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A series of 2′-fluorinated C -nucleosides were prepared and tested for anti-HCV activity. Among them, the triphosphate of 2′-fluoro-2′- C -methyl adenosine C -nucleoside ( 15 ) was a potent and selective inhibitor of the NS5B polymerase and maintained activity against the S282T resistance mutant. A number of phosphoramidate prodrugs were then prepared and evaluated leading to the identification of the 1-aminocyclobutane-1-carboxylic acid isopropyl ester variant ( 53 ) with favorable pharmacokinetic properties including efficient liver delivery in animals.

Background GS-9256 is an inhibitor of HCV NS3 protease with a macrocyclic structure and novel phosphinic acid pharmacophore. Methods Key preclinical properties of GS-9256 including in vitro antiviral activity, cross-resistance and pharmacokinetic properties were investigated in non-human species. Results In genotype (GT) 1b Huh-luc cells with a replicon encoding luciferase, GS-9256 had a mean 50% effective concentration (EC50) value of 20.0 nM, with minimal cytotoxicity. Antiviral activity was similar in a number of additional GT1b and GT1a replicon cell lines. Similar potency was observed in chimeric replicons encoding the NS3 protease of GT1 clinical isolates. GS-9256 was less active in GT2a replicon cells (14.2-fold increase in EC50). Additive to synergistic in vitro antiviral activity was observed when GS-9256 was combined with other agents including interferon-α, ribavirin, NS5B polymerase inhibitors GS-6620 and tegobuvir, as well as the NS5A inhibitor ledipasvir. GS-9256 retained wild-type activity against all tested NS5B and NS5A inhibitor resistance mutations. GS-9256 was metabolically stable in microsomes and hepatocytes of tested species, including rodents, dogs and humans. GS-9256 had high bioavailability in mice (near 100%) and moderate bioavailability in rats (14%), dogs (21%) and monkeys (14%). Elimination half-lives were approximately 2 h in mice, 0.6 h in rats, 5 h in dogs and 4 h in monkey. A study in bile duct-cannulated rats indicated that the major route of elimination is through biliary excretion of unmetabolized GS-9256. Conclusions GS-9256 showed a favourable preclinical profile supportive of clinical development for the treatment of chronic HCV infection in GT1 patients.

The first synthesis of 1'-C-CN, 2'-F, 2'-C-Me pyrimidines is described. Anti-HCV activity was assessed and compared to the 1'-C-CN, 2'-C-Me as well as the 2'-F, 2'-C-Me pyrimidines. A phosphoramidate prodrug of the cytidine derivative showed activity in the low micromolar range against HCV replicons.

The first synthesis of 1'-cyano-2'-C-methyl pyrimidine nucleosides is described. Anti-HCV activity of these nucleosides and their nucleotide phosphoramidate prodrugs was assessed and compared to the 1'-unsubstituted counterparts and to the related 1'-cyano-2'-C-methyl C-nucleoside parent of GS-6620.

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.

A sulfonamide replacement of the P2–P3 amide bond in the context of macrocyclic HCV NS3 protease inhibitors was investigated. These analogs displayed good inhibitory potency in the absence of any P3 capping group. The synthesis and preliminary SAR are described.

Conversion of diethyl 1-amino-2-vinylcyclopropanephosphonate to ethyl 1-amino-2-vinylcyclopropanephosphinate was accomplished by using either nucleophilic or electrophilic carbon reagents. Hydrolysis of the phosphonate diethyl ester to the mono acid followed by oxalyl chloride treatment provided the phosphonomonochloridate, which was treated with nucleophilic organometallic agents to afford phosphinate ethyl esters. Alternatively, the chloridate was reduced to the phosphonous ethyl ester, and then alkylated with various electrophilic alkylating agents to obtain phosphinate ethyl esters.

The discovery of GS-9451 is reported. Modification of the P3 cap and P2 quinoline with a series of solubilizing groups led to the identification of potent HCV NS3 protease inhibitors with greatly improved pharmacokinetic properties in rats, dogs and monkeys.

