor microenvironment. Such “bystander effects” may offset spatial heterogeneity in prodrug activation but the relative importance of this effect is not understood. Here, we quantify the contribution of bystander effects to antitumor activity for the first time, by developing a spatially resolved pharmacokinetic/pharmacodynamic (SR-PK/PD) model for PR-104, a phosphate ester pre-prodrug that is converted systemically to the hypoxia-activated prodrug PR-104A. Using Green’s function methods we calculated concentrations of oxygen, PR-104A and its active metabolites, and resultant cell killing, at each point of a mapped three-dimensional tumor microregion. Model parameters were determined in vitro, using single cell suspensions to determine relationships between PR-104A metabolism and clonogenic cell killing, and multicellular layer (MCL) cultures to measure tissue diffusion coefficients. LC-MS/MS detection of active metabolites in the extracellular medium following exposure of anoxic single cell suspensions and MCLs to PR-104A confirmed that metabolites can diffuse out of cells and through a tissue-like environment. The SR-PK/PD model estimated that bystander effects contribute 30 and 50% of PR-104 activity in SiHa and HCT116 tumors, respectively. Testing the model by modulating PR-104A-activating reductases and hypoxia in tumor xenografts showed overall clonogenic killing broadly consistent with model predictions. Overall, our data suggest that bystander effects are important in PR-104 antitumor activity, although their reach may be limited by macroregional heterogeneity in hypoxia and reductase expression in tumors. The reported computational and experimental techniques are broadly applicable to all targeted anticancer prodrugs and could be used to identify strategies for rational prodrug optimization. PMID

  16. Design of Fexofenadine Prodrugs Based on Tissue-Specific Esterase Activity and Their Dissimilar Recognition by P-Glycoprotein.

    PubMed

    Ohura, Kayoko; Nakada, Yuichiro; Kotani, Shunsuke; Imai, Teruko

    2015-09-01

    The aim of this study was to develop a suitable prodrug for fexofenadine (FXD), a model parent drug, that is resistant to intestinal esterase but converted to FXD by hepatic esterase. Carboxylesterases (CESs), human carboxylesterase 1 (hCE1) and human carboxylesterase 2 (hCE2), are the major esterases in human liver and intestine, respectively. These two CESs show quite different substrate specificities, and especially, hCE2 poorly hydrolyzes prodrugs with large acyl groups. FXD contains a carboxyl group and is poorly absorbed because of low membrane permeability and efflux by P-glycoprotein (P-gp). Therefore, two potential FXD prodrugs, ethyl-FXD and 2-hydroxyethyl-FXD, were synthesized by substitution of the carboxyl group in FXD. Both derivatives were resistant to intestinal hydrolysis, indicating their absorption as intact prodrugs. Ethyl-FXD was hydrolyzed by hepatic hCE1, but 2-hydroxyethyl-FXD was not. Both derivatives showed high membrane permeability in human P-gp-negative LLC-PK1 cells. In LLC-GA5-COL300 cells overexpressing human P-gp, ethyl-FXD was transported by P-gp, but its efflux was easily saturated. Whereas 2-hydroxyethyl-FXD showed more efficient P-gp-mediated transport than FXD. Although the structure of 2-hydroxyethyl-FXD only differs from ethyl-FXD by substitution of a hydroxyl group, 2-hydroxyethyl-FXD is unsuitable as a prodrug. However, ethyl-FXD is a good candidate prodrug because of good intestinal absorption and hepatic conversion by hCE1. PMID:25953731

  17. Novel prodrug-like fusion toxin with protease-sensitive bioorthogonal PEGylation for tumor targeting.

    PubMed

    Stefan, Nikolas; Zimmermann, Martina; Simon, Manuel; Zangemeister-Wittke, Uwe; Plückthun, Andreas

    2014-12-17

    Highly potent biotoxins like Pseudomonas exotoxin A (ETA) are attractive payloads for tumor targeting. However, despite replacement of the natural cell-binding domain of ETA by tumor-selective antibodies or alternative binding proteins like designed ankyrin repeat proteins (DARPins) the therapeutic window of such fusion toxins is still limited by target-independent cellular uptake, resulting in toxicity in normal tissues. Furthermore, the strong immunogenicity of the bacterial toxin precludes repeated administration in most patients. Site-specific modification to convert ETA into a prodrug-like toxin which is reactivated specifically in the tumor, and at the same time has a longer circulation half-life and is less immunogenic, is therefore appealing. To engineer a prodrug-like fusion toxin consisting of the anti-EpCAM DARPin Ec1 and a domain I-deleted variant of ETA (ETA″), we used strain-promoted azide alkyne cycloaddition for bioorthogonal conjugation of linear or branched polyethylene glycol (PEG) polymers at defined positions within the toxin moiety. Reversibility of the shielding was provided by a designed peptide linker containing the cleavage site for the rhinovirus 3C model protease. We identified two distinct sites, one within the catalytic domain and one close to the C-terminal KDEL sequence of Ec1-ETA″, simultaneous PEGylation of which resulted in up to 1000-fold lower cytotoxicity in EpCAM-positive tumor cells. Importantly, the potency of the fusion toxin was fully restored by proteolytic unveiling. Upon systemic administration in mice, PEGylated Ec1-ETA″ was much better tolerated than Ec1-ETA″; it showed a longer circulation half-life and an almost 10-fold increased area under the curve (AUC). Our strategy of engineering prodrug-like fusion toxins by bioorthogonal veiling opens new possibilities for targeting tumors with more specificity and efficacy. PMID:25350699

  18. Boronic prodrug of endoxifen as an effective hormone therapy for breast cancer.

    PubMed

    Zhang, Changde; Zhong, Qiu; Zhang, Qiang; Zheng, Shilong; Miele, Lucio; Wang, Guangdi

    2015-07-01

    As a prodrug, tamoxifen is activated by the P450 enzyme CYP2D6 that is responsible for converting it to the active metabolites, 4-hydroxytamoxifen and endoxifen. Patients with genetic polymorphisms of CYP2D6 may not receive the full benefit of tamoxifen therapy. There is increasing evidence that poor metabolizer patients have lower plasma concentrations of endoxifen and suffer worse disease outcome, although some clinical studies reported no correlation between CYP2D6 polymorphism and tamoxifen therapy outcome. Endoxifen is currently undergoing clinical trials as a potentially improved and more potent SERM (Selective Estrogen Receptor Modulator) for endocrine therapy that is independent of CYP2D6 status in patients. However, direct administration of endoxifen may present the problem of low bioavailability due to its rapid first-pass metabolism via O-glucuronidation. We have designed and synthesized ZB483, a boronic prodrug of endoxifen suitable for oral administration with greatly enhanced bioavailability by increasing the concentration of endoxifen in mouse blood. Our study demonstrated that ZB483 potently inhibited growth of ER+ breast cancer cells in vitro and was efficiently converted to endoxifen in cell culture media by oxidative deboronation. This metabolic conversion is equally efficient in vivo as indicated in the pharmacokinetic study in mice. Moreover, when administered at the same dose, oral ZB483 afforded a 30- to 40-fold higher plasma level of endoxifen in mice than oral administration of endoxifen. The significantly enhanced bioavailability of endoxifen conferred by the boronic prodrug was further validated in an in vivo efficacy study. ZB483 was demonstrated to be more efficacious than endoxifen in inhibiting xenograft tumor growth in mice at equal dosage but more so at lower dosage. Together, these preclinical studies demonstrate that ZB483 is a promising endocrine therapy agent with markedly enhanced bioavailability in systemic circulation and

  19. Synthesis and in vivo evaluation of prodrugs of 9-[2-(phosphonomethoxy)ethoxy]adenine.

    PubMed

    Serafinowska, H T; Ashton, R J; Bailey, S; Harnden, M R; Jackson, S M; Sutton, D

    1995-04-14

    A number of esters and amides of the anti-HIV nucleotide analogue 9-[2-(phosphonomethoxy)-ethoxy]adenine (1) have been synthesized as potential prodrugs and evaluated for oral bioavailability in mice. Dialkyl esters 17-20 were prepared via a Mitsunobu coupling of alcohols 8-11 with 9-hydroxypurine 12 whereas (acyloxy)alkyl esters 25-33 and bis-[(alkoxycarbonyl)methyl] and bis(amidomethyl) esters 34-39 were obtained by reaction of 1 with a suitable alkylating agent. Phosphonodichloridate chemistry was employed for the preparation of dialkyl and diaryl esters 42-65, and bis(phosphonoamidates) 66 and 67. Following oral administration to mice, most of the dialkyl esters 17-20 were well-absorbed and then converted to the corresponding monoesters, but minimal further metabolism to 1 occurred. Bis[(pivaloyloxy)methyl] ester 25 displayed an oral bioavailability of 30% that was 15-fold higher than the bioavailability observed after dosing of 1. Methyl substitution at the alpha carbon of the bis[(pivaloyloxy)methyl] ester 25 (33) increased the oral bioavailability of 1 to 74%. Some of the diaryl esters also showed improved absorption properties in comparison with that of 1. In particular, the crystalline hydrochloride salt of diphenyl ester 55 was well-absorbed and efficiently converted to the parent compound with an oral bioavailability of 50%. On the basis of these results as well as the physicochemical properties of the prodrugs and their stability in mouse duodenal contents, the hydrochloride salt of diphenyl ester 55 was identified as the preferred prodrug of 1. PMID:7731022

  20. Leveraging Hypoxia-Activated Prodrugs to Prevent Drug Resistance in Solid Tumors.

    PubMed

    Lindsay, Danika; Garvey, Colleen M; Mumenthaler, Shannon M; Foo, Jasmine

    2016-08-01

    Experimental studies have shown that one key factor in driving the emergence of drug resistance in solid tumors is tumor hypoxia, which leads to the formation of localized environmental niches where drug-resistant cell populations can evolve and survive. Hypoxia-activated prodrugs (HAPs) are compounds designed to penetrate to hypoxic regions of a tumor and release cytotoxic or cytostatic agents; several of these HAPs are currently in clinical trial. However, preliminary results have not shown a survival benefit in several of these trials. We hypothesize that the efficacy of treatments involving these prodrugs depends heavily on identifying the correct treatment schedule, and that mathematical modeling can be used to help design potential therapeutic strategies combining HAPs with standard therapies to achieve long-term tumor control or eradication. We develop this framework in the specific context of EGFR-driven non-small cell lung cancer, which is commonly treated with the tyrosine kinase inhibitor erlotinib. We develop a stochastic mathematical model, parametrized using clinical and experimental data, to explore a spectrum of treatment regimens combining a HAP, evofosfamide, with erlotinib. We design combination toxicity constraint models and optimize treatment strategies over the space of tolerated schedules to identify specific combination schedules that lead to optimal tumor control. We find that (i) combining these therapies delays resistance longer than any monotherapy schedule with either evofosfamide or erlotinib alone, (ii) sequentially alternating single doses of each drug leads to minimal tumor burden and maximal reduction in probability of developing resistance, and (iii) strategies minimizing the length of time after an evofosfamide dose and before erlotinib confer further benefits in reduction of tumor burden. These results provide insights into how hypoxia-activated prodrugs may be used to enhance therapeutic effectiveness in the clinic. PMID

  1. Preparation and characterization of microemulsion formulations of nicotinic acid and its prodrugs for transdermal delivery.

    PubMed

    Tashtoush, Bassam M; Bennamani, Amina N; Al-Taani, Bashar M

    2013-01-01

    At pharmacological doses, nicotinic acid has a lipid-regulating effect and is in use clinically for that purpose. However, despite of all features, its utility is strongly limited by several disadvantages such as, extensive hepatic metabolism and flushing. Transdermal delivery of nicotinic acid may, therefore, be the solution to reducing side effects associated with oral administration, and to maintaining constant therapeutic blood levels for longer duration. The aim of this investigation was to develop a suitable formulation or select a suitable vehicle for the transdermal delivery of highly lipophilic prodrugs of nicotinic acid (dodecyl and myristyl nicotinate) designed to deliver nicotinic acid through skin without causing vasodilatation and flushing and optimizing its delivery to the blood stream. A microemulsion system and penetration enhancers have been attempted in this study. The microemulsion system was composed of isopropyl myristate (IPM), water and a 4:1 (w/w) mixture of Labrasol and Peceol where a pseudoternary phase diagram was constructed. Furthermore, the microemulsion formulations with different component ratios were characterized by determination of conductivity, pH, particle size, viscosity and refractive index. According to the particle size analysis, conductivity and viscosity measurements, the microemulsion formulations that formed were of oil-in-water type. The transdermal permeability of nicotinic acid and its prodrugs was evaluated in vitro using Franz diffusion cells fitted with mice skin and nicotinic acid concentration was analyzed by high performance liquid chromatography. A theoretical design of percutaneous penetration optimization in which prodrugs derivation and enhancer application are combined based on the skin diffusion model was experimentally verified. The selected formulations seemed promising for developing a transdermal drug delivery system of nicotinic acid from dodecyl nicotinate that would offer advantages like possible

  2. Leveraging Hypoxia-Activated Prodrugs to Prevent Drug Resistance in Solid Tumors

    PubMed Central

    Lindsay, Danika; Garvey, Colleen M.; Mumenthaler, Shannon M.; Foo, Jasmine

    2016-01-01

    Experimental studies have shown that one key factor in driving the emergence of drug resistance in solid tumors is tumor hypoxia, which leads to the formation of localized environmental niches where drug-resistant cell populations can evolve and survive. Hypoxia-activated prodrugs (HAPs) are compounds designed to penetrate to hypoxic regions of a tumor and release cytotoxic or cytostatic agents; several of these HAPs are currently in clinical trial. However, preliminary results have not shown a survival benefit in several of these trials. We hypothesize that the efficacy of treatments involving these prodrugs depends heavily on identifying the correct treatment schedule, and that mathematical modeling can be used to help design potential therapeutic strategies combining HAPs with standard therapies to achieve long-term tumor control or eradication. We develop this framework in the specific context of EGFR-driven non-small cell lung cancer, which is commonly treated with the tyrosine kinase inhibitor erlotinib. We develop a stochastic mathematical model, parametrized using clinical and experimental data, to explore a spectrum of treatment regimens combining a HAP, evofosfamide, with erlotinib. We design combination toxicity constraint models and optimize treatment strategies over the space of tolerated schedules to identify specific combination schedules that lead to optimal tumor control. We find that (i) combining these therapies delays resistance longer than any monotherapy schedule with either evofosfamide or erlotinib alone, (ii) sequentially alternating single doses of each drug leads to minimal tumor burden and maximal reduction in probability of developing resistance, and (iii) strategies minimizing the length of time after an evofosfamide dose and before erlotinib confer further benefits in reduction of tumor burden. These results provide insights into how hypoxia-activated prodrugs may be used to enhance therapeutic effectiveness in the clinic. PMID

  3. Esterase-activatable β-lapachone prodrug micelles for NQO1-targeted lung cancer therapy

    PubMed Central

    Ma, Xinpeng; Huang, Xiumei; Moore, Zachary; Huang, Gang; Kilgore, Jessica A.; Wang, Yiguang; Hammer, Suntrea; Williams, Noelle S.; Boothman, David A.; Gao, Jinming

    2016-01-01

    Lung cancer is one of the most lethal forms of cancer and current chemotherapeutic strategies lack broad specificity and efficacy. Recently, β-lapachone (β-lap) was shown to be highly efficacious in killing non-small cell lung cancer (NSCLC) cells regardless of their p53, cell cycle and caspase status. Pre-clinical and clinical use of β-lap (clinical form, ARQ501 or 761) is hampered by poor pharmacokinetics and toxicity due to hemolytic anemia. Here, we report the development and preclinical evaluation of β-lap prodrug nanotherapeutics consisting of diester derivatives of β-lap encapsulated in biocompatible and biodegradable poly(ethylene glycol)-b-poly(d,l-lactic acid) (PEG-b-PLA) micelles. Compared to the parent drug, diester derivatives of β-lap showed higher drug loading densities inside PEG-b-PLA micelles. After esterase treatment, micelle-delivered β-lap-dC3 and -dC6 prodrugs were converted to β-lap. Cytotoxicity assays using A549 and H596 lung cancer cells showed that both micelle formulations maintained NAD(P)H:quinone oxidoreductase 1 (NQO1)-dependent cytotoxicity. However, antitumor efficacy study of β-lap-dC3 micelles against orthotopic A549 NSCLC xenograft-bearing mice showed significantly greater long-term survival over β-lap-dC6 micelles or β-lap-HPβCD complexes. Improved therapeutic efficacy of β-lap-dC3 micelles correlated with higher area under the concentration-time curves of β-lap in tumors, and enhanced pharmacodynamic endpoints (e.g., PARP1 hyperactivation, γH2AX, and ATP depletion). β-Lap-dC3 prodrug micelles provide a promising strategy for NQO1-targeted therapy of lung cancer with improved safety and antitumor efficacy. PMID:25542645

  4. Lipophilic nalmefene prodrugs to achieve a one-month sustained release.

    PubMed

    Gaekens, Tim; Guillaume, Michel; Borghys, Herman; De Zwart, Loeckie L; de Vries, Ronald; Embrechts, Roger C A; Vermeulen, An; Megens, Anton A H P; Leysen, Josée E; Herdewijn, Piet; Annaert, Pieter P; Atack, John R

    2016-06-28

    Nalmefene is an opioid antagonist which as a once-a-day tablet formulation has recently been approved for reducing ethanol intake in alcoholic subjects. In order to address the compliance issue in this patient population, a number of potential nalmefene prodrugs were synthesized with the aim of providing a formulation that could provide plasma drug concentrations in the region of 0.5-1.0ng/mL for a one-month period when dosed intramuscular to dogs or minipigs. In an initial series of studies, three different lipophilic nalmefene derivatives were evaluated: the palmitate (C16), the octadecyl glutarate diester (C18-C5) and the decyl carbamate (CB10). They were administered intramuscularly to dogs in a sesame oil solution at a dose of 1mg-eq. nalmefene/kg. The decyl carbamate was released relatively quickly from the oil depot and its carbamate bond was too stable to be used as a prodrug. The other two derivatives delivered a fairly constant level of 0.2-0.3ng nalmefene/mL plasma for one month and since there was no significant difference between these two, the less complex palmitate monoester was chosen to demonstrate that dog plasma nalmefene concentrations were dose-dependent at 1, 5 and 20mg-eq. nalmefene/kg. In a second set of experiments, the effect of the chain length of the fatty acid monoester promoieties was examined. The increasingly lipophilic octanoate (C8), decanoate (C10) and dodecanoate (C12) derivatives were evaluated in dogs and in minipigs, at a dose of 5mg-eq. nalmefene/kg and plasma nalmefene concentrations were measured over a four-week period. The pharmacokinetic profiles were very similar in both species with Cmax decreasing and Tmax increasing with increasing fatty acid chain length and the target plasma concentrations (0.5-1.0ng/mL over a month-long period) were achieved with the dodecanoate (C12) prodrug. These data therefore demonstrate that sustained plasma nalmefene concentrations can be achieved in both dog and minipig using nalmefene

  5. Synthesis and evaluation of 18F labeled FET prodrugs for tumor imaging

    PubMed Central

    Wang, Limin; Lieberman, Brian P.; Ploessl, Karl; Kung, Hank F.

    2013-01-01

    Introduction O-(2-[18F]fluoroethyl)-L-tyrosine (FET, [18F]1) is a useful amino-acid-based imaging agent for brain tumors. This paper reports the synthesis and evaluation of three FET prodrugs, O-(2-[18F]fluoroethyl)-L-tyrosyl-L-glycine (FET-Gly, [18F]2), O-(2-[18F]fluoroethyl)-L-tyrosyl-L-alanine (FET-Ala, [18F]3) and N-acetyl O-(2-[18F]fluoroethyl)-L-tyrosine (AcFET, [18F]4), which could be readily hydrolyzed to FET in vivo for tumor imaging. We investigated their metabolism in the blood and imaging properties in comparison to FET ([18F]1). Methods Three new [18F]FET derivatives, 2 – 4, were prepared from their corresponding tosylate-precursors through nucleophilic fluorination and subsequent deprotection reactions. In vitro uptake studies were carried out in 9L glioma cancer cell lines. In vitro and in vivo hydrolysis studies were conducted to evaluate the hydrolysis of FET prodrugs in blood and in Fisher 344 rats. Biodistribution and PET imaging studies were then performed in rats bearing 9L tumors. Results New FET prodrugs were prepared with 3 – 28 % decay corrected radiochemical yields, good enantiomeric purity (> 95 %) and high radiochemical purity (> 95 %). FET-Gly ([18F]2), FET-Ala ([18F]3), and AcFET ([18F]4) exhibited negligible uptake in comparison to the high uptake of FET ([18F]1) in 9L cells. Metabolism studies of FET-Gly ([18F]2), FET-Ala ([18F]3), and AcFET ([18F]4) in rat and human blood showed that FET-Ala ([18F]3) was hydrolyzed to FET ([18F]1) faster than FET-Gly ([18F]2) or AcFET ([18F]4). Most of the FET-Ala (79 %) was converted to FET ([18F]1) within 5 min in blood in vivo. Biodistribution studies demonstrated that FET-Ala ([18F]3) displayed the highest tumor uptake. The tumor-to-background ratios of FET-Ala ([18F]3) and FET ([18F]1) were comparable and appeared to be better than those of FET-Gly ([18F]2) and AcFET ([18F]4). PET imaging studies showed that both FET ([18F]1) and FET-Ala ([18F]3) could visualize tumors effectively, and that

  6. Controllable synthesis of polymerizable ester and amide prodrugs of acyclovir by enzyme in organic solvent.

    PubMed

    Li, Xia; Wu, Qi; Lv, De-shui; Lin, Xian-fu

    2006-05-15

    A facile control of the acylation position at the primary hydroxyl and amino of acyclovir, respectively, was achieved and five polymerizable acyclovir prodrugs were synthesized. Various reaction conditions were studied in detail. Thus, lipase acrylic resin from Candida antarctica (CAL-B) in pyridine or acetone showed high chemo-selectivity toward the primary hydroxyl of acyclovir. However, lipase PS 'Amano' (PS) in DMSO selectively acylated the amino group. The selectivity of PS could be adjusted by changing reaction solvents. The acyclovir vinyl derivatives obtained would be important monomers used for the preparation of macromolecular nucleoside drugs. PMID:16431120

  7. Double ester prodrugs of FR900098 display enhanced in-vitro antimalarial activity.

    PubMed

    Wiesner, Jochen; Ortmann, Regina; Jomaa, Hassan; Schlitzer, Martin

    2007-12-01

    Fosmidomycin and FR900098 are inhibitors of the 1-deoxy-D-xylulose 5-phosphate reductoisomerase (DXR; IspC), a key enzyme of the mevalonate-independent isoprenoid biosynthesis pathway. We have determined the in-vitro antimalarial activity of two double ester prodrugs 2, 3 in direct comparison with the unmodified FR900098 1 against intraerythrocytic forms of Plasmodium falciparum. Temporarily masking the polar properties of the phosphonate moiety of the DXR inhibitor FR900098 1 enhanced not only its oral bioavailability but also the intrinsic activity of this series against the parasites. PMID:17994601

  8. Synthesis/biological evaluation of hydroxamic acids and their prodrugs as inhibitors for Botulinum neurotoxin A light chain

    PubMed Central

    Seki, Hajime; Pellett, Sabine; Šilhár, Peter; Stowe, G. Neil; Blanco, Beatriz; Lardy, Matthew; Johnson, Eric A.; Janda, Kim D.

    2014-01-01

    Botulinum neurotoxin A (BoNT/A) is the most potent toxin known. Unfortunately, it is also a potential bioweapon in terrorism, which is without an approved therapeutic treatment once cellular intoxication takes place. Previously, we reported how hydroxamic acid prodrug carbamates increased cellular uptake, which translated to successful inhibition of this neurotoxin. Building upon this research, we detail BoNT/A protease molecular modeling studies accompanied by the construction of small library of hydroxamic acids based on 2,4-dichlorocinnamic hydroxamic acid scaffold and their carbamate prodrug derivatization along with the evaluation of these molecules in both enzymatic and cellular models. PMID:24360826

  9. Synthesis and biological properties of prodrugs of (S)-3-(adenin-9-yl)-2-(phosphonomethoxy)propanoic acid.

    PubMed

    Kaiser, Martin Maxmilian; Poštová-Slavětínská, Lenka; Dračínský, Martin; Lee, Yu-Jen; Tian, Yang; Janeba, Zlatko

    2016-01-27

    The lack of antiviral activity of recently described (S)-3-(adenin-9-yl)-2-(phosphonomethoxy)propanoic acid, or (S)-CPMEA in brief, has been speculated to possibly be due to the increased hydrophilicity of the molecule and, thus, by its limited cellular permeability. Efficient syntheses of novel lipophilic prodrugs of (S)-CPMEA masking either the carboxylic group or preferably both the phosphonate and carboxylic moieties, have been developed in order to increase bioavailability of the parent compound. Two prodrugs of (S)-CPMEA, namely phosphonate bis-amidate 15 and phenyloxy amidate 16, exhibited pan-genotypic anti-HCV activity at submicromolar concentrations. PMID:26706348

  10. Synthesis and evaluation of N-acylsulfonamide and N-acylsulfonylurea prodrugs of a prostacyclin receptor agonist.

    PubMed

    Nakamura, Akio; Yamada, Tetsuhiro; Asaki, Tetsuo

    2007-12-15

    N-Acylsulfonamide and N-acylsulfonylurea derivatives of the carboxylic acid prostacyclin receptor agonist 1 were synthesized and their potential as prodrug forms of the carboxylic acid was evaluated in vitro and in vivo. These compounds were converted to the active compound 1 by hepatic microsomes from rats, dogs, monkeys, and humans, and some of the compounds were shown to yield sustained plasma concentrations of 1 when they were orally administered to monkeys. These types of analogues, including NS-304 (2a), are potentially useful prodrugs of 1. PMID:17881233

  11. Design, synthesis, and evaluation of water-soluble morpholino-decorated paclitaxel prodrugs with remarkably decreased toxicity.

