Efficacies of Entecavir against Lamivudine-Resistant Hepatitis B Virus Replication and Recombinant Polymerases In Vitro

Abstract

Entecavir (ETV) is a potent and selective inhibitor of hepatitis B virus (HBV) replication in vitro and in vivo that is currently in clinical trials for the treatment of chronic HBV infections. A major limitation of the current HBV antiviral therapy, lamivudine (3TC), is the emergence of drug-resistant HBV in a majority of treated patients due to specific mutations in the nucleotide binding site of HBV DNA polymerase (HBV Pol). To determine the effects of 3TC resistance mutations on inhibition by ETV triphosphate (ETV-TP), a series of in vitro studies were performed. The inhibition of wild-type and 3TC-resistant HBV Pol by ETV-TP was measured using recombinant HBV nucleocapsids, and compared to that of 3TC-TP. These enzyme inhibition studies demonstrated that ETV-TP is a highly potent inhibitor of wild-type HBV Pol and is 100-to 300-fold more potent than 3TC-TP against 3TC-resistant HBV Pol. Cell culture assays were used to gauge the potential for antiviral cross-resistance of 3TC-resistant mutants to ETV. Results demonstrated that ETV inhibited the replication of 3TC-resistant HBV, but 20-to 30-fold higher concentrations were required. To gain further perspective regarding the potential therapeutic use of ETV, its phosphorylation was examined in hepatoma cells treated with extracellular concentrations representative of drug levels in plasma in ETV-treated patients. At these concentrations, intracellular ETV-TP accumulated to levels expected to inhibit the enzyme activity of both wild-type and 3TC-resistant HBV Pol. These findings are predictive of potent antiviral activity of ETV against both wild-type and 3TC-resistant HBV.Infection with hepatitis B virus (HBV) is a medical problem of global proportions. Despite the existence of a safe vaccine, approximately 5% of the world population is infected with HBV. Approved therapies for chronic HBV infection are treatment with alpha interferon or lamivudine (3TC). Drawbacks to treatment with alpha interferon include a low sustained response rate, undesirable side effects, the need for parenteral administration, and high cost. Therapy with 3TC is less costly and more convenient to use, but it also suffers from a low sustained response rate. Of more fundamental concern is that while initial treatment of patients with 3TC results in a rapid lowering of HBV DNA levels in the blood, its efficacy is severely compromised in most patients by the development of antiviral resistance after prolonged therapy.The inhibition of HBV replication by nucleoside analogs results from the recognition of nucleoside analog triphosphates (TPs) by the RNA-dependent DNA polymerase of HBV (HBV Pol). Clinical resistance to 3TC results from amino acid substitutions at position 550 (methionine [M]) in the highly conserved YMDD motif of HBV Pol (31). Variants with the double mutation M550V/L526M or the single mutation M550I predominate (reviewed in reference 14). Molecular modeling studies suggest that these mutations alter the nucleotide binding site of HBV Pol to cause steric hindrance of 3TC-TP ...

The HBV Drug Entecavir — Effects on HIV-1 Replication and Resistance

“…Rather, the appearance of the M184V mutation likely reflects the fact that it abolishes the anti-HIV activity of entecavir ( Figures 2C and 2D). Interestingly, selection of the M184V mutation in HIV RT could be anticipated since structural models show that the M184V corresponds to the HBV pol mutation, rtM204V (23), which decreases HBV susceptibility to entecavir (24). In both cases, the targeted methionine is in the YMDD motif at the active site.…”

Mechanism of viral persistence and resistance to nucleoside and nucleotide analogs in chronic Hepatitis B virus infection

“…Interestingly, it was shown that lamivudine resistant mutants are not sensitive to L-pyrimidine analogs such as emtricitabine, clevudine, elvucitabine, telbivudine while they remain susceptible to purine analogs such as adefovir, tenofovir, and to some extent to entecavir (Delaney et al, 2001;Fu and Cheng, 2000;Levine et al, 2002;Menne et al, 2002;Seigneres et al, 2002). The adefovir resistant strains are also sensitive to lamivudine, emtricitabine, and entecavir, and to some extent to tenofovir (Angus et al, 2003;Villeneuve et al, 2003).…”

Entecavir for the treatment of lamivudine-refractory chronic hepatitis B patients in China

“…In vitro studies have shown that entecavir effectively suppresses the replication of lamivudine-resistant HBV with an EC 50 of 0.026 lM [22]. The selection of the 1 mg dose of entecavir for evaluation in the current study in lamivudine-refractory patients resulted from a multinational phase II dose-ranging study in which entecavir 1 mg demonstrated superior efficacy to continued lamivudine 100 mg, and yielded the greatest viral load reduction compared with the other doses of entecavir studied (0.5 and 0.1 mg) [23].…”