Inhibition of Hepatitis B Virus Polymerase by Entecavir

Abstract

Entecavir (ETV; Baraclude) is a novel deoxyguanosine analog with activity against hepatitis B virus (HBV).ETV differs from the other nucleoside/tide reverse transcriptase inhibitors approved for HBV therapy, lamivudine (LVD) and adefovir (ADV), in several ways: ETV is >100-fold more potent against HBV in culture and, at concentrations below 1 M, displays no significant activity against human immunodeficiency virus (HIV). Additionally, while LVD and ADV are obligate DNA chain terminators, ETV halts HBV DNA elongation after incorporating a few additional bases. Three-dimensional homology models of the catalytic center of the HBV reverse transcriptase (RT)-DNA-deoxynucleoside triphosphate (dNTP) complex, based on the HIV RT-DNA structure, were used with in vitro enzyme kinetic studies to examine the mechanism of action of ETV against HBV RT. A novel hydrophobic pocket in the rear of the RT dNTP binding site that accommodates the exocyclic alkene moiety of ETV was predicted, establishing a basis for the superior potency observed experimentally. HBV DNA chain termination by ETV was accomplished through disfavored energy requirements as well as steric constraints during subsequent nucleotide addition. Validation of the model was accomplished through modeling of LVD resistance substitutions, which caused an eightfold decrease in ETV susceptibility and were predicted to reduce, but not eliminate, the ETV-binding pocket, in agreement with experimental observations. ADV resistance changes did not affect the ETV docking model, also agreeing with experimental results. Overall, these studies explain the potency, mechanism, and cross-resistance profile of ETV against HBV and account for the successful treatment of naive and LVD-or ADV-experienced chronic HBV patients.More than 350 million people worldwide are chronically infected with hepatitis B virus (HBV), and a significant proportion of them will ultimately develop severe liver disease, including cirrhosis, hepatocellular carcinoma, and other severe complications (42). Entecavir (ETV; formerly called BMS-200475), lamivudine (LVD or 3TC; ␤-L-2Ј,3Ј-dideoxy-3Ј-thiacytidine), adefovir-dipivoxil prodrug [ADV or PMEA; 9-(2-phosphonylmethoxyethyl) adenine], and most recently, telbivudine (LdT) are oral HBV nucleoside/ tide reverse transcriptase inhibitors (NRTIs) approved for the treatment of chronic HBV infection. Upon entry into the cell, all of these inhibitors require subsequent phosphorylation by cellular enzymes to generate their active moieties. The combination of intrinsic potency, exposure level, efficiency of intracellular phosphorylation, and genetic barriers to resistance all contribute to the initial and long-term efficacy of these molecules.NRTIs inhibit the only known enzymatic target of HBV, the viral polymerase (Pol), which is characterized by several unique biological features (reviewed in reference 39). The mRNA encoding Pol serves as the template for synthesis of genomic virion DNA through reverse transcriptase (RT) activity. The RNA template is the mRNA that...

Computational model of hepatitis B virus DNA polymerase: Molecular dynamics and docking to understand resistant mutations

“…This pocket is lined with hydrophobic residues like rtA87, rtF88, rtP177, rtL180, and rtM204, along with the last nucleotide of the primer DNA strand. These findings are consistent with those reported by Langley et al 9 …”

“…Langley and coworkers have reported a similar binding pose for entecavir, and studied in detail the mechanism of action of entecavir in chain termination. 9 The sulfur atom of the oxathiolane ring in lamivudine occupied the same pocket. The increased hydrophobic interactions in the pocket caused a higher docking score and decreased DG for lamivudine for postdynamics docking.…”

“…6 Good sequence homology between HDP and HIV-1 RT has enabled several researchers to build 3D homology models of the structure of the HDP catalytic domain, [7][8][9] which were used to provide insights about the binding mode of various inhibitors and developing resistance. Das et al aligned the sequence of HDP to HIV-1 RT and used an HIV-1 RT X-ray structure 1RTD.pdb for building a tertiary structure of HDP, which was used to reveal the mechanism of HDP resistance to lamivudine and emtricitabine.…”

“…Those authors used the same sequence alignment as Das et al and prepared their 3D model using 1RTD as a template. 9 In this article, we report a different homology model of HDP based on a better alignment and a more recent HIV-1 RT X-ray structure. Our model can be used to explain all mutations in HDP reported to date as well as the inactivity of some known HIV-1 RT inhibitors against HDP.…”

The saga of entecavir

“…Entecavir is a deoxyguanosine analogue that is rapidly phosphorylated intracellularly into its active 5 0 -triphosphate form. It inhibits HBV replication at three important steps: protein-linked priming of the HBV polymerase, synthesis of the first negative strand of the HBV DNA by reverse transcription, and synthesis of the second positive strand [9]. Its efficient phosphorylation and its triple action in the inhibition of HBV replication may explain its potency in the suppression of HBV DNA in small doses of 0.5-1 mg licensed for use.…”