    PubMed

    Feng, Siliang; Chen, Kuncheng; Wang, Chenhong; Jiang, Xifeng; Dong, Huajin; Gong, Zehui; Liu, Keliang

    2016-08-01

    Novel water-soluble paclitaxel prodrugs were designed and synthesized by introducing morpholino groups through different linkers. These derivatives showed 400-20,000-times greater water solubility than paclitaxel as well as comparable activity in MCF-7 and HeLa cell lines. The prodrug PM4 was tested in the S-180 tumor mouse model, with paclitaxel as the positive control. The results showed that PM4 had comparable antitumor activity as paclitaxel, with tumor inhibition of 54% versus 56%, and remarkably decreased toxicity. The survival rate of treated mice was 8/8 in the PM4 group, compared to 3/8 in the paclitaxel group. PMID:27311893

  12. First-In-Human Study Demonstrating Tumor-Angiogenesis by PET/CT Imaging with 68Ga-NODAGA-THERANOST, a High-Affinity Peptidomimetic for αvβ3 Integrin Receptor Targeting

    PubMed Central

    Baum, Richard P.; Kulkarni, Harshad R.; Müller, Dirk; Danthi, Narasimhan; Kim, Young-Seung; Brechbiel, Martin W.

    2015-01-01

    Abstract 68Ga-NODAGA-THERANOST™ is an αvβ3 integrin antagonist and the first radiolabeled peptidomimetic to reach clinical development for targeting integrin receptors. In this first-in-human study, the feasibility of integrin receptor peptidomimetic positron emission tomography/computed tomography (PET/CT) imaging was confirmed in patients with non-small-cell lung cancer and breast cancer. Methods: Patients underwent PET/CT imaging with 68Ga NODAGA-THERANOST. PET images were analyzed qualitatively and quantitatively and compared to 2-deoxy-2-(18F) fluoro-d-glucose (18F-FDG) findings. Images were obtained 60 minutes postinjection of 300–500 MBq of 68Ga-NODAGA-THERANOST. Results: 68Ga-NODAGA-THERANOST revealed high tumor-to-background ratios (SUVmax=4.8) and uptake at neoangiogenesis sites. Reconstructed fused images distinguished cancers with high malignancy potential and enabled enhanced bone metastasis detection. 18F-FDG-positive lung and lymph node metastases did not show uptake, indicating the absence of neovascularization. Conclusions: 68Ga-NODAGA-THERANOST was found to be safe and effective, exhibiting in this study rapid blood clearance, stability, rapid renal excretion, favorable biodistribution and PK/PD, low irradiation burden (μSv/MBq/μg), and convenient radiolabeling. This radioligand might enable theranostics, that is, a combination of diagnostics followed by the appropriate therapeutics, namely antiangiogenic therapy, image-guided presurgical assessment, treatment response evaluation, prediction of pathologic response, neoadjuvant-peptidomimetic-radiochemotherapy, and personalized medicine strategies. Further clinical trials evaluating 68Ga-NODAGA-THERANOST are warranted. PMID:25945808

  13. Dissociable effects of the prodrug phendimetrazine and its metabolite phenmetrazine at dopamine transporters

    PubMed Central

    Solis, Ernesto; Suyama, Julie A.; Lazenka, Matthew F.; DeFelice, Louis J.; Negus, S. Stevens; Blough, Bruce E.; Banks, Matthew L.

    2016-01-01

    Phendimetrazine (PDM) is a clinically available anorectic and a candidate pharmacotherapy for cocaine addiction. PDM has been hypothesized to function as a prodrug that requires metabolism to the amphetamine-like monoamine transporter substrate phenmetrazine (PM) to produce its pharmacological effects; however, whether PDM functions as an inactive prodrug or has pharmacological activity on its own remains unclear. The study aim was to determine PDM pharmacological mechanisms using electrophysiological, neurochemical, and behavioral procedures. PDM blocked the endogenous basal hDAT (human dopamine transporter) current in voltage-clamped (−60 mV) oocytes consistent with a DAT inhibitor profile, whereas its metabolite PM induced an inward hDAT current consistent with a DAT substrate profile. PDM also attenuated the PM-induced inward current during co-application, providing further evidence that PDM functions as a DAT inhibitor. PDM increased nucleus accumbens dopamine levels and facilitated electrical brain stimulation reinforcement within 10 min in rats, providing in vivo evidence supporting PDM pharmacological activity. These results demonstrate that PDM functions as a DAT inhibitor that may also interact with the pharmacological effects of its metabolite PM. Overall, these results suggest a novel mechanism for PDM therapeutic effects via initial PDM DAT inhibition followed by PM DAT substrate-induced dopamine release. PMID:27514281

  14. Encapsulation of lipophilic kiteplatin Pt(iv) prodrugs in PLGA-PEG micelles.

    PubMed

    Margiotta, Nicola; Savino, Salvatore; Denora, Nunzio; Marzano, Cristina; Laquintana, Valentino; Cutrignelli, Annalisa; Hoeschele, James D; Gandin, Valentina; Natile, Giovanni

    2016-08-16

    Biodegradable, PEG-coated, nanoparticles (NPs) have gained therapeutic application as injectable colloidal systems for the controlled and site-specific release of drugs. In this paper, encapsulation in PLGA-PEG polymer NPs has been exploited to lower the toxicity and to increase the antitumor activity of kiteplatin ([PtCl2(cis-1,4-DACH)]). Kiteplatin contains an isomeric form of the diamine ligand present in oxaliplatin and proved to be particularly active against ovarian and colon cancers. To favor encapsulation of the platinum drug in the hydrophobic core of the polymeric micelles, Pt(iv) prodrugs having hydrophobic carboxylic ligands at the axial positions were used in place of hydrophilic Pt(ii) complexes (compounds 1-4). The size, size distribution, and zeta potential (ZP) were measured by dynamic light scattering (DLS) and laser Doppler velocimetry (LDV), and drug encapsulation efficiency (EE) correlated to the alkyl chain length of the different Pt(iv) prodrugs. The number of the Pt atoms per NP (in the range of 1.3-2.4 × 10(6)) is comparable to that of polysilsesquioxane-based NPs and higher than that found for other nanoparticle platforms. The platinum-loaded PLGA-PEG NPs, tested in vivo in a syngeneic murine solid tumor (LLC), had a higher antitumor effect and, most importantly, were markedly less toxic than kiteplatin. PMID:27094010

  15. A Prodrug-type, MMP-2-targeting Nanoprobe for Tumor Detection and Imaging

    PubMed Central

    Wang, Yaping; Lin, Tingting; Zhang, Wenyuan; Jiang, Yifan; Jin, Hongyue; He, Huining; Yang, Victor C.; Chen, Yi; Huang, Yongzhuo

    2015-01-01

    Tumor-associated proteases (TAPs) have been intensively studied because of their critical roles in cancer development. As a case in point, expression of matrix metalloproteases (MMP) is significantly up-regulated in tumorigenesis, invasion, and metastasis among a majority of cancers. Here we present a prodrug-type, MMP-2-responsive nanoprobe system with high efficiency and low toxicity for detecting MMP-2-overexpressed tumors. The nanoprobe system is featured by its self-assembled fabrication and FRET effect. This prodrug-type nanoprobe is selectively activated by MMP-2, and thus useful for detection of the MMP-2-overexpressed cells and tumors. The nanoprobe system works successfully in various animal tumor models, including human fibrosarcoma and subcutaneous glioma xenograft. Furthermore, in order to overcome the blood brain barrier (BBB) and achieve brain tumor targeting, a transferrin-receptor targeting peptide (T7 peptide) is strategically incorporated into the nanoprobe. The T7-functionalized nanoprobe is capable of detecting the orthotopic brain tumor, with clear, real-time in vivo imaging. This method is promising for in vivo detection of brain tumor, and real-time monitor of a TAP (i.e., MMP-2). PMID:26000052

  16. Formation Rate-Limited Pharmacokinetics of Biologically Active Epoxy Transformers of Prodrug Treosulfan.

    PubMed

    Romański, Michał; Kasprzyk, Anna; Karbownik, Agnieszka; Szałek, Edyta; Główka, Franciszek K

    2016-05-01

    A prodrug treosulfan (TREO) is being evaluated in clinical trials as a myeloablative agent before hematopoietic stem cell transplantation. The active derivatives of TREO, monoepoxide (EBDM), and diepoxide (DEB) are formed in a pH-dependent nonenzymatic reaction. The aim of the study was to investigate pharmacokinetics of the TREO epoxy transformers in a rabbit model and explain the causes of low plasma concentrations of EBDM and DEB observed in patients receiving high-dose TREO before hematopoietic stem cell transplantation. New Zealand white rabbits (n = 5 per cohort) received an intravenous infusion of TREO (group I), injection of DEB (group II), and injection of a solution containing EBDM (group III). When EBDM and DEB were administered to the rabbits, they underwent a very rapid elimination (half-life 0.069 and 0.046 h) associated with a high systemic clearance (10.0 and 14.0 L h(-1) kg(-1)). After administration of TREO, the t1/2 of EBDM was statistically equal to the t1/2 of the prodrug (1.6 h). To conclude, after administration of TREO, its epoxy transformers demonstrate a formation-limited elimination. Then EBDM and DEB have the same elimination half-life as TREO, but the levels of EBDM and DEB in the body, including plasma, are much lower than TREO on account of their inherently high clearance. PMID:27044946

  17. Dissociable effects of the prodrug phendimetrazine and its metabolite phenmetrazine at dopamine transporters.

    PubMed

    Solis, Ernesto; Suyama, Julie A; Lazenka, Matthew F; DeFelice, Louis J; Negus, S Stevens; Blough, Bruce E; Banks, Matthew L

    2016-01-01

    Phendimetrazine (PDM) is a clinically available anorectic and a candidate pharmacotherapy for cocaine addiction. PDM has been hypothesized to function as a prodrug that requires metabolism to the amphetamine-like monoamine transporter substrate phenmetrazine (PM) to produce its pharmacological effects; however, whether PDM functions as an inactive prodrug or has pharmacological activity on its own remains unclear. The study aim was to determine PDM pharmacological mechanisms using electrophysiological, neurochemical, and behavioral procedures. PDM blocked the endogenous basal hDAT (human dopamine transporter) current in voltage-clamped (-60 mV) oocytes consistent with a DAT inhibitor profile, whereas its metabolite PM induced an inward hDAT current consistent with a DAT substrate profile. PDM also attenuated the PM-induced inward current during co-application, providing further evidence that PDM functions as a DAT inhibitor. PDM increased nucleus accumbens dopamine levels and facilitated electrical brain stimulation reinforcement within 10 min in rats, providing in vivo evidence supporting PDM pharmacological activity. These results demonstrate that PDM functions as a DAT inhibitor that may also interact with the pharmacological effects of its metabolite PM. Overall, these results suggest a novel mechanism for PDM therapeutic effects via initial PDM DAT inhibition followed by PM DAT substrate-induced dopamine release. PMID:27514281

  18. Bioconversion of naltrexone and its 3-O-alkyl-ester prodrugs in a human skin equivalent.

    PubMed

    Hammell, Dana C; Stolarczyk, Elzbieta I; Klausner, Mitch; Hamad, Mohamed O; Crooks, Peter A; Stinchcomb, Audra L

    2005-04-01

    The purpose of this study was to compare the percutaneous absorption and bioconversion of naltrexone (NTX), naltrexone-3-O-valerate (VAL), and naltrexone-3-O-(2'-ethylbutyrate) (ETBUT) in a human skin equivalent model (EpiDerm) and in fresh human skin in vitro. NTX diffusion and metabolism to 6-beta-naltrexol (NTXol) were quantitated and compared in the EpiDerm and in excised fresh human skin. VAL and ETBUT diffusion and bioconversion studies were also completed in EpiDerm. Naltrexone bioconverted to levels of 3+/-2% NTXol in the EpiDerm and 1+/-0.5% in fresh human skin. VAL hydrolyzed rapidly in the EpiDerm and mainly (93+/-4%) NTX was found in the receiver compartment, similar to human skin. More intact ETBUT permeated the EpiDerm tissue compared to VAL, and only 15+/-11% NTX was found in the receiver. Significantly higher fluxes of NTX and the prodrugs were observed with the EpiDerm compared to human skin. A similar flux enhancement level was observed for VAL, compared to NTX base, in the EpiDerm and the human skin. Metabolically active human epidermal models like EpiDerm are useful as an alternative experimental system to human skin for prediction of topical/transdermal drug/prodrug bioconversion. PMID:15736197

  19. Colon-specific prodrugs of 4-aminosalicylic acid for inflammatory bowel disease.

    PubMed

    Dhaneshwar, Suneela S

    2014-04-01

    Despite the advent of biological products, such as anti-tumor necrosis factor-α monoclonal antibodies (infliximab and adalimumab), for treatment of moderate to severe cases of inflammatory bowel disease (IBD), most patients depend upon aminosalicylates as the conventional treatment option. In recent years, the increased knowledge of complex pathophysiological processes underlying IBD has resulted in development of a number of newer pharmaceutical agents like low-molecular-weight heparin, omega-3 fatty acids, probiotics and innovative formulations such as high-dose, once-daily multi-matrix mesalamine, which are designed to minimize the inflammatory process through inhibition of different targets. Optimization of delivery of existing drugs to the colon using the prodrug approach is another attractive alternative that has been utilized and commercialized for 5-aminosalicylic acid (ASA) in the form of sulfasalazine, balsalazide, olsalazine and ipsalazine, but rarely for its positional isomer 4-ASA - a well-established antitubercular drug that is twice as potent as 5-ASA against IBD, and more specifically, ulcerative colitis. The present review focuses on the complete profile of 4-ASA and its advantages over 5-ASA and colon-targeting prodrugs reported so far for the management of IBD. The review also emphasizes the need for reappraisal of this promising but unexplored entity as a potential treatment option for IBD. PMID:24707139

  20. Propyphenazone-based analogues as prodrugs and selective cyclooxygenase-2 inhibitors.

    PubMed

    Radwan, Mohamed F; Dalby, Kevin N; Kaoud, Tamer S

    2014-09-11

    Improving the gastrointestinal safety profile of nonsteroidal anti-inflammatory drugs (NSAIDs) is an important goal. Herein, we report two strategies, using the nonacidic propyphenazone structure, with potential to overcome the side effects of NSAIDs. Propyphenazone was employed to temporarily mask the free acid group of the widely used NSAIDs ibuprofen, diclofenac, and ketoprofen to develop three mutual prodrugs hypothesized to have minimal GI irritation. The three prodrugs exhibit in vivo anti-inflammatory and analgesic activities with improved potency over each parent drug when compared to a nonhydrolyzable control betahistine-propyphenazone (BET-MP). Additionally, ANT-MP formed by the irreversible coupling of propyphenazone and 4-aminoantipyrine, displayed exceptional COXII selectivity (COXII IC50 of 0.97 ± 0.04 μM, compared to no observed inhibition of COXI at 160 μM). Inhibition of COXII suppresses inflammatory diseases without affecting COXI-mediated GI tract events. ANT-MP exhibited maximal analgesic effect when tested in vivo in an abdominal writhing assay (100% protection) and its anti-inflammatory activity showed a peak at 2 h in a carrageenan-induced paw edema model. Its unique selectivity toward the COXII enzyme was investigated using molecular modeling techniques. PMID:25221653

  1. Propyphenazone-Based Analogues as Prodrugs and Selective Cyclooxygenase-2 Inhibitors

    PubMed Central

    2014-01-01

    Improving the gastrointestinal safety profile of nonsteroidal anti-inflammatory drugs (NSAIDs) is an important goal. Herein, we report two strategies, using the nonacidic propyphenazone structure, with potential to overcome the side effects of NSAIDs. Propyphenazone was employed to temporarily mask the free acid group of the widely used NSAIDs ibuprofen, diclofenac, and ketoprofen to develop three mutual prodrugs hypothesized to have minimal GI irritation. The three prodrugs exhibit in vivo anti-inflammatory and analgesic activities with improved potency over each parent drug when compared to a nonhydrolyzable control betahistine–propyphenazone (BET–MP). Additionally, ANT–MP formed by the irreversible coupling of propyphenazone and 4-aminoantipyrine, displayed exceptional COXII selectivity (COXII IC50 of 0.97 ± 0.04 μM, compared to no observed inhibition of COXI at 160 μM). Inhibition of COXII suppresses inflammatory diseases without affecting COXI-mediated GI tract events. ANT–MP exhibited maximal analgesic effect when tested in vivo in an abdominal writhing assay (100% protection) and its anti-inflammatory activity showed a peak at 2 h in a carrageenan-induced paw edema model. Its unique selectivity toward the COXII enzyme was investigated using molecular modeling techniques. PMID:25221653

  2. Lipid-based drug carriers for prodrugs to enhance drug delivery.

    PubMed

    Zaro, Jennica L

    2015-01-01

    The combination of lipid drug delivery systems with prodrugs offers several advantages including improved pharmacokinetics, increased absorption, and facilitated targeting. Lipidization and use of lipid carriers can increase the pharmacological half-life of the drug, thus improving pharmacokinetics and allowing less frequent dosing. Lipids also offer advantages such as increased absorption through the intestines for oral drug absorption and to the CNS for brain delivery. Furthermore, the use of lipid delivery systems can enhance drug targeting. Endogenous proteins bind lipids in the blood and carry them to the liver to enable targeting of this organ. Drugs with significant side effects in the stomach can be specifically delivered to enterocytes by exploiting lipases for prodrug activation. Finally, lipids can be used to target the lymphatic system, thus bypassing the liver and avoiding first-pass metabolism. Lymphatic targeting is also important for antiviral drugs in the protection of B and T lymphocytes. In this review, both lipid-drug conjugates and lipid-based carriers will be discussed. An overview, including the chemistry and assembly of the systems, as well as examples from the clinic and in development, will be provided. PMID:25269430

  3. Colon-specific prodrugs of 4-aminosalicylic acid for inflammatory bowel disease

    PubMed Central

    Dhaneshwar, Suneela S

    2014-01-01

    Despite the advent of biological products, such as anti-tumor necrosis factor-α monoclonal antibodies (infliximab and adalimumab), for treatment of moderate to severe cases of inflammatory bowel disease (IBD), most patients depend upon aminosalicylates as the conventional treatment option. In recent years, the increased knowledge of complex pathophysiological processes underlying IBD has resulted in development of a number of newer pharmaceutical agents like low-molecular-weight heparin, omega-3 fatty acids, probiotics and innovative formulations such as high-dose, once-daily multi-matrix mesalamine, which are designed to minimize the inflammatory process through inhibition of different targets. Optimization of delivery of existing drugs to the colon using the prodrug approach is another attractive alternative that has been utilized and commercialized for 5-aminosalicylic acid (ASA) in the form of sulfasalazine, balsalazide, olsalazine and ipsalazine, but rarely for its positional isomer 4-ASA - a well-established antitubercular drug that is twice as potent as 5-ASA against IBD, and more specifically, ulcerative colitis. The present review focuses on the complete profile of 4-ASA and its advantages over 5-ASA and colon-targeting prodrugs reported so far for the management of IBD. The review also emphasizes the need for reappraisal of this promising but unexplored entity as a potential treatment option for IBD. PMID:24707139

  4. Design of enzymatically cleavable prodrugs of a potent platinum-containing anticancer agent.

    PubMed

    Ding, Song; Pickard, Amanda J; Kucera, Gregory L; Bierbach, Ulrich

    2014-12-01

    Using a versatile synthetic approach, a new class of potential ester prodrugs of highly potent, but systemically too toxic, platinum-acridine anticancer agents was generated. The new hybrids contain a hydroxyl group, which has been masked with a cleavable lipophilic acyl moiety. Both butanoic (butyric) and bulkier 2-propanepentanoic (valproic) esters were introduced. The goals of this design were to improve the drug-like properties (e.g., logD) and to reduce the systemic toxicity of the pharmacophore. Two distinct pathways by which the target compounds undergo effective ester hydrolysis, the proposed activating step, have been confirmed: platinum-assisted, self-immolative ester cleavage in a low-chloride environment (LC-ESMS, NMR spectroscopy) and enzymatic cleavage by human carboxylesterase-2 (hCES-2) (LC-ESMS). The valproic acid ester derivatives are the first example of a metal-containing agent cleavable by the prodrug-converting enzyme. They show excellent chemical stability and reduced systemic toxicity. Preliminary results from screening in lung adenocarcinoma cell lines (A549, NCI-H1435) suggest that the mechanism of the valproic esters may involve intracellular deesterification. PMID:25303639

  5. Hydrogen peroxide-activatable antioxidant prodrug as a targeted therapeutic agent for ischemia-reperfusion injury

    PubMed Central

    Lee, Dongwon; Park, Seunggyu; Bae, Soochan; Jeong, Dahee; Park, Minhyung; Kang, Changsun; Yoo, Wooyoung; Samad, Mohammed A.; Ke, Qingen; Khang, Gilson; Kang, Peter M.

    2015-01-01

    Overproduction of hydrogen peroxide (H2O2) causes oxidative stress and is the main culprit in the pathogenesis of ischemia/reperfusion (I/R) injury. Suppression of oxidative stress is therefore critical in the treatment of I/R injury. Here, we report H2O2-activatable antioxidant prodrug (BRAP) that is capable of specifically targeting the site of oxidative stress and exerting anti-inflammatory and anti-apoptotic activities. BRAP with a self-immolative boronic ester protecting group was designed to scavenge H2O2 and release HBA (p-hydroxybenzyl alcohol) with antioxidant and anti-inflammatory activities. BRAP exerted potent antioxidant and anti-inflammatory activity in lipopolysaccharide (LPS)- and H2O2-stimulated cells by suppressing the generation of ROS and pro-inflammatory cytokines. In mouse models of hepatic I/R and cardiac I/R, BRAP exerted potent antioxidant, anti-inflammatory and anti-apoptotic activities due to the synergistic effects of H2O2-scavenging boronic esters and therapeutic HBA. In addition, administration of high doses of BRAP daily for 7 days showed no renal or hepatic function abnormalities. Therefore BRAP has tremendous therapeutic potential as H2O2-activatable antioxidant prodrug for the treatment of I/R injuries. PMID:26563741

  6. Nanoparticle-mediated delivery of a rapidly activatable prodrug of SN-38 for neuroblastoma therapy

    PubMed Central

    Alferiev, Ivan S.; Iyer, Radhika; Croucher, Jamie L.; Adamo, Richard F.; Zhang, Kehan; Mangino, Jennifer L.; Kolla, Venkatadri; Fishbein, Ilia; Brodeur, Garrett M.; Levy, Robert J.; Chorny, Michael

    2015-01-01

    Nanomedicine-based strategies have the potential to improve therapeutic performance of a wide range of anticancer agents. However, the successful implementation of nanoparticulate delivery systems requires the development of adequately sized nanocarriers delivering their therapeutic cargo to the target in a protected, pharmacologically active form. The present studies focused on a novel nanocarrier-based formulation strategy for SN-38, a topoisomerase I inhibitor with proven anticancer potential, whose clinical application is compromised by toxicity, poor stability and incompatibility with conventional delivery vehicles. SN-38 encapsulated in biodegradable sub-100 nm sized nanoparticles (NP) in the form of its rapidly activatable prodrug derivative with tocopherol succinate potently inhibited the growth of neuroblastoma cells in a dose- and exposure time-dependent manner, exhibiting a delayed response pattern distinct from that of free SN-38. In a xenograft model of neuroblastoma, prodrug-loaded NP caused rapid regression of established large tumors, significantly delayed tumor regrowth after treatment cessation and markedly extended animal survival. The NP formulation strategy enabled by a reversible chemical modification of the drug molecule offers a viable means for SN-38 delivery achieving sustained intratumoral drug levels and contributing to the potency and extended duration of antitumor activity, both prerequisites for effective treatment of neuroblastoma and other cancers. PMID:25770994

  7. Magnetic nanoparticle hyperthermia induced cytosine deaminase expression in microencapsulated E. coli for enzyme-prodrug therapy

    PubMed Central

    Nemani, Krishnamurthy V.; Ennis, Riley C.; Griswold, Karl E.; Gimi, Barjor

    2015-01-01

    Engineered bacterial cells that are designed to express therapeutic enzymes under the transcriptional control of remotely inducible promoters can mediate the de novo conversion of non-toxic prodrugs to their cytotoxic forms. In situ cellular expression of enzymes provides increased stability and control of enzyme activity as compared to isolated enzymes. We have engineered Escherichia coli (E. coli), designed to express cytosine deaminase at elevated temperatures, under the transcriptional control of thermo-regulatory λpL-cI857 promoter cassette which provides a thermal switch to trigger enzyme synthesis. Enhanced cytosine deaminase expression was observed in cultures incubated at 42 °C as compared to 30 °C, and enzyme expression was further substantiated by spectrophotometric assays indicating enhanced conversion of 5-fluorocytosine to 5-fluorouracil. The engineered cells were subsequently co-encapsulated with magnetic iron oxide nanoparticles in immunoprotective alginate microcapsules, and cytosine deaminase expression was triggered remotely by alternating magnetic field-induced hyperthermia. The combination of 5-fluorocytosine with AMF-activated microcapsules demonstrated tumor cell cytotoxicity comparable to direct treatment with 5-fluorouracil chemotherapy. Such enzyme-prodrug therapy, based on engineered and immunoisolated E. coli, may ultimately yield an improved therapeutic index relative to monotherapy, as AMF mediated hyperthermia might be expected to pre-sensitize tumors to chemotherapy under appropriate conditions. PMID:25820125

  8. An Essential Type I Nitroreductase from Leishmania major Can Be Used to Activate Leishmanicidal Prodrugs

    PubMed Central

    Voak, Andrew A.; Gobalakrishnapillai, Vithurshaa; Seifert, Karin; Balczo, Edina; Hu, Longqin; Hall, Belinda S.; Wilkinson, Shane R.

