Characterization of the Anti-Hepatitis C Virus Activity of New Nonpeptidic Small-Molecule Cyclophilin Inhibitors with the Potential for Broad Anti-Flaviviridae Activity

Abstract

Although members of the Flaviviridae display high incidence, morbidity, and mortality rates, the development of specific antiviral drugs for each virus is unlikely. Cyclophilins, a family of host peptidyl-prolyl cis-trans isomerases (PPIases), play a pivotal role in the life cycles of many viruses and therefore represent an attractive target for broad-spectrum antiviral development. We report here the pangenotypic anti-hepatitis C virus (HCV) activity of a small-molecule cyclophilin inhibitor (SMCypI). Mechanistic and modeling studies revealed that the SMCypI bound to cyclophilin A in competition with cyclosporine (CsA), inhibited its PPIase activity, and disrupted the CypA-nonstructural protein 5A (NS5A) interaction. Resistance selection showed that the lead SMCypI hardly selected amino acid substitutions conferring low-level or no resistance in vitro. Interestingly, the SMCypI selected D320E and Y321H substitutions, located in domain II of the NS5A protein. These substitutions were previously associated with low-level resistance to cyclophilin inhibitors such as alisporivir. Finally, the SMCypI inhibited the replication of other members of the Flaviviridae family with higher 50% effective concentrations (EC50s) than for HCV. Thus, because of its chemical plasticity and simplicity of synthesis, our new family of SMCypIs represents a promising new class of drugs with the potential for broad-spectrum anti-Flaviviridae activity as well as an invaluable tool to explore the role of cyclophilins in viral life cycles.

Small-Molecule Inhibitors of Cyclophilins Block Opening of the Mitochondrial Permeability Transition Pore and Protect Mice From Hepatic Ischemia/Reperfusion Injury

“…Our aim was to develop CypA inhibitors with antiviral activity against the hepatitis C virus. We showed that our SMCypIs bind CypA and potently inhibit both CypA PPIase activity and the replication of hepatitis C virus and related viruses in cell culture 19,33 . Because the different cyclophilins are structurally very close (essentially differing by their cellular localizations and functions), we assessed whether the new SMCypIs inhibit CypD activity.…”

Fitness‐associated substitutions following failure of direct‐acting antivirals assessed by deep sequencing of full‐length hepatitis C virus genomes

“…To study the combined effect of Y93H and the suspected fitness‐associated substitutions, consensus amino acids in the genotype 3a replicon were reverted to the baseline polymorphism observed in each patient's sample prior to therapy, in the presence of the Y93H substitution. Huh7.5 cells were transfected with wild‐type or mutated DBN3acc, as previously described 18 . Increasing concentrations of daclatasvir (10 to 5000 pM) were added 4 h after transfection, whereas control cells were treated with 1% DMSO.…”

Cyclophilins and Their Roles in Hepatitis C Virus and Flavivirus Infections: Perspectives for Novel Antiviral Approaches

“…SMCypI have potent PPIase-inhibitory activity, and were found to have potent antiviral activity against HCV genotype 1b replication [30]. The lead SMCypI, termed C31, was subsequently shown to disrupt the CypA-NS5A interaction [101], similar to what was shown previously for CsA. Further suggesting that SMCypI acts by the same mechanisms as CsA, C31 treatment selected for resistance mutations in NS5A-D2 [101] (genotype 1b D320E/Y321H; analogous to the D316E/Y317N mutation in genotype 2a selected by CsA treatment [76]).…”