Entecavir Resistance Research Articles

Detection of Resistance Mutations in Chronic Hepatitis B Patients Receiving Antiviral Therapy for Over One Year

Primary mutations conferring hepatitis B virus (HBV) antiviral resistance and associated secondary/compensatory mutations were investigated in this study by DNA sequencing (SEQ) and two commercial Line Probe Assays (LiPAs) (Inno-Lipa HBV DRv2 and Inno-Lipa HBV DRv3, Innogenetics, Belgium). A total of 71 subjects under follow-up for chronic hepatitis B and receiving lamivudine (LVD) therapy for one year or longer at the Hacettepe University Faculty of Medicine, Department of Internal Medicine, Gastroenterology Unit were included in the study with informed consent. Male to female ratio and mean age was noted as 47/24 and 43 ± 15.8 (age range: 13-71) years, respectively. Twenty samples with low HBV DNA levels (mean: 204.6 IU/ml) could not be interpreted by SEQ due to insufficient amplification. All samples with a positive consensus PCR were further analysed via LiPAs, as directed by the manufacturer. Thus a total of 51 and 56 samples could be evaluated via SEQ and LiPA assays, respectively. In 13 of the 51 (25.5%) samples by SEQ and in 9 of 56 (16%) samples by LiPAs, primary and compensatory mutations associated with resistance were identified. Multiple mutations that comprise L180M + M204I in four and L180M + M204V in one sample and single mutations (M204I) in three samples were identified by SEQ. In one sample which had multiple mutations associated with LVD resistance single mutations (S202G, N236T) associated with entecavir resistance and in two other such samples mutations associated with adefovir resistance were detected by SEQ. Also, in three samples amino acid substitution at position rt215 (QÆS) as alone and in one sample with multiple mutations were observed via SEQ. In five of nine samples primary and compensatory multiple mutations (L180M + M204I in one sample, L80I + L180M + M204I in two samples, L80I/V + M204I in one sample) and primary single mutations associated with LVD resistance (M204I/V) in four samples were detected by Inno-Lipa HBV DRv2. Two different mutations (G202, ILFM184) were observed in two samples with multiple mutations associated LVD resistance via Inno-Lipa HBV DRv3. However, mutation at position rt184 was evaluated as a weak positive. Any mutation associated with adefovir resistance was not detected by LiPA. As a result, SEQ and LiPAs displayed comparable performances for the detection of HBV drug resistance mutations. We suggested that for the evaluation of discordant results, it should be better to test consecutive serum samples.

Dynamics of lamivudine-resistant hepatitis B virus strains in patients with entecavir rescue therapy

Entecavir rescue therapy is frequently used in patients with lamivudine-resistant hepatitis B virus (HBV) strains. The aim of this study was to investigate evolutionary patterns of HBV quasispecies during entecavir rescue therapy and evaluate their impacts on therapeutic efficacy. We enrolled 21 chronic hepatitis B patients who failed to respond to lamivudine therapy and were switched to entecavir treatment. Measurement of serum HBV DNA and sequence analysis of HBV reverse transcriptase were done up to 144 weeks. Four patients of this series showed a reversion to wild-type HBV after entecavir treatment and in three of them, a complete viral response (<2.6 log10 copies/ml) was achieved. An additional five patients developed entecavir genotypic resistance, with prior occurrence of lamivudine-resistant mutation (L180 M ± M204 V/I). A viral breakthrough was observed in four of the five patients with entecavir-resistant mutants. The remaining 12 patients of this series showed dominance of lamivudine-resistant mutants throughout the entecavir rescue therapy, and five of them achieved a complete viral response at the end of follow-up. The average HBV DNA level was significantly lower in patients with a reversion to wild-type HBV than in those without it (P < 0.05). In conclusion, reversion to wild-type HBV is a favorable indicator for response to entecavir rescue therapy in lamivudine-refractory patients with chronic hepatitis B. The presence of lamivudine-resistant mutations contributes to the development of entecavir resistance.