    2013-01-01

    Nitroaromatic prodrugs are used to treat a range of microbial infections with selectivity achieved by specific activation reactions. For trypanosomatid parasites, this is mediated by type I nitroreductases. Here, we demonstrate that the causative agent of leishmaniasis, Leishmania major, expresses an FMN-containing nitroreductase (LmNTR) that metabolizes a wide range of substrates, and based on electron donor and acceptor preferences, it may function as an NADH:quinone oxidoreductase. Using gene deletion approaches, we demonstrate that this activity is essential to L. major promastigotes, the parasite forms found in the insect vector. Intriguingly, LmNTR+/− heterozygote promastigote parasites could readily differentiate into infectious metacyclic cells but these were unable to establish infections in cultured mammalian cells and caused delayed pathology in mice. Furthermore, we exploit the LmNTR activity evaluating a library of nitrobenzylphosphoramide mustards using biochemical and phenotypic screens. We identify a subset of compounds that display significant growth inhibitory properties against the intracellular parasite form found in the mammalian hosts. The leishmanicidal activity was shown to be LmNTR-specific as the LmNTR+/− heterozygote promastigotes displayed resistance to the most potent mustards. We conclude that LmNTR can be targeted for drug development by exploiting its prodrug activating property or by designing specific inhibitors to block its endogenous function. PMID:23946481

  9. Involvement of Carboxylesterase in Hydrolysis of Propranolol Prodrug during Permeation across Rat Skin

    PubMed Central

    Imai, Teruko; Takase, Yuko; Iwase, Harunobu; Hashimoto, Mitsuru

    2013-01-01

    The use of a prodrug, a conjugate of an active drug with a lipophilic substituent, is a good way of increasing the cutaneous absorption of a drug. However, the activity of dermal hydrolases has rarely been investigated in humans, or experimental animals. In the present study, we focused on the identification of rat dermal esterases and the hydrolysis of a prodrug during permeation across rat skin. We found that carboxylesterase (CES), especially the rat CES1 isozyme, Hydrolase A, is expressed in rat skin and that the hydrolysis of p-nitrophenyl acyl derivatives and caproyl-propranolol (PL) was 20-fold lower in the 9000g supernatant fraction of skin homogenate than in liver microsomes. A permeation study of caproyl-PL was performed in rat full-thickness and stripped skin using a flow-through diffusion cell. Caproyl-PL was easily partitioned into the stratum corneum and retained, not only in the stratum corneum, but also in viable epidermis and dermis. Caproyl-PL could barely be detected in the receptor fluid after application to either full-thickness or stripped skin. PL, derived from caproyl-PL, was, however, detected in receptor fluid after extensive hydrolysis of caproyl-PL in viable skin. Permeation of PL was markedly decreased under CES inhibition, indicating that the net flux of caproyl-PL is dependent on its conversion rate to PL. PMID:24300511

  10. Development and bioorthogonal activation of palladium-labile prodrugs of gemcitabine.

    PubMed

    Weiss, Jason T; Dawson, John C; Fraser, Craig; Rybski, Witold; Torres-Sánchez, Carmen; Bradley, Mark; Patton, E Elizabeth; Carragher, Neil O; Unciti-Broceta, Asier

    2014-06-26

    Bioorthogonal chemistry has become one of the main driving forces in current chemical biology, inspiring the search for novel biocompatible chemospecific reactions for the past decade. Alongside the well-established labeling strategies that originated the bioorthogonal paradigm, we have recently proposed the use of heterogeneous palladium chemistry and bioorthogonal Pd(0)-labile prodrugs to develop spatially targeted therapies. Herein, we report the generation of biologically inert precursors of cytotoxic gemcitabine by introducing Pd(0)-cleavable groups in positions that are mechanistically relevant for gemcitabine's pharmacological activity. Cell viability studies in pancreatic cancer cells showed that carbamate functionalization of the 4-amino group of gemcitabine significantly reduced (>23-fold) the prodrugs' cytotoxicity. The N-propargyloxycarbonyl (N-Poc) promoiety displayed the highest sensitivity to heterogeneous palladium catalysis under biocompatible conditions, with a reaction half-life of less than 6 h. Zebrafish studies with allyl, propargyl, and benzyl carbamate-protected rhodamines confirmed N-Poc as the most suitable masking group for implementing in vivo bioorthogonal organometallic chemistry. PMID:24867590

  11. Doxorubicin/heparin composite nanoparticles for caspase-activated prodrug chemotherapy.

    PubMed

    Khaliq, Nisar Ul; Sandra, Febrina Carolina; Park, Dal Yong; Lee, Jae Young; Oh, Keun Sang; Kim, Dongkyu; Byun, Youngro; Kim, In-San; Kwon, Ick Chan; Kim, Sang Yoon; Yuk, Soon Hong

    2016-09-01

    Caspase-activated prodrug chemotherapy is introduced and demonstrated using the composite nanoparticles (NPs), which deliver doxorubicin (DOX) and DEVD-S-DOX together to the tumor tissue. DEVD-S-DOX, DOX linked to a peptide moiety (DEVD), is a prodrug that is cleaved into free DOX by caspase-3 upon apoptosis. DEVD-S-DOX has no therapeutic efficacy, but it changes into free DOX with the expression of caspase-3. With the accumulation of the composite NPs in the tumor tissue by the enhanced permeation and retention (EPR) effect, a small exposure of DOX in the tumor cells initiated apoptosis in a localized area of the tumor tissue, which induced caspase-3 activation. Cleavage of DEVD-S-DOX into free DOX by caspase-3 continued with repetitive activation of caspase-3 and cleavage of DEVD-S-DOX at the tumor site. The composite NPs were characterized with transmittance electron microscopy (TEM) and particle size analyzer. We then evaluated the nanoparticle drug release, therapeutic efficacy, and in vivo biodistribution for tumor targeting using a non-invasive live animal imaging technology and the quantification of DOX with high performance liquid chromatography. DOX-induced apoptosis-targeted chemotherapy (DIATC) was verified by in vitro/in vivo DEVD-S-DOX response to free DOX and cellular uptake behavior of the composite NPs with flow cytometry analysis. Significant antitumor efficacy with minimal cardiotoxicity was also observed, which supported DIATC for improved chemotherapy. PMID:27286189

  12. Absorption, distribution, metabolism, and excretion of the novel antibacterial prodrug tedizolid phosphate.

    PubMed

    Ong, Voon; Flanagan, Shawn; Fang, Edward; Dreskin, Howard J; Locke, Jeffrey B; Bartizal, Kenneth; Prokocimer, Philippe

    2014-08-01

    Tedizolid phosphate is a novel antibacterial prodrug with potent activity against Gram-positive pathogens. In vitro and in vivo studies demonstrated that the prodrug is rapidly converted by nonspecific phosphatases to the biologically active moiety tedizolid. Single oral dose radiolabeled (14)C-tedizolid phosphate kinetic studies in human subjects (100 µCi in 204 mg tedizolid phosphate free acid) confirmed a rapid time to maximum tedizolid concentration (Tmax, 1.28 hours), a long terminal half-life (10.6 hours), and a Cmax of 1.99 µg/ml. Metabolite analysis of plasma, fecal, and urine samples from rats, dogs, and humans confirmed that tedizolid is the only measurable metabolite in plasma after intravenous (in animals only) or oral administration and that tedizolid sulfate is the major metabolite excreted from the body. Excellent mass balance recovery was achieved and demonstrated that fecal excretion is the predominant (80-90%) route of elimination across species, primarily as tedizolid sulfate. Urine excretion accounted for the balance of drug elimination but contained a broader range of minor metabolites. Glucuronidation products were not detected. Similar results were observed in rats and dogs after both intravenous and oral administration. The tedizolid metabolites showed less potent antibacterial activity than tedizolid. The observations from these studies support once daily dosing of tedizolid phosphate and highlight important metabolism and excretion features that differentiate tedizolid phosphate from linezolid. PMID:24875463

  13. Magnetic nanoparticle hyperthermia induced cytosine deaminase expression in microencapsulated E. coli for enzyme-prodrug therapy.

    PubMed

    Nemani, Krishnamurthy V; Ennis, Riley C; Griswold, Karl E; Gimi, Barjor

    2015-06-10

    Engineered bacterial cells that are designed to express therapeutic enzymes under the transcriptional control of remotely inducible promoters can mediate the de novo conversion of non-toxic prodrugs to their cytotoxic forms. In situ cellular expression of enzymes provides increased stability and control of enzyme activity as compared to isolated enzymes. We have engineered Escherichia coli (E. coli), designed to express cytosine deaminase at elevated temperatures, under the transcriptional control of thermo-regulatory λpL-cI857 promoter cassette which provides a thermal switch to trigger enzyme synthesis. Enhanced cytosine deaminase expression was observed in cultures incubated at 42°C as compared to 30°C, and enzyme expression was further substantiated by spectrophotometric assays indicating enhanced conversion of 5-fluorocytosine to 5-fluorouracil. The engineered cells were subsequently co-encapsulated with magnetic iron oxide nanoparticles in immunoprotective alginate microcapsules, and cytosine deaminase expression was triggered remotely by alternating magnetic field-induced hyperthermia. The combination of 5-fluorocytosine with AMF-activated microcapsules demonstrated tumor cell cytotoxicity comparable to direct treatment with 5-fluorouracil chemotherapy. Such enzyme-prodrug therapy, based on engineered and immunoisolated E. coli, may ultimately yield an improved therapeutic index relative to monotherapy, as AMF mediated hyperthermia might be expected to pre-sensitize tumors to chemotherapy under appropriate conditions. PMID:25820125

  14. Synthesis and anticancer activities of amphiphilic 5-fluoro-2'-deoxyuridylic acid prodrugs.

    PubMed

    Ludwig, Peter S; Schwendener, Reto A; Schott, Herbert

    2005-05-01

    Amphiphilic anticancer prodrugs of 5'-fluoro-2'-deoxyuridine-5'-monophosphate (5-FdUMP) were synthesized according to the hydrogen phosphonate method by coupling lipophilic cytosine derivatives or a phospholipid with 5-fluoro-2'-deoxyuridine (5-FdU). Studies within the in vitro Anticancer Screen Program of the National Cancer Institute have demonstrated high anticancer activities of the heterodinucleoside phosphates: N4-palmitoyl-2'-deoxycytidylyl-(3' --> 5')-3'-O-acetyl-5-fluoro-2'-deoxyuridine (dC(pam)-5-FdU(Ac), N4-palmitoyl-2',3'-dideoxycytidylyl-(5' --> 5')-3'-O-acetyl-5-fluoro-2'-deoxyuridine (ddC(pam)-(5' --> 5')-5-FdU(Ac), 5-fluoro-2'-deoxyuridylyl-(3' --> 5')-5-fluoro-N4-hexadecyl-2'-deoxycytidine (5-FdU-5-FdC(hex)), and of the new liponucleotide 1-O-octadecyl-rac-glycerylyl-(3 --> 5')-5-fluoro-2'-deoxyuridine (Oct1Gro-(3 --> 5')-5-FdU). The anticancer activities of these prodrugs are comparable to those of 5-FdU and the tumor specificities are modulated by their structures. The highest cytotoxic activity being even superior to 5-FdU was expressed by the dimer 5-FdU-5-FdC(hex). PMID:15893023

  15. Overcoming tumor resistance to cisplatin by cationic lipid-assisted prodrug nanoparticles.

    PubMed

    Cao, Zhi-Ting; Chen, Zhi-Yao; Sun, Chun-Yang; Li, Hong-Jun; Wang, Hong-Xia; Cheng, Qin-Qin; Zuo, Zu-Qi; Wang, Ji-Long; Liu, Yang-Zhong; Wang, Yu-Cai; Wang, Jun

    2016-07-01

    Chemotherapy resistance has become a major challenge in the clinical treatment of lung cancer which is the leading cancer type for the estimated deaths. Recent studies have shown that nanoparticles as drug carriers can raise intracellular drug concentration by achieving effectively cellular uptake and rapid drug release, and therefore reverse the acquired chemoresistance of tumors. In this context, nanoparticles-based chemotherapy represents a promising strategy for treating malignancies with chemoresistance. In the present study, we developed cationic lipid assisted nanoparticles (CLAN) to deliver polylactide-cisplatin prodrugs to drug resistant lung cancer cells. The nanoparticles were formulated through self-assembly of a biodegradable poly(ethylene glycol)-block-poly(lactide) (PEG-PLA), a hydrophobic polylactide-cisplatin prodrug, and a cationic lipid. The cationic nanoparticles were proven to significantly improve cell uptake of cisplatin, leading to an increased DNA-Pt adduct and significantly promoted DNA damage in vitro. Moreover, our study reveals that cationic nanoparticles, although are slightly inferior in blood circulation and tumor accumulation, are more effective in blood vessel extravasation. The CLANs ultimately enhances the cellular drug availability and leads to the reversal of cisplatin resistance. PMID:27088406

  16. Ester Prodrugs of Ketoprofen: Synthesis, In Vitro Stability, In Vivo Biological Evaluation and In Silico Comparative Docking Studies Against COX-1 and COX-2.

    PubMed

    Ahmed, Musa; Azam, Faizul; Gbaj, Abdul; Zetrini, Abdulmottaleb E; Abodlal, Amna Salem; Rghigh, Abir; Elmahdi, Eman; Hamza, Amel; Salama, Mabruk; Bensaber, Salah M

    2016-01-01

    Prompted by the ineptness of the currently used non-steroidal antiinflammatory drugs (NSAIDs) to control gastric mucosal and renal adverse reactions, several ester prodrugs of ketoprofen were synthesized and characterized by IR, 1H NMR and mass spectral data. Physicochemical properties such as aqueous solubility, octanol-water partition coefficient log P, chemical stability and enzymatic hydrolysis of the synthesized molecules have been studied to assess their potential as prodrugs. The obtained results confirmed that all ester prodrugs are chemically stable, possess increased lipophilicity compared to their parent compounds and converted to the active drugs in vivo. All of the tested ester prodrugs exhibited marked anti-inflammatory activity ranging from 91.8% to 113.3% in comparison with the parent drug, ketoprofen. A mutual prodrug obtained from two antiinflammatory molecules, ketoprofen and salicylic acid has been noted to potentiate the activity making it most active molecule of the series. The ulcerogenic index of the ester prodrugs was significantly lower than the parent drug, ketoprofen. Comparative docking studies against X-ray crystal structures of COX-1 and COX-2 further provided understanding of their interaction with the cyclooxygenases that will facilitate design of better inhibitors (or prodrugs) with sufficient specificity for COX-2 against COX-1. The study offers an innovative strategy for finding a molecule with safer therapeutic profile for longterm treatment of inflammatory diseases. PMID:26785683

  17. Lipophilic prodrug conjugates allow facile and rapid synthesis of high loading capacity liposomes without the need for post-assembly purification

    PubMed Central

    Mikhalin, Alexander A.; Evdokimov, Nikolai M.; Frolova, Liliya V.; Magedov, Igor V.; Kornienko, Alexander; Johnston, Robert; Rogelj, Snezna; Tartis, Michaelann S.

    2014-01-01

    Dihydropyridopyrazoles are simplified synthetic analogues of podophyllotoxin that can effectively mimic its molecular scaffold and act as potent mitotic spindle poisons in dividing cancer cells. However, despite nanomolar potencies and ease of synthetic preparation, further clinical development of these promising anticancer agents is hampered due to their poor aqueous solubility. In this paper, we developed a prodrug strategy that enables incorporation of dihydropyridopyrazoles into liposome bilayers to overcome the solubility issues. The active drug was covalently connected to either myristic or palmitic acid anchor via carboxylesterase hydrolyzable linkage. The resulting prodrugs were self assembled into liposome bilayers from hydrated lipid films using ultrasound without the need for post-assembly purification. The average particle size of the prodrug-loaded liposomes was about 90 nm. The prodrug incorporation was verified by differential scanning calorimetry, spectrophotometry and gel filtration reaching maximum at 0.3 and 0.35 prodrug/lipid molar ratios for myristic and palmitic conjugates, respectively. However, the ratio of 0.2 was used in the particle size and biological activity experiments to maintain long-term stability of the prodrug-loaded liposomes against phase separation during storage. Antiproliferative activity was tested against HeLa and Jurkat cancer cell lines in vitro showing that the liposomal prodrug retained antitubulin activity of the parent drug and induced apoptosis mediated cancer cell death. Overall, the established data provide a powerful platform for further clinical development of dihydropyridopyrazoles using liposomes as the drug delivery system. PMID:25534989

  18. Synthesis, Chemical and Enzymatic Hydrolysis, and Aqueous Solubility of Amino Acid Ester Prodrugs of 3-Carboranyl Thymidine Analogues for Boron Neutron Capture Therapy of Brain Tumors

    PubMed Central

    Hasabelnaby, Sherifa; Goudah, Ayman; Agarwal, Hitesh K.; Abd alla, Mosaad S. M.; Tjarks, Werner

    2012-01-01

    Various water-soluble L-valine-, L-glutamate-, and glycine ester prodrugs of two 3-Carboranyl Thymidine Analogues (3-CTAs), designated N5 and N5-2OH, were synthesized for Boron Neutron Capture Therapy (BNCT) of brain tumors since the water solubilities of the parental compounds proved to be insufficient in preclinical studies. The amino acid ester prodrugs were prepared and stored as hydrochloride salts. The water solubilities of these amino acid ester prodrugs, evaluated in phosphate buffered saline (PBS) at pH 5, pH 6 and pH 7.4, improved 48 to 6600 times compared with parental N5 and N5-2OH. The stability of the amino acid ester prodrugs was evaluated in PBS at pH 7.4, Bovine serum, and Bovine cerebrospinal fluid (CSF). The rate of the hydrolysis in all three incubation media depended primarily on the amino acid promoiety and, to a lesser extend, on the site of esterification at the deoxyribose portion of the 3-CTAs. In general, 3'-amino acid ester prodrugs were less sensitive to chemical and enzymatic hydrolysis than 5'-amino acid ester prodrugs and the stabilities of the latter decreased in the following order: 5'-valine > 5'-glutamate > 5'-glycine. The rate of the hydrolysis of the 5'-amino acid ester prodrugs in Bovine CSF was overall higher than in PBS and somewhat lower than in Bovine serum. Overall, 5'-glutamate ester prodrug of N5 and the 5'-glycine ester prodrugs of N5 and N5-2OH appeared to be the most promising candidates for preclinical BNCT studies. PMID:22889558

  19. Contact-facilitated drug delivery with Sn2 lipase labile prodrugs optimize targeted lipid nanoparticle drug delivery.

    PubMed

    Pan, Dipanjan; Pham, Christine T N; Weilbaecher, Katherine N; Tomasson, Michael H; Wickline, Samuel A; Lanza, Gregory M

    2016-01-01

    Sn2 lipase labile phospholipid prodrugs in conjunction with contact-facilitated drug delivery offer an important advancement in Nanomedicine. Many drugs incorporated into nanosystems, targeted or not, are substantially lost during circulation to the target. However, favorably altering the pharmacokinetics and volume of distribution of systemic drug delivery can offer greater efficacy with lower toxicity, leading to new prolonged-release nanoexcipients. However, the concept of achieving Paul Erhlich's inspired vision of a 'magic bullet' to treat disease has been largely unrealized due to unstable nanomedicines, nanosystems achieving low drug delivery to target cells, poor intracellular bioavailability of endocytosed nanoparticle payloads, and the substantial biological barriers of extravascular particle penetration into pathological sites. As shown here, Sn2 phospholipid prodrugs in conjunction with contact-facilitated drug delivery prevent premature drug diffusional loss during circulation and increase target cell bioavailability. The Sn2 phospholipid prodrug approach applies equally well for vascular constrained lipid-encapsulated particles and micelles the size of proteins that penetrate through naturally fenestrated endothelium in the bone marrow or thin-walled venules of an inflamed microcirculation. At one time Nanomedicine was considered a 'Grail Quest' by its loyal opposition and even many in the field adsorbing the pains of a long-learning curve about human biology and particles. However, Nanomedicine with innovations like Sn2 phospholipid prodrugs has finally made 'made the turn' toward meaningful translational success. PMID:26296541

  20. Utilization of peptide carrier system to improve intestinal absorption: targeting prolidase as a prodrug-converting enzyme

    NASA Technical Reports Server (NTRS)

    Bai, J. P.; Hu, M.; Subramanian, P.; Mosberg, H. I.; Amidon, G. L.

    1992-01-01

    The feasibility of targeting prolidase as a peptide prodrug-converting enzyme has been examined. The enzymatic hydrolysis by prolidase of substrates for the peptide transporter L-alpha-methyldopa-pro and several dipeptide analogues without an N-terminal alpha-amino group (phenylpropionylproline, phenylacetylproline, N-benzoylproline, and N-acetylproline) was investigated. The Michaelis-Menten parameters Km and Vmax for L-alpha-methyldopa-pro are 0.09 +/- 0.02 mM and 3.98 +/- 0.25 mumol/min/mg protein, respectively. However, no hydrolysis of the dipeptide analogues without an N-terminal alpha-amino group is observed, suggesting that an N-terminal alpha-amino group is required for prolidase activity. These results demonstrate that prolidase may serve as a prodrug-converting enzyme for the dipeptide-type prodrugs, utilizing the peptide carrier for transport of prodrugs into the mucosal cells and prolidase, a cytosolic enzyme, to release the drug. However, a free alpha-amino group appears to be necessary for prolidase hydrolysis.

  1. Identification of GS 4104 as an Orally Bioavailable Prodrug of the Influenza Virus Neuraminidase Inhibitor GS 4071

    PubMed Central

    Li, Weixing; Escarpe, Paul A.; Eisenberg, Eugene J.; Cundy, Kenneth C.; Sweet, Clive; Jakeman, Kenneth J.; Merson, James; Lew, Willard; Williams, Matt; Zhang, Lijun; Kim, Choung U.; Bischofberger, Norbert; Chen, Ming S.; Mendel, Dirk B.

    1998-01-01

    GS 4071 is a potent carbocyclic transition-state analog inhibitor of influenza virus neuraminidase with activity against both influenza A and B viruses in vitro. GS 4116, the guanidino analog of GS 4071, is a 10-fold more potent inhibitor of influenza virus replication in tissue culture than GS 4071. In this study we determined the oral bioavailabilities of GS 4071, GS 4116, and their respective ethyl ester prodrugs in rats. Both parent compounds and the prodrug of the guanidino analog exhibited poor oral bioavailability (2 to 4%) and low peak concentrations in plasma (Cmaxs; Cmax <0.06 μg/ml). In contrast, GS 4104, the ethyl ester prodrug of GS 4071, exhibited good oral bioavailability (35%) as GS 4071 and high Cmaxs of GS 4071 (Cmax = 0.47 μg/ml) which are 150 times the concentration necessary to inhibit influenza virus neuraminidase activity by 90%. The bioavailability of GS 4104 as GS 4071 was also determined in mice (30%), ferrets (11%), and dogs (73%). The plasma of all four species exhibited high, sustained concentrations of GS 4071 such that at 12 h postdosing the concentrations of GS 4071 in plasma exceeded those necessary to inhibit influenza virus neuraminidase activity by 90%. These results demonstrate that GS 4104 is an orally bioavailable prodrug of GS 4071 in animals and that it has the potential to be an oral agent for the prevention and treatment of influenza A and B virus infections in humans. PMID:9517946

  2. Targeting (cellular) lysosomal acid ceramidase by B13: Design, synthesis and evaluation of novel DMG-B13 ester prodrugs

    PubMed Central

    Bai, Aiping; Szulc, Zdzislaw, M.; Bielawski, Jacek; Pierce, Jason S.; Rembisa, Barbara; Terzieva, Silva; Mao, Cungui; Xu, Ruijuan; Wu, Bill; Clarke, Christopher J.; Newcomb, Benjamin; Liu, Xiang; Norris, James; Hannun, Yusuf A.; Bielawska, Alicja

    2015-01-01

    Acid ceramidase (ACDase) is being recognized as a therapeutic target for cancer. B13 represents a moderate inhibitor of ACDase. The present study concentrates on the lysosomal targeting of B13 via its N, N-dimethylglycine (DMG) esters (DMG-B13 prodrugs). Novel analogs, the isomeric mono-DMG-B13, LCL522 (3-O-DMG-B13•HCl) and LCL596 (1-O-DMG-B13•HCl) and di-DMG-B13, LCL521 (1,3-O, O-DMG-B13•2HCl) conjugates, were designed and synthesized through N, N-dimethyl glycine (DMG) esterification of the hydroxyl groups of B13. In MCF7 cells, DMG-B13 prodrugs were efficiently metabolized to B13. The early inhibitory effect of DMG-B13 prodrugs on cellular ceramidases was ACDase specific by their lysosomal targeting. The corresponding dramatic decrease of cellular Sph (80-97% Control/1h) by DMG-B13 prodrugs was mainly from the inhibition of the lysosomal ACDase. PMID:25456083

  3. Phosphonooxymethyl Prodrug of Triptolide: Synthesis, Physicochemical Characterization, and Efficacy in Human Colon Adenocarcinoma and Ovarian Cancer Xenografts.