A potent and novel class of product-like inhibitors of the HCV NS3 protease was discovered by employing a phosphinic acid as a carboxylate isostere. The replicon activity and pharmacokinetic profile of this series of compounds was optimized by exploring the substitution of the phosphinic acid, as well as conformationally constraining these compounds through macrocyclization. The syntheses and preliminary biological evaluation of these phosphinic acids is described.

A series of N1-alkyl pyrimidinediones were designed, synthesized and evaluated as HIV-1 non-nucleoside reverse transcriptase inhibitors (NNRTIs). Our efforts identified compound 10b, which represents the lead compound in this series with pharmacokinetics and antiviral potency that may support once-daily dosing.

A novel class of phosphonate derivatives was designed to mimic the interaction of product-like carboxylate based inhibitors of HCV NS3 protease. A phosphonic acid (compound 2) was demonstrated to be a potent HCV NS3 protease inhibitor, and a potential candidate for treating HCV infection. The syntheses and preliminary biological evaluation of this phosphonate class of inhibitor are described.

GS-9160 is a novel and potent inhibitor of human immunodeficiency virus type 1 (HIV-1) integrase (IN) that specifically targets the process of strand transfer. It is an authentic inhibitor of HIV-1 integration, since treatment of infected cells results in an elevation of two-long terminal repeat circles and a decrease of integration junctions. GS-9160 has potent and selective antiviral activity in primary human T lymphocytes producing a 50% effective concentration (EC50) of ∼2 nM, with a selectivity index (50% cytotoxic concentration/EC50) of ∼2,000. The antiviral potency of GS-9160 decreased by 6- to 10-fold in the presence of human serum. The antiviral activity of GS-9160 is synergistic in combination with representatives from three different classes of antiviral drugs, namely HIV-1 protease inhibitors, nonnucleoside reverse transcriptase inhibitors, and nucleotide reverse transcriptase inhibitors. Viral resistance selections performed with GS-9160 yielded a novel pattern of mutations within the catalytic core domain of IN; E92V emerged initially, followed by L74M. While E92V as a single mutant conferred 12-fold resistance against GS-9160, L74M had no effect as a single mutant. Together, these mutations conferred 67-fold resistance to GS-9160, indicating that L74M may potentiate the resistance caused by E92V. The pharmacokinetic profile of GS-9160 in healthy human volunteers revealed that once-daily dosing was not likely to achieve antiviral efficacy; hence, the clinical development of this compound was discontinued.

A diphosphate of a novel cyclopentyl based nucleoside phosphonate with potent inhibition of HIV reverse transcriptase (RT) (20, IC(50)=0.13 microM) has been discovered. In cell culture the parent phosphonate diacid 9 demonstrated antiviral activity EC(50)=16 microM, within two-fold of GS-9148, a prodrug of which is currently under clinical investigation, and within 5-fold of tenofovir (PMPA). In vitro cellular metabolism studies using 9 confirmed that the active diphosphate metabolite is produced albeit at a lower efficiency relative to GS-9148.

A series of C3 halobenzyl-substituted tricyclic HIV integrase inhibitors was prepared. Improvement in cell-based inhibitor potency was observed in comparison to previously disclosed tricyclic pyrroloquinolines carrying the ‘halobenzyl tail’ at the lactam nitrogen. Animal PK for several of the C3-substituted inhibitors was examined, with a dihaloaryl analog achieving good balance in protein-shifted EC50 and t1/2 in animal PK studies.

GS-9148 (2'-Fd4AP, 4) has been identified as a nucleoside phosphonate reverse transcriptase (RT) inhibitor with activity against wild-type HIV (EC(50)=12 microM). Unlike many clinical RT inhibitors, relevant reverse transcriptase mutants (M184V, K65R, 6-TAMs) maintain a susceptibility to 2'-Fd4AP that is similar to wild-type virus. The 2'-fluorine group was rationally designed into the molecule to improve the selectivity profile and in preliminary studies using HepG2 cells, compound 4 showed no measurable effect on mitochondrial DNA content indicating a low potential for mitochondrial toxicity.

