Resistant Hepatitis Research Articles

20-fold increase in risk of lamivudine resistance in hepatitis B virus subtype adw

We investigated subtype-dependent development of lamivudine resistance in hepatitis B virus (HBV) longitudinally in 26 consecutive patients (13 adw and 13 ayw carriers) during antiviral treatment of chronic hepatitis B. Lamivudine resistance developed in seven adw carriers and one ayw carrier. Risk of lamivudine resistance was significantly higher for adw carriers than for ayw carriers (p=0.03). We believe that the adw subtype of HBV is associated with a high risk of lamivudine resistance, which might be linked to simultaneous changes of the HBsAg that occurs with the emergence of resistance.

Molecular anatomy and pathophysiologic implications of drug resistance in hepatitis B virus infection

Synthesis of the hepatitis B virus (HBV) DNA genome occurs within the viral nucleocapsid in a mechanistically ordered fashion. The nucleocapsid contains small pores that permit influx of nucleotide triphosphates and metabolites of nucleoside analogues such as lamivudine for DNA synthesis. Lamivudine is a potent inhibitor of HBV and human immunodeficiency virus (HIV) reverse transcriptases, but substitutions of isoleucine or valine for methionine within the tyrosine-methionine-aspartate-aspartate (YMDD) motif are associated with virologic and clinical resistance to lamivudine therapy. Under lamivudine selection pressure, the high viral production rate and the low fidelity viral polymerase contribute to frequent development of the YMDD mutants. However, the pattern and dynamics of emergence of the mutant viruses over the wild-type virus are determined by multiple factors including replication efficiency, host immune response, and availability of replication space. Structural modeling of HIV reverse transcriptase has permitted key insights into the molecular basis of lamivudine resistance of HBV based on evolutionary relatedness of HIV and HBV. The side groups of isoleucine and valine of the YMDD mutants sterically prevent lamivudine from appropriately configuring into the nucleotide binding site of the reverse transcriptase. Aminotransferase flares are associated with lamivudine therapy and may signify clinical resistance with emergence of YMDD mutants. They may also herald the recovery phase with seroconversion and viral clearance. Reconstitution of the endogenous anti-HBV immune response may be equally important in the control of viral replication by lamivudine and other nucleoside analogues.GASTROENTEROLOGY 2001;120:1000-1008

In vivo dynamics and pathogenicity of wild-type and resistant Hepatitis B virus during long-term lamivudine monotherapy - a clinical note.

Genotypic resistance of Hepatitis B virus (HBV) against lamivudine evolves within months after onset of therapy. To determine the longitudinal order in which resistance mutations appear and to compare the kinetics and pathogenicity of wild-type and resistant HBV. In a longitudinal study, consecutive samples were drawn over a period of 28 months from a patient with chronic hepatitis B, and resistance mutations were followed by sequencing a part of the polymerase region of HBV. These data were compared with HBV copy numbers, HBsAg and ALT levels, and results of consecutive liver biopsies. After 21 weeks of treatment, a silent mutation at codon 528 (CTG to TTG) occurred. Significant genotypic resistance was detectable after 68 weeks, indicated by a substitution of isoleucine for methionine at residue 552 (M552I). Nineteen weeks later, the virus exhibited additional resistance-associated mutations (L528M and I552V). The resulting high-level resistance was reflected by an increase of serum HBV copies of 4.7 log(10). The turnover of wild-type and resistant HBV was 2.6x10(6) and 1.8x10(6) virions/day, respectively. HBsAg and ALT levels were lower within the period when resistant HBV was detectable. During treatment the progress of liver fibrosis was arrested. The in vivo replicative capacities and dynamics of wild-type and resistant HBV were similar. However, resistant HBV seemed to exhibit reduced pathogenicity.

Monitoring the virus load can predict the emergence of drug-resistant hepatitis B virus strains in renal transplantation patients during lamivudine therapy.