Resistance is no Longer a Problem with Entecavir and Tenofovir

The treatment of chronic hepatitis B has been transformed with the advent of nucleos(t)ide analogues. With this came the recognition of viral resistance which was defined as rebound of viral HBV DNA >1 log from nadir associated with genotypic mutations of viral resistance. Resistance was associated with loss of efficacy, with the clinical consequences of viral relapse, reactivation of chronic hepatitis B, increased risk of hepatitis B flares, liver decompensation, and increased mortality especially in cirrhotic patients. Resistance rates varied between different nucleos(t)ide analogues and were generally classified as low or high genetic barrier to resistance. Different pathways to development of viral resistance were recognized, with therapeutic options for rescue therapy determined by such pathways. The two high genetic barrier drugs, entecavir and tenofovir showed remarkably low or absence of viral resistance in treatment naive patients over 5 years or longer of therapy. However in treatment-experienced patients, particularly those with viral resistance, results were less optimal. For entecavir, prior lamivudine resistance conferred an entecavir resistance rate of 43 % over 5 years while with tenofovir, although resistance did not seem apparent, patients with prior adefovir resistance had suboptimal responses to tenofovir therapy. In cases of multiresistant hepatitis B virus defined as resistance to at least two nucleos(t)ide analogues in the same viral strain, rescue with a combination of tenofovir and entecavir appeared highly efficacious. Consequently, the optimistic view that these new drugs lead to absence of resistance needs to be tempered by the larger burden of drug experienced and drug resistant patients compared to treatment naive patients. Furthermore, it is unclear whether resistance may be a concern with high genetic barrier drugs after one or two decades of therapy. In time, it is possible that with the increasing use of high genetic barrier drugs, viral resistance will be a lesser problem, but unlikely to disappear completely.

Efficacy and safety of telbivudine plus adefovir dipivoxil combination therapy and entecavir monotherapy for HBeAg‐positive chronic hepatitis B patients with resistance to adefovir dipivoxil

The objective of this study was to compare the efficacy and safety of two rescue strategies for hepatitis B e antigen (HBeAg)-positive chronic hepatitis B (CHB) patients with resistance to adefovir dipivoxil (ADV). This prospective study included 58 HBeAg-positive CHB patients with resistance to ADV; 30 patients underwent telbivudine (LdT) plus ADV combination therapy and 28 patients switched to entecavir (ETV) monotherapy. After 48weeks of treatment, the rates of hepatitis B virus (HBV) DNA <3 log10 copies/mL in the LdT plus ADV group and the ETV group were not significantly different (73.3% vs 57.1%, P=0.195). Six patients receiving LdT plus ADV had HBeAg seroconversion, while none of the patients receiving ETV alone had HBeAg seroconversion (20% vs 0%, P=0.039). During the 48-week treatment period, two patients in the ETV monotherapy group had viral breakthrough and the strains were confirmed to be of a variant associated with ETV resistance (rtM204V+ rtL180M+ rtT184G), while one patient receiving LdT plus ADV had viral breakthrough and an LdT-associated resistance mutation (rtM204I) was detected. For the majority of the patients, both LdT plus ADV combination treatment or ETV monotherapy were generally well tolerated, and no serious side effects were observed. Both LdT plus ADV combination therapy and ETV monotherapy led to significant decreases in serum HBV DNA in HBeAg-positive CHB patients with resistance to ADV, and LdT plus ADV combination therapy exhibited a significantly higher rate of HBeAg seroconversion compared with ETV monotherapy.

