Early detection and quantification of lamivudine-resistant hepatitis B virus mutants by fluorescent biprobe hybridization assay in lamivudine-treated patients

Fumi Umeoka, Yoshiaki Iwasaki, Masayuki Matsumura, Akinobu Takaki, Haruhiko Kobashi, Masashi Tatsukawa, Hidenori Shiraha, Shin Ichi Fujioka, Kohsaku Sakaguchi, Yasushi Shiratori

Research output: Contribution to journalArticle

4 Citations (Scopus)

Abstract

Background: Long-term lamivudine treatment induces the emergence of lamivudine-resistant hepatitis B virus (HBV). The objective of this study was to develop a fluorescent biprobe hybridization (FBH) assay for the detection and quantification of HBV mutants in the clinical course of lamivudine-treated patients and to evaluate its clinical usefulness. Methods: We developed an FBH assay to detect mutations in the HBV DNA polymerase gene. The assay's detection sensitivity was determined using a dilution series of wild-type/mutant plasmid DNA. Blood samples obtained from 27 lamivudine-treated patients were analyzed. Results: Mutant DNA levels as low as 10% of total HBV DNA were detected (sensitivity = 100%, specificity = 80%). HBV mutants were detected in five of the 27 patients during an average follow-up of 20 months after lamivudine administration. In one of the five patients, the YIDD mutant was detected at the initiation of lamivudine treatment, while the remaining four patients were identified as having YIDD mutants within 3 months after beginning lamivudine administration. Of the five patients with an HBV mutant, four developed breakthrough hepatitis more than 10 months after the detection of HBV mutants, following the reappearance or a re-increase of HBV DNA, characterized by a predominance of the mutant. The YIDD mutant was detected in one patient, even when the titer of the serum HBV DNA was below the detection limit of commercially available quantitative polymerase chain reaction. Conclusions: The FBH assay is an efficient method for detecting and quantifying HBV mutants, as early as 3 months after lamivudine administration.

Original languageEnglish
Pages (from-to)693-701
Number of pages9
JournalJournal of Gastroenterology
Volume41
Issue number7
DOIs
Publication statusPublished - Jun 2006

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Lamivudine
Hepatitis B virus
DNA
DNA-Directed DNA Polymerase
Hepatitis
Limit of Detection
Plasmids
Sensitivity and Specificity
Polymerase Chain Reaction
Mutation

Keywords

  • Fluorescent biprobe hybridization assay
  • Hepatitis B virus
  • Lamivudine
  • Mutation

ASJC Scopus subject areas

  • Gastroenterology

Cite this

Early detection and quantification of lamivudine-resistant hepatitis B virus mutants by fluorescent biprobe hybridization assay in lamivudine-treated patients. / Umeoka, Fumi; Iwasaki, Yoshiaki; Matsumura, Masayuki; Takaki, Akinobu; Kobashi, Haruhiko; Tatsukawa, Masashi; Shiraha, Hidenori; Fujioka, Shin Ichi; Sakaguchi, Kohsaku; Shiratori, Yasushi.

In: Journal of Gastroenterology, Vol. 41, No. 7, 06.2006, p. 693-701.

Research output: Contribution to journalArticle

Umeoka, Fumi ; Iwasaki, Yoshiaki ; Matsumura, Masayuki ; Takaki, Akinobu ; Kobashi, Haruhiko ; Tatsukawa, Masashi ; Shiraha, Hidenori ; Fujioka, Shin Ichi ; Sakaguchi, Kohsaku ; Shiratori, Yasushi. / Early detection and quantification of lamivudine-resistant hepatitis B virus mutants by fluorescent biprobe hybridization assay in lamivudine-treated patients. In: Journal of Gastroenterology. 2006 ; Vol. 41, No. 7. pp. 693-701.
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abstract = "Background: Long-term lamivudine treatment induces the emergence of lamivudine-resistant hepatitis B virus (HBV). The objective of this study was to develop a fluorescent biprobe hybridization (FBH) assay for the detection and quantification of HBV mutants in the clinical course of lamivudine-treated patients and to evaluate its clinical usefulness. Methods: We developed an FBH assay to detect mutations in the HBV DNA polymerase gene. The assay's detection sensitivity was determined using a dilution series of wild-type/mutant plasmid DNA. Blood samples obtained from 27 lamivudine-treated patients were analyzed. Results: Mutant DNA levels as low as 10{\%} of total HBV DNA were detected (sensitivity = 100{\%}, specificity = 80{\%}). HBV mutants were detected in five of the 27 patients during an average follow-up of 20 months after lamivudine administration. In one of the five patients, the YIDD mutant was detected at the initiation of lamivudine treatment, while the remaining four patients were identified as having YIDD mutants within 3 months after beginning lamivudine administration. Of the five patients with an HBV mutant, four developed breakthrough hepatitis more than 10 months after the detection of HBV mutants, following the reappearance or a re-increase of HBV DNA, characterized by a predominance of the mutant. The YIDD mutant was detected in one patient, even when the titer of the serum HBV DNA was below the detection limit of commercially available quantitative polymerase chain reaction. Conclusions: The FBH assay is an efficient method for detecting and quantifying HBV mutants, as early as 3 months after lamivudine administration.",
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AU - Umeoka, Fumi