A series of C5-aza tricyclic HIV integrase inhibitors was prepared. A highly potent and orally bioavailable compound (compound 9) was identified and selected for development.

A series of nucleoside phosphonate reverse transcriptase (RT) inhibitors have been synthesized and their anti-HIV activity and resistance profiles evaluated. The most potent analog [5-(6-amino-purin-9-yl)-2,5-dihydro-furan-2-yloxymethyl]-phosphonic acid (d4AP) demonstrated a HIV EC50 = 2.1 μM, and the most favorable resistance profile against HIV-1 variants with K65R, M184V or multiple thymidine analog mutations in RT.

Cyclic phosphonomethoxy pyrimidine nucleosides that are bioisosteres of the monophosphate metabolites of HIV reverse transcriptase (RT) inhibitors AZT, d4T, and ddC have been synthesized. The RT inhibitory activities of the phosphonates were reduced for both dideoxy (dd) and dideoxydidehydro (d4) analogs compared to the nucleosides. Bis-isopropyloxymethylcarbonyl (BisPOC) prodrugs were prepared on selected compounds and provided > 150-fold improvements in antiviral activity.

We report a study of the influence of Lewis acids upon the regioselectivity of ring opening of quinolinic[2,3]anhydrides to provide 2-(isopropoxycarbonyl)-nicotinic acids. In the presence of stoichiometric amounts of indium trifluoromethanesulfonate or lanthanum trifluoromethanesulfonate, the desired 2-position ester was generated with greater than 95:5 regioselectivity. This methodology was also applied to 6-methyl-[2,3]-quinoline to provide similar results.

AN1057A/B has shown potent activity against MRSA. A novel and concise route to the synthesis of its heterocycle core 2,5-diamino-5,6-dihydro-1H-pyrimidine-4-one is described. This methodology allows the synthesis of an array of analogs with different amine substitutions at the 2-position.

Graphical abstract A series of 1′-substituted analogs of 4-aza-7,9-dideazaadenosine C-nucleoside were prepared and evaluated for the potential as antiviral agents. These compounds showed a broad range of inhibitory activity against various RNA viruses. In particular, the whole cell potency against HCV when R = CN was attributed to inhibition of HCV NS5B polymerase and intracellular concentration of the corresponding nucleoside triphosphate., A series of 1′-substituted analogs of 4-aza-7,9-dideazaadenosine C-nucleoside were prepared and evaluated for the potential as antiviral agents. These compounds showed a broad range of inhibitory activity against various RNA viruses. In particular, the whole cell potency against HCV when R = CN was attributed to inhibition of HCV NS5B polymerase and intracellular concentration of the corresponding nucleoside triphosphate.

Several 1′-substituted analogs of Tubercidin C-nucleosides were prepared using a highly convergent synthesis. Good to high diastereoselectivity was achieved using a variety of nucleophiles targeting the 1′-position. The source for this stereoselectivity is herein proposed. It is thought to be attributed to a temperature-dependent chelation of the incoming nucleophile to either the 2′- or 3′-benzyloxy ether of the ribose core.

We report a study of the influence of Lewis acids upon the regioselectivity of ring opening of quinolinic[2,3]anhydrides to provide 2-(isopropoxycarbonyl)-nicotinic acids. In the presence of stoichiometric amounts of indium trifluoromethanesulfonate or lanthanum trifluoromethanesulfonate, the desired 2-position ester was generated with greater than 95:5 regioselectivity. This methodology was also applied to 6-methyl-[2,3]-quinoline to provide similar results.