The development of resistant hepatitis B virus (HBV) strains during lamivudine treatment has been described repeatedly. To investigate whether the development of such resistant HBV strains can be predicted in an early phase of therapy, the HBV loads of 11 renal transplantation patients were screened at 3-month intervals by a quantitative HBV polymerase chain reaction (PCR) assay. Lamivudine resistance was detected by sequence analysis. Five patients developed resistance to lamivudine in the 12-15-month follow-up period. In all of them, a virus load of 1x103 HBV DNA copies still was detectable after 3 months of therapy. This was statistically significantly different from those patients who did not develop lamivudine resistance within the observation period, all of whom had no HBV DNA detectable after 3 months of treatment (P=.0022). Thus, virus load testing by use of a sensitive PCR assay allows the early prediction of the emergence of lamivudine-resistant HBV strains.

Line Probe Assay for Monitoring Drug Resistance in Hepatitis B Virus-Infected Patients during Antiviral Therapy

Since the introduction of antiviral compounds such as lamivudine and famciclovir in the treatment schedules of patients with chronic hepatitis B virus (HBV) infection, the accumulation of a variety of mutations in the HBV polymerase gene has been observed. The selection of these mutations is generally considered the cause of viral nonresponsiveness and treatment failure. Therefore, the detection of these mutations is of clinical importance. Previously genotyped HBV strains isolated from treated and untreated patients were amplified with primers specific for the HBV polymerase region from amino acids 465 to 562. Amplified products were cloned into plasmid vectors. The clones were used as reference strains. A set of 38 highly specific oligonucleotide probes covering three different codon positions, L528M, M552V/I, and V/L/M555I, were selected. These probes were applied as 19 different lines on a membrane strip. The strips were then hybridized with PCR fragments from the reference panel, revealing the amino acids at the three codon positions simultaneously for each clone. PCR products generated from two patients infected with HBV genotypes A and C, respectively, and treated with nucleoside analogs were analyzed on these strips. A gradual increase in genetic HBV polymerase complexity was observed in follow-up samples compared to that in pretreatment samples. Additional analysis of HBV polymerase DNA fragments in recombinant plasmid clones demonstrated the existence of (i) clones with double mutations, (ii) clones with single mutations at either codon 528, 552, or 555, and (iii) the simultaneous occurrence of two or more viral populations within one sample. This line probe assay detected the complex quasispecies nature of HBV and provided some insight into the dynamics of resistance mutations.

Outcome of lamivudine resistant hepatitis B virus infection in the liver transplant recipient

BACKGROUNDIn many transplant centres lamivudine is an important component of prophylaxis against, and treatment of, hepatitis B virus (HBV) graft infection. Drug resistant HBV species with specific polymerase mutations may...

Susceptibility of lamivudine-resistant hepatitis B virus to other reverse transcriptase inhibitors.

The emergence of resistant hepatitis B virus (HBV), with mutations in the YMDD motif of the polymerase gene after treatment with lamivudine, is becoming an important clinical problem. In this study, susceptibility of wild-type and lamivudine-resistant HBV M552I, M552V, and L528M/M552V mutants to other reverse transcriptase inhibitors was investigated by transient transfection of full-length HBV DNA into human hepatoma cells. HBV DNA replication was monitored by Southern blot hybridization, which showed the presence of a single-stranded band (representative of the HBV replicative intermediates) in the drug-free, wild-type HBV-transfected cells. This band was diminished in the samples of wild-type HBV DNA treated with either lamivudine, adefovir, or lobucavir. The band intensities from the lamivudine-resistant mutants were not decreased by treatment with lamivudine, but were decreased by the treatments with adefovir or lobucavir. In contrast, penciclovir and nevirapine did not diminish the intensity of the single-stranded band of wild-type HBV or the lamivudine-resistant mutants. These results demonstrate that lamivudine-resistant HBV is susceptible to adefovir and lobucavir. Lamivudine-resistant HBV should be treated with adefovir or lobucavir, and combination therapy with lamivudine and adefovir/lobucavir may prevent the emergence of lamivudine-resistant HBV.