    PubMed

    Patil, Satish; Lis, Lev G; Schumacher, Robert J; Norris, Beverly J; Morgan, Monique L; Cuellar, Rebecca A D; Blazar, Bruce R; Suryanarayanan, Raj; Gurvich, Vadim J; Georg, Gunda I

    2015-12-10

    A disodium phosphonooxymethyl prodrug of the antitumor agent triptolide was prepared from the natural product in three steps (39% yield) and displayed excellent aqueous solubility at pH 7.4 (61 mg/mL) compared to the natural product (17 μg/mL). The estimated shelf life (t90) for hydrolysis of the prodrug at 4 °C and pH 7.4 was found to be two years. In a mouse model of human colon adenocarcinoma (HT-29), the prodrug administered intraperitoneally was effective in reducing or eliminating xenograft tumors at dose levels as low as 0.3 mg/kg when given daily and at 0.9 mg/kg when given less frequently. When given via intraperitoneal and oral routes at daily doses of 0.6 and 0.9 mg/kg, the prodrug was also effective and well tolerated in a mouse model of human ovarian cancer (A2780). PMID:26596892

  4. Co-delivery of doxorubicin and curcumin by pH-sensitive prodrug nanoparticle for combination therapy of cancer

    NASA Astrophysics Data System (ADS)

    Zhang, Yumin; Yang, Cuihong; Wang, Weiwei; Liu, Jinjian; Liu, Qiang; Huang, Fan; Chu, Liping; Gao, Honglin; Li, Chen; Kong, Deling; Liu, Qian; Liu, Jianfeng

    2016-02-01

    Ample attention has focused on cancer drug delivery via prodrug nanoparticles due to their high drug loading property and comparatively lower side effects. In this study, we designed a PEG-DOX-Cur prodrug nanoparticle for simultaneous delivery of doxorubicin (DOX) and curcumin (Cur) as a combination therapy to treat cancer. DOX was conjugated to PEG by Schiff’s base reaction. The obtained prodrug conjugate could self-assemble in water at pH 7.4 into nanoparticles (PEG-DOX NPs) and encapsulate Cur into the core through hydrophobic interaction (PEG-DOX-Cur NPs). When the PEG-DOX-Cur NPs are internalized by tumor cells, the Schiff’s base linker between PEG and DOX would break in the acidic environment that is often observed in tumors, causing disassembling of the PEG-DOX-Cur NPs and releasing both DOX and Cur into the nuclei and cytoplasma of the tumor cells, respectively. Compared with free DOX, free Cur, free DOX-Cur combination, or PEG-DOX NPs, PEG-DOX-Cur NPs exhibited higher anti-tumor activity in vitro. In addition, the PEG-DOX-Cur NPs also showed prolonged blood circulation time, elevated local drug accumulation and increased tumor penetration. Enhanced anti-tumor activity was also observed from the PEG-DOX-Cur-treated animals, demonstrating better tumor inhibitory property of the NPs. Thus, the PEG-DOX-Cur prodrug nanoparticle system provides a simple yet efficient approach of drug delivery for chemotherapy.

  5. Synthesis, pH-Dependent, and Plasma Stability of Meropenem Prodrugs for Potential Use Against Drug-Resistant Tuberculosis

    PubMed Central

    Teitelbaum, Aaron M.; Meissner, Anja; Harding, Ryan A.; Wong, Christopher A.; Aldrich, Courtney C.; Remmel, Rory P.

    2013-01-01

    Meropenem, a broad-spectrum parenteral β-lactam antibiotic, in combination with clavulanate has recently shown efficacy in patients with extensively drug-resistant tuberculosis. As a result of meropenem’s short half-life and lack of oral bioavailability, the development of an oral therapy is warranted for TB treatment in underserved countries where chronic parenteral therapy is impractical. To improve the oral absorption of meropenem, several alkyloxycarbonyloxyalkyl ester prodrugs with increased lipophilicity were synthesized and their stability in physiological aqueous solutions and guinea pig as well as human plasma was evaluated. The stability of prodrugs in aqueous solution at pH 6.0 and 7.4 was significantly dependent on the ester promoiety with the major degradation product identified as the parent compound meropenem. However, in simulated gastrointestinal fluid (pH 1.2) the major degradation product identified was ring-opened meropenem with the promoiety still intact, suggesting the gastrointestinal environment may reduce the absorption of meropenem prodrugs in vivo unless administered as an enteric-coated formulation. Additionally, the stability of the most aqueous stable prodrugs in guinea pig or human plasma was short, implying a rapid release of parent meropenem. PMID:23845282

  6. Solid dispersions of the penta-ethyl ester prodrug of diethylenetriaminepentaacetic acid (DTPA): Formulation design and optimization studies

    PubMed Central

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

    2015-01-01

    The penta-ethyl ester prodrug of diethylenetriaminepentaacetic acid (DTPA), which exists as an oily liquid, was incorporated into a solid dispersion for oral administration by the solvent evaporation method using blends of polyvinylpyrrolidone (PVP), Eudragit® RL PO and α-tocopherol. D-optimal mixture design was used to optimize the formulation. Formulations that had a high concentration of both Eudragit® RL PO and α-tocopherol exhibited low water absorption and enhanced stability of the DTPA prodrug. Physicochemical properties of the optimal formulation were evaluated using Fourier transform infrared (FTIR) spectroscopy and differential scanning calorimetry (DSC). In vitro release of the prodrug was evaluated using the USP Type II apparatus dissolution method. DSC studies indicated that the matrix had an amorphous structure, while FTIR spectrometry showed that DTPA penta-ethyl ester and excipients did not react with each other during formation of the solid dispersion.. Dissolution testing showed that the optimized solid dispersion exhibited a prolonged release profile, which could potentially result in a sustained delivery of DTPA penta-ethyl to enhance bioavailability. In conclusion, DTPA penta-ethyl ester was successfully incorporated into a solid matrix with high drug loading and improved stability compared to prodrug alone. PMID:24047113

  7. A safe, simple and efficient doxorubicin prodrug hybrid micelle for overcoming tumor multidrug resistance and targeting delivery.

    PubMed

    Bao, Yuling; Yin, Mingxing; Hu, Xiaomeng; Zhuang, Xiangting; Sun, Yu; Guo, Yuanyuan; Tan, Songwei; Zhang, Zhiping

    2016-08-10

    A pH-sensitive prodrug, TPGS-CHN-DOX, was introduced by conjugating anticancer drug, doxorubicin (DOX), onto d-α-tocopherol polyethylene glycol 1000 succinate (TPGS) via a cleavable Schiff base linkage. The prodrug was mixed with a PEGylated lipid to form a simple but multifunctional hybrid micelle system, which can realize high drug loading capability and biocompatibility, extended blood circulation time, inhibited drug resistance in cancer cells, improved therapeutic response, reduced side effects, and easy functionalities for targeting delivery. The hybrid micelles exhibited in vitro pH-sensitive drug release, enhanced cellular uptake and strengthened cytotoxicity on both drug-sensitive human breast cancer MCF-7 and resistant MCF-7/ADR cells. P-glycoprotein functional inhibition and mitochondria-associated cell apoptosis induced by TPGS were thought to play an important role in overcoming the multidrug resistance. As a result, the hybrid micelles demonstrated good anticancer efficacy in MCF-7/ADR xenograft model. Additionally, after modifying with a tumor-specific targeting peptic ligand, cRGD, the tumor growth/metastasis inhibition was further evidenced in integrin receptor overexpressed melanoma cancer B16F10 and even murine hepatocarcinoma H22 models. This TPGS-based pH-sensitive prodrug provides a safe and "Molecular economical" way in the rational design of prodrugs for overcoming multidrug resistance and targeting delivery, which can improve the potency for clinical use. PMID:27264552

  8. Amino Acid Derivatives as Palmitoylethanolamide Prodrugs: Synthesis, In Vitro Metabolism and In Vivo Plasma Profile in Rats

    PubMed Central

    Vacondio, Federica; Bassi, Michele; Silva, Claudia; Castelli, Riccardo; Carmi, Caterina; Scalvini, Laura; Lodola, Alessio; Vivo, Valentina; Flammini, Lisa; Barocelli, Elisabetta; Mor, Marco; Rivara, Silvia

    2015-01-01

    Palmitoylethanolamide (PEA) has antinflammatory and antinociceptive properties widely exploited in veterinary and human medicine, despite its poor pharmacokinetics. Looking for prodrugs that could progressively release PEA to maintain effective plasma concentrations, we prepared carbonates, esters and carbamates at the hydroxyl group of PEA. Chemical stability (pH 7.4) and stability in rat plasma and liver homogenate were evaluated by in vitro assays. Carbonates and carbamates resulted too labile and too resistant in plasma, respectively. Ester derivatives, prepared by conjugating PEA with various amino acids, allowed to modulate the kinetics of PEA release in plasma and stability in liver homogenate. L-Val-PEA, with suitable PEA release in plasma, and D-Val-PEA, with high resistance to hepatic degradation, were orally administered to rats and plasma levels of prodrugs and PEA were measured at different time points. Both prodrugs showed significant release of PEA, but provided lower plasma concentrations than those obtained with equimolar doses of PEA. Amino-acid esters of PEA are a promising class to develop prodrugs, even if they need further chemical optimization. PMID:26053855

  9. Contact-facilitated drug delivery with Sn2 lipase labile prodrugs optimize targeted lipid nanoparticle drug delivery

    PubMed Central

    Pan, Dipanjan; Pham, Christine TN; Weilbaecher, Katherine N; Tomasson, Michael H; Wickline, Samuel A; Lanza, Gregory M

    2016-01-01

    Sn2 lipase labile phospholipid prodrugs in conjunction with contact-facilitated drug delivery offer an important advancement in Nanomedicine. Many drugs incorporated into nanosystems, targeted or not, are substantially lost during circulation to the target. However, favorably altering the pharmacokinetics and volume of distribution of systemic drug delivery can offer greater efficacy with lower toxicity, leading to new prolonged-release nanoexcipients. However, the concept of achieving Paul Erhlich's inspired vision of a ‘magic bullet’ to treat disease has been largely unrealized due to unstable nanomedicines, nanosystems achieving low drug delivery to target cells, poor intracellular bioavailability of endocytosed nanoparticle payloads, and the substantial biological barriers of extravascular particle penetration into pathological sites. As shown here, Sn2 phospholipid prodrugs in conjunction with contact-facilitated drug delivery prevent premature drug diffusional loss during circulation and increase target cell bioavailability. The Sn2 phospholipid prodrug approach applies equally well for vascular constrained lipid-encapsulated particles and micelles the size of proteins that penetrate through naturally fenestrated endothelium in the bone marrow or thin-walled venules of an inflamed microcirculation. At one time Nanomedicine was considered a ‘Grail Quest’ by its loyal opposition and even many in the field adsorbing the pains of a long-learning curve about human biology and particles. However, Nanomedicine with innovations like Sn2 phospholipid prodrugs has finally made ‘made the turn’ toward meaningful translational success. PMID:26296541

  10. Co-delivery of doxorubicin and curcumin by pH-sensitive prodrug nanoparticle for combination therapy of cancer

    PubMed Central

    Zhang, Yumin; Yang, Cuihong; Wang, Weiwei; Liu, Jinjian; Liu, Qiang; Huang, Fan; Chu, Liping; Gao, Honglin; Li, Chen; Kong, Deling; Liu, Qian; Liu, Jianfeng

    2016-01-01

    Ample attention has focused on cancer drug delivery via prodrug nanoparticles due to their high drug loading property and comparatively lower side effects. In this study, we designed a PEG-DOX-Cur prodrug nanoparticle for simultaneous delivery of doxorubicin (DOX) and curcumin (Cur) as a combination therapy to treat cancer. DOX was conjugated to PEG by Schiff’s base reaction. The obtained prodrug conjugate could self-assemble in water at pH 7.4 into nanoparticles (PEG-DOX NPs) and encapsulate Cur into the core through hydrophobic interaction (PEG-DOX-Cur NPs). When the PEG-DOX-Cur NPs are internalized by tumor cells, the Schiff’s base linker between PEG and DOX would break in the acidic environment that is often observed in tumors, causing disassembling of the PEG-DOX-Cur NPs and releasing both DOX and Cur into the nuclei and cytoplasma of the tumor cells, respectively. Compared with free DOX, free Cur, free DOX-Cur combination, or PEG-DOX NPs, PEG-DOX-Cur NPs exhibited higher anti-tumor activity in vitro. In addition, the PEG-DOX-Cur NPs also showed prolonged blood circulation time, elevated local drug accumulation and increased tumor penetration. Enhanced anti-tumor activity was also observed from the PEG-DOX-Cur-treated animals, demonstrating better tumor inhibitory property of the NPs. Thus, the PEG-DOX-Cur prodrug nanoparticle system provides a simple yet efficient approach of drug delivery for chemotherapy. PMID:26876480

  11. Phosphonooxymethyl Prodrug of Triptolide: Synthesis, Physicochemical Characterization, and Efficacy in Human Colon Adenocarcinoma and Ovarian Cancer Xenografts

    PubMed Central

    2015-01-01

    A disodium phosphonooxymethyl prodrug of the antitumor agent triptolide was prepared from the natural product in three steps (39% yield) and displayed excellent aqueous solubility at pH 7.4 (61 mg/mL) compared to the natural product (17 μg/mL). The estimated shelf life (t90) for hydrolysis of the prodrug at 4 °C and pH 7.4 was found to be two years. In a mouse model of human colon adenocarcinoma (HT-29), the prodrug administered intraperitoneally was effective in reducing or eliminating xenograft tumors at dose levels as low as 0.3 mg/kg when given daily and at 0.9 mg/kg when given less frequently. When given via intraperitoneal and oral routes at daily doses of 0.6 and 0.9 mg/kg, the prodrug was also effective and well tolerated in a mouse model of human ovarian cancer (A2780). PMID:26596892

  12. Structure-Based Design and Synthesis of Nω-Nitro-L-Arginine-Containing Peptidomimetics as Selective Inhibitors of Neuronal Nitric Oxide Synthase. Displacement of the Heme Structural Water

    PubMed Central

    Seo, Jiwon; Igarashi, Jotato; Li, Huiying; Martásek, Pavel; Roman, Linda J.; Poulos, Thomas L.; Silverman, Richard B.

    2008-01-01

    The neuronal isoform of nitric oxide synthase (nNOS), the enzyme responsible for the production of nitric oxide in the central nervous system, represents an attractive target for the treatment of various neurodegenerative disorders. X-ray crystal structures of complexes of nNOS with two nNOS-selective inhibitors, (4S)-N-{4-amino-5-[(2-aminoethylamino]pentyl}-N′-nitroguanidine (1) and 4-N-(Nω-nitro-L-argininyl)-trans-4-amino-L-proline amide (2), led to the discovery of a conserved structural water molecule that was hydrogen bonded between the two heme propionates and the inhibitors (Figure 2). Based on this observation, we hypothesized that by attaching a hydrogen bond donor group to the amide nitrogen of 2 or to the secondary amine nitrogen of 1, the inhibitor molecules could displace the structural water molecule and obtain a direct interaction with the heme cofactor. To test this hypothesis, peptidomimetic analogues 3–5, which have either an N-hydroxyl (3 and 5) or N-amino (4) donor group, were designed and synthesized. X-ray crystal structures of nNOS with inhibitors 3 and 5 bound verified that the N-hydroxyl group had, indeed, displaced the structural water molecule and provided a direct interaction with the heme propionate moiety (Figures 4 and 5). Surprisingly, in vitro activity assay results indicated that the addition of a hydroxyl group (3) only increased the potency slightly against the neuronal isoform over the parent compound (1). Rationalizations for the small increase in potency are consistent with other changes in the crystal structures. PMID:17425297

  13. Design, synthesis and evaluation of mutual prodrug of 4-biphenylacetic acid and quercetin tetramethyl ether (BPA-QTME) as gastrosparing NSAID.

    PubMed

    Madhukar, Mamta; Sawraj, Shruti; Sharma, Pritam Dev

    2010-06-01

    A novel mutual prodrug consisting of 4-biphenylacetic acid (BPA) and quercetin tetramethyl ether (QTME) has been synthesized as a gastrosparing NSAID, devoid of ulcerogenic side effects. The physicochemical properties, including aqueous solubility, partition coefficient, chemical stability and enzymatic hydrolysis of synthesized derivative have been studied to assess its prodrug potential. Its antiinflammatory, antiulcer and analgesic activities were also evaluated. The results indicated that BPA-QTME derivative is chemically stable, biolabile and possesses optimum lipophilicity. The synthesized compound also exhibited retention of antiinflammatory activity with reduced ulcerogenicity. Based on these observations, the therapeutic potential of this mutual prodrug is discussed. PMID:20227799

  14. Synthesis and Biological Evaluation of a Novel Pentagastrin-Toxin Conjugate Designed for a Targeted Prodrug Mono-therapy of Cancer

    PubMed Central

    Tietze, Lutz F.; Panknin, Olaf; Krewer, Birgit; Major, Felix; Schuberth, Ingrid

    2008-01-01

    A novel carbamate prodrug 2 containing a pentagastrin moiety was synthesized. 2 was designed as a detoxified analogue of the highly cytotoxic natural antibiotic duocarmycin SA (1) for the use in a targeted prodrug monotherapy of cancers expressing cholecystokinin (CCK-B)/gastrin receptors. The synthesis of prodrug 2 was performed using a palladium-catalyzed carbonylation of bromide 6, followed by a radical cyclisation to give the pharmacophoric unit 10, coupling of 10 to the DNA-binding subunit 15 and transformation of the resulting seco-drug 3b into the carbamate 2 via addition of a pentagastrin moiety. PMID:19325786

  15. Potential of Host Defense Peptide Prodrugs as Neutrophil Elastase-Dependent Anti-Infective Agents for Cystic Fibrosis

    PubMed Central

    Humphreys, Hilary; Greene, Catherine M.; Fitzgerald-Hughes, Deirdre; Devocelle, Marc

    2014-01-01

    Host defense peptides (HDPs) are short antimicrobial peptides of the innate immune system. Deficiencies in HDPs contribute to enhanced susceptibility to infections, e.g., in cystic fibrosis (CF). Exogenous HDPs can compensate for these deficiencies, but their development as antimicrobials is limited by cytotoxicity. Three HDP prodrugs were designed so their net positive charge is masked by a promoiety containing a substrate for the enzyme neutrophil elastase (NE). This approach can confine activation to sites with high NE levels. Enzyme-labile peptides were synthesized, and their activation was investigated using purified NE. Susceptibilities of Pseudomonas aeruginosa to parent and prodrug peptides in the presence and absence of NE-rich CF human bronchoalveolar lavage (BAL) fluid and different NaCl concentrations were compared. The effect of the HDP promoiety on cytotoxicity was determined with cystic fibrosis bronchial epithelial (CFBE41o-) cells. NE in CF BAL fluids activated the HDP prodrugs, restoring bactericidal activity against reference and clinical isolates of P. aeruginosa. However, activation also required the addition of 300 mM NaCl. Under these conditions, the bactericidal activity levels of the HDP prodrugs differed, with pro-P18 demonstrating the greatest activity (90% to 100% of that of the parent, P18, at 6.25 μg/ml). Cytotoxic effects on CFBE41o- cells were reduced by the addition of the promoiety to HDPs. We demonstrate here for the first time the selective activation of novel HDP prodrugs by a host disease-associated enzyme at in vivo concentrations of the CF lung. This approach may lead to the development of novel therapeutic agents with low toxicity that are active under the challenging conditions of the CF lung. PMID:24277028

  16. Efficacy of low-dose oral metronomic dosing of the prodrug of gemcitabine, LY2334737, in human tumor xenografts.

    PubMed

    Pratt, Susan E; Durland-Busbice, Sara; Shepard, Robert L; Donoho, Gregory P; Starling, James J; Wickremsinhe, Enaksha R; Perkins, Everett J; Dantzig, Anne H

    2013-04-01

    LY2334737, an oral prodrug of gemcitabine, is cleaved in vivo, releasing gemcitabine and valproic acid. Oral dosing of mice results in absorption of intact prodrug with slow systemic hydrolysis yielding higher plasma levels of LY2334737 than gemcitabine and prolonged gemcitabine exposure. Antitumor activity was evaluated in human colon and lung tumor xenograft models. The dose response for efficacy was examined using 3 metronomic schedules, once-a-day dosing for 14 doses, every other day for 7 doses, and once a day for 7 doses, 7 days rest, followed by an additional 7 days of once-a-day dosing. These schedules gave significant antitumor activity and were well tolerated. Oral gavage of 6 mg/kg LY2334737 daily for 21 days gave equivalent activity to i.v. 240 mg/kg gemcitabine. HCl administered once a week for 3 weeks to mice bearing a patient mesothelioma tumor PXF 1118 or a non-small cell lung cancer tumor LXFE 937. The LXFE 397 tumor possessed elevated expression of the equilibrative nucleoside transporter-1 (ENT1) important for gemcitabine uptake but not prodrug uptake and responded significantly better to treatment with LY2334737 than gemcitabine (P ≤ 0.001). In 3 colon xenografts, antitumor activity of LY2334737 plus a maximally tolerated dose of capecitabine, an oral prodrug of 5-fluorouracil, was significantly greater than either monotherapy. During treatment, the expression of carboxylesterase 2 (CES2) and concentrative nucleoside transporter-3 was induced in HCT-116 tumors; both are needed for the activity of the prodrugs. Thus, metronomic oral low-dose LY2334737 is efficacious, well tolerated, and easily combined with capecitabine for improved efficacy. Elevated CES2 or ENT1 expression may enhance LY2334737 tumor response. PMID:23371859

  17. Role of glutamate 64 in the activation of the prodrug 5-fluorocytosine by yeast cytosine deaminase.

    PubMed

    Wang, Jifeng; Sklenak, Stepan; Liu, Aizhuo; Felczak, Krzysztof; Wu, Yan; Li, Yue; Yan, Honggao

    2012-01-10

    Yeast cytosine deaminase (yCD) catalyzes the hydrolytic deamination of cytosine to uracil as well as the deamination of the prodrug 5-fluorocytosine (5FC) to the anticancer drug 5-fluorouracil. In this study, the role of Glu64 in the activation of the prodrug 5FC was investigated by site-directed mutagenesis, biochemical, nuclear magnetic resonance (NMR), and computational studies. Steady-state kinetics studies showed that the mutation of Glu64 causes a dramatic decrease in k(cat) and a dramatic increase in K(m), indicating Glu64 is important for both binding and catalysis in the activation of 5FC. (19)F NMR experiments showed that binding of the inhibitor 5-fluoro-1H-pyrimidin-2-one (5FPy) to the wild-type yCD causes an upfield shift, indicating that the bound inhibitor is in the hydrated form, mimicking the transition state or the tetrahedral intermediate in the activation of 5FC. However, binding of 5FPy to the E64A mutant enzyme causes a downfield shift, indicating that the bound 5FPy remains in an unhydrated form in the complex with the mutant enzyme. (1)H and (15)N NMR analysis revealed trans-hydrogen bond D/H isotope effects on the hydrogen of the amide of Glu64, indicating that the carboxylate of Glu64 forms two hydrogen bonds with the hydrated 5FPy. ONIOM calculations showed that the wild-type yCD complex with the hydrated form of the inhibitor 1H-pyrimidin-2-one is more stable than the initial binding complex, and in contrast, with the E64A mutant enzyme, the hydrated inhibitor is no longer favored and the conversion has a higher activation energy, as well. The hydrated inhibitor is stabilized in the wild-type yCD by two hydrogen bonds between it and the carboxylate of Glu64 as revealed by (1)H and (15)N NMR analysis. To explore the functional role of Glu64 in catalysis, we investigated the deamination of cytosine catalyzed by the E64A mutant by ONIOM calculations. The results showed that without the assistance of Glu64, both proton transfers before and

  18. Role of Glutamate 64 in the Activation of the Prodrug 5-fluorocytosine by Yeast Cytosine Deaminase†

    PubMed Central

    Wang, Jifeng; Sklenak, Stepan; Liu, Aizhuo; Felczak, Krzysztof; Wu, Yan; Li, Yue; Yan, Honggao

    2012-01-01

    Yeast cytosine deaminase catalyzes the hydrolytic deamination of cytosine to uracil as well as the deamination of the prodrug 5-fluorocytosine (5FC) to the anticancer drug 5-fluorouracil. In this study, the role of Glu64 in the activation of the prodrug 5FC was investigated by site-directed mutagenesis, biochemical, NMR, and computational studies. Steady-state kinetics studies showed that the mutation of Glu64 causes a dramatic decrease in kcat and a dramatic increase in Km, indicating Glu64 is important for both binding and catalysis in the activation of 5FC. 19F-NMR experiments showed that binding of the inhibitor 5-fluoro-1H-pyrimidin-2-one (5FPy) to the wild type yCD causes an upfield shift, indicating that the bound inhibitor is in the hydrated form, mimicking the transition state or the tetrahedral intermediate in the activation of 5FC. However, binding of 5FPy to the E64A mutant enzyme causes a downfield shift, indicating that the bound 5FPy remains in an unhydrated form in the complex with the mutant enzyme. 1H and 15N NMR analysis revealed trans-hydrogen-bond D/H isotope effects on the hydrogen of the amide of Glu64, indicating that the carboxylate of Glu64 forms two hydrogen bonds with the hydrated 5FPy. ONIOM calculations showed that the wild type yCD complex with the hydrated form of the inhibitor 1H-pyrimidin-2-one is more stable than the initial binding complex, and in contrast, with the E64A mutant enzyme, the hydrated inhibitor is no longer favored and the conversion has higher activation energy as well. The hydrated inhibitor is stabilized in the wild-type yCD by two hydrogen bonds between it and the carboxylate of Glu64 as revealed by 1H and 15N NMR analysis. To explore the functional role of Glu64 in catalysis, deamination of cytosine catalyzed by the E64A mutant was investigated by ONIOM calculations. The results showed that without the assistance of Glu64, both proton transfers before and after the formation of the tetrahedral reaction

  19. Targeted delivery of a cisplatin prodrug for safer and more effective prostate cancer therapy in vivo

    PubMed Central

    Dhar, Shanta; Kolishetti, Nagesh; Lippard, Stephen J.; Farokhzad, Omid C.

