117 230

Cited 32 times in

Comparison of multiplex restriction fragment mass polymorphism and sequencing analyses for detecting entecavir resistance in chronic hepatitis B.

Authors
Han, KH; Hong, SP; Choi, SH; Shin, SK; Cho, SW; Ahn, SH; Hahn, JS; Kim, SO
Citation
Antiviral therapy, 16(1):77-87, 2011
Journal Title
Antiviral therapy
ISSN
1359-65352040-2058
Abstract
BACKGROUND: Drug resistance is a major limitation to the long-term efficacy of controlling chronic hepatitis B (CHB). There is a growing need to analyse multiple mutations associated with drug resistance because sequential or combinational use of antivirals is increasingly being used in treatment. In this study, we introduced a multiplex restriction fragment mass polymorphism (RFMP) assay for detecting mutations conferring entecavir and lamivudine resistance, and compared its performance with direct or clonal sequencing assays.
METHODS: Multiplex PCR was performed with mixed primers designed to interrogate rt184, rt202, rt204 and rt250. The PCR products were digested with restriction enzymes and the resulting fragments were analysed by mass spectrometry. A total of 251 serum samples, taken serially from 45 patients who received entecavir treatment after confirmed diagnosis of lamivudine resistance and inadequate adefovir dipivoxil response, were analysed by the multiplex RFMP assay.
RESULTS: The multiplex RFMP assay correctly identified known viral sequences with sufficient analytical sensitivity to detect as few as 100 IU/ml of HBV and with superior ability to determine haplotypes composed of neighbouring variations. Complex mutational patterns and relative abundances determined by multiplex RFMP assay were in good concordance with results obtained by direct or clonal sequencing analyses. Defined mixtures were successfully and consistently identified at 2% relative concentration of mutant versus wild-type virus by the assay.
CONCLUSIONS: The multiplex RFMP assay is an accurate and sensitive means to detect entecavir and lamivudine resistance mutations, simultaneously. The method is expected to enable early and efficient diagnosis of multiple drug resistance mutations for optimal management of CHB.
MeSH terms
Adenine/administration & dosage/analogs & derivatives/therapeutic useAntiviral Agents/administration & dosage/therapeutic useDrug Resistance, Viral/*geneticsGenotypeGuanine/administration & dosage/analogs & derivatives/therapeutic useHepatitis B virus/drug effects/geneticsHepatitis B, Chronic/*diagnosis/drug therapy/*genetics/virologyHumansLamivudine/administration & dosage/therapeutic useMass Spectrometry/methodsMutationPhosphonic Acids/administration & dosage/therapeutic usePolymerase Chain Reaction/*methodsPolymorphism, Genetic*Polymorphism, Restriction Fragment LengthSequence Analysis, DNA/methodsViral Load
DOI
10.3851/IMP1702
PMID
21311111
Appears in Collections:
Journal Papers > School of Medicine / Graduate School of Medicine > Gastroenterology
AJOU Authors
조, 성원
Files in This Item:
fulltext not available.txtDownload
Export
RIS (EndNote)
XLS (Excel)
XML

qrcode

해당 아이템을 이메일로 공유하기 원하시면 인증을 거치시기 바랍니다.

Items in DSpace are protected by copyright, with all rights reserved, unless otherwise indicated.

Browse