As a class, nucleotide inhibitors (NIs) of the hepatitis C virus (HCV) nonstructural protein 5B (NS5B) RNA-dependent RNA polymerase offer advantages over other direct-acting antivirals, including properties, such as pangenotype activity, a high barrier to resistance, and reduced potential for drug-drug interactions. We studied the in vitro pharmacology of a novel C -nucleoside adenosine analog monophosphate prodrug, GS-6620. It was found to be a potent and selective HCV inhibitor against HCV replicons of genotypes 1 to 6 and against an infectious genotype 2a virus (50% effective concentration [EC 50 ], 0.048 to 0.68 μM). GS-6620 showed limited activities against other viruses, maintaining only some of its activity against the closely related bovine viral diarrhea virus (EC 50 , 1.5 μM). The active 5′-triphosphate metabolite of GS-6620 is a chain terminator of viral RNA synthesis and a competitive inhibitor of NS5B-catalyzed ATP incorporation, with K i / K m values of 0.23 and 0.18 for HCV NS5B genotypes 1b and 2a, respectively. With its unique dual substitutions of 1′-CN and 2′- C -Me on the ribose ring, the active triphosphate metabolite was found to have enhanced selectivity for the HCV NS5B polymerase over host RNA polymerases. GS-6620 demonstrated a high barrier to resistance in vitro . Prolonged passaging resulted in the selection of the S282T mutation in NS5B that was found to be resistant in both cellular and enzymatic assays (>30-fold). Consistent with its in vitro profile, GS-6620 exhibited the potential for potent anti-HCV activity in a proof-of-concept clinical trial, but its utility was limited by the requirement of high dose levels and pharmacokinetic and pharmacodynamic variability.

Rational drug design utilizing available X-ray crystal structures of sialic acid analogues bound to the active site of influenza virus neuraminidase has led to the discovery of a series of potent carbocyclic influenza neuraminidase inhibitors. From this series, GS 4104 (oseltamivir, TAMIFLU) has emerged as a promising antiviral for the treatment and prophylaxis of human influenza infection. This article will summarize the design, discovery, and development of oseltamivir as an oral therapeutic to treat influenza infection.

We herein describe an efficient synthesis of optically active diethyl 1-amino-2-vinylcyclopropane-1-phosphonate (analogous to 1-amino-2-vinylcyclopropane-1-carboxylate). The racemic phosphonate diethyl ester was obtained from an imine derived from aminomethylphosphonate diester and trans-1,4-dibromo-2-butene. Crystallizations of the dibenzoyl- l -tartaric acid salt allowed for separation of enantiomers. The enantiomerically pure material was used to synthesize an extremely potent tripeptide phosphonate inhibitor of HCV NS3 protease. X-ray crystal structure of the inhibitor bound to the HCV NS3 protease confirmed the absolute stereochemistry of the title compound.

Influenza virus neuraminidase (NA) catalyses the cleavage of sialic acid residues terminally linked to glycoproteins and glycolipids and plays an important role in the replication of the virus. Recently, several potent NA inhibitors have been synthesized based on the rational design of mimicking the transition state of the sialic acid cleavage. Zanamivir and oseltamivir (GS 4104, the prodrug of GS 4071) have emerged as promising influenza NA inhibitors for the treatment and prophylaxis of human influenza virus infection. This review describes the recent work toward the discovery and development of influenza NA inhibitors.

A facile construction of the 3,6-diamino-1,2,3,4-tetrahydropyridine-4-one class of compounds is described. Compound 2, a carbon analog at the N-3 position of TAN-1057A/B, was synthesized using this approach.

We have recently described GS 4071, a carbocyclic transition-state analog inhibitor of the influenza virus neuraminidase, which has potent inhibitory activity comparable to that of 4-guanidino-Neu5Ac2en (GG167; zanamivir) when tested against influenza A virus replication and neuraminidase activity in vitro. We now report that GS 4071 is active against several strains of influenza A and B viruses in vitro and that oral GS 4104, an ethyl ester prodrug which is converted to GS 4071 in vivo, is active in the mouse and ferret models of influenza virus infection. Oral administration of 10 mg of GS 4104 per kg of body weight per day caused a 100-fold reduction in lung homogenate viral titers and enhanced survival in mice infected with influenza A or B viruses. In ferrets, a 25-mg/kg dose of GS 4104 given twice daily reduced peak viral titers in nasal washings and eliminated constitutional responses to influenza virus infection including fever, increased nasal signs (sneezing, nasal discharge, mouth breathing), and decreased activity. Consistent with our demonstration that the parent compound is highly specific for influenza virus neuraminidases, no significant drug-related toxicity was observed after the administration of oral dosages of GS 4104 of up to 800 mg/kg/day for 14 days in nonclinical toxicology studies with rats. These results indicate that GS 4104 is a novel, orally active antiviral agent with the potential to be used for the prophylaxis and treatment of influenza A and B virus infections.

