Entecavir allows an unexpectedly high residual replication of HBV mutants resistant to lamivudine

Background: Entecavir is an efficient inhibitor of HBV reverse transcriptase (RT) and widely used for therapy of chronic hepatitis B. Entecavir treatment of HBV patients with lamivudine-resistant viral strains, however, often fails, but the mechanism of cross-resistance development is not fully understood. Methods: Using non-linear regression models, dose-response curves of cloned HBV strains from patients pre-treated with RT inhibitors were established in human hepatoma cell lines after transfection with HBV genomes containing HBV polymerase genes from patient isolates. 50% and 90% inhibitory concentrations (IC₅₀ and IC₉₀) and corresponding antiviral resistance factors (RF₅₀ and RF₉₀) were calculated. Results: The entecavir dose-response curve of lamivudine-resistant HBV RT mutants rtM204 for the replication of HBV decreased less than expected with increasing drug dose. Remarkably, due to the flat dose-response curves, RF₉₀ values against entecavir of samples with rtM204 substitutions were up to 30x higher than their RF₅₀ values. Conclusions: The unexpectedly high IC₉₀ indicates a strong residual replication capacity of lamivudine-resistant HBV rtM204 variants under entecavir therapy, although IC₅₀ values are initially within the therapeutic range of entecavir. This characteristic favours rapid selection of additional mutants with overt resistance against entecavir. Thus, the current phenotypic resistance assays should include determination of IC₉₀.