Sensitivity of l-(-)2′,3′-dideoxythiacytidine resistant hepatitis B virus to other antiviral nucleoside analogues

l-(-)2′,3′-Dideoxythiacytidine ( l(-)SddC, Lamivudine) resistant hepatitis B virus (HBV) develops in patients after prolonged treatment. Point mutations detected in the viral genome from these patients have been shown to be responsible for l(-)SddC resistance. Therefore, new drugs active against l(-)SddC resistant HBV are needed. Using a transient transfection system, we studied the sensitivity of l(-)SddC resistant HBV to other anti-HBV nucleoside analogues. It was found that the L526M mutation alone caused greater resistance to penciclovir (PCV) than did the V553I mutation alone. Both mutations also caused the virus to be less sensitive to l(-)SddC and 2′-fluoro-5-methyl-β- l-arabinofuranosyluracil ( l-FMAU), although the degree of resistance was much less than that to PCV. The A546V mutation had no impact on the sensitivity to l(-)SddC, l-FMAU, and PCV. When these single mutations were coupled with the M550V/I mutation, all the double mutants were resistant to those drugs. Although 2′,3′-dideoxy-2′,3′-didehydro-β- l(-)-5-fluorocytidine ( l(-)Fd4C) was also less active, the ic 50 of l(-)Fd4C against the l(-)SddC resistant mutant was at least fifty times lower than that against cell growth in culture. DNA polymerase associated with l(-)SddC resistant virions was also found to be less sensitive than that with wild-type HBV to those l-nucleoside triphosphates. All the l(-)SddC resistant mutants were still sensitive to 9-(2-phosphonylmethoxyethyl)-adenine (PMEA). These results suggest that different mutations in the HBV genome have a different impact on its sensitivity to those compounds, and l(-)SddC resistant HBV may also be resistant to PCV, l-FMAU, and l(-)Fd4C. A nucleoside analogue less toxic than PMEA could be developed against l(-)SddC resistant HBV.

Interferon-α plus ribavirin in chronic hepatitis C resistant to previous interferon-α course: results of a randomized multicenter trial

Background/Aims: Interferon-α plus ribavirin seem to be more efficacious than interferon monotherapy in chronic hepatitis C. In a multicenter randomized trial, we evaluated the efficacy of this association for interferon-α resistant chronic hepatitis C. Methods: Fifty patients who were non-responders to recombinant or lymphoblastoid interferon-α were randomized to receive either ribavirin (800 mg/day) plus leucocytic interferon-α (3 mega units thrice weekly) or the same dose of interferon-α alone, for 6 months. Effects of therapy were evaluated by serum aminotransferase and hepatitis C virus RNA levels and control liver biopsies. Results: At the end of treatment, aminotransferase levels become normal in 9/26 patients receiving combination therapy (35% [confidence interval, 16% to 53%]) and in 2/24 receiving interferon-α alone (8% [confidence interval, −3% to 19%]) ( p=0.03). Aminotransferase normalization was never associated with hepatitis C virus RNA clearance. All patients with normal aminotransferase relapsed after discontinuation of therapy. At the end of treatment, mean hepatitis C virus RNA levels significantly decreased only in the group receiving combination therapy, but returned to pretreatment values 6 months thereafter. No histological improvement was observed in either group. Conclusions: There is no indication for treatment with interferon-α at the dose of 3 mega units thrice weekly plus 800 mg/day of ribavirin for 6 months in chronic hepatitis C resistant to interferon-α.

Functional analysis of mutations conferring lamivudine resistance on hepatitis B virus.