Virological response and antiviral resistance mutations in chronic hepatitis B subjects experiencing entecavir therapy: An Indian subcontinent perspective

Entecavir is one of the therapeutic options currently available for the management of chronic hepatitis B. In this study, we aimed to analyse the virological response and antiviral resistance mutations in chronic hepatitis B subjects experiencing entecavir therapy from the Indian subcontinent. A total of 45 chronic hepatitis B subjects were studied at baseline and were followed up on entecavir treatment. Among these subjects, 25 (56%) were HBeAg-positive at baseline. Virological response was measured by hepatitis B virus (HBV) DNA levels. HBV reverse transcriptase (rt) domains were sequenced for the identification of resistance mutations. Three-Dimensional (3D) model of HBV polymerase/rt protein, docking and molecular dynamics simulation (MDS) studies were performed for characterization of antiviral resistance mutations. At the median treatment duration of 6 (IQR 6–11) months, 38 (84%) showed virological response. Subjects who showed anti-HBe response demonstrated significant association with virological response (p=0.034). On sequence analysis, none of the subjects were identified with signature entecavir resistance mutations. However, one subject was exclusively detected with rtV173L mutation. Molecular modeling, docking and MDS studies revealed that the rtV173L mutation cannot confer resistance to entecavir independently. Our findings also showed that the prevailing HBV genotypes, subgenotypes and HBsAg subtypes in this population does not influence treatment outcome to entecavir therapy. In conclusion, entecavir is a potent drug in terms of viral DNA suppression. In addition, none of the subjects developed antiviral resistance mutations to entecavir. Therefore entecavir is a suitable drug of choice in the management of chronic HBV.

High frequency of complex mutational patterns in lamivudine resistant hepatitis B virus isolates

The evolution of hepatitis-B virus (HBV) drug resistance is characterized by the emergence of resistance conferring mutations and compensatory mutations. Therefore HBV drug-resistant isolates often harbor multiple mutations in the reverse transcriptase (RT) and corresponding mutations in the hepatitis B surface antigen (HBsAg). In this study mutational patterns of 60 HBV isolates harboring drug resistance mutations rtM204V or rtM204I were retrospectively analyzed. Both mutations, especially mutation rtM204V, were most often accompanied by compensatory mutations rtV173L and rtL180M but also by mutations conferring entecavir (n = 5) or adefovir resistance (n = 4). In addition, 22 (36.7%) drug resistant HBV isolates carried mutations related to immune escape in the HBsAg. In seven cases premature stop codons in HBsAg were detected resulting in the expression of truncated HBsAg. Clonal analysis of these seven quasispecies even disclosed the presence of HBV isolates carrying further stop codons and in one case the occurrence of resistance mutation rtA181T without the stop codon mutation sW172*. Interestingly, only one HBV clone carried the resistance mutations rtM204V and rtA181T. HBV drug resistant isolates frequently harbored HBsAg mutations associated with immune escape or disease progression pointing to a complex interaction of both proteins. HBV genotypic resistance tests based on population sequencing methods seemed to be inappropriate for determining the clinical relevance of stop codons in the HBsAg.

Two‐step evolution of the hepatitis B drug‐resistant mutations in a patient who developed primary entecavir resistance

Few cases of primary entecavir resistance in chronic hepatitis B patients have been reported to date. The serial profiling of the HBV polymerase gene mutations from a treatment-naive patient who developed drug resistance after 32months of entecavir therapy is presented here. Serum samples were collected at multiple time points from before the start of therapy to virological and biochemical breakthrough. The evolution of the hepatitis B virus polymerase gene mutations was analysed with commercial line probe assay and pyrosequencing. Drug resistance mutation analysis by pyrosequencing revealed a two-step process in the selection of drug resistance. The patient had a good initial response to entecavir 0.5mg/day. A partially resistant HBV strain first emerged as the predominant species from as early as 2weeks. After a period of non-compliance to therapy, there was virological breakthrough, which resolved on restarting entecavir. Shortly after, there was secondary failure of entecavir therapy, caused by a new resistant strain carrying all three mutations required. In this patient, pre-existence of minor population of partially resistant viral strains and treatment non-compliance probably contributed to his development of primary entecavir resistance.