    2011-01-01

    Targeted delivery and controlled release of inactive platinum (Pt) prodrugs may offer a new approach to improve the efficacy and tolerability of the Pt family of drugs, which are used to treat 50% of all cancers today. Using prostate cancer (PCa) as a model disease, we previously described the engineering of aptamer (Apt)-targeted poly(D,L-lactic-co-glycolic acid)-b-poly(ethylene glycol) (PLGA-b-PEG) nanoparticles (NPs) encapsulating a Pt(IV) prodrug c,t,c[Pt(NH3)2-(O2CCH2CH2CH2CH2CH3)2Cl2] (1) (Pt-PLGA-b-PEG-Apt-NP), which target the extracellular domain of the prostate specific membrane antigen (PSMA), for enhanced in vitro cytotoxicity. Here we demonstrate enhanced in vivo pharmacokinetics (PK), biodistribution, tolerability, and efficacy of Pt-PLGA-b-PEG-Apt-NP (150±15 nm encapsulating ∼5% wt/wt Pt(IV) prodrug) when compared to cisplatin administered in its conventional form in normal Sprague Dawley rats, Swiss Albino mice, and the PSMA-expressing LNCaP subcutaneous xenograft mouse model of PCa, respectively. The 10-d maximum tolerated dose following a single i.v. injection of Pt-PLGA-b-PEG-NP in rats and mice was determined at 40 mg/kg and 5 mg/kg, respectively. PK studies with Pt-PLGA-b-PEG-NP revealed prolonged Pt persistence in systemic blood circulation and decreased accumulation of Pt in the kidneys, a major target site of cisplatin toxicity. Pt-PLGA-b-PEG-Apt-NPs further displayed the significant dose-sparing characteristics of the drug, with equivalent antitumor efficacy in LNCaP xenografts at 1/3 the dose of cisplatin administered in its conventional form (0.3 mg/kg vs. 1 mg/kg). When considering the simultaneous improvement in tolerability and efficacy, the Pt-PLGA-b-PEG-Apt NP provides a remarkable improvement in the drug therapeutic index. PMID:21233423

  20. Post-injury administration of NAAG peptidase inhibitor prodrug, PGI-02776, in experimental TBI.

    PubMed

    Feng, Jun-Feng; Van, Ken C; Gurkoff, Gene G; Kopriva, Christina; Olszewski, Rafal T; Song, Minsoo; Sun, Shifeng; Xu, Man; Neale, Joseph H; Yuen, Po-Wai; Lowe, David A; Zhou, Jia; Lyeth, Bruce G

    2011-06-13

    Traumatic brain injury (TBI) leads to a rapid and excessive increase in glutamate concentration in the extracellular milieu, which is strongly associated with excitotoxicity and neuronal degeneration. N-acetylaspartylglutamate (NAAG), a prevalent peptide neurotransmitter in the vertebrate nervous system, is released along with glutamate and suppresses glutamate release by actions at pre-synaptic metabotropic glutamate autoreceptors. Extracellular NAAG is hydrolyzed to N-acetylaspartate and glutamate by peptidase activity. In the present study PGI-02776, a newly designed di-ester prodrug of the urea-based NAAG peptidase inhibitor ZJ-43, was tested for neuroprotective potential when administered intraperitoneally 30 min after lateral fluid percussion TBI in the rat. Stereological quantification of hippocampal CA2-3 degenerating neurons at 24 h post injury revealed that 10 mg/kg PGI-02776 significantly decreased the number of degenerating neurons (p<0.05). Both average latency analysis of Morris water maze performance and assessment of 24-hour memory retention revealed significant differences between sham-TBI and TBI-saline. In contrast, no significant difference was found between sham-TBI and PGI-02776 treated groups in either analysis indicating an improvement in cognitive performance with PGI-02776 treatment. Histological analysis on day 16 post-injury revealed significant cell death in injured animals regardless of treatment. In vitro NAAG peptidase inhibition studies demonstrated that the parent compound (ZJ-43) exhibited potent inhibitory activity while the mono-ester (PGI-02749) and di-ester (PGI-02776) prodrug compounds exhibited moderate and weak levels of inhibitory activity, respectively. Pharmacokinetic assays in uninjured animals found that the di-ester (PGI-02776) crossed the blood-brain barrier. PGI-02776 was also readily hydrolyzed to both the mono-ester (PGI-02749) and the parent compound (ZJ-43) in both blood and brain. Overall, these findings

  1. Azido- and chlorido-cobalt complex as carrier-prototypes for antitumoral prodrugs.

    PubMed

    Pires, Bianca M; Giacomin, Letícia C; Castro, Frederico A V; Cavalcanti, Amanda dos S; Pereira, Marcos D; Bortoluzzi, Adailton J; Faria, Roberto B; Scarpellini, Marciela

    2016-04-01

    Cobalt(III) complexes are well-suited systems for cytotoxic drug release under hypoxic conditions. Here, we investigate the effect of cytotoxic azide release by cobalt-containing carrier-prototypes for antitumoral prodrugs. In addition, we study the species formed after reduction of Co(3+) → Co(2+) in the proposed models for these prodrugs. Three new complexes, [Co(III)(L)(N3)2]BF4(1), [{Co(II)(L)(N3)}2](ClO4)2(2), and [Co(II)(L)Cl]PF6(3), L=[(bis(1-methylimidazol-2-yl)methyl)(2-(pyridyl-2-yl)ethyl)amine], were synthesized and studied by several spectroscopic, spectrometric, electrochemical, and crystallographic methods. Reactivity and spectroscopic data reveal that complex 1 is able to release N3(-) either after reduction with ascorbic acid, or by ambient light irradiation, in aqueous phosphate buffer (pH6.2, 7.0 and 7.4) and acetonitrile solutions. The antitumoral activities of compounds 1-3 were tested in normoxia on MCF-7 (human breast adenocarcinoma), PC-3 (human prostate) and A-549 (human lung adenocarcinoma epithelial) cell lines, after 24h of exposure. Either complexes or NaN3 presented IC50 values higher than 200 μM, showing lower cytotoxicity than the clinical standard antitumoral complex cisplatin, under the same conditions. Complexes 1-3 were also evaluated in hypoxia on A-549 and results indicate high IC50 data (>200 μM) after 24h of exposure. However, an increase of cancer cell susceptibility to 1 and 2 was observed at 300 μM. Regarding complex 3, no cytotoxic activity was observed in the same conditions. The data presented here indicate that the tridentate ligand L is able to stabilize both oxidation states of cobalt (+3 and +2). In addition, the cobalt(III) complex generates the low cytotoxic cobalt(II) species after reduction, which supports their use as as carrier prototypes for antitumoral prodrugs. PMID:26881993

  2. Excited‐State Dynamics of a Two‐Photon‐Activatable Ruthenium Prodrug

    PubMed Central

    Greenough, Simon E.; Horbury, Michael D.; Smith, Nichola A.; Sadler, Peter J.; Paterson, Martin J.

    2016-01-01

    Abstract We present a new approach to investigate how the photodynamics of an octahedral ruthenium(II) complex activated through two‐photon absorption (TPA) differ from the equivalent complex activated through one‐photon absorption (OPA). We photoactivated a RuII polypyridyl complex containing bioactive monodentate ligands in the photodynamic therapy window (620–1000 nm) by using TPA and used transient UV/Vis absorption spectroscopy to elucidate its reaction pathways. Density functional calculations allowed us to identify the nature of the initially populated states and kinetic analysis recovers a photoactivation lifetime of approximately 100 ps. The dynamics displayed following TPA or OPA are identical, showing that TPA prodrug design may use knowledge gathered from the more numerous and easily conducted OPA studies. PMID:26632426

  3. Evaluation of fluorinated biphenyl ether pro-drug scaffolds employing the chemical-microbial approach.

    PubMed

    Hampton, Alex S; Mikulski, Lena; Palmer-Brown, William; Murphy, Cormac D; Sandford, Graham

    2016-05-01

    Incorporation of fluorine in a drug can dramatically affect its metabolism and methods to assess the effect of fluorine substitution on drug metabolism are required for effective drug design. Employing a previously developed chemical-microbial method the metabolism of a series of fluorinated biphenyl ethers was determined. The substrates were synthesized via Ullmann-type condensation reactions between bromotoluene and fluorophenol. The ethers were incubated with the fungus Cunninghamella elegans, which oxidises xenobiotics in an analogous fashion to mammals, generating a number of hydroxylated biphenyl ethers and acids. The propensity of the fluorinated ring to be hydroxylated depended upon the position of the fluorine atom, and the oxidation of the methyl group was observed when it was meta to the oxygen. The experiments demonstrate the applicability of the method to rapidly determine the effect of fluorine substitution on CYP-catalysed biotransformation of pro-drug molecules. PMID:27020303

  4. Recent clinical experience with famciclovir--a "third generation" nucleoside prodrug.

    PubMed

    Chakrabarty, Arun; Tyring, Stephen K; Beutner, Karl; Rauser, Michael

    2004-09-01

    The herpesviruses continue to produce considerable morbidity in man. Once infected with herpes simplex (HSV), the virus remains dormant within the nervous system and may reactivate if provoked by stress, trauma and/or other factors. To date, there is no cure, but antiviral medication can reduce duration and severity of symptoms and prophylaxis can suppress recurrent episodes of disease. The second-generation guanosine nucleosides, acyclovir and penciclovir, are effective inhibitors with low toxicity; both, however, have relatively low oral bioavailability. Subsequently, the orally bioavailable prodrugs valaciclovir and famciclovir have been introduced. These compounds offer high oral bioavailabilty and deliver acyclovir and penciclovir, respectively, to the target cells by means of more convenient dosing schedules. This short review points to recent experience with famciclovir in the management of HSV and varicella-zoster virus. PMID:15535046

  5. [Pharmacokinetics of cefixime in volunteers and a literature comparison with the new ester prodrug cephalosporins].

    PubMed

    Kees, F; Naber, K G

    1990-01-01

    The pharmacokinetic parameters of cefixime were determined in healthy volunteers following oral administration of 200 mg cefixime as tablet, syrup and dry suspension, respectively. All three galenic formulations showed reliable absorption. Mean peak plasma concentrations amounted to 2.4-3.4 mg/l and were reached after 3.3-3.5 h. Mean terminal half-lives were 2.9-3.1 h. The mean areas under the plasma concentration-time curves ranged between 18 and 26 mg/l.h; 18-24% of the dose administered were recovered unchanged in the urine. The best bioavailability was obtained with the dry suspension followed by the tablet and the syrup. With respect to the ester pro-drug cephalosporins, cefuroxime axetil, cefetamet pivoxyl and cefotiam hexetil, cefixime exhibits higher plasma half-life and area under the curve as well as, comparable absolute bioavailability but consistently lower urinary recovery which indicates higher non-renal clearance. PMID:2079377

  6. Synthesis and in vitro evaluation of bisphosphonated glycopeptide prodrugs for the treatment of osteomyelitis.

    PubMed

    Tanaka, Kelly S E; Dietrich, Evelyne; Ciblat, Stéphane; Métayer, Claude; Arhin, Francis F; Sarmiento, Ingrid; Moeck, Gregory; Parr, Thomas R; Far, Adel Rafai

    2010-02-15

    As therapeutic agents of choice in the treatment of complicated infections, glycopeptide antibiotics are often preferentially used in cases of osteomyelitis, an infection located in bone and notoriously difficult to successfully manage. Yet frequent and heavy doses of these systemically administered antibiotics are conventionally prescribed to obtain higher antibiotic levels in the bone and reduce the high recurrence rates. Targeting antibiotics to the bone after systemic administration would present at least three potential advantages: (i) greater efficacy, by concentrating the therapeutic agent in bone; (ii) greater convenience, through a reduction in the frequency of administration; and (iii) greater safety, by reducing the levels of systemic drug exposure. We present here the design, synthesis and in vitro evaluation of eight prodrugs of the glycopeptide antibacterial agents vancomycin and oritavancin taking advantage of the affinity of the bisphosphonate group for bone for delivery to osseous tissues. PMID:20097069

  7. Discovery of an L-alanine ester prodrug of the Hsp90 inhibitor, MPC-3100.

    PubMed

    Kim, Se-Ho; Tangallapally, Rajendra; Kim, In Chul; Trovato, Richard; Parker, Daniel; Patton, J Scott; Reeves, Leslie; Bradford, Chad; Wettstein, Daniel; Baichwal, Vijay; Papac, Damon; Bajji, Ashok; Carlson, Robert; Yager, Kraig M

    2015-11-15

    Various types of Hsp90 inhibitors have been and continue to undergo clinical investigation. One development candidate is the purine-based, synthetic Hsp90 inhibitor 1 (MPC-3100), which successfully completed a phase I clinical study. However, further clinical development of 1 was hindered by poor solubility and consequent formulation issues and promoted development of a more water soluble prodrug. Towards this end, numerous pro-moieties were explored in vitro and in vivo. These studies resulted in identification of L-alanine ester mesylate, 2i (MPC-0767), which exhibited improved aqueous solubility, adequate chemical stability, and rapid bioconversion without the need for solubilizing excipients. Based on improved physical characteristics and favorable PK and PD profiles, 2i mesylate was selected for further development. A convergent, scalable, chromatography-free synthesis for 2i mesylate was developed to support further clinical evaluation. PMID:26483201

  8. Lipophilic prodrugs of nucleoside triphosphates as biochemical probes and potential antivirals

    PubMed Central

    Gollnest, Tristan; de Oliveira, Thiago Dinis; Schols, Dominique; Balzarini, Jan; Meier, Chris

    2015-01-01

    The antiviral activity of nucleoside reverse transcriptase inhibitors is often limited by ineffective phosphorylation. We report on a nucleoside triphosphate (NTP) prodrug approach in which the γ-phosphate of NTPs is bioreversibly modified. A series of TriPPPro-compounds bearing two lipophilic masking units at the γ-phosphate and d4T as a nucleoside analogue are synthesized. Successful delivery of d4TTP is demonstrated in human CD4+ T-lymphocyte cell extracts by an enzyme-triggered mechanism with high selectivity. In antiviral assays, the compounds are potent inhibitors of HIV-1 and HIV-2 in CD4+ T-cell (CEM) cultures. Highly lipophilic acyl residues lead to higher membrane permeability that results in intracellular delivery of phosphorylated metabolites in thymidine kinase-deficient CEM/TK− cells with higher antiviral activity than the parent nucleoside. PMID:26503889

  9. Engineering a Prostate-Specific Membrane Antigen–Activated Tumor Endothelial Cell Prodrug for Cancer Therapy

    PubMed Central

    Denmeade, Samuel R.; Mhaka, Annastasiah M.; Rosen, D. Marc; Brennen, W. Nathaniel; Dalrymple, Susan; Dach, Ingrid; Olesen, Claus; Gurel, Bora; DeMarzo, Angelo M.; Wilding, George; Carducci, Michael A.; Dionne, Craig A.; Møller, Jesper V.; Nissen, Poul; Christensen, S. Brøgger; Isaacs, John T.

    2013-01-01

    Heterogeneous expression of drug target proteins within tumor sites is a major mechanism of resistance to anticancer therapies. We describe a strategy to selectively inhibit, within tumor sites, the function of a critical intracellular protein, the sarcoplasmic/endoplasmic reticulum calcium adenosine triphosphatase (SERCA) pump, whose proper function is required by all cell types for viability. To achieve targeted inhibition, we took advantage of the unique expression of the carboxypeptidase prostate-specific membrane antigen (PSMA) by tumor endothelial cells within the microenvironment of solid tumors. We generated a prodrug, G202, consisting of a PSMA-specific peptide coupled to an analog of the potent SERCA pump inhibitor thapsigargin. G202 produced substantial tumor regression against a panel of human cancer xenografts in vivo at doses that were minimally toxic to the host. On the basis of these data, a phase 1 dose-escalation clinical trial has been initiated with G202 in patients with advanced cancer. PMID:22745436

  10. Colon-specific drug delivery based on a cyclodextrin prodrug: release behavior of biphenylylacetic acid from its cyclodextrin conjugates in rat intestinal tracts after oral administration.

    PubMed

    Minami, K; Hirayama, F; Uekama, K

    1998-06-01

    An antiinflammatory drug biphenylylacetic acid (BPAA) as a model drug was selectively conjugated onto one of the primary hydroxyl groups of alpha-, beta-, and gamma-cyclodextrins (CyDs) through an ester or amide linkage, and the in vivo drug release behavior of these prodrugs in rat gastrointestinal tracts after oral administration was investigated. The CyD prodrugs were stable in rat stomach and small intestine and negligibly absorbed from these tracts. Three to six hours after the oral administration, most of the prodrugs had moved to the cecum and colon. The alpha- and gamma-CyD amide prodrugs were hydrolyzed to the maltose conjugate in the cecum and colon, and these prodrugs and the conjugate were negligibly absorbed. On the other hand, the alpha- and gamma-CyD ester prodrugs produced BPAA in the cecum and colon, and BPAA appeared in the blood after 3-6 h. Both beta-CyD amide and ester prodrugs released only small or negligible amounts of the maltose conjugate or BPAA in the cecum and colon, within 24 h, probably due to the low solubility in the biological media. Further, the antiinflammatory effect of the gamma-CyD ester prodrug was evaluated using the model of carageenan-induced acute edema in rat paw and compared with those of BPAA alone and the BPAA/beta-CyD complex prepared by the kneading method in a molar ratio of 1:1. In the case of the beta-CyD complex, a rapid antiinflammatory response was observed, compared to BPAA alone, because the drug is mainly absorbed from the small intestine after a fast dissolution of the complex. In sharp contrast, the gamma-CyD ester prodrug needed a fairly long lag time to exhibit the drug activity, because BPAA is produced after the prodrug had reached the cecum and colon. The present results clearly suggest that the CyD prodrug approach can provide a versatile means for constructions of not only colon-specific delivery systems but also delayed-release system of certain drugs. PMID:9607948

  11. Targeted enzyme prodrug therapy for metastatic prostate cancer – a comparative study of L-methioninase, purine nucleoside phosphorylase, and cytosine deaminase

    PubMed Central

    2014-01-01

    Background Enzyme prodrug therapy shows promise for the treatment of solid tumors, but current approaches lack effective/safe delivery strategies. To address this, we previously developed three enzyme-containing fusion proteins targeted via annexin V to phosphatidylserine exposed on the tumor vasculature and tumor cells, using the enzymes L-methioninase, purine nucleoside phosphorylase, or cytosine deaminase. In enzyme prodrug therapy, the fusion protein is allowed to bind to the tumor before a nontoxic drug precursor, a prodrug, is introduced. Upon interaction of the prodrug with the bound enzyme, an anticancer compound is formed, but only in the direct vicinity of the tumor, thereby mitigating the risk of side effects while creating high intratumoral drug concentrations. The applicability of these enzyme prodrug systems to treating prostate cancer has remained unexplored. Additionally, target availability may increase with the addition of low dose docetaxel treatment to the enzyme prodrug treatment, but this effect has not been previously investigated. To this end, we examined the binding strength and the cytotoxic efficacy (with and without docetaxel treatment) of these enzyme prodrug systems on the human prostate cancer cell line PC-3. Results All three fusion proteins exhibited strong binding; dissociation constants were 0.572 nM for L-methioninase-annexin V (MT-AV), 0.406 nM for purine nucleoside phosphorylase-annexin V (PNP-AV), and 0.061 nM for cytosine deaminase-annexin V (CD-AV). MT-AV produced up to 99% cell death (p < 0.001) with limited cytotoxicity of the prodrug alone. PNP-AV with docetaxel created up to 78% cell death (p < 0.001) with no cytotoxicity of the prodrug alone. CD-AV with docetaxel displayed up to 60% cell death (p < 0.001) with no cytotoxicity of the prodrug alone. Docetaxel treatment created significant increases in cytotoxicity for PNP-AV and CD-AV. Conclusions Strong binding of fusion proteins to the prostate cancer cells

  12. Hypoxia-dependent retinal toxicity of bioreductive anticancer prodrugs in mice.

    PubMed

    Lee, A E; Wilson, W R

    2000-02-15

    The bioreductive anticancer prodrug CI-1010 ((2R)-1-[(2-bromoethyl)amino]-3-(2-nitro-1H-imidazol-1-yl)-2-propanol hydrobromide) is an alkylating nitroimidazole which shows selective toxicity against hypoxic cells in murine tumors, but causes extensive apoptosis in the outer retina in rodents and monkeys. This irreversible retinal toxicity has terminated preclinical development of CI-1010. We have investigated whether such toxicity is due to physiological hypoxia in the retina, and whether it is a general feature of hypoxia-selective bioreductive drugs. Retinal damage was quantified by morphometric analysis of histological sections following treatment of female C57Bl6 mice. Both CI-1010 and tirapazamine (TPZ, 1,2,4-benzotriazin-3-amine 1,4-dioxide), a bioreductive drug in Phase III clinical trial, caused a time and dose-dependent loss of photoreceptor cells of the outer retina following administration of single intraperitoneal doses. The lesion caused by TPZ was qualitatively similar to that with CI-1010, but was less severe at equivalent fractions of the maximum tolerated dose (as defined by lethality). With both bioreductive drugs, lesion severity was increased if animals breathed 10% O(2) for 3 h after drug administration, while breathing 95% O(2)/5% CO(2) was protective. Other hypoxia-selective bioreductive drugs tested (the quinone porfiromycin, the anthraquinone N-oxide AQ4N and the nitrogen mustard prodrugs SN 23816 and SN 25341) did not cause retinal damage at their maximum tolerated doses. This study suggests that the retinal toxicity of bioreductive drugs might be avoided by manipulation of tissue hypoxia using 95% O(2)/5% CO(2), although this intervention could suppress antitumor activity. The finding that not all bioreductive drugs cause retinal toxicity suggests this toxicity can be avoided through appropriate drug design. PMID:10662604

  13. Identification of activating enzymes of a novel FBPase inhibitor prodrug, CS-917

    PubMed Central

    Kubota, Kazuishi; Inaba, Shin-ichi; Nakano, Rika; Watanabe, Mihoko; Sakurai, Hidetaka; Fukushima, Yumiko; Ichikawa, Kimihisa; Takahashi, Tohru; Izumi, Takashi; Shinagawa, Akira

    2015-01-01

    CS-917 (MB06322) is a selective small compound inhibitor of fructose 1,6-bisphosphatase (FBPase), which is expected to be a novel drug for the treatment of type 2 diabetes by inhibiting gluconeogenesis. CS-917 is a bisamidate prodrug and activation of CS-917 requires a two-step enzyme catalyzed reaction. The first-step enzyme, esterase, catalyzes the conversion of CS-917 into the intermediate form (R-134450) and the second-step enzyme, phosphoramidase, catalyzes the conversion of R-134450 into the active form (R-125338). In this study, we biochemically purified the CS-917 esterase activity in monkey small intestine and liver. We identified cathepsin A (CTSA) and elastase 3B (ELA3B) as CS-917 esterases in the small intestine by mass spectrometry, whereas we found CTSA and carboxylesterase 1 (CES1) in monkey liver. We also purified R-134450 phosphoramidase activity in monkey liver and identified sphingomyelin phosphodiesterase, acid-like 3A (SMPADL3A), as an R-134450 phosphoramidase, which has not been reported to have any enzyme activity. Recombinant human CTSA, ELA3B, and CES1 showed CS-917 esterase activity and recombinant human SMPDL3A showed R-134450 phosphoramidase activity, which confirmed the identification of those enzymes. Identification of metabolic enzymes responsible for the activation process is the requisite first step to understanding the activation process, pharmacodynamics and pharmacokinetics of CS-917 at the molecular level. This is the first identification of a phosphoramidase other than histidine triad nucleotide-binding protein (HINT) family enzymes and SMPDL3A might generally contribute to activation of the other bisamidate prodrugs. PMID:26171222

  14. Readily adaptable release kinetics of prodrugs using protease-dependent reversible PEGylation.

    PubMed

    Böttger, Roland; Knappe, Daniel; Hoffmann, Ralf

    2016-05-28

    Protein and peptide therapeutics with good in vitro activities often fail due to poor bioavailability, circulation lifetime, and immunogenicity. PEGylation, i.e. conjugation of polyethylene glycol (PEG), significantly improves serum stability and renal clearance besides reducing the immunogenicity and thus enhances pharmacokinetics and tolerance in vivo. Several PEGylated drugs are marketed including several top-selling blockbusters. However, PEGylation can mask the binding site, especially in peptides, and thereby reduce the activity drastically, which is only rarely compensated by the improved bioavailability. Prodrug strategies using temporary PEGylation, i.e. the authentic drug is released from a PEG-linked precursor by hydrolysis or enzymatic degradation, can overcome these weaknesses. Recently, we reported a strategy coupling PEG via a peptide linker cleaved C-terminally by trypsin-like proteases in blood to release the unmasked therapeutic peptide. Here, we designed twelve short peptide linkers (four or five residues) to tune the release-rates of oncocin Onc112, a proline-rich antimicrobial peptide. In 25% aqueous mouse serum, Onc112 was released with half-life times from 0.5 to 12h. When elongated N-terminally with 5kDa ɑ-methoxy-ω-mercapto PEG as thioether, the half-life times of the prodrugs ranged from 7 to 42h in full mouse serum. Conjugation of a 20kDa instead of the 5kDa PEG increased the half-life times more than twofold, whereas longer peptide linkers up to twelve residues increased them only slightly. In all cases, Onc112 was released continuously providing stable peptide levels for at least 16h. The kinetics will allow the specific design of PEG-linker-drug-combinations for optimizing the pharmacokinetics of promising peptide therapeutics. PMID:27067364

  15. A chimeric platelet-targeted urokinase prodrug selectively blocks new thrombus formation

    PubMed Central

    Fuentes, Rudy E.; Zaitsev, Sergei; Ahn, Hyun Sook; Hayes, Vincent; Kowalska, M. Anna; Lambert, Michele P.; Wang, Yuhuan; Siegel, Donald L.; Bougie, Daniel W.; Aster, Richard H.; Myers, Daniel D.; Stepanova, Victoria; Cines, Douglas B.; Muzykantov, Vladimir R.; Poncz, Mortimer

    2015-01-01

    The use of fibrinolytic agents to prevent new thrombus formation is limited by an increased risk of bleeding due to lysis of hemostatic clots that prevent hemorrhage in damaged blood vessels. We sought to develop an agent that provides thromboprophylaxis without carrying a significant risk of causing systemic fibrinolysis or disrupting hemostatic clots. We previously showed that platelet (PLT) α granule–delivered urokinase plasminogen activator (uPA) is highly effective in preventing thrombosis, while being associated with little systemic fibrinolysis or bleeding. Here, we generated a chimeric prodrug composed of a single-chain version of the variable region of an anti-αIIbβ3 mAb fused to a thrombin-activatable, low-molecular-weight pro-uPA (PLT/uPA-T). PLT/uPA-T recognizes human αIIbβ3 on both quiescent and activated platelets and is enzymatically activated specifically by thrombin. We found that this prodrug binds tightly to human platelets even after gel filtration, has a prolonged half-life in mice transgenic for human αIIb compared with that of uPA-T, and prevents clot formation in a microfluidic system. Importantly, in two murine injury models, PLT/uPA-T did not lyse preexisting clots, even when administration was delayed by as little as 10 minutes, while it concurrently prevented the development of nascent thrombi. Thus, PLT/uPA-T represents the prototype of a platelet-targeted thromboprophylactic agent that selectively targets nascent over preexisting thrombi. PMID:26690701

  16. An FtsZ-Targeting Prodrug with Oral Antistaphylococcal Efficacy In Vivo

    PubMed Central

    Kaul, Malvika; Mark, Lilly; Zhang, Yongzheng; Parhi, Ajit K.; LaVoie, Edmond J.