Two patterns of mutation are commonly observed in the polymerase gene of lamivudine [(-)2'-deoxy-3'-thiacytidine]-resistant hepatitis B virus (HBV). The M539I substitution in the conserved YMDD motif occurs independently of other changes, whereas the M539V substitution is associated with an additional upstream change (L515M). These mutations were introduced into a common background and their effects on HBV DNA replication and lamivudine resistance studied. The L515M and M539V mutations provided only partial resistance while the M539I mutation conferred a high degree of lamivudine resistance. The combination of the L515M and M539V mutations gave an intermediate level of replication competence, compared with either mutation alone, and increased resistance to lamivudine. This probably accounts for these two mutations always being observed together. The M539I mutation reduced replication competence.

Cyclosporine therapy in patients with steroid resistant autoimmune hepatitis

Autoimmune hepatitis is a form of chronic liver disease characterized by progressive hepatocellular inflammation, which usually responds to treatment with corticosteroids. However, 10% of patients with autoimmune hepatitis are refractory to corticosteroids and develop progressive liver disease and cirrhosis. We describe five patients with autoimmune hepatitis who did not respond to conventional corticosteroids and azathioprine therapy who were then treated with cyclosporine A. Cyclosporine A was started at 2–3 mg/kg/day and induced biochemical remission in four of five patients within 3 months. One of the four responders relapsed within 1 month of discontinuing cyclosporine on two occasions. Each time, liver tests promptly normalized after reinitiation of cyclosporine. Two responders were managed with cyclosporine alone. The single patient who did not respond to cyclosporine developed progressive liver failure, underwent orthotopic liver transplantation, and subsequently died of disseminated cytomegalovirus infection. Cyclosporine was generally well tolerated and none of the patients developed renal insufficiency. These data and review of 11 cases in the literature show that cyclosporine can induce remission of liver disease in patients with autoimmune hepatitis who are refractory to corticosteroids.

Digestive system disease Recent treatment. 3 Interferon resistant chronic hepatitis C and reatment.

Digestive system disease Recent treatment. 3 Interferon resistant chronic hepatitis C and reatment.

Infections in solid organ transplant recipients

Delineation of the pathogenic potential and assessment of the efficacy of newer therapeutic modalities for a number of viruses have been among the more notable developments in solid organ transplant infections within the past year. Infection caused by the novel herpesvirus, human herpesvirus-6, was proposed to be a significant cause of fever of unknown origin and a predictor of subsequent cytomegalovirus infection. Interleukin-2-primed isologous T lymphocytes led to the complete regression of Epstein-Barr virus-positive post-transplant lymphoproliferative disorder. Although interferon-alpha alone was shown to be largely ineffective as prophylaxis and treatment for hepatitis C virus hepatitis, the combination of interferon-alpha and ribavirin appeared promising. Documentation within the past year, however, of lamivudine and famciclovir resistance in hepatitis B virus, ganciclovir resistance in cytomegalovirus, and azole resistance in Candida, is a sobering reminder that antimicrobial prophylaxis must be utilized rationally and selectively.

Infections in solid-organ transplant recipients

Delineation of the pathogenic potential and assessment of the efficacy of newer therapeutic modalities for a number of viruses have been among the more notable developments in solid organ transplant infections within the past year. Infection caused by the novel herpesvirus, human herpesvirus-6, was proposed to be a significant cause of fever of unknown origin and a predictor of subsequent cytomegalovirus infection. Interleukin-2-primed isologous T lymphocytes led to the complete regression of Epstein-Barr virus-positive post-transplant lymphoproliferative disorder. Although interferon-alpha alone was shown to be largely ineffective as prophylaxis and treatment for hepatitis C virus hepatitis, the combination of interferon-alpha and ribavirin appeared promising. Documentation within the past year, however, of lamivudine and famciclovir resistance in hepatitis B virus, ganciclovir resistance in cytomegalovirus, and azole resistance in Candida, is a sobering reminder that antimicrobial prophylaxis must be utilized rationally and selectively.