Clonal Analysis of the Quasispecies of Antiviral-Resistant HBV Genomes in Patients with Entecavir Resistance during Rescue Treatment and Successful Treatment of Entecavir Resistance with Tenofovir

Clonal analysis of quasispecies of resistant HBV genomes in patients with entecavir (ETV) resistance receiving lamivudine (3TC) plus adefovir (ADV) rescue therapy has never been performed. A sample of 10 patients with ETV resistance who were switched to 3TC+ADV treatment were analysed for changes in viral quasispecies. Serum samples at baseline, and at months 3 and 6 of 3TC+ADV treatment could be clonally analysed in 7 of 10 patients; 3-82 clones per sample (total 1,068 clones, mean 63) were sequenced. 3TC+ADV therapy led to a modest decline in HBV DNA. Almost all clones had L180M and M204V 3TC resistance mutations before and during combination therapy. All clones had ≥1 of the S202G, T184F, T184A, T184L, T184I and M250V ETV resistance mutations. The percentages of detected clones bearing 3TC (rtL180M and rtM204V) and ETV mutations did not change with rescue 3TC+ADV therapy. In 7 of 8 patients with detectable HBV DNA (median 5.17 log(10) copies/ml) after a median 24 months of ADV therapy, HBV DNA became undetectable with 3TC plus tenofovir after 6 months of treatment. In patients with ETV resistance tenofovir is effective. Clonal analysis data indicate no selection of specific HBV mutants during rescue 3TC+ADV.

Performance evaluation of the HepB Typer-Entecavir kit for detection of entecavir resistance mutations in chronic hepatitis B

Background/AimsMolecular diagnostic methods have enabled the rapid diagnosis of drug-resistant mutations in hepatitis B virus (HBV) and have reduced both unnecessary therapeutic interventions and medical costs. In this study we evaluated the analytical and clinical performances of the HepB Typer-Entecavir kit (GeneMatrix, Korea) in detecting entecavir-resistance-associated mutations.MethodsThe HepB Typer-Entecavir kit was evaluated for its limit of detection, interference, cross-reactivity, and precision using HBV reference standards made by diluting high-titer viral stocks in HBV-negative human serum. The performance of the HepB Typer-Entecavir kit for detecting mutations related to entecavir resistance was compared with direct sequencing for 396 clinical samples from 108 patients.ResultsUsing the reference standards, the detection limit of the HepB Typer-Entecavir kit was found to be as low as 500 copies/mL. No cross-reactivity was observed, and elevated levels of various interfering substances did not adversely affect its analytical performance. The precision test conducted by repetitive analysis of 2,400 replicates with reference standards at various concentrations showed 99.9% agreement (2398/2400). The overall concordance rate between the HepB Typer-Entecavir kit and direct sequencing assays in 396 clinical samples was 99.5%.ConclusionsThe HepB Typer-Entecavir kit showed high reliability and precision, and comparable sensitivity and specificity for detecting mutant virus populations in reference and clinical samples in comparison with direct sequencing. Therefore, this assay would be clinically useful in the diagnosis of entecavir-resistance-associated mutations in chronic hepatitis B.

Do different lamivudine‐resistant hepatitis B genotypes carry the same risk of entecavir resistance?

Entecavir switch is one of the treatment options for lamivudine-resistant hepatitis B (HBV) patients in Asia. This study examined the outcome of patients with different baseline resistance genotypes in a cohort study. In this study, 14 patients with chronic HBV were treated with entecavir 1 mg/day for 5 years. Enrolment criteria include: documented lamivudine resistant mutations, treatment with adefovir 10 mg/day for at least 24 weeks, and Child-Pugh score <7. Most had previous failed adefovir therapy and compensated cirrhosis of the liver. Clinical outcomes, liver biochemistries, and HBV DNA were monitored regularly. Patients with virologic breakthrough were rescued with add-on adefovir. At the end of the treatment period, the mean HBV DNA fell from 5.92 × 10(6) (baseline) to 3.67 × 10(1) IU/ml. The presence of a HBV polymerase rtM204V mutation at the baseline was found to be the major risk factor for adverse outcomes. Compared to the patients with the rtM204I mutant, patients with the rtM204V mutant had increased risk of virologic breakthrough (80% vs. 0%, P = 0.010) requiring add-on adefovir, slower virologic responses (log rank test, P = 0.0011), failure to reach undetectable HBV DNA levels (60% vs. 0%, P = 0.045), and higher risk of entecavir-resistance (60% vs. 0%, P = 0.045). All the patients with rtM204I and rtA181 mutants had undetectable HBV DNA from 18th month. In summary, lamivudine-resistant HBV patients with the rtM204V mutation have the highest risk of developing entecavir resistance, and entecavir monotherapy should be avoided. Those with the rtM204I and rtA181V mutations may have lower risks, but regular surveillance for viral breakthrough is required.