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 ( C max s; C max C max s of GS 4071 (C max = 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.

The carbocyclic transition state sialic acid analog GS4071 ([3R,4R,5S]-4-acetamido-5-amino-3-[1-ethylpropoxy]-1-cyclohexane-1 -carboxylic acid), a potent influenza virus neuraminidase inhibitor, was highly inhibitory to influenza A/NWS/33 (H1N1), A/Victoria/3/75 (H3N2), A/Shangdong/09/93 (H3N2) and B/Hong Kong/5/72 viruses in Madin Darby canine kidney (MDCK) cells. The 50% effective concentrations in these experiments ranged from 1.8 to 59.5 microM, with no cytotoxicity evident at 1000 microM, using inhibition of viral cytopathic effect determined visually and by neutral red dye uptake. The ethyl ester prodrug of GS4071, GS4104, administered by oral gavage (p.o.), had significant inhibitory effects on infections in mice induced by these viruses. Antiviral effects were seen as prevention of death, increase in mean day to death, inhibition of decline of arterial oxygen saturation, lessened lung consolidation and inhibition of infectious virus recovered from the lungs. No toxicity was seen in dosages up to 100 mg/kg/day (highest evaluated). Comparison experiments run versus the influenza A (H1N1) virus-induced infection using GS4104, GS4071 and the neuraminidase inhibitor zanamivir (GG167, 4-guanidino-Neu5Ac2en), all administered p.o., indicated a 10-fold or greater potency for inhibiting the infection by GS4104. The minimum effective dosage for GS4104 was 0.1 mg/kg/day, with the compound administered twice daily for 5 days beginning 4 h pre-virus exposure. Oral therapy with GS4104 could be delayed from 48 to at least 60 h after exposure of mice to influenza A (H1N1) virus and still render a significant antiviral effect, the time of delay being dependent on the viral challenge dose. Intranasal instillation of GS4071 and GG167 to mice infected with influenza virus was highly inhibitory to the infection, the minimum effective dosages to significantly prevent death being 0.01 mg/kg/day for GS4071 and 0.1 mg/kg/day for GG167. Caging of infected mice treated with 10 mg/kg/day of GS4104 with infected saline-treated animals did not transfer any influenza-inhibitory effect to the latter animals. These data provide strong evidence of the potential of orally administered GS4104 for treatment of influenza A and B virus infections in humans.

Potentially orally bioavailable prodrugs of the antiretroviral agent 9-[2-(phosphonomethoxy)propyl]adenine (PMPA) were evaluated. Alkyl methyl carbamates were synthesized by alkylation of PMPA with the corresponding alkyl chloromethyl carbonate and N-alkyl chloromethyl carbamate reagents. The prodrugs were evaluated for in vitro antiviral activity in addition to chemical and enzymic stability. The inhibition of human immunodeficiency virus type 1 (HIV-1) strain IIIB replication in MT-2 cells by the carbonate prodrugs was found to be 2.5-500-fold increased compared to PMPA. The alkyl methyl carbonates, except t-butyl methyl carbonate, had reasonable chemical stability at pH 2.2 and 7.4, but were rapidly converted to the corresponding monoester of PMPA in the presence of dog plasma. The alkyl methyl carbamate prodrugs such as N-t-butyl methyl carbamate were found to have high stability in vitro. Based on its chemical stability and good oral bioavailability, bis(POC)PMPA (isopropyl methyl carbonate) was chos...