    2013-01-01

    The bacterial cell division protein FtsZ represents a novel antibiotic target that has yet to be exploited clinically. The benzamide PC190723 was among the first FtsZ-targeting compounds to exhibit in vivo efficacy in a murine infection model system. Despite its initial promise, the poor formulation properties of the compound have limited its potential for clinical development. We describe here the development of an N-Mannich base derivative of PC190723 with enhanced drug-like properties and oral in vivo efficacy. The N-Mannich base derivative (TXY436) is ∼100-fold more soluble than PC190723 in an acidic aqueous vehicle (10 mM citrate, pH 2.6) suitable for oral in vivo administration. At physiological pH (7.4), TXY436 acts as a prodrug, converting to PC190723 with a conversion half-life of 18.2 ± 1.6 min. Pharmacokinetic analysis following intravenous administration of TXY436 into mice yielded elimination half-lives of 0.26 and 0.96 h for the TXY436 prodrug and its PC190723 product, respectively. In addition, TXY436 was found to be orally bioavailable and associated with significant extravascular distribution. Using a mouse model of systemic infection with methicillin-sensitive Staphylococcus aureus or methicillin-resistant S. aureus, we show that TXY436 is efficacious in vivo upon oral administration. In contrast, the oral administration of PC190723 was not efficacious. Mammalian cytotoxicity studies of TXY436 using Vero cells revealed an absence of toxicity up to compound concentrations at least 64 times greater than those associated with antistaphylococcal activity. These collective properties make TXY436 a worthy candidate for further investigation as a clinically useful agent for the treatment of staphylococcal infections. PMID:24041882

  17. Sulforaphane Preconditioning Sensitizes Human Colon Cancer Cells towards the Bioreductive Anticancer Prodrug PR-104A

    PubMed Central

    Erzinger, Melanie M.; Bovet, Cédric; Hecht, Katrin M.; Senger, Sabine; Winiker, Pascale; Sobotzki, Nadine; Cristea, Simona; Beerenwinkel, Niko; Shay, Jerry W.; Marra, Giancarlo; Wollscheid, Bernd; Sturla, Shana J.

    2016-01-01

    The chemoprotective properties of sulforaphane (SF), derived from cruciferous vegetables, are widely acknowledged to arise from its potent induction of xenobiotic-metabolizing and antioxidant enzymes. However, much less is known about the impact of SF on the efficacy of cancer therapy through the modulation of drug-metabolizing enzymes. To identify proteins modulated by a low concentration of SF, we treated HT29 colon cancer cells with 2.5 μM SF. Protein abundance changes were detected by stable isotope labeling of amino acids in cell culture. Among 18 proteins found to be significantly up-regulated, aldo-keto reductase 1C3 (AKR1C3), bioactivating the DNA cross-linking prodrug PR-104A, was further characterized. Preconditioning HT29 cells with SF reduced the EC50 of PR-104A 3.6-fold. The increase in PR-104A cytotoxicity was linked to AKR1C3 abundance and activity, both induced by SF in a dose-dependent manner. This effect was reproducible in a second colon cancer cell line, SW620, but not in other colon cancer cell lines where AKR1C3 abundance and activity were absent or barely detectable and could not be induced by SF. Interestingly, SF had no significant influence on PR-104A cytotoxicity in non-cancerous, immortalized human colonic epithelial cell lines expressing either low or high levels of AKR1C3. In conclusion, the enhanced response of PR-104A after preconditioning with SF was apparent only in cancer cells provided that AKR1C3 is expressed, while its expression in non-cancerous cells did not elicit such a response. Therefore, a subset of cancers may be susceptible to combined food-derived component and prodrug treatments with no harm to normal tissues. PMID:26950072

  18. Phosphoryloxymethyl carbamates and carbonates--novel water-soluble prodrugs for amines and hindered alcohols.

    PubMed

    Safadi, M; Oliyai, R; Stella, V J

    1993-09-01

    Phosphoryloxymethyl carbonates and carbamates of the type R1R2X-CO-O-CH2-O-PO3(-2) (X = O or N) were evaluated as potentially novel water-soluble collapsible prodrugs for alcohols and amines. These were prepared by reaction of alpha-chloromethyl chloroformate with the starting alcohol or amine to give the corresponding alpha-chloromethyl carbonate or carbamate, respectively. Reaction with silver dibenzyl phosphate followed by debenzylation by hydrogenolysis gave the desired products. The aqueous chemical stability of the phosphoryloxymethylcarbonyl derivatives of 2-indanol (3a), beta-(3,4-dimethoxyphenyl)ethylamine (3b), and benzocaine (3c) were evaluated. The aqueous hydrolysis of 3a-3c resulted in regeneration of the parent alcohol or amines. As expected, the hydrolytic behaviors of these derivatives were found to differ from that of simple alkyl and aryl phosphomonoesters. The rates of hydrolysis were extremely rapid, with the dianionic phosphate species possessing a higher reactivity than the monoanionic species. This was attributed to the proximity of the phosphate group to the carbonyl moiety. The carbamate derivatives, 3b and 3c, displayed greater chemical stability compared to the carbonate derivative, 3a. Alkaline phosphatases-mediated hydrolysis of the phosphate ester bond in 3c led to a rapid cascade reaction resulting in regeneration of the parent amine, benzocaine. Although the alcohol derivative described here appeared to be too chemically unstable to be ideal as a prodrug, the derivatives of the amines might have some use. They are expected to be cleaved in vivo by alkaline phosphatases. PMID:8234176

  19. Sulforaphane Preconditioning Sensitizes Human Colon Cancer Cells towards the Bioreductive Anticancer Prodrug PR-104A.

    PubMed

    Erzinger, Melanie M; Bovet, Cédric; Hecht, Katrin M; Senger, Sabine; Winiker, Pascale; Sobotzki, Nadine; Cristea, Simona; Beerenwinkel, Niko; Shay, Jerry W; Marra, Giancarlo; Wollscheid, Bernd; Sturla, Shana J

    2016-01-01

    The chemoprotective properties of sulforaphane (SF), derived from cruciferous vegetables, are widely acknowledged to arise from its potent induction of xenobiotic-metabolizing and antioxidant enzymes. However, much less is known about the impact of SF on the efficacy of cancer therapy through the modulation of drug-metabolizing enzymes. To identify proteins modulated by a low concentration of SF, we treated HT29 colon cancer cells with 2.5 μM SF. Protein abundance changes were detected by stable isotope labeling of amino acids in cell culture. Among 18 proteins found to be significantly up-regulated, aldo-keto reductase 1C3 (AKR1C3), bioactivating the DNA cross-linking prodrug PR-104A, was further characterized. Preconditioning HT29 cells with SF reduced the EC50 of PR-104A 3.6-fold. The increase in PR-104A cytotoxicity was linked to AKR1C3 abundance and activity, both induced by SF in a dose-dependent manner. This effect was reproducible in a second colon cancer cell line, SW620, but not in other colon cancer cell lines where AKR1C3 abundance and activity were absent or barely detectable and could not be induced by SF. Interestingly, SF had no significant influence on PR-104A cytotoxicity in non-cancerous, immortalized human colonic epithelial cell lines expressing either low or high levels of AKR1C3. In conclusion, the enhanced response of PR-104A after preconditioning with SF was apparent only in cancer cells provided that AKR1C3 is expressed, while its expression in non-cancerous cells did not elicit such a response. Therefore, a subset of cancers may be susceptible to combined food-derived component and prodrug treatments with no harm to normal tissues. PMID:26950072

  20. Colistin Methanesulfonate Is an Inactive Prodrug of Colistin against Pseudomonas aeruginosa

    PubMed Central

    Bergen, Phillip J.; Li, Jian; Rayner, Craig R.; Nation, Roger L.

    2006-01-01

    There is a dearth of information on the pharmacodynamics of “colistin,” despite its increasing use as a last line of defense for treatment of infections caused by multidrug-resistant gram-negative organisms. The antimicrobial activities of colistin and colistin methanesulfonate (CMS) were investigated by studying the time-kill kinetics of each against a type culture of Pseudomonas aeruginosa in cation-adjusted Mueller-Hinton broth. The appearance of colistin from CMS spiked at 8.0 and 32 mg/liter was measured by high-performance liquid chromatography, which generated colistin concentration-time profiles. These concentration-time profiles were subsequently mimicked in other incubations, independent of CMS, by incrementally spiking colistin. When the cultures were spiked with CMS at either concentration, there was a substantial delay in the onset of the killing effect which was not evident until the concentrations of colistin generated from the hydrolysis of CMS had reached approximately 0.5 to 1 mg/liter (i.e., ∼0.5 to 1 times the MIC for colistin). The time course of the killing effect was similar when colistin was added incrementally to achieve the same colistin concentration-time course observed from the hydrolysis of CMS. Given that the killing kinetics of CMS can be accounted for by the appearance of colistin, CMS is an inactive prodrug of colistin with activity against P. aeruginosa. This is the first study to demonstrate the formation of colistin in microbiological media containing CMS and to demonstrate that CMS is an inactive prodrug of colistin. These findings have important implications for susceptibility testing involving “colistin,” in particular, for MIC measurement and for microbiological assays and pharmacokinetic and pharmacodynamic studies. PMID:16723551

  1. Modification of existing antibiotics in the form of precursor prodrugs that can be subsequently activated by nitroreductases of the target pathogen.

    PubMed

    Çelik, Ayhan; Yetiş, Gülden; Ay, Mehmet; Güngör, Tuğba

    2016-08-15

    The use of existing antibiotics in the form of prodrug followed by activation using enzymes of pathogenic origin could be a useful approach for antimicrobial therapy. To investigate this idea, a common antibiotic, sulfamethoxazole has been redesigned in the form of a prodrug by simple functional group replacement. Upon reductive activation by a type I nitroreductase from a pathogen, the drug displayed enhanced antimicrobial capacity. This strategy could improve the efficacy and selectively of antibiotics and reduce the incidence of resistance. PMID:27390065

  2. Comparison of two self-assembled macromolecular prodrug micelles with different conjugate positions of SN38 for enhancing antitumor activity

    PubMed Central

    Liu, Yi; Piao, Hongyu; Gao, Ying; Xu, Caihong; Tian, Ye; Wang, Lihong; Liu, Jinwen; Tang, Bo; Zou, Meijuan; Cheng, Gang

    2015-01-01

    7-Ethyl-10-hydroxycamptothecin (SN38), an active metabolite of irinotecan (CPT-11), is a remarkably potent antitumor agent. The clinical application of SN38 has been extremely restricted by its insolubility in water. In this study, we successfully synthesized two macromolecular prodrugs of SN38 with different conjugate positions (chitosan-(C10-OH)SN38 and chitosan-(C20-OH)SN38) to improve the water solubility and antitumor activity of SN38. These prodrugs can self-assemble into micelles in aqueous medium. The particle size, morphology, zeta potential, and in vitro drug release of SN38 and its derivatives, as well as their cytotoxicity, pharmacokinetics, and in vivo antitumor activity in a xenograft BALB/c mouse model were studied. In vitro, chitosan-(C10-OH)SN38 (CS-(10s)SN38) and chitosan-(C20-OH) SN38 (CS-(20s)SN38) were 13.3- and 25.9-fold more potent than CPT-11 in the murine colon adenocarcinoma cell line CT26, respectively. The area under the curve (AUC)0–24 of SN38 after intravenously administering CS-(10s)SN38 and CS-(20s)SN38 to Sprague Dawley rats was greatly improved when compared with CPT-11 (both P<0.01). A larger AUC0–24 of CS-(20s)SN38 was observed when compared to CS-(10s)SN38 (P<0.05). Both of the novel self-assembled chitosan-SN38 prodrugs demonstrated superior anticancer activity to CPT-11 in the CT26 xenograft BALB/c mouse model. We have also investigated the differences between these macromolecular prodrug micelles with regards to enhancing the antitumor activity of SN38. CS-(20s)SN38 exhibited better in vivo antitumor activity than CS-(10s)SN38 at a dose of 2.5 mg/kg (P<0.05). In conclusion, both macromolecular prodrug micelles improved the in vivo conversion rate and antitumor activity of SN38, but the prodrug in which C20-OH was conjugated to macromolecular materials could be a more promising platform for SN38 delivery. PMID:25848251

  3. Genetically engineered pre-microRNA-34a prodrug suppresses orthotopic osteosarcoma xenograft tumor growth via the induction of apoptosis and cell cycle arrest

    PubMed Central

    Zhao, Yong; Tu, Mei-Juan; Wang, Wei-Peng; Qiu, Jing-Xin; Yu, Ai-Xi; Yu, Ai-Ming

    2016-01-01

    Osteosarcoma (OS) is the most common primary malignant bone tumor in children, and microRNA-34a (miR-34a) replacement therapy represents a new treatment strategy. This study was to define the effectiveness and safety profiles of a novel bioengineered miR-34a prodrug in orthotopic OS xenograft tumor mouse model. Highly purified pre-miR-34a prodrug significantly inhibited the proliferation of human 143B and MG-63 cells in a dose dependent manner and to much greater degrees than controls, which was attributed to induction of apoptosis and G2 cell cycle arrest. Inhibition of OS cell growth and invasion were associated with release of high levels of mature miR-34a from pre-miR-34a prodrug and consequently reduction of protein levels of many miR-34a target genes including SIRT1, BCL2, c-MET, and CDK6. Furthermore, intravenous administration of in vivo-jetPEI formulated miR-34a prodrug significantly reduced OS tumor growth in orthotopic xenograft mouse models. In addition, mouse blood chemistry profiles indicated that therapeutic doses of bioengineered miR-34a prodrug were well tolerated in these animals. The results demonstrated that bioengineered miR-34a prodrug was effective to control OS tumor growth which involved the induction of apoptosis and cell cycle arrest, supporting the development of bioengineered RNAs as a novel class of large molecule therapeutic agents. PMID:27216562

  4. Synthesis and pharmaceutical properties of N-acyloxymethyl prodrugs of Allop with potential anti-trypanosomal activity.

    PubMed

    Gualdesi, M S; Ortiz, C S; Raviolo, M A

    2016-04-01

    We report herein the synthesis, and the physicochemical and pharmacokinetic properties of N-acyloxymethyl prodrugs of allopurinol (Allop) (2a-f). Allop is a compound with activity against Trypanosoma cruzi, a causative agent of Chagas disease. Its pathology leads to a huge number of infections and deaths per year, because in addition to many sufferers only having limited access to health services only an inefficient chemotherapy is available. Relevant pharmaceutical properties (pKa, stability, solubility, lipophilicity, in vitro permeability, binding protein, xanthine oxidase binding) were also determined. The results obtained showed that derivatives behave as prodrugs of Allop, since they exhibit improved physicochemical and pharmacokinetic properties relative to their precursor. This behavior turns these compounds into active reservoirs of Allop, and reduces its unfavorable characteristics, so 2a-f compounds are excellent candidates for the treatment of Chagas disease. This work is therefore an important contribution leading to the suppression of Chagas disease. PMID:26181622

  5. Ring-Opening Polymerization of Prodrugs: A Versatile Approach to Prepare Well-Defined Drug Loaded Nanoparticles**

    PubMed Central

    Liu, Jinyao; Liu, Wenge; Weitzhandler, Isaac; Bhattacharyya, Jayanta; Li, Xinghai; Wang, Jing; Qi, Yizhi; Bhattacharjee, Somnath; Chilkoti, Ashutosh

    2014-01-01

    We report a new methodology for the synthesis of polymer-drug conjugates from “compound”—all in one—prodrug monomers that consist of a cyclic polymerizable group that is appended to a drug through a cleavable linker. We show that organocatalyzed ring-opening polymerization can polymerize these monomers into well-defined polymer prodrugs that are designed to self-assemble into nanoparticles and release drug in response to a physiologically relevant stimulus. This method is compatible with structurally diverse drugs and allows different drugs to be copolymerized with quantitative conversion of the monomers. The drug loading can be controlled by adjusting the monomer(s) to initiator feed ratio and drug release can be encoded into the polymer by the choice of linker. Initiating these monomers from a polyethylene glycol macroinitiator yields amphiphilic diblock copolymers that spontaneously self-assemble into micelles with a long plasma circulation, which is useful for systemic therapy. PMID:25427831

  6. Oxidative Halogenation of Cisplatin and Carboplatin: Synthesis, Spectroscopy, and Crystal and Molecular Structures of Pt(IV) Prodrugs

    PubMed Central

    Johnstone, Timothy C.; Alexander, Sarah M.; Wilson, Justin J.; Lippard, Stephen J.

    2014-01-01

    A series of Pt(IV) prodrugs has been obtained by oxidative halogenation of either cisplatin or carboplatin. Iodobenzene dichloride is a general reagent that cleanly provides prodrugs bearing axial chlorides without the need to prepare intervening Pt(IV) intermediates or handle chlorine gas. Elemental bromine and iodine afford Pt(IV) compounds as well, although in the case of the iodine-mediated oxidation of carboplatin, an amido-bridged Pt(IV) side product also formed. A detailed analysis of the changes in spectroscopic and structural parameters induced by varying the axial halide is presented. A number of recurring motifs are observed in the solid state structures of these compounds. PMID:25367395

  7. Asymmetric Synthesis of a CBI-Based Cyclic N-Acyl O-Amino Phenol Duocarmycin Prodrug

    PubMed Central

    2015-01-01

    A short, asymmetric synthesis of a cyclic N-acyl O-amino phenol duocarmycin prodrug subject to reductive activation based on the simplified 1,2,9,9a-tetrahydrocyclopropa[c]benz[e]indol-4-one (CBI) DNA alkylation subunit is described. A key element of the approach entailed treatment of iodo-epoxide 7, prepared by N-alkylation of 6 with (S)-glycidal 3-nosylate, with EtMgBr at room temperature to directly provide the optically pure alcohol 8 in 78% yield (99% ee) derived from an effective metal–halogen exchange and subsequent regioselective intramolecular 6-endo-tet cyclization. Following O-debenzylation, introduction of a protected N-methylhydroxamic acid, direct trannannular spirocyclization, and subsequent stereoelectronically controlled acid-catalyzed cleavage of the resulting cyclopropane (HCl), further improvements in a unique intramolecular cyclization with N–O bond formation originally introduced for formation of the reductively labile prodrug functionality are detailed. PMID:25247380

  8. Phospholipid-Based Prodrugs for Drug Targeting in Inflammatory Bowel Disease: Computational Optimization and In-Vitro Correlation.

    PubMed

    Dahan, Arik; Ben-Shabat, Shimon; Cohen, Noa; Keinan, Shahar; Kurnikov, Igor; Aponick, Aaron; Zimmermann, Ellen M

    2016-01-01

    In inflammatory bowel disease (IBD) patients, the enzyme phospholipase A2 (PLA2) is overexpressed in the inflamed intestinal tissue, and hence may be exploited as a prodrug-activating enzyme allowing drug targeting to the site(s) of gut inflammation. The purpose of this work was to develop powerful modern computational approaches, to allow optimized a-priori design of phospholipid (PL) based prodrugs for IBD drug targeting. We performed simulations that predict the activation of PL-drug conjugates by PLA2 with both human and bee venom PLA2. The calculated results correlated well with in-vitro experimental data. In conclusion, a-priori drug design using a computational approach complements and extends experimentally derived data, and may improve resource utilization and speed drug development. PMID:27086789

  9. Growth inhibition of Mycobacterium smegmatis by prodrugs of deoxyxylulose phosphate reducto-isomerase inhibitors, promising anti-mycobacterial agents.

    PubMed

    Ponaire, Sarah; Zinglé, Catherine; Tritsch, Denis; Grosdemange-Billiard, Catherine; Rohmer, Michel

    2012-05-01

    Since Mycobacterium tuberculosis sets up several multiple anti-tuberculosis drug resistance mechanisms, development of new drugs with innovative target is urgent. The methylerythritol phosphate pathway (MEP) involved in the biosynthesis of essential metabolites for the survival of mycobacteria, represents such a target. Fosmidomycin 1a and FR900098 1b, two inhibitors of DXR, do not affect the viability of M. tuberculosis cells, due to a lack of uptake. To overcome the absence of the mycobacterial cell wall crossing of these compounds, we synthesized and tested the inhibition potency of acyloxymethyl phosphonate esters as prodrugs of fosmidomycin 1a, FR900098 1b and their analogs 2a and 2b on Mycobacterium smegmatis. Only the prodrugs 4b-6b inhibit the bacterial growth and could be effective anti-mycobacterial agents. PMID:22405649

  10. The prodrug DHED selectively delivers 17β-estradiol to the brain for treating estrogen-responsive disorders

    PubMed Central

    Prokai, Laszlo; Nguyen, Vien; Szarka, Szabolcs; Garg, Puja; Sabnis, Gauri; Bimonte-Nelson, Heather A.; McLaughlin, Katie J.; Talboom, Joshua S.; Conrad, Cheryl D.; Shughrue, Paul J.; Gould, Todd D.; Brodie, Angela; Merchenthaler, Istvan; Koulen, Peter; Prokai-Tatrai, Katalin

    2015-01-01

    Many neurological and psychiatric maladies originate from the deprivation of the human brain from estrogens. However, current hormone therapies cannot be used safely to treat these conditions commonly associated with menopause because of detrimental side-effects in the periphery. The latter also prevents the use of the hormone for neuroprotection. Here we show that a small-molecule bioprecursor prodrug, 10β,17β-dihydroxyestra-1,4-dien-3-one (DHED), converts to 17β-estradiol in the brain after systemic administration, but remains inert in the rest of the body. The localized and rapid formation of estrogen from the prodrug was revealed by a series of in vivo bioanalytical assays and through in vivo imaging in rodents. DHED treatment efficiently alleviated symptoms originated from brain estrogen deficiency in animal models of surgical menopause and provided neuroprotection in a rat stroke model. Concomitantly, we determined that 17β-estradiol formed in the brain from DHED elicited changes in gene expression and neuronal morphology identical to those obtained after direct 17β-estradiol treatment. Altogether, complementary functional and mechanistic data show that our approach is highly relevant therapeutically, because administration of the prodrug selectively produces estrogen in the brain independently from the route of administration and treatment regimen. Therefore, peripheral responses associated with the use of systemic estrogens, such as stimulation of the uterus and estrogen-responsive tumor growth, were absent. Collectively, our brain-selective prodrug approach may safely provide estrogen neuroprotection and medicate neurological and psychiatric symptoms developing from estrogen deficiency, particularly those encountered after surgical menopause, without the adverse side-effects of current hormone therapies. PMID:26203081

  11. Dual Stimuli-Responsive Polymer Prodrugs Quantitatively Loaded by Nanoparticles for Enhanced Cellular Internalization and Triggered Drug Release.

