Viral quasispecies of hepatitis B virus in patients with YMDD mutation and lamivudine resistance may not predict the efficacy of lamivudine/adefovir rescue therapy

Abstract

The association between hepatitis B virus (HBV) quasispecies (QS) and the efficacy of nucleos(t)ide analog therapy is currently not well defined, particularly in the case of lamivudine (LAM)/adefovir (ADV) combination rescue therapy for patients with chronic HBV infection (CHB) presenting with LAM resistance. In the present study, 16 CHB patients with the rtM204I/V mutation in the tyrosine-methionine-aspartate-aspartate motif of the C domain of the polymerase gene who switched to LAM/ADV treatment due to LAM resistance were assessed. HBV DNA was isolated from these patients and the reverse transcriptase (RT) region was sequenced. The QS heterogeneity and distribution was analyzed, the mutation sites were recorded and the phylogenetic trees were constructed. The results indicated that QS heterogeneity and distribution in the RT and S regions were not significantly different between responders (RS) and non-RS (NRS) at baseline (P>0.05), except for the higher frequency of a dominant strain in the RT region at the nucleotide level in the RS group (P=0.039). In addition, in NRS, no significant difference in QS heterogeneity or distribution in these regions was identified at six months vs. the baseline. Furthermore, although in the non-responder group the frequency of the LAM resistance-associated mutations (rtM204V/I) decreased at 6 months compared with the baseline, it did not disappear in any of the patients after six months of treatment. Analysis of individual patients did not indicate any consistent selection of specific HBV mutants during LAM/ADV rescue therapy. In conclusion, the baseline HBV QS within the RT and S regions may not be a valid predictor of the response to LAM/ADV rescue treatment in CHB patients with LAM resistance.

Keywords: adefovir; hepatitis B virus; lamivudine; quasispecies; resistance mutation.

Figure 1.

QS composition within the reverse transcription region of hepatitis B virus during LAM/adefovir combination rescue therapy against LAM resistance. The vertical bars represent the number and proportion of viral QS in each sample. Each color represents one specific QS. (A) QS composition at the nucleotide level and (B) QS composition at the amino acid level at M0 and M6. QS, quasispecies; LAM, lamivudine; NRS, non-responders; M0, baseline; M6, 6 months post-treatment.

Figure 2.

QS distribution within the reverse transcription region of hepatitis B virus during LAM/adefovir combination rescue therapy against LAM resistance. (A and B) Number of QS at (A) the nucleotide level and (B) the amino acid level at M0 and M6. (C and D) Frequency of dominant strains at (C) the nucleotide level and (D) the amino acid level at M0 and M6. The horizontal lines indicate mean ± standard error of mean in every group and each point represents the number of QS in each patient. QS, quasispecies; LAM, lamivudine; NRS, non-responders; M0, baseline; M6, 6 months post-treatment.

Figure 3.

QS complexity and diversity in the reverse transcription region of hepatitis B virus during lamivudine/adefovir combination rescue therapy. (A and B) QS complexity at (A) the nucleotide level and (B) the amino acid level at M0 and M6; (C and D) d at (C) the nucleotide level and (D) the amino acid level at M0 and M6. (E and F) Number of (E) dS and (F) dN at M0 and M6. The horizontal lines indicate mean ± standard error of mean in every group and each point represents the number of QS in each patient. NRS, non-response; Sn, Shannon entropy; d, mean genetic distance; dS, number of synonymous substitutions per synonymous site; dN, number of non-synonymous substitutions per non-synonymous site; QS, quasispecies; NRS, non-responders; M0, baseline; M6, 6 months post-treatment.

Figure 4.

QS distribution in the S region of hepatitis B virus during lamivudine/adefovir combination rescue therapy. QS number at (A) the nucleotide level and (B) the amino acid level at M0 and M6. Frequency of dominant strains at (C) the nucleotide level and (D) at the amino acid level at M0 and M6. The horizontal lines indicate mean ± standard error of mean in every group and each point represents the number of QS in each patient. QS, quasispecies; NRS, non-responders; M0, baseline; M6, 6 months post-treatment.

Figure 5.

QS complexity and diversity in the S region of hepatitis B virus during lamivudine/adefovir combination rescue therapy. (A and B) QS complexity at (A) the nucleotide level and (B) the amino acid level at M0 and M6; (C and D) d at (C) the nucleotide level and (D) the amino acid level at M0 and M6. (E and F) Number of (E) dS and (F) dN at M0 and M6. The horizontal lines indicate mean ± standard error of mean in every group and each point represents the number of QS in each patient. NRS, non-response; Sn, Shannon entropy; d, mean genetic distance; dS, number of synonymous substitutions per synonymous site; dN, number of non-synonymous substitutions per non-synonymous site; QS, quasispecies; NRS, non-responders; M0, baseline; M6, 6 months post-treatment.