• Korean Journal of Clinical Laboratory Science
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• v.37 no.3
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• pp.173-177
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• 2005

YMDD motif mutants of the hepatitis B virus(HBV) emerge in chronic hepatitis B patients after prolonged lamivudine treatment. HBV DNA breakthrough may be accompained by the emergence of YMDD mutants. We compared the performance of the sequencing method with that of RFMP method in chronic hepatitis B patients who had suffered the HBV DNA breakthrough after lamivudine treatment. Both sequencing and RFMP methods were used to detect YMDD variants in 20 chronic hepatitis B patients. YMDD mutants were detected in 17 samples (85.0%) by both methods. Among them, no mutants were detected in two samples(10.0%), while they were detected in the other sample(5.0%) with the RFMP method. The concordance rate between both methods was 95.0%. There was inconsistency in one sample showing mutants detected by the RFMP method, but not by sequencing method. In the sequencing method, the mutants was detected in the major type virus, but not in the minor type virus. However, both sequencing and RFMP methods were highly concordant except in one sample, so it is suggested that both methods are useful to detect YMDD mutants of chronic hepatitis B patients.

• Asian Pacific Journal of Cancer Prevention
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• v.14 no.9
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• pp.5489-5494
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• 2013

In the current study we aimed to show the common YMDD motif mutations in viral polymerase gene in chronic hepatitis B patients during lamivudine and adefovir therapy. Forty-one serum samples obtained from chronic hepatitis B patients (24 male, 17 female; age range: 34-68 years) were included in the study. HBV-DNA was extracted from the peripheral blood of the patients using an extraction kit (Invisorb, Instant Spin DNA/RNA Virus Mini Kit, Germany). A line probe assay and direct sequencing analyses (INNO-LIPA HBV DR v2; INNOGENETICS N.V, Ghent, Belgium) were applied to determine target mutations of the viral polymerase gene in positive HBV-DNA samples. A total of 41 mutations located in 21 different codons were detected in the current results. In 17 (41.5%) patients various point mutations were detected leading to lamivudin, adefovir and/or combined drug resistance. Wild polymerase gene profiles were detected in 24 (58.5%) HBV positive patients of the current cohort. Eight of the 17 samples (19.5%) having rtM204V/I/A missense transition and/or transversion point mutations and resistance to lamivudin. Six of the the mutated samples (14.6%) having rtL180M missense transversion mutation and resistance to combined adefovir and lamivudin. Three of the mutated samples (7.5%) having rtG215H by the double base substituation and resistance to adefovir. Three of the mutated samples (7.5%) having codon rtL181W due to the missense transversion point mutations and showed resistance to combined adefovir and lamivudin. Unreported novel point mutations were detected in the different codons of polymerase gene region in the current HBV positive cohort fromTurkish population. The current results provide evidence that rtL180M and rtM204V/I/A mutations of HBV-DNA may be associated with a poor antiviral response and HBV chronicity during conventional therapy in Turkish patients.

• Korean Journal of Clinical Laboratory Science
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• v.38 no.3
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• pp.196-202
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• 2006

Treatment of hepatitis B virus (HBV) with lamivudine is effective in suppressing virus replication and results in reduced inflammatory activity. However the most troublesome problem of lamivudine treatment is the emergence of lamivudine-resistant strains with amino acid substitution in the YMDD motif of DNA polymerase gene during the treatment. The aim of this study was to determine the mutation of YMDD motif (codon 552) and codon 528 in chronic HBV patients with lamivudine therapy using PCR-direct sequencing and to investigate the relationship between lamivudine mediated HBV mutation and HBeAg. HBV DNA was extracted from serum samples of HBV patients and amplified by nested PCR with two sets of primer pairs selected in HBV DNA polymerase gene. Amplified PCR product was analyzed by 2% agarose gel electrophoresis and direct sequencing. HBV mutation was detected in 124 out of 207 samples (60%). Single mutation was 50.8% for M552I, 43.5% for M552V, 5.7% for M552I/V and the L528M mutation was 67.0%. Double mutation was 43.6% for M552V/L528M, 33.1% for M552I/L528(wild type), 17.7% for M552I/L528M and 5.6% for M552I/V/L528M. Serine mutation at YMDD motif (M552S) was not found and the L528M mutation frequently accompanied M552V type. In this study, the typical difference of frequencies for HBV mutation depending on HBeAg was not found. Moreover, the PCR-direct sequencing method used in this study might be a powerful tool for the mutation study in clinical reference laboratories with high volume.