Efficacy of Entecavir With or Without Tenofovir Disoproxil Fumarate for Nucleos(t)ide-Naïve Patients With Chronic Hepatitis B

Entecavir (ETV) and tenofovir disoproxil fumarate (TDF) are potent antiviral agents that might have additive or synergistic antiviral activity in treatment of patients with chronic hepatitis B (CHB). We compared the efficacy and safety of ETV monotherapy with those of a combination of ETV and TDF. We performed a randomized, open-label, multicenter, superiority study of 379 nucleos(t)ide-naïve patients with hepatitis B e antigen (HBeAg)-positive (n = 264) or HBeAg-negative (n = 115) CHB. Subjects were given ETV 0.5 mg (n = 182) or a combination of ETV 0.5 mg and TDF 300 mg (n = 197) for 100 weeks. At week 96, comparable proportions of patients in each study arm achieved the primary end point of a level of hepatitis B virus (HBV) DNA <50 IU/mL (83.2% vs 76.4%; P = .088). Among HBeAg-positive patients, a greater proportion given combination therapy achieved levels of HBV DNA <50 IU/mL than those given ETV alone (80.4% vs 69.8%; P = .046). However, this difference was observed only in patients with baseline levels of HBV DNA ≥ 10(8) IU/mL (79% vs 62%) and not in those with baseline levels of HBV DNA <10(8) IU/mL (83% in both arms). Rates of HBeAg loss and HBeAg seroconversion were comparable between groups, whereas the rate of alanine aminotransferase normalization was greater in the ETV monotherapy group. No HBV variants associated with ETV or TDF resistance were detected. Safety profiles were consistent with previous reports of ETV or TDF monotherapy. The antiviral efficacy of ETV monotherapy is comparable to that of ETV plus TDF in a mixed population of nucleos(t)ide-naïve patients with CHB (70% HBeAg positive). The combination therapy could provide an incremental benefit to HBeAg-positive patients with baseline levels of HBV DNA ≥ 10(8) IU/mL.

High efficacy of adefovir and entecavir combination therapy in patients with nucleoside-refractory hepatitis B

Background/AimsNewly developed and potent antiviral agents suffer from the problem of drug resistance. Multidrug resistance is a major impediment in the treatment of patients with chronic hepatitis B (CHB). In line with American Association for the Study of Liver Diseases guidelines, adefovir dipivoxil (ADV) add-on therapy is recommended in the case of lamivudine resistance, while tenofovir disoproxil fumarate (TDF) is recommended for ADV or entecavir (ETV) resistance. TDF is currently not available in Korea. ADV+ETV combination therapy may be a viable alternative to TDF in patients with either ADV or ETV resistance. However, the efficacy of ADV+ETV combination therapy in patients with CHB and multidrug resistance is unclear. This study investigated the efficacy of ADV+ETV combination therapy in patients with multidrug resistance.MethodsTwenty-five patients were enrolled and were administered ADV+ETV combination therapy for at least 6 months. Blood was drawn at baseline and at 3, 6, 9, and 12 months after commencing treatment, and the following blood parameters were analyzed: alanine transaminase, hepatitis B e-antigen (HBeAg), anti-hepatitis B e-antigen, and hepatitis B virus (HBV) DNA levels. The initial virological response (IVR) was defined as an HBV DNA level of <4 log10 copies/mL after 6 months of combination therapy.ResultsThe IVR rate was 76%. The proportion of patients with a high viral load (≥5.0 log) dropped from 76% at baseline to only 5% after 6 months of treatment. The biochemical response rate during the first 6 months was 71%. HBeAg was lost in 2 patients (10%).ConclusionsADV+ETV combination therapy induced a good IVR in CHB patients who were refractory to more than 2 antiviral agents. This regimen may be a good alternative to TDF in Korea, where that drug is not available.

