Combinations of 2'-C-Methylcytidine Analogues with Interferon-α2b and Triple Combination with Ribavirin in the Hepatitis C Virus Replicon System

Abstract

Background: Hepatitis C virus (HCV) polymerase is an essential enzyme for HCV replication and has multiple inhibitor binding sites making it a major target for antiviral intervention. It is apparent that no single drug can inhibit HCV replication in humans. Hence, combinations of nucleoside analogues β-d-2'-C-methylcytidine (2'-C-MeC; NM-107) or β-d-2'-deoxy-2'-fluoro-2'-C-methylcytidine (2'-F-C-MeC; PSI-6130) with interferon-a2b (IFN-a2b) or triple combination with ribavirin (RBV) were evaluated. Methods: Huh-7 cells containing the self-replicating subgenomic HCV replicon (Clone B) were used for drug combination studies. After drug treatment for 5 days, total cellular RNA was then extracted and both ribosomal RNA and HCV replicon RNA were amplified in a singlestep multiplex real-time PCR assay. Drug interaction analyses were performed using the CalcuSyn program. The current standard of care for chronic hepatitis C virus (HCV) infection is a combination of pegylated interferon (IFN)-a and ribavirin (RBV), which results in a sustained virological response in about a half of infected individuals; however, this combined modality is associated with considerable side effects [1,2]. Several new HCV inhibitors have been identified and some are being evaluated in controlled clinical trials [3]. Recently, promising 2′-modified nucleoside analogues with activity against HCV have been reported; for instance, valopicitabine (NM-283), the 3′-valine ester of β-d-2′-Cmethylcytidine (2′-C-MeC; NM-107), is a prodrug nucleoside that after intracellular phosphorylation to its 5′-triphosphate metabolite, selectively inhibits HCV RNA-dependent RNA polymerase [4,5]. Although valopicitabine was recently evaluated in clinical trials and the combination of valopicitabine with IFN-a (plus RBV) has led to successful viral RNA suppression in a number of HCV-infected individuals, this drug was found to have gastrointestinal toxicity leading to its discontinuation in its present form [6]. Another novel cytidine analogue, β-d-2′-deoxy-2′-fluoro-2′-C-methylcytidine (PSI-6130, 2′-F-C-MeC has demonstrated potent and specific in vitro anti-HCV activity with no apparent cytotoxicity [7,8]. Recently, in a 14 day Phase I monotherapy study, a new prodrug of 2′-F-C-MeC named R7128 was shown to be highly potent at lowering levels of HCV RNA, with a 2.7-log reduction in the viral load of individuals infected with HCV genotype 1 who had failed prior IFN therapy [9,10]. These nucleoside analogues have demonstrated excellent anti-HCV activity in vitro in both antireplicon and infectious HCV systems [11]. Furthermore, nucleoside analogue prodrugs can be administered in combination with IFN, with or without Results: Double combinations of 2'-C-MeC or 2'-F-C-

Intracellular Effects of the Hepatitis C Virus Nucleoside Polymerase Inhibitor RO5855 (Mericitabine Parent) and Ribavirin in Combination

“…The inhibitory activities (50% effective concentration [EC 50 ] and EC 90 ) of each compound, together with the percent inhibition observed with the maximal concentrations in the cell viability assay, were first determined (see Table S2 in the supplemental material). The inhibitory activities observed in the replicon assays were in agreement with previously published data, and all concentration ranges chosen for the drug-drug interaction analyses were below the cytotoxic concentrations for the compounds (47)(48)(49).…”

In Vitro Drug Combination Studies of Letermovir (AIC246, MK-8228) with Approved Anti-Human Cytomegalovirus (HCMV) and Anti-HIV Compounds in Inhibition of HCMV and HIV Replication

“…CIs were computed at the EC 50 , EC 75 , and EC 90 levels using CalcuSyn software (Biosoft). Since large effects are thought to be more therapeutically relevant than small effects, the CI wt (see Materials and Methods) was calculated, in addition (36,39).…”

Inhibitory effects of Pycnogenol® on hepatitis C virus replication

“…To evaluate the synergistic or antagonistic effects of combinations of PYC and IFN-alpha or PYC and telaprevir, we calculated the combination index (CI) using CalcuSyn (Biosoft, Ferguson, MO, USA), where CI < 1 indicates synergism, CI = 1 indicates an additive effect, and CI > 1 indicates antagonism. The weighted average CI was calculated using the formula: CI = [CI 50 + 2CI 75 + 3CI 90 + 4CI 95 ]/ 10, where CI 50 , CI 75 , CI 90 , and CI 95 are the CI values at 50%, 75%, 90% and 95% inhibition, respectively (Bassit et al, 2008;Chou and Talalay, 1984).…”