COMBINATION OF THE NS3/4A PROTEASE INHIBITOR ITMN-191 WITH THE ALLOSTERIC NS5B POLYMERASE INHIBITOR ITMN-8020 ENHANCES REPLICON CLEARANCE AND REDUCES THE EMERGENCE OF DRUG RESISTANT VARIANTS
H. Tan, G. Wang, C. Moy, K. Kossen, R. Rajagopalan, S. Misialek, D. Ruhrmund, L. Hooi, N. Snarskaya, A. Stoycheva, B. Buckman, S. Seiwert, L. Beigelman Intermune, Inc., Brisbane, USA
Background: ITMN-191 is an inhibitor of the HCV NS3/4A protease, and ITMN-8020 is an allosteric inhibitor of the HCV NS5B polymerase. ITMN-191 is being evaluated in phase 1b studies in combination with the current SOC and in the INFORM-1 clinical study in combination with R7128, a nucleoside inhibitor of NS5B. Due to the potential for drug resistance with NS3 protease inhibitors and presumably with allosteric NS5B polymerase inhibitors, we examined the antiviral effect of ITMN-191 and ITMN-8020 alone and in combination. Methods: In the HCV clearance assay, cells harboring an HCV genotype-1b replicon were treated for 2 weeks with ITMN-191, ITMN-8020, or a combination of inhibitors in the absence of G418 selection for replicon retention. Cells were counted and aliquots harvested for RT-PCR-based quantification of replicon RNA levels under G418 selection over 3 subsequent weeks. In the colony formation assay, cells were treated with ITMN-191 at ~1.1X, ~11X, or ~17X its EC50 either alone or in the presence of ITMN-8020 at ~1X, ~10X, or ~15X its EC50. After 3 weeks in culture with G418, cells were fixed and stained or total cellular RNA extracted. Results: In an HCV clearance assay, ITMN-191 at 15X its EC50 eliminated HCV replicon in the absence of G418 selective pressure for replicon retention while 15X the EC50 of ITMN-8020 did not. Addition of the lowest tested concentration of ITMN-191 (1.9X EC50) to ITMN-8020 at 15X its EC50 resulted in replicon clearance, demonstrating significant combined antiviral effect. In the colony formation assay in the presence of G418 selective pressure for replicon retention, ITMN-8020 at 15X its EC50 reduced but did not eliminate cell colonies resulting from HCV replicon persistence. However, addition of ITMN-191 at 11X its EC50 prevented formation of any cell colonies, indicating the combined antiviral effects of the two agents forced replicon elimination in a dose-dependent fashion. Conclusions: The combination of ITMN-191 with ITMN-8020 in vitro results in enhanced antiviral activity and suppression of drug-resistant variants. These findings suggest that the combination of these two agents may represent a viable therapeutic approach, resulting in antiviral activity greater than that observed with ITMN-191 alone.