Patterns of circulating hepatitis B virus serum nucleic acids during lamivudine therapy

Abstract

We examined whether lamivudine treatment, in addition to the rapid decline of HBV serum DNA described in a large number of laboratories, causes changes in composition and amount of discernable circulating viral DNA and RNA. Nucleic acids were extracted from serial serum samples of a patient infected chronically and treated with lamivudine for 14 weeks. Three sequence segments of the HBV genome synthesized successively during replication, X, C, and X-preC, were analyzed by competitive PCR and RT/PCR. In addition, RNA was examined for differential polyadenylation. Before treatment, identical DNA copy numbers (10(9)/ml) were found in all three segments. C segment DNA displayed the expected rapid decline. X-preC, a target contiguous only on plus-strand DNA behaved similarly. In contrast, the X segment DNA copy numbers showed a less pronounced decrease remaining at higher values (10(7)/ml) than the C and X-preC segment (both about 2 x 10(5)/ml) at the end of therapy. X segment RNA displayed a persisting copy number of about 10(7)/ml, while C and X-preC RNA decreased to about 10(5) copies/ml. Polyadenylated HBV RNA, full-length and truncated, persisted initially at 10(5) but decreased to 10(4) to 10(3) copies/ml at the end of treatment. The major conclusions are the actual numbers of virus particles during lamivudine therapy can only be assessed via X segment DNA, since it is reverse transcribed first, and Lamivudine induced coexistence of DNA and RNA for the C and X segment at similar levels indicates drug-arrested intermediates of reverse transcribed HBV DNA minus-strand. Packaged RNA lacks a poly(A) tail whereas polyadenylated RNA is likely not packaged.

Serum HBV RNA is a predictor of early emergence of the YMDD mutant in patients treated with lamivudine†

“…The presence of HBV RNA in serum samples of patients with HBV infection has been previously reported. [19][20][21] Because the TMA-HPA uses RNA transcription and amplification of transcripts by T7 RNA polymerase, 22 we assumed that the discrepancy was a result of the presence of HBV RNA in the serum of LAM-and ETV-treated patients. The presence of HBV RNA in a patient treated with LAM was reported previously.…”

“…The presence of HBV RNA in a patient treated with LAM was reported previously. 21 In that report, the authors mainly analyzed truncated HBV RNA, which they assumed was transcribed from the integrated genome. 20,21 They showed a large difference between HBV DNA and truncated HBV RNA, which did not decrease during LAM therapy.…”

“…21 In that report, the authors mainly analyzed truncated HBV RNA, which they assumed was transcribed from the integrated genome. 20,21 They showed a large difference between HBV DNA and truncated HBV RNA, which did not decrease during LAM therapy. We also detected HBV DNA and HBV nucleic acid by realtime PCR and real-time RT-PCR.…”

“…Because the former method detects both HBV DNA and HBV RNA, we thought that the difference in measurement by the 2 methods was a result of the presence of a large amount of HBV RNA. [19][20][21] We thus studied patients with chronic HBV infection who were being treated with LAM or entecavir (ETV) for the presence of HBV RNA. We also assumed that the presence of a large amount of HBV RNA would indicate that transcription and virus particle formation were still active in such patients.…”

Small-Molecule Effectors of Hepatitis B Virus Capsid Assembly Give Insight into Virus Life Cycle

“…The predominant antiviral strategy against HBV is to attack the reverse transcriptase. Unsurprisingly, HBV reverse transcriptase inhibitors lead to drug-resistant mutants in HBV (7,24,30) and also human immunodeficiency virus (14). Resistance can have broader consequences because of the extensive gene overlap in HBV, since some reverse transcriptase mutations lead to surface protein that is insensitive to antibodies generated by the HBV vaccine (25).…”