Molecular characterization of a novel entecavir mutation pattern isolated from a multi-drug refractory patient with chronic hepatitis B infection

Background Prolonged antiviral treatment results in selection and accumulation of resistant strains in quasispecies pool in hepatitis B virus (HBV) infection. Objectives The aim of this study was to characterise a novel HBV pattern which shows resistance to lamivudine, adefovir dipivoxil and entecavir using in vitro phenoyping assay. Study design A male 36 years old patient diagnosed with anti HBe-positive chronic hepatitis B (CHB) had received lamivudine treatment for 7 years following an initial unsuccessfull interferon treatment. The therapy had been switched to adefovir and then to entecavir when breakthrough occcured during each treatment. This led only to a temporary HBV DNA decline which soon was followed by viral breakthrough despite the lack of known entecavir resistance mutations. Patient died after 9 months of entecavir treatment from liver failure. A total of 434 clones from 6 different serum samples were analysed retrospectively. HBV genomes bearing mutation patterns suggestive of antiviral resistance were analysed by in vitro phenotyping assay. Results Dominance of a clone carrying L80LV, L91I, M204I, S219A, N238D, Y245H changes was detected in the last serum sample of the patient just before his death. This pattern displayed 30.4 fold resistance to entecavir when compared with the wild type HBV by in vitro phenotyping assay. Conclusion A novel mutation pattern showing a high degree of resistance to entecavir was documented. In this pattern, the S219A and Y245H mutations mainly seem to contribute to the emergence of ETV resistance.

Entecavir plus tenofovir combination as rescue therapy in pre-treated chronic hepatitis B patients: An international multicenter cohort study

Long-term viral suppression is a major goal to prevent disease progression in patients with HBV. Aim of this study was to investigate the efficacy and safety of entecavir plus tenofovir combination in 57 CHB partial responders or multidrug resistant patients. Investigator-initiated open-label cohort study. Quantitative HBV-DNA measurement and resistance testing (line-probe-assays and direct-sequencing) at baseline and every 3 months. Fifty seven patients (37 HBeAg+), median age 45 years, previously treated with a median of three lines of antiviral therapy (range 1-6), 24/57 with advanced liver disease, were included. Median ALT at baseline was 1.0 ULN (range 0.3-22) and HBV-DNA 1.5 × 10(4)IU/ml (range 500-1 × 10(11)IU/ml). Median treatment duration of combination therapy was 21 months. HBV-DNA level dropped 3 logs (median, range 0-8 log; p<0.0001), 51/57 patients became HBV-DNA undetectable, median after 6 months (95% CI, 4.6-7). The probability for HBV DNA suppression was not reduced in patients with adefovir or entecavir resistance or in patients with advanced liver disease. Viral suppression led to decline in ALT (median 0.7 ULN; range 0.2-2.4; p=0.001). Five patients lost HBeAg (after 15, 18, 20, 21, and 27 months, respectively), one patient showed HBs-seroconversion. Patients with advanced disease did not show clinical decompensation, two patients with cirrhosis and undetectable HBV DNA developed HCCs. No death, newly induced renal impairment or lactic acidosis were reported. Rescue therapy with entecavir and tenofovir in CHB patients harboring viral resistance patterns or showing only partial antiviral responses to preceding therapies was efficient, safe, and well tolerated in patients with and without advanced liver disease (249).

