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Abstract: Pyrimidine analogs have long found use over a broad chemotherapeutic spectrum. In an effort to further explore the antiviral potential of several uracil derivatives previously synthesized in our laboratories, a series of benzylated pyrimidines were designed and synthesized. Introduction of the benzyl residue onto the 5-phenylaminouracil scaffold was carried out using 2,4-bis(trimethylsilyloxy)pyrimidine with the corresponding benzyl bromides. Similarly, 1-benzyl-5-(benzylamino)- and 1-benzyl-5-(phenethylamino)uracils were obtained via amination of 1-benzyl-5-bromouracils with benzylamine or phenylethylamine. The results of the broad screen antiviral studies revealed that compounds 5 and 11 exhibit promising inhibitory activity against HIV-1 in CEM-SS culture. A 50% protective effect was observed at concentrations of 11.9 and 9.5μМ, respectively. Moreover, compounds 8 and 3 exhibited good inhibitory effects against EBV in АKАТА cell culture with EC50 values of 2.3 and 12μM, respectively. The synthesis and biological studies are detailed herein. [ABSTRACT FROM AUTHOR]

Abstract: Three nucleoside analogues, 3′-fluoro-2′,3′-dideoxythymidine (FLT), 3′-azido-2′,3′-dideoxythymidine (AZT), and 2′,3′-dideoxy-3′-thiacytidine (3TC) were conjugated with three different dicarboxylic acids to afford the long chain dicarboxylate esters of nucleosides. In general, dinucleoside ester conjugates of FLT and 3TC with long chain dicarboxylic acids exhibited higher anti-HIV activity than their parent nucleosides. Dodecanoate and tetradecanoate dinucleoside ester derivatives of FLT were found to be the most potent compounds with EC50 values of 0.8–1.0nM and 3–4nM against HIV-1US/92/727 and HIV-1IIIB cells, respectively. The anti-HIV activity of the 3TC conjugates containing long chain dicarboxylate diester (EC50 =3–60nM) was improved by 1.5–66 fold when compared to 3TC (EC50 =90–200nM). This study reveals that the symmetrical ester conjugation of dicarboxylic acids with a number of nucleosides results in conjugates with improved anti-HIV profile. [Copyright &y& Elsevier]

Mercaptobenzamide thioesters and thioethers are chemically simple HIV-1 maturation inhibitors with a unique mechanism of action, low toxicity, and a high barrier to viral resistance. A structure-activity relationship (SAR) profile based on 39 mercaptobenzamide prodrug analogs exposed divergent activity/toxicity roles for the internal and terminal amides. To probe the relationship between antiviral activity and toxicity, we generated an improved computational model for the binding of mercaptobenzamide thioesters (SAMTs) to the HIV-1 NCp7 C-terminal zinc finger, revealing the presence of a second low-energy binding orientation, hitherto undisclosed. Finally, using NMR-derived thiol–thioester exchange equilibrium constants, we propose that thermodynamics plays a role in determining the antiviral activity observed in the SAR profile. Image 1 • Synthesis of 39 mercaptobenzamide prodrug analogs. • Generation of improved mercaptobenzamide–NCp7 computational model. • Calculation of an activity limit. [ABSTRACT FROM AUTHOR]

Objective: Users' sensory perceptions and experiences of intravaginal products can inform acceptability and adherence. Focusing on the meanings women derive from formulation/device characteristics facilitates developers' design iterations toward optimizing user experience. We investigated how users of long-acting gels and intravaginal rings (IVRs) impute meaning to characteristics that may affect future product use.Study Design: Focus groups were conducted with contraceptive IVR and vaginal lubricant users. Current perceptibility science and historical theory on the cultural acceptability of fertility regulating methods informed the analysis.Results: A total of 21 IVR users and 29 lubricant users attended focus groups in which they manipulated products in their hands and discussed reactions to product characteristics. Participants used prior product experiences and sensory perceptions of prototype manipulations to inform meanings about product properties and performance for pregnancy, disease prevention, comfort, and perceived efficacy. The meanings derived from product characteristics depended on why the product would be used; a characteristic deemed problematic in one risk context may be considered preferable in another.Conclusions: Intravaginal product users create narratives that ascribe influence or causality to product characteristics. These meanings, whether correct or incorrect biologically, will shape vaginal product acceptability, use, and effectiveness.Implications: Long-acting and sustained-release drug delivery systems will be part of the multipurpose prevention continuum. Developers must consider how sensory experiences and culturally salient assumptions shape the meanings users make of product design characteristics. Those meanings will ultimately impact use and effectiveness. [ABSTRACT FROM AUTHOR]

