Christian Voitenleitner

Discovery of a Potent Boronic Acid Derived Inhibitor of the HCV RNA-Dependent RNA Polymerase.

A boronic acid moiety was found to be a critical pharmacophore for enhanced in vitro potency against wild-type hepatitis C replicons and known clinical polymorphic and resistant HCV mutant replicons. The synthesis, optimization, and structure-activity relationships associated with inhibition of HCV replication in a subgenomic replication system for a series of non-nucleoside boron-containing HCV RNA-dependent RNA polymerase (NS5B) inhibitors are described. A summary of the discovery of 3 (GSK5852), a molecule which entered clinical trials in subjects infected with HCV in 2011, is included.

Preclinical Characterization of GSK2336805, a Novel Inhibitor of Hepatitis C Virus Replication That Selects for Resistance in NS5A

ABSTRACT GSK2336805 is an inhibitor of hepatitis C virus (HCV) with picomolar activity on the standard genotype 1a, 1b, and 2a subgenomic replicons and exhibits a modest serum shift. GSK2336805 was not active on 22 RNA and DNA viruses that were profiled. We have identified changes in the N-terminal region of NS5A that cause a decrease in the activity of GSK2336805. These mutations in the genotype 1b replicon showed modest shifts in compound activity (<13-fold), while mutations identified in the genotype 1a replicon had a more dramatic impact on potency. GSK2336805 retained activity on chimeric replicons containing NS5A patient sequences from genotype 1 and patient and consensus sequences for genotypes 4 and 5 and part of genotype 6. Combination and cross-resistance studies demonstrated that GSK2336805 could be used as a component of a multidrug HCV regimen either with the current standard of care or in combination with compounds with different mechanisms of action that are still progressing through clinical development.

In Vitro Characterization of GSK2485852, a Novel Hepatitis C Virus Polymerase Inhibitor

ABSTRACT GSK2485852 (referred to here as GSK5852) is a hepatitis C virus (HCV) NS5B polymerase inhibitor with 50% effective concentrations (EC50s) in the low nanomolar range in the genotype 1 and 2 subgenomic replicon system as well as the infectious HCV cell culture system. We have characterized the antiviral activity of GSK5852 using chimeric replicon systems with NS5B genes from additional genotypes as well as NS5B sequences from clinical isolates of patients infected with HCV of genotypes 1a and 1b. The inhibitory activity of GSK5852 remained unchanged in these intergenotypic and intragenotypic replicon systems. GSK5852 furthermore displays an excellent resistance profile and shows a <5-fold potency loss across the clinically important NS5B resistance mutations P495L, M423T, C316Y, and Y448H. Testing of a diverse mutant panel also revealed a lack of cross-resistance against known resistance mutations in other viral proteins. Data from both the newer 454 sequencing method and traditional population sequencing showed a pattern of mutations arising in the NS5B RNA-dependent RNA polymerase in replicon cells exposed to GSK5852. GSK5852 was more potent than HCV-796, an earlier inhibitor in this class, and showed greater reductions in HCV RNA during long-term treatment of replicons. GSK5852 is similar to HCV-796 in its activity against multiple genotypes, but its superior resistance profile suggests that it could be an attractive component of an all-oral regimen for treating HCV.

Hepatitis C genotype 1a replicon improved through introduction of fitness mutations

The use of subgenomic replicon systems has long been a valuable screening tool for the discovery of small molecule antivirals against Hepatitis C virus. While genotype 1a replicon systems have been widely used in stable systems, use in transient assays has been hampered by low signal. Here we describe the generation of a more robust genotype 1a (H77) replicon through the introduction of two fitness mutations, NS4A-K1691R and NS4B-E1726G, for use in transient transfections. While these mutations significantly improved the signal to noise ratio, leading to more robust data, they have no effect on the potency of tool compounds against various targets of HCV, thereby making this new system a powerful tool for screening of compounds against the genotype 1a replicon.

818 PRECLINICAL CHARACTERIZATION OF GSK2485852, A NOVEL HCV POLYMERASE INHIBITOR

The search for direct acting antiviral (DAA) compounds that act against targets on the Hepatitis C virus or host proteins that play a crucial role in the replication of this virus is of high importance since the current standard of care does not achieve a sustained virological response (SVR) in many patients even after an extended treatment period. GSK2485852 is an inhibitor of the HCV NS5B polymerase and is highly active with EC50 values in the low nanomolar range in the genotype 1 and 2 subgenomic replicon system as well as the infectious HCV cell-culture system. In an attempt to elucidate potency of GSK2485852 beyond the standard replicon assays, we used chimeric replicon systems with NS5B genes from different genotypes as well as NS5B sequences from clinical isolates of patients infected with HCV of genotype 1a and 1b. The inhibitory activity of GSK2485852 remained unchanged on these intergenotypic and intragenotypic replicon systems. GSK2485852 furthermore displays an excellent resistance profile and shows less than a five-fold potency loss across clinically important NS5B resistance mutations like S282T, P495L, M423T, C316Y or Y448H. A diverse mutant panel tested also revealed a lack of cross resistance against known resistance mutations against other viral proteins. Data from both 454 and population sequencing showed a pattern of mutations arising in the NS5B RNA dependent RNA polymerase. GSK2485852 shows superior viral RNA reductions of up to five logs in genotype 1b replicons.

A randomized, double blind, dose escalation, first time in human study to assess the safety, tolerability, pharmacokinetics, and antiviral activity of single doses of GSK2485852 in chronically infected hepatitis C subjects

This first‐time‐in‐human, randomized, double‐blind, placebo‐controlled, dose‐escalation study assessed the safety, tolerability, pharmacokinetics, and antiviral activity of GSK2485852, a hepatitis C virus (HCV) NS5B inhibitor, in 27 chronically infected HCV genotype‐1 subjects. Subjects received GSK2485852 70, 420, and 70 mg with a moderate fat/caloric meal. Safety, pharmacokinetics, antiviral activity, HCV genotype/phenotype, and interleukin 28B genotype were evaluated. A statistically significant reduction in HCV ribonucleic acid (RNA) was observed after a single dose of 420 mg GSK2485852 (−1.33 log10 IU/mL) compared with placebo (−0.09 log10 IU/mL) at 24 hours post‐dose. Subjects receiving 70 mg GSK2485852 were exposed to concentrations above the protein‐adjusted 90% effective concentration for a short time; none experienced a significant decline in HCV RNA (−0.47 log10 copies/mL). GSK2485852 was readily absorbed; however, the observed geometric mean maximum plasma concentration (Cmax) and area under the curve (AUC) values were significantly lower than expected due to a higher‐than‐predicted‐oral clearance. Co‐administration with food reduced the AUC and Cmax of GSK2485852 by 40% and 70%, respectively. Two metabolites were detected in human blood with one having approximately 50% higher concentrations than those of the parent. GSK2485852 was well‐tolerated and exhibited antiviral activity after a single 420 mg dose in HCV subjects.