• Asian Pacific Journal of Cancer Prevention
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• v.13 no.8
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• pp.3657-3661
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• 2012

HBV infection is contagious and may be transmitted vertically or horizontally by blood products and body secretions. Over 50% of Iranian carriers have contracted the infection prenatally, making this the most likely route of transmission of HBV in Iran. This study assesses the resistance to Lamivudine in patients with chronic hepatitis B infection using a new ZNA probe Real Time PCR method. To evaluate the effectiveness of Lamivudine therapy for chronic hepatitis B infection, a study was conducted on 70 patients (63 men and 7women), who had received the drug first line. All patients were tested for the presence of HBsAg and HBeAg, the serum ALT level and the HBV DNA load before and after treatment. In all samples resistance to Lamivudine was tested with the ZNA Probe. Our results showed that ZNA Probe Real Time PCR method could detect wild type,YMDD, and its mutants, tyrosine-isoleucine-aspartate-aspartate and tyrosine-valine-aspartate-Aspartate. Among an estimated seventy patients with chronic hepatitis B infection, 18 (25.7%) were resistant to lamivudine. Only one patient was negative for presence of HBS-Ag (5.6%) and two patients were negative for HBe-Ag (11.1%). Real-time PCR with Zip nucleic acid probes is a sensitive, specific and rapid detection method for mutations in the YMDD motif, which will be essential for monitoring patients undergoing Lamivudine antiviral therapy.

• Asian Pacific Journal of Cancer Prevention
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• v.15 no.2
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• pp.1065-1065
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• 2014

• Asian Pacific Journal of Cancer Prevention
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• v.13 no.11
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• pp.5463-5467
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• 2012

Hepatitis B virus (HBV) infection is contagious with transmissiobn vertically or horizontally by blood products and body secretions. Over 50% of Iranian carriers contracted the infection prenatally, making this the most likely route of transmission of HBV in Iran. To evaluate the resistance to adefovir (ADV) therapy in patients with chronic hepatitis B infection, a study was conducted on 70 patients (63 males and 7 females), who had received in first line lamivudine and second line adefovir. All were tested for the presence of hepatitis B surface antigen (HBsAg), hepatitis B envelope antigen (HBeAg), serum alanine amino transferase (ALT) level and HBV DNA load before and after treatment with ADV. In all samples, resistance to lamivudine and ADV was tested with real time PCR. Among seventy patients with chronic hepatitis B infection, 18 (25.7%) were resistant to LAM and 8 (11.4%) were resistant to ADV. Only one patient was negative for the presence of HBS-Ag (5.6%) and two were negative for HBe-Ag (11.1%). In this study we used a new method (ALLGIO probe assay) that has high sensitivity in detection of adefovir resistance mutants, which we recommend to other researchers. Mutant strains of the YMDD motif of HBV polymerase can be found in some patients under treatment with lamivudine and ADV. ADV has been demonstrated to be efficient in patients with lamivudine resistant HBV.

• Pediatric Gastroenterology, Hepatology & Nutrition
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• v.11 no.2
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• pp.130-136
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• 2008

Purpose: Lamivudine is known to be effective for the treatment of chronic hepatitis B in adults. However, data on lamivudine therapy in pediatrics is limited. The aim of this study was to evaluate the efficacy and durability of lamivudine therapy for chronic hepatitis B in Korean children. Methods: A total of 44 children (27 males and 17 females, ages 6 months to 14.8 years, mean age 6.7 years) with chronic hepatitis B who received lamivudine (3 mg/kg/day, max 100 mg) for at least 12 months were enrolled. We evaluated the serum AST, ALT and serological HBV markers (HBsAg and anti-HBs, HBeAg and anti HBe, and HBV DNA) periodically. Predictive three year cumulative seroconversion rates were obtained using the Kaplan-Meier method. Results: Twenty one (48%) of 44 children achieved seroconversion of HBeAg by three years, while 23 (42%) children did not. HBV DNA was cleared in 34 (77%) children and the serum ALT levels were normalized in 41 children (93%). The three year cumulative seroconversion rates were 60% for HBeAg, and the clearance rates were 76% for HBV DNA. Eighteen children who discontinued lamivudine after HBeAg seroconversion maintained the therapeutic response for three years (treatment duration 13~58 months mean 24 months). Viral breakthrough developed in 12 children (27%) during the therapy and the YMDD mutation was documented in 11 children (25%). The mean duration for the development of a mutation was 22.7 months. Loss of HBsAg occurred in 6 children (14%). The pretreatment ALT levels were higher in responders; however, the differences were not statistically significant (p>0.05). Conclusion: The results of this study showed that lamivudine treatment had a favorable effect and durable therapeutic response in children with chronic hepatitis B. Long term follow-up and alternative therapy are warranted for those patients who do not respond to this treatment.