No abstract

GS-9669 is a highly optimized thumb site II nonnucleoside inhibitor of the hepatitis C virus (HCV) RNA polymerase, with a binding affinity of 1.35 nM for the genotype (GT) 1b protein. It is a selective inhibitor of HCV RNA replication, with a mean 50% effective concentration (EC 50 ) of <11 nM in genotype 1 and 5 replicon assays, but lacks useful activity against genotypes 2 to 4. The M423T mutation is readily generated clinically upon monotherapy with the thumb site II inhibitors filibuvir and lomibuvir, and it is notable that GS-9669 exhibited only a 3-fold loss in potency against this variant in the genotype 1b replicon. Rather than M423T, resistance predominantly tracks to residues R422K and L419M and residue I482L in GT 1b and 1a replicons, respectively. GS-9669 exhibited at least additive activity in combination with agents encompassing four other direct modes of action (NS3 protease, NS5A, NS5B via an alternative allosteric binding site, and NS5B nucleotide) as well as with alpha interferon or ribavirin in replicon assays. It exhibited high metabolic stability in in vitro human liver microsomal assays, which, in combination with its pharmacokinetic profiles in rat, dog, and two monkey species, is predictive of good human pharmacokinetics. GS-9669 is well suited for combination with other orally active, direct-acting antiviral agents in the treatment of genotype 1 chronic HCV infection. (This study has been registered at ClinicalTrials.gov under registration number NCT01431898.)

Investigation of thiophene-2-carboxylic acid HCV NS5B site II inhibitors, guided by measurement of cell culture medium binding, revealed the structure-activity relationships for intrinsic cellular potency. The pharmacokinetic profile was enhanced through incorporation of heterocyclic ethers on the N-alkyl substituent. Hydroxyl groups were incorporated to modulate protein binding. Intrinsic potency was further improved through enantiospecific introduction of an olefin in the N-acyl motif, resulting in the discovery of the phase 2 clinical candidate GS-9669. The unexpected activity of this compound against the clinically relevant NS5B M423T mutant, relative to the wild type, was shown to arise from both the N-alkyl substituent and the N-acyl group.

Because no currently available vaccine can prevent HIV infection, pre-exposure prophylaxis (PrEP) with antiretrovirals (ARVs) is an important tool for combating the HIV pandemic1,2. Long-acting ARVs promise to build on the success of current PrEP strategies, which must be taken daily, by reducing the frequency of administration3. GS-CA1 is a small-molecule HIV capsid inhibitor with picomolar antiviral potency against a broad array of HIV strains, including variants resistant to existing ARVs, and has shown long-acting therapeutic potential in a mouse model of HIV infection4. Here we show that a single subcutaneous administration of GS-CA1 provides long-term protection against repeated rectal simian–human immunodeficiency virus (SHIV) challenges in rhesus macaques. Whereas all control animals became infected after 15 weekly challenges, a single 300 mg kg−1 dose of GS-CA1 provided per-exposure infection risk reduction of 97% for 24 weeks. Pharmacokinetic analysis showed a correlation between GS-CA1 plasma concentration and protection from SHIV challenges. GS-CA1 levels greater than twice the rhesus plasma protein-adjusted 95% effective concentration conferred 100% protection in this model. These proof-of-concept data support the development of capsid inhibitors as a novel long-acting PrEP strategy in humans.

BackgroundGS-CA2, an analog of GS-CA1, is a novel and selective inhibitor of HIV-1 capsid function. Safety and pharmacokinetics (PK) of GS-CA2 is currently being evaluated in healthy human subjects. Herein, we present the anti-HIV activity and nonclinical PK of GS-CA2, demonstrating its potential as a first-in-class long-acting antiretroviral agent.MethodsGS-CA2 antiviral activity was evaluated in MT-4 cells and in human peripheral blood mononuclear cells (PBMCs) acutely infected with HIV-1 (IIIb) and clinical HIV-1 isolates, respectively. Standard in vitro methods were used to characterize compound lipophilicity (LogD), solubility and relative binding to cell culture and plasma proteins. Metabolic stability was assessed in cryopreserved hepatocytes. GS-CA2 PK parameters following intravenous and subcutaneous (SC) administration were assessed in rat and dog. GS-CA2 plasma concentrations were determined by HPLC-MS/MS.ResultsGS-CA2 showed potent and selective anti-HIV activity in MT-4 cells (EC50 = 0.1 nM; CC50 = 26.6 µM). In PBMCs, GS-CA2 displayed a mean EC50 of 0.05 nM (0.02–0.16 nM) against 23 HIV-1 clinical isolates representing all major subtypes. GS-CA2 is highly lipophilic (LogD of 3.7) with low aqueous solubility (<0.01 mg/mL) and low predicted clearance (CL) in human hepatocytes (0.01 L/h/kg). In rat and dog, GS-CA2 demonstrated low CL (<4% of liver blood flow). GS-CA2 PK in rat and dog exhibited sustained and slow drug release following a single SC administration. Factors including species, formulation, concentration, dose, volume, and number of injections were examined for the effect on systemic exposure over time. GS-CA2 plasma concentrations in dogs (Figure 1) were maintained above the human plasma protein binding-adjusted EC95 (4 nM) for the entire study duration (16 weeks).ConclusionGS-CA2 is a selective and first-in-class HIV capsid inhibitor with picomolar potency and potential to be clinically effective against a broad range of HIV-1 strains. In animals following a single SC injection, GS-CA2 maintained therapeutically relevant concentrations for >3 months. These nonclinical data support clinical development of GS-CA2 as a novel long-acting antiretroviral agent suitable for the treatment of HIV-1 infection. Disclosures J. Zheng, Gilead Sciences, Inc.: Employee, Salary. S. R. Yant, Gilead Sciences, Inc.: Employee, Salary. S. Ahmadyar, Gilead Sciences, Inc.: Employee, Salary. T. Y. Chan, Gilead Sciences, Inc.: Employee, Salary. A. Chiu, Gilead Sciences, Inc.: Employee, Salary. T. Cihlar, Gilead Sciences, Inc.: Employee, Salary. J. O. Link, Gilead Sciences, Inc.: Employee, Salary. B. Lu, Gilead Sciences, Inc.: Employee, Salary. J. Mwangi, Gilead Sciences, Inc.: Employee, Salary. W. Rowe, Gilead Sciences, Inc.: Employee, Salary. S. D. Schroeder, Gilead Sciences, Inc.: Employee, Salary. G. J. Stepan, Gilead Sciences, Inc.: Employee, Salary. K. W. Wang, Gilead Sciences, Inc.: Employee, Salary. R. Subramanian, Gilead Sciences, Inc.: Employee, Salary. W. C. Tse, Gilead Sciences, Inc.: Employee, ...