Analysis of evolution and virological characteristics of rtI233V mutations in the hepatitis B virus reverse transcriptase domain / 解放军医学杂志

ABSTRACT

Objective To analyze the evolution of rtI233V mutation in the reverse transcriptase domain of hepatitis B virus (HBV) and its association with adefovir dipivoxil (ADV) resistance. Methods The rate of detection of rtI233V mutation in 9830 patients with chronic HBV infection was analyzed. HBV reverse transcriptase genes isolated from serial serum samples of two patients were amplified by nested PCR, and clonal sequencing (>20 clones/sample) was performed to analyze the evolution of rtI233V mutations. The replica of pTriEx-HBV1.1 vectors harboring wild-type and mutant strains (rtI233V, rtN236T, rtI233V+rtN236T) were respectively constructed and transiently transfected into HepG2 cells. Media containing serial concentrations of lamivudine (LAM), ADV, entecavir (ETV), or tenofovir (TDF) were used to treat the cells. Then HBV DNA in the supernatants was quantitatively determined by real-time PCR in order to analyze HBV mutants' replication competence and phenotypic characteristics under the drug pressures. Results The detection rate of rtI233V mutation in 9830 nucleos(t)ide analogues-treated patients was 0.28% (28/9830), including 0.19% (19 patients) with rtI233V individual mutation and 0.09% (9 patients) with rtI233V mutation combining with rtN236T or other mutations. All of the patients had rtI233V had ADV exposure history 16 (57.1%) of them received ADV monotherapy for over six months, and 12(42.9%) of them received ADV combined sequential therapy for over 12 months. Replication competence and phenotypic resistance analysis showed rtI233V and wild-type strains had similar viral replication competence, while rtN236T exhibited significantly lower replication competence compared with wild-type strains. rtI233V strains remained sensitive to LAM, ADV, ETV, and TDF and showed little influence on drug resistance when combined with rtN236T, but it showed ability to restore the defected replication capacity of rtN236T strains. Conclusions The rtI233V mutation is associated with sub-optimal response to ADV. Though it does not directly reduce virus sensitivity to ADV, rtI233V mutation enhances replication competence of ADV resistant strains, and it is a complementary mutation.