    PubMed

    Huang, Mingming; Zhao, Kaijie; Wang, Lei; Lin, Shanqing; Li, Junjie; Chen, Jingbo; Zhao, Chengai; Ge, Zhishen

    2016-05-11

    Direct encapsulation of hydrophobic drugs into amphiphilic block copolymer micelles is frequently subjected to low drug loading efficiency (DLE) and loading content (DLC), as well as lower micellar stability and uncontrollable drug release. In this report, we prepare the copolymer prodrugs (PPEMA-co-PCPTM) via reversible addition-fragmentation chain transfer (RAFT) polymerization of 2-(piperidin-1-yl)ethyl methacrylate (PEMA) and reduction-responsive CPT monomer (CPTM), which were quantitatively encapsulated into poly(ethylene glycol)-block-poly(ε-caprolactone) (PEG-b-PCL) micelles. The polymer prodrug-loaded nanoparticles showed high stability for a long time in aqueous solution or blood serum and even maintain similar size after a lyophilization-dissolution cycle. The tumoral pH (∼6.8)-responsive properties of PPEMA segments endow the micellar cores with triggered transition from neutral to positively charged and swellable properties. The PEG-b-PCL nanoparticles loading polymer prodrugs (PPEMA-b-PCPTM) eliminated burst drug release. Simultaneously, CPT drug release can be triggered by reductive agents and solution pH. At pH 6.8, efficient cellular internalization was achieved due to positively charged cores of the nanoparticles. As compared with nanoparticles loading PCPTM, higher cytotoxicity was observed by the nanoparticles loading PPEMA-b-PCPTM at pH 6.8. Further multicellular tumor spheroid (MCTs) penetration and growth suppression studies demonstrated that high-efficiency penetration capability and significant size shrinkage of MCTs were achieved after treatment by PPEMA-b-PCPTM-loaded nanoparticles at pH 6.8. Therefore, the responsive polymer prodrug encapsulation strategy represents an effective method to overcome the disadvantages of common hydrophobic drug encapsulation approaches by amphiphilic block copolymer micelles and simultaneously endows the nanoparticles with responsive drug release behaviors as well as enhanced cellular internalization and

  12. Pros and cons of bifunctional platinum(IV) antitumor prodrugs: two are (not always) better than one.

    PubMed

    Gabano, Elisabetta; Ravera, Mauro; Osella, Domenico

    2014-07-14

    This article evaluates the efficacy and applicability of bifunctional prodrugs consisting of a six-coordinate Pt(iv) octahedral core and one or more bioactive molecules. The platinum(iv) complexes release upon reduction the corresponding cytotoxic Pt(ii) agents and the bioactive molecules, able to inhibit some biochemical mechanisms of cancer growth and/or prevent the deactivation of the Pt(ii) metabolites. PMID:24874896

  13. Direct retroviral delivery of human cytochrome P450 2B6 for gene-directed enzyme prodrug therapy of cancer.

    PubMed

    Kan, O; Griffiths, L; Baban, D; Iqball, S; Uden, M; Spearman, H; Slingsby, J; Price, T; Esapa, M; Kingsman, S; Kingsman, A; Slade, A; Naylor, S

    2001-07-01

    Human cytochrome P450 2B6 (CYP2B6) metabolizes the prodrug cyclophosphamide (CPA) to produce phosphoramide mustard that cross-links DNA leading to cell death. We have constructed a novel retroviral vector encoding CYP2B6 (designated "MetXia-P450") and used it to transduce the human tumor cell lines HT29 and T47D. MetXia-P450 transduction sensitised these cells to the cytotoxic effects of the prodrug CPA. Results from in vitro experiments demonstrated adverse effects on the clonogenic survival of cyclophosphamide-treated cells transduced with MetXia-P450. Cytotoxic activity accompanied by bystander effect was particularly evident in 3-D multicellular spheroid models suggesting that this in vitro system may be a more appropriate model for assessing the efficacy of gene directed-enzyme prodrug therapy (GDEPT). We have applied this approach in a clinically relevant gene therapy protocol on established subcutaneous tumor xenografts. These studies show for the first time the efficacy of a P450-based GDEPT strategy mediated by a direct retroviral gene transfer in vivo. PMID:11498768

  14. AS-924, a novel, orally active, bifunctional prodrug of ceftizoxime: physicochemical properties, oral absorption in animals, and antibacterial activity.

    PubMed

    Mori, N; Kodama, T; Sakai, A; Suzuki, T; Sugihara, T; Yamaguchi, S; Nishijima, T; Aoki, A; Toriya, M; Kasai, M; Hatano, S; Kitagawa, M; Yoshimi, A; Nishimura, K

    2001-11-01

    AS-924 is an oral prodrug of the antibiotic ceftizoxime (CTIZ), a parenteral use cephalosporin. This novel prodrug, produced by esterifying CTIZ with a lipophilic pivaloyloxymethyl (POM) group and introducing a water soluble L-alanyl group, is expected to increase the bioavailability and thereby, augment the antibacterial activity of CTIZ in vivo compared with existing prodrugs. To study the effect of the L-alanyl group in AS-924 on its bioavailability, the plasma concentration profiles of CTIZ in dogs were examined following the dosing of AS-924 and CTIZ-POM, in powder form, after pretreatment with the antacid ranitidine, and following the dosing of AS-924 after pretreatment with a gastrointestinal motility stimulant metoclopramide or suppressant scopolamine butylbromide. The absorption rate of AS-924 was constant under these different conditions due to its unique balance of lipophilicity and water solubility. CTIZ is as antibacterially active as pre-existing oral cephalosporins against Gram-positive clinical isolates, while being more active against all Gram-negative isolates-particularly Enterobacteriaceae and Haemophilus influenzae. A simulation model for the eradication profile of bacteria in computer programmed pharmacokinetic (PK) system was carried out to study the antibacterial action of CTIZ in human. CTIZ was proven to eradicate Streptococcus pneumoniae and H. influenzae effectively, while cefpodoxime (CPOD), the active moiety of CPOD proxetil, eradicated S. pneumoniae, but not H. influenzae. These results confirm that, AS-924 is a potent oral antibiotic and would be expected to be clinically effective and efficient. PMID:11711261

  15. Enzyme-functionalized vascular grafts catalyze in-situ release of nitric oxide from exogenous NO prodrug.

    PubMed

    Wang, Zhihong; Lu, Yaxin; Qin, Kang; Wu, Yifan; Tian, Yingping; Wang, Jianing; Zhang, Jimin; Hou, Jingli; Cui, Yun; Wang, Kai; Shen, Jie; Xu, Qingbo; Kong, Deling; Zhao, Qiang

    2015-07-28

    Nitric oxide (NO) is an important signaling molecule in cardiovascular system, and the sustained release of NO by endothelial cells plays a vital role in maintaining patency and homeostasis. In contrast, lack of endogenous NO in artificial blood vessel is believed to be the main cause of thrombus formation. In this study, enzyme prodrug therapy (EPT) technique was employed to construct a functional vascular graft by immobilization of galactosidase on the graft surface. The enzyme-functionalized grafts exhibited excellent catalytic property in decomposition of the exogenously administrated NO prodrug. Localized and on-demand release of NO was demonstrated by in vitro release assay and fluorescent probe tracing in an ex vivo model. The immobilized enzyme retained catalytic property even after subcutaneous implantation of the grafts for one month. The functional vascular grafts were implanted into the rat abdominal aorta with a 1-month monitoring period. Results showed effective inhibition of thrombus formation in vivo and enhancement of vascular tissue regeneration and remodeling on the grafts. Thus, we create an enzyme-functionalized vascular graft that can catalyze prodrug to release NO locally and sustainably, indicating that this approach may be useful to develop new cell-free vascular grafts for treatment of vascular diseases. PMID:26004323

  16. Self-assembled nanoparticles from hyaluronic acid-paclitaxel prodrugs for direct cytosolic delivery and enhanced antitumor activity.

    PubMed

    Xu, Chaoran; He, Wei; Lv, Yaqi; Qin, Chao; Shen, Lingjia; Yin, Lifang

    2015-09-30

    A prodrug-based nanosystem obtained by formulating prodrug and nanotechnology into a system is one of the most promising strategies to enhance drug delivery for disease treatment. Herein, we report a new nanosystem based on HA-PTX conjugates (HA-PTX Ns), which penetrated across cell membranes into cytosol, thus enhancing paclitaxel (PTX) delivery. HA-PTX Ns were successfully obtained based on HA-PTX, and their average particle size was approximately 200 nm. Importantly, unlike other prodrug-based nanosystems, HA-PTX Ns obtained cellular entry without entrapment within the lysosomal-endosomal system by using pathways including clathrin-mediated endocytosis, microtubule-associated internalization, macropinocytosis and cholesterol-dependence. Due to significant accumulation in tumors, HA-PTX Ns had more than a 4-fold decrease in tumor volume on day 14 in contrast with PTX alone. In conclusion, HA-PTX Ns could enter cells, bypass the lysosomal-endosomal system and improve PTX delivery. PMID:26232702

  17. Adipose Tissue–derived Mesenchymal Stem Cells Expressing Prodrug-converting Enzyme Inhibit Human Prostate Tumor Growth

    PubMed Central

    Cavarretta, Ilaria T; Altanerova, Veronika; Matuskova, Miroslava; Kucerova, Lucia; Culig, Zoran; Altaner, Cestmir

    2009-01-01

    The ability of human adipose tissue–derived mesenchymal stem cells (AT-MSCs), engineered to express the suicide gene cytosine deaminase::uracil phosphoribosyltransferase (CD::UPRT), to convert the relatively nontoxic 5-fluorocytosine (5-FC) into the highly toxic antitumor 5-fluorouracil (5-FU) together with their ability to track and engraft into tumors and micrometastases makes these cells an attractive tool to activate prodrugs directly within the tumor mass. In this study, we tested the feasibility and efficacy of these therapeutic cells to function as cellular vehicles of prodrug-activating enzymes in prostate cancer (PC) therapy. In in vitro migration experiments we have shown that therapeutic AT-MSCs migrated to all the prostate cell lines tested. In a pilot preclinical study, we observed that coinjections of human bone metastatic PC cells along with the transduced AT-MSCs into nude mice treated with 5-FC induced a complete tumor regression in a dose dependent manner or did not even allow the establishment of the tumor. More importantly, we also demonstrated that the therapeutic cells were effective in significantly inhibiting PC tumor growth after intravenous administration that is a key requisite for any clinical application of gene-directed enzyme prodrug therapies. PMID:19844197

  18. Preparation and characterization of poly(lactic-co-glycolic acid) microspheres loaded with a labile antiparkinson prodrug.

    PubMed

    D'Aurizio, E; van Nostrum, C F; van Steenbergen, M J; Sozio, P; Siepmann, F; Siepmann, J; Hennink, W E; Di Stefano, A

    2011-05-16

    L-dopa-α-lipoic acid (LD-LA) is a new multifunctional prodrug for the treatment of Parkinson's disease. In human plasma, LD-LA catechol esters and amide bonds are chemically and enzymatically cleaved, respectively, resulting in a half-life time of about fifty minutes. In the present work, the unstable LD-LA was entrapped into biodegradable poly(lactic-co-glycolic acid) (PLGA) microspheres designed as depot systems to protect this prodrug against degradation and to obtain a sustained release of the intact compound. The microspheres were prepared by an oil-in-water emulsion/solvent evaporation technique and the effect of formulation and processing parameters (polymer concentration in the organic solvent, volumes ratio of the phases, rate of the organic solvent evaporation) on microspheres characteristics (size, loading, morphology, release) was investigated. Also emphasis was given on the stability of the drug before and after release as well as on the underlying mass transport mechanisms controlling LD-LA release. Interestingly, when encapsulated in appropriate conditions into PLGA microspheres, the labile prodrug was stabilized and released via Fickian diffusion up to more than one week. PMID:21356295

  19. Bi-functional prodrugs of 5-aminolevulinic acid and butyric acid increase erythropoiesis in anemic mice in an erythropoietin-independent manner.

    PubMed

    Rephaeli, Ada; Tarasenko, Nataly; Fibach, Eitan; Rozic, Gabriela; Lubin, Ido; Lipovetsky, Julia; Furman, Svetlana; Malik, Zvi; Nudelman, Abraham

    2016-08-25

    Anemia is a major cause of morbidity and mortality worldwide resulting from a wide variety of pathological conditions. In severe cases it is treated by blood transfusions or injection of erythroid stimulating agents, e.g., erythropoietin (Epo), which can be associated with serious adverse effects. Therefore, there is a need to develop new treatment modalities. We recently reported that treatment of erythroleukemic cells with the novel the bi-functional prodrugs of 5-aminolevulinic acid (ALA) and butyric acid (BA), AN233 and AN908, enhanced hemoglobin (Hb) synthesis to a substantially higher level than did ALA and BA individually or their mixture. Herein, we describe that these prodrugs when given orally to mice induced histone deacetylase inhibition in the kidneys, bone marrow and spleen, thus, indicating good penetrability to the tissues. In mice where anemia was chemically induced, treatment with the prodrugs increased the Hb, the number of red blood cells (RBCs) and the percentage of reticulocytes to normal levels. The prodrugs had no adverse effects even after repeated treatment at 100-200mg/kg for 50days. The lack of increased levels of Epo in the blood of mice that were treated with the prodrugs suggests that AN233 and AN908 affected the Hb and RBC levels in an Epo-independent manner. Taken together with our previous studies, we propose that the prodrugs increase globin expression by BA inhibition of histone deacetylase and elevation heme synthesis by ALA. These results support an Epo-independent approach for treating anemia with these prodrugs. PMID:27283485

  20. Biotransformations of Antidiabetic Vanadium Prodrugs in Mammalian Cells and Cell Culture Media: A XANES Spectroscopic Study

    PubMed Central

    2016-01-01

    The antidiabetic activities of vanadium(V) and -(IV) prodrugs are determined by their ability to release active species upon interactions with components of biological media. The first X-ray absorption spectroscopic study of the reactivity of typical vanadium (V) antidiabetics, vanadate ([VVO4]3–, A) and a vanadium(IV) bis(maltolato) complex (B), with mammalian cell cultures has been performed using HepG2 (human hepatoma), A549 (human lung carcinoma), and 3T3-L1 (mouse adipocytes and preadipocytes) cell lines, as well as the corresponding cell culture media. X-ray absorption near-edge structure data were analyzed using empirical correlations with a library of model vanadium(V), -(IV), and -(III) complexes. Both A and B ([V] = 1.0 mM) gradually converged into similar mixtures of predominantly five- and six-coordinate VV species (∼75% total V) in a cell culture medium within 24 h at 310 K. Speciation of V in intact HepG2 cells also changed with the incubation time (from ∼20% to ∼70% VIV of total V), but it was largely independent of the prodrug used (A or B) or of the predominant V oxidation state in the medium. Subcellular fractionation of A549 cells suggested that VV reduction to VIV occurred predominantly in the cytoplasm, while accumulation of VV in the nucleus was likely to have been facilitated by noncovalent bonding to histone proteins. The nuclear VV is likely to modulate the transcription process and to be ultimately related to cell death at high concentrations of V, which may be important in anticancer activities. Mature 3T3-L1 adipocytes (unlike for preadipocytes) showed a higher propensity to form VIV species, despite the prevalence of VV in the medium. The distinct V biochemistry in these cells is consistent with their crucial role in insulin-dependent glucose and fat metabolism and may also point to an endogenous role of V in adipocytes. PMID:25906315

  1. Biotransformations of Antidiabetic Vanadium Prodrugs in Mammalian Cells and Cell Culture Media: A XANES Spectroscopic Study.

    PubMed

    Levina, Aviva; McLeod, Andrew I; Pulte, Anna; Aitken, Jade B; Lay, Peter A

    2015-07-20

    The antidiabetic activities of vanadium(V) and -(IV) prodrugs are determined by their ability to release active species upon interactions with components of biological media. The first X-ray absorption spectroscopic study of the reactivity of typical vanadium (V) antidiabetics, vanadate ([V(V)O4](3-), A) and a vanadium(IV) bis(maltolato) complex (B), with mammalian cell cultures has been performed using HepG2 (human hepatoma), A549 (human lung carcinoma), and 3T3-L1 (mouse adipocytes and preadipocytes) cell lines, as well as the corresponding cell culture media. X-ray absorption near-edge structure data were analyzed using empirical correlations with a library of model vanadium(V), -(IV), and -(III) complexes. Both A and B ([V] = 1.0 mM) gradually converged into similar mixtures of predominantly five- and six-coordinate V(V) species (∼75% total V) in a cell culture medium within 24 h at 310 K. Speciation of V in intact HepG2 cells also changed with the incubation time (from ∼20% to ∼70% V(IV) of total V), but it was largely independent of the prodrug used (A or B) or of the predominant V oxidation state in the medium. Subcellular fractionation of A549 cells suggested that V(V) reduction to V(IV) occurred predominantly in the cytoplasm, while accumulation of V(V) in the nucleus was likely to have been facilitated by noncovalent bonding to histone proteins. The nuclear V(V) is likely to modulate the transcription process and to be ultimately related to cell death at high concentrations of V, which may be important in anticancer activities. Mature 3T3-L1 adipocytes (unlike for preadipocytes) showed a higher propensity to form V(IV) species, despite the prevalence of V(V) in the medium. The distinct V biochemistry in these cells is consistent with their crucial role in insulin-dependent glucose and fat metabolism and may also point to an endogenous role of V in adipocytes. PMID:25906315

  2. The Rab GTPase-Activating Protein TBC1D4/AS160 Contains an Atypical Phosphotyrosine-Binding Domain That Interacts with Plasma Membrane Phospholipids To Facilitate GLUT4 Trafficking in Adipocytes

    PubMed Central

    Tan, Shi-Xiong; Ng, Yvonne; Burchfield, James G.; Ramm, Georg; Lambright, David G.; Stöckli, Jacqueline

    2012-01-01

    The Rab GTPase-activating protein TBC1D4/AS160 regulates GLUT4 trafficking in adipocytes. Nonphosphorylated AS160 binds to GLUT4 vesicles and inhibits GLUT4 translocation, and AS160 phosphorylation overcomes this inhibitory effect. In the present study we detected several new functional features of AS160. The second phosphotyrosine-binding domain in AS160 encodes a phospholipid-binding domain that facilitates plasma membrane (PM) targeting of AS160, and this function is conserved in other related RabGAP/Tre-2/Bub2/Cdc16 (TBC) proteins and an AS160 ortholog in Drosophila. This region also contains a nonoverlapping intracellular GLUT4-containing storage vesicle (GSV) cargo-binding site. The interaction of AS160 with GSVs and not with the PM confers the inhibitory effect of AS160 on insulin-dependent GLUT4 translocation. Constitutive targeting of AS160 to the PM increased the surface GLUT4 levels, and this was attributed to both enhanced AS160 phosphorylation and 14-3-3 binding and inhibition of AS160 GAP activity. We propose a model wherein AS160 acts as a regulatory switch in the docking and/or fusion of GSVs with the PM. PMID:23045393

  3. The tyrosine kinase BceF and the phosphotyrosine phosphatase BceD of Burkholderia contaminans are required for efficient invasion and epithelial disruption of a cystic fibrosis lung epithelial cell line.

    PubMed

    Ferreira, Ana S; Silva, Inês N; Fernandes, Fábio; Pilkington, Ruth; Callaghan, Máire; McClean, Siobhán; Moreira, Leonilde M

    2015-02-01

    Bacterial tyrosine kinases and their cognate protein tyrosine phosphatases are best known for regulating the biosynthesis of polysaccharides. Moreover, their roles in the stress response, DNA metabolism, cell division, and virulence have also been documented. The aim of this study was to investigate the pathogenicity and potential mechanisms of virulence dependent on the tyrosine kinase BceF and phosphotyrosine phosphatase BceD of the cystic fibrosis opportunistic pathogen Burkholderia contaminans IST408. The insertion mutants bceD::Tp and bceF::Tp showed similar attenuation of adhesion and invasion of the cystic fibrosis lung epithelial cell line CFBE41o- compared to the parental strain B. contaminans IST408. In the absence of bceD or bceF genes, B. contaminans also showed a reduction in the ability to translocate across polarized epithelial cell monolayers, demonstrated by a higher transepithelial electrical resistance, reduced flux of fluorescein isothiocyanate-labeled bovine serum albumin, and higher levels of tight junction proteins ZO-1, occludin, and claudin-1 present in monolayers exposed to these bacterial mutants. Furthermore, bceD::Tp and bceF::Tp mutants induced lower levels of interleukin-6 (IL-6) and IL-8 release than the parental strain. In conclusion, although the mechanisms of pathogenicity dependent on BceD and BceF are not understood, these proteins contribute to the virulence of Burkholderia by enhancement of cell attachment and invasion, disruption of epithelial integrity, and modulation of the proinflammatory response. PMID:25486990

  4. The Tyrosine Kinase BceF and the Phosphotyrosine Phosphatase BceD of Burkholderia contaminans Are Required for Efficient Invasion and Epithelial Disruption of a Cystic Fibrosis Lung Epithelial Cell Line

    PubMed Central

    Ferreira, Ana S.; Silva, Inês N.; Fernandes, Fábio; Pilkington, Ruth; Callaghan, Máire; McClean, Siobhán

    2014-01-01

    Bacterial tyrosine kinases and their cognate protein tyrosine phosphatases are best known for regulating the biosynthesis of polysaccharides. Moreover, their roles in the stress response, DNA metabolism, cell division, and virulence have also been documented. The aim of this study was to investigate the pathogenicity and potential mechanisms of virulence dependent on the tyrosine kinase BceF and phosphotyrosine phosphatase BceD of the cystic fibrosis opportunistic pathogen Burkholderia contaminans IST408. The insertion mutants bceD::Tp and bceF::Tp showed similar attenuation of adhesion and invasion of the cystic fibrosis lung epithelial cell line CFBE41o- compared to the parental strain B. contaminans IST408. In the absence of bceD or bceF genes, B. contaminans also showed a reduction in the ability to translocate across polarized epithelial cell monolayers, demonstrated by a higher transepithelial electrical resistance, reduced flux of fluorescein isothiocyanate-labeled bovine serum albumin, and higher levels of tight junction proteins ZO-1, occludin, and claudin-1 present in monolayers exposed to these bacterial mutants. Furthermore, bceD::Tp and bceF::Tp mutants induced lower levels of interleukin-6 (IL-6) and IL-8 release than the parental strain. In conclusion, although the mechanisms of pathogenicity dependent on BceD and BceF are not understood, these proteins contribute to the virulence of Burkholderia by enhancement of cell attachment and invasion, disruption of epithelial integrity, and modulation of the proinflammatory response. PMID:25486990

  5. Targeting of pegylated liposomal mitomycin-C prodrug to the folate receptor of cancer cells: Intracellular activation and enhanced cytotoxicity.

    PubMed

    Patil, Yogita; Amitay, Yasmine; Ohana, Patricia; Shmeeda, Hilary; Gabizon, Alberto

    2016-03-10

    Mitomycin C (MMC) is a powerful anti-bacterial, anti-fungal and anti-tumor antibiotic, often active against multidrug resistant cells. Despite a broad spectrum of antitumor activity, MMC clinical use is relatively limited due to its fast clearance and dose-limiting toxicity. To exploit the potential antitumor activity of MMC and reduce its toxicity we have previously developed a formulation of pegylated liposomes with a lipophilic prodrug of MMC (PL-MLP), activated by endogenous reducing agents which are abundant in the tumor cell environment in the form of different thiols. PL-MLP has minimal in vitro cytotoxicity unless reducing agents are added to the cell culture to activate the prodrug. In the present study, we hypothesized that targeting PL-MLP via folate receptors will facilitate intracellular activation of prodrug and enhance cytotoxic activity without added reducing agents. We grafted a lipophilic folate conjugate (folate-PEG(5000)-DSPE) to formulate folate targeted liposomes (FT-PL-MLP) and examined in vitro cell uptake and cytotoxic activity in cancer cell lines with high folate receptors (HiFR). 3H-cholesterol-hexadecyl ether (3H-Chol)-radiolabeled liposomes were prepared to study liposome-cell binding in parallel to cellular uptake of prodrug MLP. 3H-Chol and MLP cell uptake levels were 4-fold and 9-fold greater in KB HiFR cells when FT-PL-MLP is compared to non-targeted PL-MLP liposomes. The cytotoxic activity of FT-PL-MLP liposomes was significantly increased up to ~5-fold compared with PL-MLP liposomes in all tested HiFR expressing cell lines. The enhanced uptake and intracytoplasmic liposome delivery was confirmed by confocal fluorescence studies with Rhodamine-labeled liposomes. In vivo, no significant differences in pharmacokinetics and biodistribution were observed when PL-MLP was compared to FT-PL-MLP by the intravenous route. However, when liposomes were directly injected into the peritoneal cavity of mice with malignant ascites of J6456 Hi

  6. Carbon-carbon bond cleavage in activation of the prodrug nabumetone.

    PubMed

    Varfaj, Fatbardha; Zulkifli, Siti N A; Park, Hyoung-Goo; Challinor, Victoria L; De Voss, James J; Ortiz de Montellano, Paul R

    2014-05-01

    Carbon-carbon bond cleavage reactions are catalyzed by, among others, lanosterol 14-demethylase (CYP51), cholesterol side-chain cleavage enzyme (CYP11), sterol 17β-lyase (CYP17), and aromatase (CYP19). Because of the high substrate specificities of these enzymes and the complex nature of their substrates, these reactions have been difficult to characterize. A CYP1A2-catalyzed carbon-carbon bond cleavage reaction is required for conversion of the prodrug nabumetone to its active form, 6-methoxy-2-naphthylacetic acid (6-MNA). Despite worldwide use of nabumetone as an anti-inflammatory agent, the mechanism of its carbon-carbon bond cleavage reaction remains obscure. With the help of authentic synthetic standards, we report here that the reaction involves 3-hydroxylation, carbon-carbon cleavage to the aldehyde, and oxidation of the aldehyde to the acid, all catalyzed by CYP1A2 or, less effectively, by other P450 enzymes. The data indicate that the carbon-carbon bond cleavage is mediated by the ferric peroxo anion rather than the ferryl species in the P450 catalytic cycle. CYP1A2 also catalyzes O-demethylation and alcohol to ketone transformations of nabumetone and its analogs. PMID:24584631

  7. Tunable phosphatase-sensitive stable prodrugs of 5-aminolevulinic acid for tumor fluorescence photodetection.

    PubMed

    Babič, Andrej; Herceg, Viktorija; Ateb, Imène; Allémann, Eric; Lange, Norbert

    2016-08-10

    5-Aminolevulinic acid (5-ALA) has been at the forefront of small molecule based fluorescence-guided tumor resection and photodynamic therapy. 5-ALA and two of its esters received marketing authorization but suffer from several major limitations, namely low stability and poor pharmacokinetic profile. Here, we present a new class of 5-ALA derivatives aiming at the stabilization of 5-ALA by incorporating a phosphatase sensitive group, with or without self-cleavable linker. Compared to 5-ALA hexyl ester (5-ALA-Hex), these compounds display an excellent stability under acidic, basic and physiological conditions. The activation and conversion into the 5-ALA is controlled and can be structure-tailored. The prodrugs display reduced acute toxicity compared to 5-ALA-Hex with superior dose response profiles of protoporphyrin IX synthesis and fluorescence intensity in human glioblastoma cells in vitro. Clinically relevant fluorescence kinetics in vivo shown in U87MG glioblastoma spheroid tumor model in chick embryos provide a solid basis for their further development and translation to clinical fluorescence guided tumor resection and photodynamic therapy. PMID:27235981