Frequency and Mutation Patterns of Resistance in Patients with Chronic Hepatitis B Infection Treated with Nucleos(t)ide Analogs in Add-On and Switch Strategies

: Background: Treatment for chronic hepatitis B (CHB) has improved over the last 10 years mainly due to the development of effective oral antiviral agents [nucleoside/nucleotide analogs (NUCs)].Objectives: The aim of the present study is to identify the frequency and major patterns of resistance to the hepatitis B virus (HBV) in a Turkish population of CHB patients treated with NUCs using add-on and switch therapy strategies.Patients and Methods: The investigation involved a total of 194 patients (88 were treated using add-on therapy, and 106 were treated using switch therapy). We analyzed the HBV polymerase gene by amplification and direct sequencing procedures.Results: Primary drug-resistance mutations were detected in 84 patients (43%; 42 in add-on therapy, and 42 in switch therapy) taking lamivudine (LAM), 10 patients (5%; 6 in add-on therapy, and 4 in switch therapy) taking entecavir (ETV), and 16 patients (8%; 8 in add-on therapy, and 8 in switch therapy) taking adefovir (ADV). The most common LAM and ETV resistance mutations were rtM204I/V, rtL180M and rtT184A/I/S, respectively, while rtA181T/V and rtN236T substitutions were the most frequently observed ADV resistance mutations.Conclusions: Patients with CHB who developed NUC resistance were managed using 2 different rescue strategies. The frequency and mutation pattern of resistance were similar in patients treated with add-on and switch strategies. These findings may be helpful in the management of rescue strategies in LAM-resistant patients. Implication for health policy/practice/research/medical education:Authors revealed of major drug resistance patterns and their frequencies in add-on and switch strategies in treatment of chronic hepatitis B virus. This article is an helpful for clinicians involved in the management of rescue strategies in lamivudine-resistant patients in chronic hepatitis B. Please cite this paper as:Sayan M, Akhan SC, Senturk O. Frequency and Mutation Patterns of Resistance in Patients with Chronic Hepatitis B Infection Treated with Nucleos(t)ide Analogs in Add-On and Switch Strategies. Hepat Mon.2011;11(10):835-42. DOI: 10.5812/kowsar.1735143X.775 © 2011 Kowsar M.P.Co. All rights reserved.

Evolution of hepatitis B virus during long-term therapy in patients with chronic hepatitis B

Background. Long-term lamivudine (LAM), adefovir (ADV) and entecavir (ETV) treatment induce the emergence of drug-resistant hepatitis B virus (HBV) in patients with chronic hepatitis B infection.Aim. To evaluate the LAM, ADV and ETV resistance mutations detected in our patient group.Materials and methods. Twenty patients who had received at least two years of treatment with nucleoside/tide analogues were enrolled in this study. Patients with detectable HBV DNA were analyzed in order to detect resistance mu-tations and in this group of patients treatment was change.Results. Three patients developed LAM resistance mutations (2 presented rtM204I and one rtL180M+rtM204V/I) and one patient showed rtN236T ADV resistance mutation. During ADV and LAM treatment, one patient developed ADV plus LAM resistance mutations (rtI163V+rtL180M+rtA181V+rtN236T), in this case, HBV strains harbouring polymerase mutations did not develop LAM associated rtM204V/I primary mutation. In addition, ETV resistance mutations (rtL180M+rtT184A+rtS202G+rtM204V) were detected in one patient.Conclusions. These findings suggest that monotherapy resulted in a limited virological response and combination strategies including potent antiviral agents should be recommended for patients with resistant mutations.

Frequency and Mutation Patterns of Resistance in Patients with Chronic Hepatitis B Infection Treated with Nucleos(t)ide Analogues in Add-On and Switch Strategies

Treatment for chronic hepatitis B (CHB) has improved over the last 10 years mainly due to the development of effective oral antiviral agents [nucleoside/nucleotide analogs (NUCs)]. The aim of the present study is to identify the frequency and major patterns of resistance to the hepatitis B virus (HBV) in a Turkish population of CHB patients treated with NUCs using add-on and switch therapy strategies. The investigation involved a total of 194 patients (88 were treated using add-on therapy, and 106 were treated using switch therapy). We analyzed the HBV polymerase gene by amplification and direct sequencing procedures. Primary drug-resistance mutations were detected in 84 patients (43%; 42 in add-on therapy, and 42 in switch therapy) taking lamivudine (LAM), 10 patients (5%; 6 in add-on therapy, and 4 in switch therapy) taking entecavir (ETV), and 16 patients (8%; 8 in add-on therapy, and 8 in switch therapy) taking adefovir (ADV). The most common LAM and ETV resistance mutations were rtM204I/V, rtL180M and rtT184A/I/S, respectively, while rtA181T/V and rtN236T substitutions were the most frequently observed ADV resistance mutations. Patients with CHB who developed NUC resistance were managed using 2 different rescue strategies. The frequency and mutation pattern of resistance were similar in patients treated with add-on and switch strategies. These findings may be helpful in the management of rescue strategies in LAM-resistant patients.