The structure-activity relationships (SAR) for a new class of potent inhibitors (1) of influenza neuraminidase are described. Systematic modifications of substituents at the C-3, C-4, and C-5 positions of the carbocyclic ring were performed to establish fundamental SAR to assist in the design of potent inhibitors with activity against both of influenza A and B viruses.

The influence of C2-substitution of GS 4071 on the influenza neuraminidase inhibitory activity was investigated. The introduction of lipophilic substituents (chloro, methyl, and methylthio) at the C2 position resulted in a significant decrease of the activity. This result indicates that at the enzyme active site there is limited hydrophobic pocket for group at the C2 position of GS 4071.

A series of acyclic phosphonomethylether nucleosides were synthesized and then evaluated for inhibitory activity against respiratory viruses of clinical significance using CPE inhibition, neutral red uptake and virus yield reduction assays. Of the 20 compounds synthesized, none significantly inhibited influenza A or B viruses or respiratory syncytial virus strains A2, Long or 18537; the selective indices (SI) were less than 10. A new compound, GS-2128 (2R, 5R-9-[2,5-dihydro-5-(phosphonomethoxy)-2-furanyl]adenine; D4API), selectively inhibited adenovirus 5 (SI>10) as did GS-0577 (9-(3-hydroxy-2-phosphonylmethoxypropyl)-adenine; HPMPA) and GS-0504 [(S)-1-[3-hydroxy-2-(phosphonylmethoxypropyl)]-cytosine; HPMPC]. The 50% effective concentrations (EC50) ranged from 8–100 μg mL−1 and 50% cell inhibitory concentrations (CC50) from 40–1000 μg mL−1. All three compounds were also found to be active against laboratory strains and clinical isolates of adenovirus types 1, 2, 8 and 41 with EC50 values ranging from 0.2 to 10 μg mL−1. Two compounds, GS-438 (9-(2-phosphonylmethoxyethyl)guanine, PMEG) and GS-2542 (9-[3-phosphonomethoxy)methoxymethyl]guanine) inhibited parainfluenza virus 3 strain C243, with SI of 52 and >333, respectively. PMEG also inhibited measles virus strains CC, Halonen and Chicago with EC50 values ranging from 0.03–9 μg mL−1. These data suggest that these compounds should be considered for possible development as therapeutic agents for respiratory virus infections.

Purpose. A series of prodrugs designed to enhance the oral bioavailability of the antiretroviral agent 9-[(R)-2-(phosphonomethoxy)propyl]adenine (PMPA; 1) have been synthesized, including a bis-(acyloxymethyl) ester 2 and a series of bis-(alkoxycarbonyloxymethyl) esters 3-9. The in vitro biological stability andin vivo pharmacokinetics of these prodrugs were evaluated to support selection of a prodrug candidate for clinical evaluation.

GS-9451 is a selective hepatitis C virus (HCV) NS3 protease inhibitor in development for the treatment of genotype 1 (GT1) HCV infection. Key preclinical properties of GS-9451, including in vitro antiviral activity, selectivity, cross-resistance, and combination activity, as well as pharmacokinetic properties, were determined. In multiple GT1a and GT1b replicon cell lines, GS-9451 had mean 50% effective concentrations (EC 50 s) of 13 and 5.4 nM, respectively, with minimal cytotoxicity; similar potency was observed in chimeric replicons encoding the NS3 protease gene of GT1 clinical isolates. GS-9451 was less active in GT2a replicon cells (EC 50 = 316 nM). Additive to synergistic in vitro antiviral activity was observed when GS-9451 was combined with other agents, including alpha interferon, ribavirin, and the polymerase inhibitors GS-6620 and tegobuvir (GS-9190), as well as the NS5A inhibitor ledipasvir (GS-5885). GS-9451 retained wild-type activity against multiple classes of NS5B and NS5A inhibitor resistance mutations. GS-9451 was stable in hepatic microsomes and hepatocytes from human and three other tested species. Systemic clearance was low in dogs and monkeys but high in rats. GS-9451 showed good oral bioavailability in all three species tested. In rats, GS-9451 levels were ∼40-fold higher in liver than plasma after intravenous dosing, and elimination of GS-9451 was primarily through biliary excretion. Together, these results are consistent with the antiviral activity observed in a recent phase 1b study. The results of in vitro cross-resistance and combination antiviral assays support the ongoing development of GS-9451 in combination with other agents for the treatment of chronic HCV infection.

