Decreased infectivity of nucleoside analogs-resistant hepatitis B virus mutants

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• 作者：Gaë ; tan Billioud¹ ; ² ; Christian Pichoud¹ ; ² ; Romain Parent¹ ; ² ; Fabien Zoulim¹ ; ² ; ³ ; ^{fabien.zoulim@inserm.fr}
• 关键词：HBV ; hepatitis B virus ; RT ; reverse transcriptase ; S ; surface ; HDV ; hepatitis delta virus ; PCR ; polymerase chain reaction ; WT ; wild type ; ORF ; open reading frame ; Ag ; antigen ; SVP ; subviral particle ; cccDNA ; covalently closed circular DNA
• 刊名：Journal of Hepatology
• 出版年：2012
• 出版时间：June, 2012
• 年：2012
• 卷：56
• 期：6
• 页码：1269-1275
• 全文大小：1093 K

文摘

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Background & Aims

To understand the mechanisms of emergence and selection of HBV polymerase variants, which may also harbor mutations in the overlapping envelope protein, we analyzed the in vitro virus production and infectivity of the main viral mutants resistant to lamivudine and adefovir.

Methods

HBV-resistant mutants (rtL180M + M204V, rtV173L + L180M + M204V, rtM204I, rtL180M + M204I, rtN236T, rtA181V, rtA181V + rtN236T, rtA181T + N236T, and rtA181T) were produced in HepG2 cells permanently expressing the respective viral genomes. Viral protein expression, secretion, and viral particle production were studied by ELISA, Western blot, and transmission electron microscopy. To study only the effect of surface gene mutants on virus infectivity, HepaRG cells were inoculated with HDV pseudo-particles coated with the mutant HBV envelopes. To evaluate infectivity and replication in a global fashion, HepaRG cells were inoculated with HBV mutants.

Results

HBeAg was expressed and secreted in cell supernatants in all mutant-expressing cell lines. As expected, mutants harboring a sW196Stop mutation in the surface gene did not express small envelope proteins. All mutants expressing HBsAg were able to produce viral particles. HDV particles coated with mutant envelopes were less infectious than WT in HepaRG cells. Finally, we found that resistant mutants exhibit lower infectivity and replication ability than WT virus.

Conclusions

Based on this study, we found that envelope substitutions modulate viral protein expression, HDV coating, and viral infectivity. These envelope modifications provide novel insights into the features of emerging HBV variants during antiviral therapies and suggest that such mutants are less prone to transmission than their WT counterpart.