  8. Prodrug Strategy for PSMA-targeted Delivery of TGX-221 to Prostate Cancer Cells

    PubMed Central

    Zhao, Yunqi; Duan, Shaofeng; Zeng, Xing; Liu, Chunjing; Davies, Neal M.; Li, Benyi; Forrest, M. Laird

    2013-01-01

    TGX-221 is a potent, selective, and cell membrane permeable inhibitor of the PI3K p110β catalytic subunit. Recent studies showed that TGX-221 has anti-proliferative activity against PTEN-deficient tumor cell lines including prostate cancers. The objective of this study was to develop an encapsulation system for parenterally delivering TGX-221 to the target tissue through a prostate-specific membrane aptamer (PSMAa10) with little or no side effects. In this study, PEG-PCL micelles were formulated to encapsulate the drug, and a prodrug strategy was pursued to improve the stability of the carrier system. Fluorescence imaging studies demonstrated that the cellular uptake of both drug and nanoparticles were significantly improved by targeted micelles in a PSMA positive cell line. The area under the plasma concentration time curve of the micelle formulation in nude mice was 2.27-fold greater than the naked drug, and the drug clearance rate was 17.5-fold slower. These findings suggest a novel formulation approach for improving site-specific drug delivery of a molecular-targeted prostate cancer treatment. PMID:22494444

  9. Ocular Disposition of the Hemiglutarate Ester Prodrug of ∆9-Tetrahydrocannabinol from Various Ophthalmic Formulations

    PubMed Central

    Hingorani, Tushar; Adelli, Goutham R.; Punyamurthula, Nagendra; Gul, Waseem; ElSohly, Mahmoud A.; Repka, Michael A.; Majumdar, Soumyajit

    2013-01-01

    Purpose The overall goal of this project is to enhance ocular delivery of ∆9-Tetrahydrocannabinol (THC) through the topical route. Methods Solubility, stability and in vitro transcorneal permeability of the relatively hydrophilic hemiglutarate ester derivative, THC-HG, was studied in the presence of surfactants. The solutions were characterized with respect to micelle size, zeta potential and solution viscosity. In vivo studies were carried out in New Zealand albino rabbits. A previously reported promising THC-HG ion-pair formulation was also studied in vivo. Results Aqueous solubility and stability and in vitro transcorneal permeability of THCHG was enhanced significantly in the presence of surfactants. THC levels in the ocular tissues (except cornea) were found to be below detection limits from mineral oil, surfactant or emulsion based formulations containing THC. In contrast, micellar and ion pair based THC-HG formulations produced significantly higher total THC concentrations in the anterior ocular chamber. Conclusion In this study, although delivery of THC to the anterior chamber ocular tissues could be significantly increased through the prodrug and formulation approaches tested, further studies are needed to increase penetration to the back-of-the eye. PMID:23737345

  10. Lisdexamfetamine prodrug activation by peptidase-mediated hydrolysis in the cytosol of red blood cells.

    PubMed

    Sharman, Johannah; Pennick, Michael

    2014-01-01

    Lisdexamfetamine dimesylate (LDX) is approved as a once-daily treatment for attention-deficit/hyperactivity disorder in children, adolescents, and adults in some countries. LDX is a prodrug comprising d-amphetamine covalently linked to l-lysine via a peptide bond. Following oral administration, LDX is rapidly taken up from the small intestine by active carrier-mediated transport, probably via peptide transporter 1. Enzymatic hydrolysis of the peptide bond to release d-amphetamine has previously been shown to occur in human red blood cells but not in several other tissues. Here, we report that LDX hydrolytic activity resides in human red blood cell lysate and cytosolic extract but not in the membrane fraction. Among several inhibitors tested, a protease inhibitor cocktail, bestatin, and ethylenediaminetetra-acetic acid each potently inhibited d-amphetamine production from LDX in cytosolic extract. These results suggest that an aminopeptidase is responsible for hydrolytic cleavage of the LDX peptide bond, although purified recombinant aminopeptidase B was not able to release d-amphetamine from LDX in vitro. The demonstration that aminopeptidase-like activity in red blood cell cytosol is responsible for the hydrolysis of LDX extends our understanding of the smooth and consistent systemic delivery of d-amphetamine by LDX and the long daily duration of efficacy of the drug in relieving the symptoms of attention-deficit/hyperactivity disorder. PMID:25489246

  11. Preclinical pharmacokinetics, pharmacology and toxicology of lisdexamfetamine: a novel d-amphetamine pro-drug.

    PubMed

    Hutson, Peter H; Pennick, Michael; Secker, Roger

    2014-12-01

    Lisdexamfetamine dimesylate (LDX) is a novel pro-drug of d-amphetamine that is currently used for the treatment of attention-deficit/hyperactivity disorder in children aged ≥ 6 years and adults. LDX is enzymatically cleaved to form d-amphetamine following contact with red blood cells, which reduces the rate of appearance and magnitude of d-amphetamine concentration in the blood and hence the brain when compared with immediate-release d-amphetamine at equimolar doses. Thus, the increase of striatal dopamine efflux and subsequent increase of locomotor activity following d-amphetamine is less prominent and slower to attain maximal effect following an equimolar dose of LDX. Furthermore, unlike d-amphetamine, the pharmacodynamic effects of LDX are independent of the route of administration underlining the requirement to be hydrolyzed by contact with red blood cells. It is conceivable that these pharmacokinetic and pharmacodynamic differences may impact the psychostimulant properties of LDX in the clinic. This article reviews the preclinical pharmacokinetics, pharmacology, and toxicology of LDX. This article is part of the Special Issue entitled 'CNS Stimulants'. PMID:24594478

  12. Ticlopidine in Its Prodrug Form Is a Selective Inhibitor of Human NTPDase1

    PubMed Central

    Künzli, Beat

    2014-01-01

    Nucleoside triphosphate diphosphohydrolase-1 (NTPDase1), like other ectonucleotidases, controls extracellular nucleotide levels and consequently their (patho)physiological responses such as in thrombosis, inflammation, and cancer. Selective NTPDase1 inhibitors would therefore be very useful. We previously observed that ticlopidine in its prodrug form, which does not affect P2 receptor activity, inhibited the recombinant form of human NTPDase1 (Ki = 14 μM). Here we tested whether ticlopidine can be used as a selective inhibitor of NTPDase1. We confirmed that ticlopidine inhibits NTPDase1 in different forms and in different assays. The ADPase activity of intact HUVEC as well as of COS-7 cells transfected with human NTPDase1 was strongly inhibited by 100 µM ticlopidine, 99 and 86%, respectively. Ticlopidine (100 µM) completely inhibited the ATPase activity of NTPDase1 in situ as shown by enzyme histochemistry with human liver and pancreas sections. Ticlopidine also inhibited the activity of rat and mouse NTPDase1 and of potato apyrase. At 100 µM ticlopidine did not affect the activity of human NTPDase2, NTPDase3, and NTPDase8, nor of NPP1 and NPP3. Weak inhibition (10–20%) of NTPDase3 and -8 was observed at 1 mM ticlopidine. These results show that ticlopidine is a specific inhibitor of NTPDase1 that can be used in enzymatic and histochemistry assays. PMID:25180024

  13. Development of inhibitor-directed enzyme prodrug therapy (IDEPT) for prostate cancer.

    PubMed

    Martin, Stacy E; Ganguly, Tanushree; Munske, Gerhard R; Fulton, Melody D; Hopkins, Mark R; Berkman, Clifford E; Black, Margaret E

    2014-10-15

    Prostate cancer (PCa) is the second most common cause of cancer death among American men after lung cancer. Unfortunately, current therapies do not provide effective treatments for patients with advanced, metastatic, or hormone refractory disease. Therefore, we seek to generate therapeutic agents for a novel PCa treatment strategy by delivering a suicide enzyme (yCDtriple) to a cell membrane bound biomarker found on PCa cells (prostate-specific membrane antigen (PSMA)). This approach has resulted in a new PCa treatment strategy reported here as inhibitor-directed enzyme prodrug therapy (IDEPT). The therapeutic agents described were generated using a click chemistry reaction between the unnatural amino acid (p-azidophenylalanine (pAzF)) incorporated into yCDtriple and the dibenzylcyclooctyne moiety of our PSMA targeting agent (DBCO-PEG4-AH2-TG97). After characterization of the therapeutic agents, we demonstrate significant PCa cell killing of PSMA-positive cells. Importantly, we demonstrate that this click chemistry approach can be used to efficiently couple a therapeutic protein to a targeting agent and may be applicable to the ablation of other types of cancers and/or malignancies. PMID:25157916

  14. Studying the antiemetic effect of vitamin B6 for morning sickness: pyridoxine and pyridoxal are prodrugs.

    PubMed

    Matok, Ilan; Clark, Shannon; Caritis, Steve; Miodovnik, Menachem; Umans, Jason G; Hankins, Gary; Mattison, Donald R; Koren, Gideon

    2014-12-01

    Vitamin B6 has been known to possess antiemetic effects since 1942. This water soluble compound has several forms in the circulation including pyridoxine, pyridoxal, and pyridoxal phosphate. The active antiemetic form of vitamin B6 is unknown. This was a pre-specified substudy of a randomized, placebo-controlled trial comparing the antiemetic effect of the doxylamine-vitamin B6 combination (Diclectin®) (n = 131) to placebo (n = 126) in women with nausea and vomiting of pregnancy. Serum concentrations of pyridoxine, pyridoxal, and pyridoxal 5' phosphate (PLP) and doxylamine were measured on Days 4, 8, and 15. With Diclectin® exhibiting a significant antiemetic effect in pregnancy, serum concentrations of pyridoxine were unmeasurable in almost all patients and those of pyridoxal were undetectable in half of patients. In contrast, PLP was measurable at sustained, stable steady-state levels in all patients. Our data suggest that there is a correlation between PLP levels and PUQE score of morning sickness symptoms when pyridoxine and pyridoxal levels are undetectable, and hence they might be prodrugs of PLP, which may be the active antiemetic form of vitamin B6. PMID:25052410

  15. Manageable cytotoxicity of nanocapsules immobilizing D-amino acid oxidase via exogenous administration of nontoxic prodrug

    NASA Astrophysics Data System (ADS)

    Zhao, Yang; Zhu, Yingchun; Fu, Jingke

    2014-02-01

    D-Amino acid oxidase (DAO), which could catalyze generation of hydrogen peroxide with strong oxidbility and cytotoxicity, has become of interest as a biocatalyst for therapeutic treatments. Herein we report that amino-functional hollow mesoporous silica with large pore size (10.27 nm) and positively charged surface effectively immobilize DAO with negative charge. The adsorption, activity and stability of DAO are demonstrated to depend mainly on the amino-functionalization of surface. Significant cancer cell killing effect is observed when the cells are treated by the nanocapsules entrapping DAO together with D-alanine, showing distinct dose-dependency on concentration of the nanocapsules entrapping DAO or D-alanine. Nevertheless, the toxicity is completely neutralized by the addition of catalase, and anti-tumor effect is not observed when either the nanocapsules entrapping DAO or D-alanine is applied alone. The results indicate that cytotoxicity of the nanocapsules entrapping DAO could be managed by exogenous administration of nontoxic prodrug to tumor tissue, due to the stereoselectivity of DAO and the scarcity of its substrates in mammalian organisms. Thus, the method might be exploited as a potential treatment for cancer therapy.

  16. Molecular Basis of Prodrug Activation by Human Valacyclovirase, an [alpha]-Amino Acid Ester Hydrolase

    SciTech Connect

    Lai, Longsheng; Xu, Zhaohui; Zhou, Jiahai; Lee, Kyung-Dall; Amidon, Gordon L.

    2008-07-08

    Chemical modification to improve biopharmaceutical properties, especially oral absorption and bioavailability, is a common strategy employed by pharmaceutical chemists. The approach often employs a simple structural modification and utilizes ubiquitous endogenous esterases as activation enzymes, although such enzymes are often unidentified. This report describes the crystal structure and specificity of a novel activating enzyme for valacyclovir and valganciclovir. Our structural insights show that human valacyclovirase has a unique binding mode and specificity for amino acid esters. Biochemical data demonstrate that the enzyme hydrolyzes esters of {alpha}-amino acids exclusively and displays a broad specificity spectrum for the aminoacyl moiety similar to tricorn-interacting aminopeptidase F1. Crystal structures of the enzyme, two mechanistic mutants, and a complex with a product analogue, when combined with biochemical analysis, reveal the key determinants for substrate recognition; that is, a flexible and mostly hydrophobic acyl pocket, a localized negative electrostatic potential, a large open leaving group-accommodating groove, and a pivotal acidic residue, Asp-123, after the nucleophile Ser-122. This is the first time that a residue immediately after the nucleophile has been found to have its side chain directed into the substrate binding pocket and play an essential role in substrate discrimination in serine hydrolases. These results as well as a phylogenetic analysis establish that the enzyme functions as a specific {alpha}-amino acid ester hydrolase. Valacyclovirase is a valuable target for amino acid ester prodrug-based oral drug delivery enhancement strategies.

  17. Self-Immolative Polycations as Gene Delivery Vectors and Prodrugs Targeting Polyamine Metabolism in Cancer

    PubMed Central

    2015-01-01

    Polycations are explored as carriers to deliver therapeutic nucleic acids. Polycations are conventionally pharmacological inert with the sole function of delivering therapeutic cargo. This study reports synthesis of a self-immolative polycation (DSS-BEN) based on a polyamine analogue drug N1,N11-bisethylnorspermine (BENSpm). The polycation was designed to function dually as a gene delivery carrier and a prodrug targeting dysregulated polyamine metabolism in cancer. Using a combination of NMR and HPLC, we confirm that the self-immolative polycation undergoes intracellular degradation into the parent drug BENSpm. The released BENSpm depletes cellular levels of spermidine and spermine and upregulates polyamine catabolic enzymes spermine/spermidine N1-acetyltransferase (SSAT) and spermine oxidase (SMO). The synthesized polycations form polyplexes with DNA and facilitate efficient transfection. Taking advantage of the ability of BENSpm to sensitize cancer cells to TNFα-induced apoptosis, we show that DSS-BEN enhances the cell killing activity of TNFα gene therapy. The reported findings validate DSS-BEN as a dual-function delivery system that can deliver a therapeutic gene and improve the outcome of gene therapy as a result of the intracellular degradation of DSS-BEN to BENSpm and the subsequent beneficial effect of BENSpm on dysregulated polyamine metabolism in cancer. PMID:25153488

  18. Design of Enzymatically Cleavable Prodrugs of a Potent Platinum-Containing Anticancer Agent

    PubMed Central

    Ding, Song; Pickard, Amanda J.; Kucera, Gregory L.

    2014-01-01

    Using a versatile synthetic approach, a new class of potential ester prodrugs of highly potent, but systemically too toxic, platinum–acridine anticancer agents was generated. The new hybrids contain a hydroxyl group, which has been masked with a cleavable lipophilic acyl moiety. Both butanoic (butyric) and bulkier 2-propanepentanoic (valproic) esters were introduced. The goals of this design were to improve the drug-like properties (e.g., logD) and to reduce the systemic toxicity of the pharmacophore. Two distinct pathways by which the target compounds undergo effective ester hydrolysis, the proposed activating step, have been confirmed: platinum-assisted, self-immolative ester cleavage in a low-chloride environment (LC-ESMS, NMR spectroscopy) and enzymatic cleavage by human carboxylesterase-2 (hCES-2) (LC-ESMS). The valproic acid ester derivatives are the first example of a metal-containing agent cleavable by the pro-drug-converting enzyme. They show excellent chemical stability and reduced systemic toxicity. Preliminary results from screening in lung adenocarcinoma cell lines (A549, NCI-H1435) suggest that the mechanism of the valproic esters may involve intracellular deesterification. PMID:25303639

  19. Lisdexamfetamine prodrug activation by peptidase-mediated hydrolysis in the cytosol of red blood cells

    PubMed Central

    Sharman, Johannah; Pennick, Michael

    2014-01-01

    Lisdexamfetamine dimesylate (LDX) is approved as a once-daily treatment for attention-deficit/hyperactivity disorder in children, adolescents, and adults in some countries. LDX is a prodrug comprising d-amphetamine covalently linked to l-lysine via a peptide bond. Following oral administration, LDX is rapidly taken up from the small intestine by active carrier-mediated transport, probably via peptide transporter 1. Enzymatic hydrolysis of the peptide bond to release d-amphetamine has previously been shown to occur in human red blood cells but not in several other tissues. Here, we report that LDX hydrolytic activity resides in human red blood cell lysate and cytosolic extract but not in the membrane fraction. Among several inhibitors tested, a protease inhibitor cocktail, bestatin, and ethylenediaminetetra-acetic acid each potently inhibited d-amphetamine production from LDX in cytosolic extract. These results suggest that an aminopeptidase is responsible for hydrolytic cleavage of the LDX peptide bond, although purified recombinant aminopeptidase B was not able to release d-amphetamine from LDX in vitro. The demonstration that aminopeptidase-like activity in red blood cell cytosol is responsible for the hydrolysis of LDX extends our understanding of the smooth and consistent systemic delivery of d-amphetamine by LDX and the long daily duration of efficacy of the drug in relieving the symptoms of attention-deficit/hyperactivity disorder. PMID:25489246

  20. Synthesis, Characterization, and In Vitro Evaluation of New Ibuprofen Polymeric Prodrugs Based on 2-Hydroxypropyl Methacrylate

    PubMed Central

    Babazadeh, Mirzaagha; Sheidaei, Maryam; Abbaspour, Sara; Edjlali, Ladan

    2013-01-01

    The present research work describes the synthesis and evaluation of new acrylic-type polymeric systems having degradable ester bonds linked to ibuprofen as materials for drug delivery. Ibuprofen was linked to 2-hydroxy-propyl methacrylate by an activated ester methodology in a one-pot procedure with a high yield. The resulting material was copolymerized with either 2-hydroxyethyl methacrylate or methyl methacrylate (in 1:3 mole ratios) by the free radical polymerization method, utilizing azoisobutyronitrile at 65–70 °C. The characterization of the resulting products by FTIR, 1H NMR, 13C NMR, DSC, and elemental analysis confirmed their synthesis successfully. Ibuprofen release from the obtained polymers was preliminarily evaluated at different buffered solutions (pH 1, 7.4, and 10) into dialysis bags to show the capacity of prodrugs to release the drug under hydrolytic conditions. Detection of hydrolysis by UV spectroscopy at selected intervals showed that the drug can be released by selective hydrolysis of the ester bond at the side of the drug moiety. The release profiles indicated that the hydrolytic behavior of polymers is strongly based on the polymer hydrophilicity and the pH value of the hydrolysis solution. The results suggest that these polymers could be useful in controlled release systems. PMID:23641345

  1. Macromolecular (pro)drugs with concurrent direct activity against the hepatitis C virus and inflammation.

    PubMed

    Wohl, Benjamin M; Smith, Anton A A; Jensen, Bettina E B; Zelikin, Alexander N

    2014-12-28

    Macromolecular prodrugs (MPs) are a powerful tool to alleviate side-effects and improve the efficacy of the broad-spectrum antiviral agent ribavirin. In this work, we sought an understanding of what makes an optimal formulation within the macromolecular parameter space--nature of the polymer carrier, average molar mass, drug loading, or a good combination thereof. A panel of MPs based on biocompatible synthetic vinylic and (meth)acrylic polymers was tested in an anti-inflammatory assay with relevance to alleviating inflammation in the liver during hepatitis C infection. Pristine polymer carriers proved to have a pronounced anti-inflammatory activity, a notion which may prove significant in developing MPs for antiviral and anticancer treatments. With conjugated ribavirin, MPs revealed enhanced activity but also higher toxicity. Therapeutic windows and therapeutic indices were determined and discussed to reveal the most potent formulation and those with optimized safety. Polymers were also tested as inhibitors of replication of the hepatitis C viral RNA using a subgenomic viral replicon system. For the first time, negatively charged polymers are revealed to have an intracellular activity against hepatitis C virus replication. Concerted activity of the polymer and ribavirin afforded MPs which significantly increased the therapeutic index of ribavirin-based treatment. Taken together, the systematic investigation of the macromolecular space identified lead candidates with high efficacy and concurrent direct activity against the hepatitis C virus and inflammation. PMID:25451544

  2. Synthesis and Biological Evaluation of Sphingosine Kinase Substrates as Sphingosine-1-Phosphate Receptor Prodrugs

    PubMed Central

    Foss, Frank W.; Mathews, Thomas P.; Kharel, Yugesh; Kennedy, Perry C.; Snyder, Ashley H.; Davis, Michael D.; Lynch, Kevin R.; Macdonald, Timothy L.

    2009-01-01

    In the search for bioactive sphingosine 1-phosphate (S1P) receptor ligands, a series of 2-amino-2-heterocyclic-propanols were synthesized. These molecules were discovered to be substrates of human-sphingosine kinases 1 and 2 (SPHK1 and SPHK2). When phosphorylated, the resultant phosphates showed varied activities at the five sphingosine-1-phosphate (S1P) receptors (S1P1–5). Agonism at S1P1 was displayed in vivo by induction of lymphopenia. A stereochemical preference of the quaternary carbon was crucial for phosphorylation by the kinases and alters binding affinities at the S1P receptors. Oxazole and oxadiazole compounds are superior kinase substrates to FTY720, the prototypical prodrug immunomodulator, fingolimod (FTY720). The oxazole-derived structure was the most active for human SPHK2. Imidazole analogues were less active substrates for SPHKs, but more potent and selective agonists of the S1P1 receptor; additionally, the imidazole class of compounds rendered mice lymphopenic. PMID:19632123

  3. Adenoviral vectors for prodrug activation-based gene therapy for cancer

    PubMed Central

    Doloff, Joshua C.; Waxman, David J.

    2013-01-01

    Cancer cell heterogeneity is a common feature - both between patients diagnosed with the same cancer and within an individual patient’s tumor - and leads to widely different response rates to cancer therapies and the potential for the emergence of drug resistance. Diverse therapeutic approaches have been developed to combat the complexity of cancer, including individual treatment modalities designed to target tumor heterogeneity. This review discusses adenoviral vectors and how they can be modified to replicate in a cancer-specific manner and deliver therapeutic genes under multi-tiered regulation to target tumor heterogeneity, including heterogeneity associated with cancer stem cell-like subpopulations. Strategies that allow for combination of prodrug-activation gene therapy with a novel replication-conditional, heterogeneous tumor-targeting adenovirus are discussed, as are the benefits of using adenoviral vectors as tumor-targeting oncolytic vectors. While the anticancer activity of many adenoviral vectors has been well established in preclinical studies, only limited successes have been achieved in the clinic, indicating a need for further improvements in activity, specificity, tumor cell delivery and avoidance of immunogenicity. PMID:23869779

  4. High cell density cultivation of recombinant Escherichia coli for prodrug of recombinant human GLPs production.

    PubMed

    Zhou, Ying; Ma, Xue; Hou, Zheng; Xue, Xiaoyan; Meng, Jingru; Li, Mingkai; Jia, Min; Luo, Xiaoxing

    2012-09-01

    Glucagon-like peptide-1 (GLP-1)(2) has been attracting increasing interest on account of its prominent benefits in type 2 diabetes. However, its clinical applications are limited by the short half-life in vivo. To overcome this limitation, a new polymer of GLP-1 was developed by prodrug strategy. In this study a recombinant protein, rhGLPs, was successfully constructed, cloned into plasmid pET30a (+) and expressed in Escherichia coli ArcticExpress(DE3)RP in the form of inclusion body. The recombinant fusion protein productivity could be enhanced by high cell density culture of the recombinant strain. As a result, about 40 g wet weight cells per liter were obtained. The protein was purified by size-exclusion chromatography on a Superdex 75 column and refolded using reverse dilution and dialysis methods. SDS-PAGE, HPLC and MALDI-TOF mass spectrometry were undertaken to determine the purity and molecular weight of rhGLPs. Bioactivity assay revealed that it had glucose-lowering and insulin-releasing action in vivo. PMID:22771632

  5. Synthesis of an apionucleoside family and discovery of a prodrug with anti-HIV activity.

    PubMed

    Toti, Kiran S; Derudas, Marco; Pertusati, Fabrizio; Sinnaeve, Davy; Van den Broeck, Freya; Margamuljana, Lia; Martins, José C; Herdewijn, Piet; Balzarini, Jan; McGuigan, Christopher; Van Calenbergh, Serge

    2014-06-01

    We report the synthesis of a family of D- and L-furano-D-apionucleosides, their 3'-deoxy, as well as their 2',3'-dideoxy analogues with thymine and adenine nucleobases. Single carbon homologation of 1,2-O-isopropylidene-D-glycero-tetrafuranos-3-ulose (15) and optimized glycosylation conditions involving microwave irradiation were key to the successful synthesis of the target compounds. While all target nucleosides failed to show significant antiviral activity, we demonstrated that the triphosphate of 2',3'-deoxy-D-apio-D-furanoadenosine (1), in contrast to that of its D-apio-L-furanose epimer 2, was readily incorporated into a DNA template by HIV reverse transcriptase to act as a DNA chain terminator. This led us to convert adenine derivative 1 into two phosphoramidate prodrugs. ProTide 9b was found active against HIV-1 and HIV-2 (EC50 = 0.5-1.5 μM), indicating that the lack of activity of the parent nucleoside, and possibly also other members of the D-apio-D-furanose nucleoside family must be sought in the inefficient cellular conversion to the monophosphate. PMID:24804575