AU - Iwasaki, Yoshiaki

AU - Matsumura, Masayuki

AU - Takaki, Akinobu

AU - Kobashi, Haruhiko

AU - Tatsukawa, Masashi

AU - Shiraha, Hidenori

AU - Fujioka, Shin Ichi

AU - Sakaguchi, Kohsaku

AU - Shiratori, Yasushi

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N2 - Background: Long-term lamivudine treatment induces the emergence of lamivudine-resistant hepatitis B virus (HBV). The objective of this study was to develop a fluorescent biprobe hybridization (FBH) assay for the detection and quantification of HBV mutants in the clinical course of lamivudine-treated patients and to evaluate its clinical usefulness. Methods: We developed an FBH assay to detect mutations in the HBV DNA polymerase gene. The assay's detection sensitivity was determined using a dilution series of wild-type/mutant plasmid DNA. Blood samples obtained from 27 lamivudine-treated patients were analyzed. Results: Mutant DNA levels as low as 10% of total HBV DNA were detected (sensitivity = 100%, specificity = 80%). HBV mutants were detected in five of the 27 patients during an average follow-up of 20 months after lamivudine administration. In one of the five patients, the YIDD mutant was detected at the initiation of lamivudine treatment, while the remaining four patients were identified as having YIDD mutants within 3 months after beginning lamivudine administration. Of the five patients with an HBV mutant, four developed breakthrough hepatitis more than 10 months after the detection of HBV mutants, following the reappearance or a re-increase of HBV DNA, characterized by a predominance of the mutant. The YIDD mutant was detected in one patient, even when the titer of the serum HBV DNA was below the detection limit of commercially available quantitative polymerase chain reaction. Conclusions: The FBH assay is an efficient method for detecting and quantifying HBV mutants, as early as 3 months after lamivudine administration.

AB - Background: Long-term lamivudine treatment induces the emergence of lamivudine-resistant hepatitis B virus (HBV). The objective of this study was to develop a fluorescent biprobe hybridization (FBH) assay for the detection and quantification of HBV mutants in the clinical course of lamivudine-treated patients and to evaluate its clinical usefulness. Methods: We developed an FBH assay to detect mutations in the HBV DNA polymerase gene. The assay's detection sensitivity was determined using a dilution series of wild-type/mutant plasmid DNA. Blood samples obtained from 27 lamivudine-treated patients were analyzed. Results: Mutant DNA levels as low as 10% of total HBV DNA were detected (sensitivity = 100%, specificity = 80%). HBV mutants were detected in five of the 27 patients during an average follow-up of 20 months after lamivudine administration. In one of the five patients, the YIDD mutant was detected at the initiation of lamivudine treatment, while the remaining four patients were identified as having YIDD mutants within 3 months after beginning lamivudine administration. Of the five patients with an HBV mutant, four developed breakthrough hepatitis more than 10 months after the detection of HBV mutants, following the reappearance or a re-increase of HBV DNA, characterized by a predominance of the mutant. The YIDD mutant was detected in one patient, even when the titer of the serum HBV DNA was below the detection limit of commercially available quantitative polymerase chain reaction. Conclusions: The FBH assay is an efficient method for detecting and quantifying HBV mutants, as early as 3 months after lamivudine administration.

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