title: The YMDD and rtA194T mutations result in decreased replication capacity in wild-type HBV as well as in HBV with precore and basal core promoter mutations.
abstract: BACKGROUND
  A recent study indicated that addition of the hepatitis B e antigen (HBeAg) precore (PC) or basal core promoter (BCP) mutations to wild-type HBV offset the reduced replication of the HBV polymerase rtA194T±rtL180M+rtM204V mutations. rtA194T was reportedly associated with tenofovir resistance. We investigated these findings in genotype D HBV, where both PC and BCP naturally occur in vivo.
  
  
  METHODS
  A plasmid containing a wild-type 1.3 genome length genotype D HBV laboratory strain was used as a parent for PC, BCP, rtA194T±rtL180M+rtM204V, rtL180M+rtM204V and rtM204I mutants. Viral replication was evaluated by Southern blot analysis of intracellular HBV core DNA following transient transfection of HepG2 cells. Drug susceptibility was evaluated by quantitative PCR of intracellular HBV DNA.
  
  
  RESULTS
  PC and BCP mutations each increased HBV DNA replication by approximately 200% over wild-type. rtA194T reduced replication by <40%, whereas rtL180M+rtM204V, rtL180M+rtA194T+rtM204V or rtM204I each reduced by >75% from their respective wild-type, PC or BCP genome backbone (P<0.05). The enhanced replication by PC or BCP offset the reduction by rtA194T; however, the other reverse transcriptase (RT) mutations in PC or BCP backbones remained significantly lower than wild-type (P<0.05). Regardless of the backbone, rtA194T±rtL180M+rtM204V remained susceptible to tenofovir in vitro. rtA194T alone remained susceptible to lamivudine, while rtL180M+rtM204V and rtL180M+rtA194T+rtM204V were resistant.
  
  
  CONCLUSIONS
  PC or BCP mutations increased HBV DNA replication, offset the decreased replication by rtA194T alone, but they did not fully rescue the impaired replication conferred by other RT mutations as compared with wild-type. rtA194T±rtL180M+rtM204V did not confer tenofovir resistance.
authors.affiliation: Gilead Sciences, Inc., Foster City, CA, USA. yzhu@pharmasset.com
authors.forename: Yuao
authors.initials: Y
authors.lastname: Zhu
authors.forename: Maria
authors.initials: M
authors.lastname: Curtis
authors.forename: Katyna
authors.initials: K
authors.lastname: Borroto-Esoda
mesh_terms: D000225:Adenine / Q000031:analogs & derivatives* / Q000494:pharmacology; D000998:Antiviral Agents / Q000494:pharmacology*; D024882:Drug Resistance, Viral; D015684:Gene Products, pol / Q000235:genetics; D005838:Genotype; D056945:Hep G2 Cells; D006513:Hepatitis B e Antigens / Q000235:genetics; D006515:Hepatitis B virus / Q000187:drug effects / Q000235:genetics*; D019694:Hepatitis B, Chronic / Q000188:drug therapy* / Q000235:genetics / Q000821:virology; D006801:Humans; D019259:Lamivudine / Q000494:pharmacology*; D009154:Mutation; D063065:Organophosphonates / Q000494:pharmacology*; D000068698:Tenofovir; D014779:Virus Replication / Q000187:drug effects
chemical_list: D000998:Antiviral Agents; D015684:Gene Products, pol; D006513:Hepatitis B e Antigens; D063065:Organophosphonates; C066027:P protein, Hepatitis B virus; D019259:Lamivudine; D000068698:Tenofovir; D000225:Adenine