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

Abstract: A number of 5′-O-dicarboxylic fatty acyl monoester derivatives of 3′-azido-3′-deoxythymidine (zidovudine, AZT), 2′,3′-didehydro-2′,3′-dideoxythymidine (stavudine, d4T), and 3′-fluoro-3′-deoxythymidine (alovudine, FLT) were synthesized to improve the lipophilicity and potentially the cellular delivery of parent polar 2′,3′-dideoxynucleoside (ddN) analogs. The compounds were evaluated for their anti-HIV activity. Three different fatty acids with varying chain length of suberic acid (octanedioic acid), sebacic acid (decanedioic acid), and dodecanedioic acid were used for the conjugation with the nucleosides. The compounds were evaluated for anti-HIV activity and cytotoxicity. All dicarboxylic ester conjugates of nucleosides exhibited significantly higher anti-HIV activity than that of the corresponding parent nucleoside analogs. Among all the tested conjugates, 5′-O-suberate derivative of AZT (EC50 =0.10nM) was found to be the most potent compound and showed 80-fold higher anti-HIV activity than AZT without any significant toxicity (TC50 >500nM). [Copyright &y& Elsevier]

In light of the increasing worldwide AIDS pandemic, there is a continuing need to develop new prevention strategies to inhibit the transmission of HIV-1. In the absence of a successful vaccine, topical microbicides represent the best strategies to reduce the epidemic. Following the success of HIV therapeutic cocktail strategies, combinations of microbicides including nucleotide reverse transcriptase inhibitors (NtRTIs) and nonnucleoside reverse transcriptase inhibitors (NNRTIs) may offer significant protection from infection over single agents. We have developed a combination microbicide gel formulation for the delivery of IQP-0528, a novel NNRTI, and tenofovir (TFV), a NtRTI. Gel formulations were evaluated based on quantitative viscoelastic and physiochemical evaluations defined by a target product profile (TPP). For the majority of the evaluations, the gel formulations behaved similarly; all showed shear thinning behavior, were stable, nontoxic, and active against HIV-1 infection. Gel formulation F2759 displayed increased drug release of 289 ± 100 µg/(cm2 h1/2) and a tissue permeability of 60 times the half maximal effective concentration (EC50) of TFV and 800 times the EC50 of IQP-0528. In addition, F2759 showed osmolality within TPP and the highest performance in gel spreading. We have identified a gel formulation to deliver a combination microbicide of IQP-0528 and TFV that has significant potential to prevent infection of HIV-1. © 2012 Wiley Periodicals, Inc. and the American Pharmacists Association J Pharm Sci 101:1423-1435, 2012 [ABSTRACT FROM AUTHOR]

Abstract: This paper describes the design, construction and characterization of the first anti-HIV drug delivery system that is triggered to release its contents in the presence of human semen. Microgel particles were synthesized with a crosslinker containing a peptide substrate for the seminal serine protease prostate specific antigen (PSA) and were loaded with the HIV-1 entry inhibitor sodium poly(styrene-4-sulfonate) (pSS). The particles were composed of N-2-hydroxyproplymethacrylamide and bis-methacrylamide functionalized peptides based on the PSA substrates GISSFYSSK and GISSQYSSK. Exposure to human seminal plasma (HSP) degraded the microgel network and triggered the release of the entrapped antiviral polymer. Particles with the crosslinker composed of the substrate GISSFYSSK showed 17 times faster degradation in seminal plasma than that of the crosslinker composed of GISSQYSSK. The microgel particles containing 1mol% GISSFYSSK peptide crosslinker showed complete degradation in 30h in the presence of HSP at 37°C and pSS released from the microgels within 30min reached a concentration of 10μg/mL, equivalent to the published IC90 for pSS. The released pSS inactivated HIV-1 in the presence of HSP. The solid phase synthesis of the crosslinkers, preparation of the particles by inverse microemulsion polymerization, HSP-triggered release of pSS and inactivation of HIV-1 studies are described. [Copyright &y& Elsevier]