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Effectiveness of entecavir in chronic hepatitis B NUC-naive patients in routine clinical practice

Registration studies showed entecavir (ETV) to be effective and safe in NUC-naïve patients with chronic hepatitis B virus (HBV), but its effectiveness in routine clinical practice is unknown. Sixty-nine HBeAg positive and negative NUC naïve chronic HBV patients were treated with ETV for 110 weeks. 63% were HBeAg positive, 16% were cirrhotics, mean HBV-DNA was 7.09 log IU/ml and mean ALT was 157 IU/ml. Sixty-one (88%) patients achieved undetectable DNA, with 46%, 77% and 100% virological response rates at week 24, 48 and 96 of treatment, respectively. Thirty-seven (84%) patients in the HBeAg-positive population achieved undetectable DNA, with 67% and 100% virological response rates at week 48 and 96 of treatment, respectively. Twenty-four (96%) patients in the HBeAg-negative population achieved undetectable DNA, with 91% and 100% virological response rates at week 48 and 96 of treatment, respectively. Twenty-three (53%) patients cleared HBeAg and 19 (44%) patients seroconverted to antiHBe positive status; seven (10%) patients cleared hepatitis B surface antigen and five (7%) patients developed antiHBs. At the end of the study, 10 patients successfully stopped therapy: nine HBeAg positive (four developed antiHBs positive) and one HBeAg negative. None of the patients had primary non-response. ETV resistance was not tested. None of the patients developed hepatocellular carcinoma, underwent liver transplantation or died because of liver-related events. No serious adverse events were reported. The ETV monotherapy showed high virological response rates, a favourable safety profile for NUC-naive HBeAg-positive and negative patients treated in routine clinical practice.

Evolution of hepatitis B virus quasispecies during antiviral therapy in one chronic hepatitis B patient

To investigate the evolution of hepatitis B virus (HBV) quasispecies in one patient during lamivudine (LAM) monotherapy and switching to entecavir (ETV) rescue treatment. Serum samples were taken at seven different time points during antiviral therapy (0, 24, 48, 60, 72, 96, 152 weeks, respectively), the HBV DNA polymerase gene was amplified, cloned and sequenced to analyze the amino acid substitutions within HBV DNA polymerase gene and distribution of virus quasispecies. Quantitative detection of the HBV wild strains and total virus was performed by amplification refractory mutation system real-time PCR (ARMS-PCR). Three mutation patterns detected during antiviral therapy in the patient: rtM204V, rtM204V+rtL180M and rtM204I. The HBV quasispecies were found always in dynamic variation. The HBV populations were completely replaced with the LAM-resistant variants when the viral breakthrough was encountered during LAM monotherapy. Interestingly, the wild-type variants presented gradually dominant (79.3%) with the decline of HBV DNA load after switching to ETV rescue administration. ARMS-PCR results showed that the wild-type variants account ed for 68.55% of the HBV populations at baseline and this proportion declined to 0.21% when the viral breakthrough emerged under LAM therapy. The wild-type variants gradually increased from week 24 after switching to ETV rescue therapy and the proportion of HBV wild-type variants in the population fluctuated between 16.01% to 26.93%. The distribution of virus quasispecies were always in dynamic variation during sequential therapy with nucleotide analogs in chronic hepatitis B patients. Different patterns of dynamic HBV quasispecies may have different contribution in ETV resistance in LMV refractory patients with ETV administration.