Ribavirin plus human leucocyte interferon alpha for the treatment of interferon resistant chronic hepatitis C: a controlled trial

A controlled study was carried out on 24 patients with biopsy-proven chronic hepatitis C, who had failed to respond to a previous treatment course with human leukocyte interferon- α (LE-IFNA). The enrolled patients were randomly assigned to receive for 6 months either oral ribavirin, 800 mg daily, plus LE-IFNA, 3 MU thrice a week (Group A: 13 patients), or LE-IFNA alone (Group B: 11 patients). A follow-up period of 6 months was performed in all patients. At enrolment, the two groups showed similar biochemical and histological characteristics. At the end of treatment, normalization of ALT values was found in ten group A patients, five of whom also had undetectable serum HCV-RNA. Only one patient in group B showed such biochemical and virological response. At the end of follow-up, biochemical and virological relapse was registered in all of the Group A patients, whereas the only responding patient in group B still had normal ALT value and undetectable serum HCV-RNA. One group A patient dropped out during the second month of treatment due to development of mild anaemia. In IFN-resistant chronic hepatitis C, the combination therapy of ribavirin and IFN α can induce a complete response during treatment. However, post-treatment relapses are common and long-term benefits seem uncertain.

Failure of ketoprofen and interferon combination therapy to improve interferon-resistant chronic hepatitis C.

Preliminary reports suggest that patients with interferon (IFN)-resistant chronic hepatitis C respond better to a combination of IFN-alpha and nonsteroidal anti-inflammatory drugs than to IFN alone. The efficacy of IFN combined with ketoprofen in the treatment of patients with IFN-resistant chronic hepatitis C was evaluated. Seventeen patients, nonresponsive after at least six months of treatment with IFN-alpha 2b and subsequently treated with the combination of IFN-alpha 2b plus ketoprofen for four months, were studied. Serum aminotransferases (alanine aminotransferase [ALT] and aspartate aminotransferase [AST]) and serum hepatitis C virus (HCV) RNA were analyzed before and throughout treatment. No patient normalized serum aminotransferases after combination therapy. There were no significant differences in mean serum ALT and AST levels before and after ketoprofen intervention. Serum HCV RNA became undetectable after treatment in only one patient, but was detectable again three months after treatment cessation. These results provide no convincing evidence that the combination of IFN-alpha 2b with ketoprofen improves the response to IFN in patients nonresponsive to IFN alone.

Comparison of full-length sequences of interferon-sensitive and resistant hepatitis C virus 1b. Sensitivity to interferon is conferred by amino acid substitutions in the NS5A region.

We have previously demonstrated that sensitivity to interferon is different among hepatitis C virus (HCV) quasispecies simultaneously detected in same individuals and that interferon-resistant HCV quasispecies are selected during the treatment. To determine the genetic basis of their resistance to interferon, HCV genotype-1b was obtained from serum of three patients before and during interferon therapy, and their full-length nucleotide and deduced amino acid sequences were determined. Comparison of the pairs of interferon-resistant and interferon-sensitive HCV isolates in respective individuals demonstrated clusters of amino acid differences in the COOH-terminal half of the NS5A region (codon 2154-2383), which contained a common unique amino acid difference at codon 2218. Additional sequence data of the COOH-terminal half of the NS5A region obtained from six interferon-resistant and nine interferon-sensitive HCV confirmed the exclusive existence of missense mutations in a 40 amino acid stretch of the NS5A region around codon 2218 (from codon 2209 to 2248) in interferon-sensitive HCV. On the other hand, this region of interferon-resistant HCV was identical to that of prototype HCV genotype-1b (HCV-J, HCV-JTa, or HC-J4). We designated this region as the interferon sensitivity determining region. Thus, HCV genotype-1b with the prototype interferon sensitivity determining region appears to be interferon-resistant strains. The specific nature of these mutations might make it possible to predict prognostic effects of interferon treatment.

Mixed cryoglobulinemia may indicate resistance of hepatitis C virus response to interferon

Mixed cryoglobulinemia may indicate resistance of hepatitis C virus response to interferon