Aromatic analogues of sialic acid ( 2 and 3 ) have been synthesized as potential influenza neuraminidase inhibitors. Whereas compound 2 exhibited good neuraminidase inhibitory activity, compound 3 , possessing the glycerol side chain at the C 5 position, was not a neuraminidase inhibitor.

Among retinoic acid receptors (RARs) alpha, beta, and gamma, the messenger RNA level of RAR-gamma is the most readily detectable by Northern blotting in human and mouse skin. This observation suggests that RAR-gamma may play a critical role in the modulation of the therapeutic benefits and side effects of retinoids in skin. To test this hypothesis, 11 RAR-gamma selective retinoids were synthesized based on three related structures. Each compound was found to prefer RAR-gamma when assessed by retinoid-induced transcriptional activity (RAR-gammaRAR-betaRAR-alpha). The apparent Kd for binding to recombinant receptor protein was found to follow a similar trend. To correlate this receptor selectivity with in vivo activity, the compounds were tested topically in the Rhino mouse utriculi reduction and rabbit irritation models, two assays widely used to screen retinoids for efficacy and side effects, respectively. The results indicated that for these compounds, both efficacy in the utriculi reduction assay and irritation potential in rabbits correlated positively with the RAR-gamma transactivation activity, with r2 of 0.9 and 0.8, respectively. These data suggest that RAR-gamma is an important regulator of retinoic acid efficacy in skin and further, that the irritation associated with the use of retinoids is most likely a receptor-mediated process.

TAN-1057A/B has shown potent activity against MRSA. A new approach to the 2,5-diamino-5,6-dihydro-1 H -pyrimidine-4-one derivatives has been developed. TAN-1057A/B and its analogs were efficiently synthesized using this new approach.

A series of novel 1′-substituted analogues of 4-aza-7,9-dideazaadenosine C-nucleoside are prepared and tested in antiviral assays.

The conversions proceed via mono ester (II) by either nucleophilic or electrophilic alkylation to produce a broad spectrum of amino acid biosteres.

A series of 2′-C-methyl branched purine and pyrimidine C-nucleosides were prepared. Their anti-HCV activity and pharmacological properties were profiled, and compared with known 2′-C-Me N-nucleoside counterparts. In particular, 2′-C-Me 4-aza-7,9-dideazaadenosine C-nucleoside (2) was found to have potent and selective anti-HCV activity in vitro as well as a favorable pharmacokinetic profile and in vivo potential for enhanced potency over the corresponding N-nucleoside.

Regiospecific synthesis of alkylated A ring of arotinoids has been achieved by using Me3Al-TMSOSO2CF3 as a key reagent for conversion of carbonyl to a geminal dimethyl functionality.

A potent and novel class of phosphinic acid derived product-like inhibitors of the HCV NS3/4A protease was discovered previously. Modification of the phosphinic acid and quinoline heterocycle led to GS-9256 with potent cell-based activity and favorable pharmacokinetic parameters. Based on these attributes, GS-9256 was advanced to human clinical trial as a treatment for chronic infection with genotype 1 HCV.

A series of thymidine analogues substituted with alkoxy groups of the C-4 position of the furan ring were synthesized. Among these compounds, the methoxy analogue 9 was the most potent inhibitor of herpes simplex virus type 1 thymidine kinase.

A novel synthetic methodology for the hydroxymethyl substituted acyclic acetal functionality was developed, The rationally designed phosphonate analogue 3 of ganciclovir monophosphate was highly active against human cytomegalovirus.