Abstract: We describe the development of a new type of scaffold to target RNA structures. Multivalent binding oligomers (MBOs) are molecules in which multiple sidechains extend from a polyamine backbone such that favorable RNA binding occurs. We have used this strategy to develop MBO-based inhibitors to prevent the association of a protein–RNA complex, Tat–TAR, that is essential for HIV replication. In vitro binding assays combined with model cell-based assays demonstrate that the optimal MBOs inhibit Tat–TAR binding at low micromolar concentrations. Antiviral studies are also consistent with the in vitro and cell-based assays. MBOs provide a framework for the development of future RNA-targeting molecules. [Copyright &y& Elsevier]

Abstract: As a continuation of efforts to replace the metabolically labile methyl esters of lead alkenyldiarylmethanes (ADAMs) with stable bioisosteres, compounds bearing benzo[d]isoxazole and oxazolidine-2-one rings were designed and evaluated as a new series of potent HIV-1 non-nucleoside reverse transcriptase inhibitors with anti-HIV activity. All of the resulting ADAMs were found to inhibit HIV-1 RT with poly(rC)·oligo(dG) as the template primer. The most promising compound in this series was ADAM 3, with EC50 values of 40nM (vs HIV-1RF) and 20nM (vs HIV-1IIIB). Compound 3 also inhibited HIV-1 reverse transcriptase with an IC50 of 0.91μM. ADAM 4 has an antiviral EC50 of 0.6μM in CEM-SS cells and a plasma half-life of 51.4min. [Copyright &y& Elsevier]

Abstract: The alkenyldiarylmethanes (ADAMs) are currently being investigated as non-nucleoside HIV-1 reverse transcriptase inhibitors (NNRTIs) of potential value in the treatment of HIV infection and AIDS. During the course of these studies, a number of ADAM analogues have been identified that protect HIV-infected cells from the cytopathic effects of the virus by an unknown, HIV-1 RT-independent mechanism. Since the phosphodiesterase 4 family is required for HIV infection, the effect of various ADAMs on the activity of PDE4B2 was investigated in an effort to determine if the ADAMs could possibly be targeting phosphodiesterases. Six compounds representative of the ADAM class were tested for inhibition of cAMP hydrolysis by PDE4B2 enzymatic activity. Four ADAMs were found to be weak inhibitors of PDE4B2 and two of them were inactive. The experimental results are consistent with an antiviral mechanism that does not include inhibition of PDE4 isoforms. [Copyright &y& Elsevier]

Abstract: During studies on the alkenyldiarylmethane (ADAM) class of non-nucleoside reverse transcriptase inhibitors (NNRTIs), analogues were discovered that exhibit low micromolar and submicromolar cytotoxicities. Since the ADAMs are structurally related to the tubulin polymerization inhibitor CC-5079, a set of 14 ADAMs were tested for inhibition of tubulin polymerization in an attempt to identify the biological target responsible for their cytotoxicity. The results indicate that, overall, the ADAMs are poor inhibitors of tubulin polymerization. However, the two most cytotoxic compounds, 15 and 16, are in fact active as inhibitors of tubulin assembly with IC50 values of 3.7±0.3 and 2.8±0.2μM, respectively, and they both inhibit the binding of colchicine to tubulin. Both compounds were investigated for anticancer activity in the National Cancer Institute’s panel of 60 human cancer cell lines, and both compounds consistently displayed submicromolar cytotoxicities with mean-graph midpoint (MGM) values of 0.31±0.08 and 0.47±0.09μM, respectively. [Copyright &y& Elsevier]

Abstract: The HIV-1 non-nucleoside reverse transcriptase inhibitors (NNRTIs) constitute a large and structurally diverse set of compounds, several of which are currently used in the treatment of AIDS. A series of novel alkenyldiarylmethanes (ADAMs) were designed and synthesized as part of an ongoing investigation to replace the metabolically labile methyl ester moieties found in the ADAM pharmacophore with stable modifications that retain the potent anti-HIV activity of the parent compounds. Unsurprisingly, the rat plasma half-lives of the new ADAMs were not improved when compared to the parent compounds, but all of the synthesized ADAMs inhibited the cytopathic effect of HIV-1 in cell culture. The most potent compound identified was (E)-5-[1-(3,7-dimethyl-2-oxo-2,3-dihydro-benzoxazol-5-yl)-5-methoxycarbonyl-pent-1-enyl]-2-methoxy-3-methylbenzoic acid methyl ester (7), which inhibited the cytopathic effects of both HIV-1RF and HIV-1IIIB strains in cell cultures with EC50 values of 30 and 90nM, respectively, and inhibited HIV-1 reverse transcriptase with an IC50 of 20nM. [Copyright &y& Elsevier]