Application of a newly developed high-sensitivity HBsAg chemiluminescent enzyme immunoassay for hepatitis B patients with HBsAg seroclearance.

We modified and automated a highly sensitive chemiluminescent enzyme immunoassay (CLEIA) for surface antigen (HBsAg) detection using a combination of monoclonal antibodies, each for a specific epitope of HBsAg, and by improving an earlier conjugation technique. Of 471 hepatitis B virus (HBV) carriers seen in our hospital between 2009 and 2012, 26 were HBsAg seronegative as determined by the Abbott Architect assay. The Lumipulse HBsAg-HQ assay was used to recheck those 26 patients who demonstrated seroclearance by the Abbott Architect assay. The performance of the Lumipulse HBsAg-HQ assay was compared with that of a quantitative HBsAg detection system (Abbott Architect) and the Roche Cobas TaqMan HBV DNA assay (CTM) (lower limit of detection, 2.1 log copies/ml) using blood serum samples from patients who were determined to be HBsAg seronegative by the Abbott Architect assay. Ten patients had spontaneous HBsAg loss. Of 8 patients treated with nucleotide analogues (NAs), two were HBsAg seronegative after stopping lamivudine therapy and 6 were HBsAg seronegative during entecavir therapy. Eight acute hepatitis B (AH) patients became HBsAg seronegative. Of the 26 patients, 16 were HBsAg positive by the Lumipulse HBsAg-HQ assay but negative by the Abbott Architect assay. The differences between the two assays in terms of detectable HBsAg persisted over the long term in the spontaneous loss group (median, 10 months), the NA-treated group (2.5 months), and the AH group (0.5 months). In 9 patients, the Lumipulse HBsAg-HQ assay detected HBsAg when HBV DNA was negative by the CTM assay. HBsAg was also detected by the Lumipulse HBsAg-HQ assay in 4 patients with an anti-HBs concentration of >10 mIU/ml, 3 of whom had no HBsAg escape mutations. The automatic, highly sensitive HBsAg CLEIA Lumipulse HBsAg-HQ is a convenient and precise assay for HBV monitoring.

Hepatitis C virus kinetics by administration of pegylated interferon-α in human and chimeric mice carrying human hepatocytes with variants of the IL28B gene.

OBJECTIVE: Recent studies have demonstrated that genetic polymorphisms near the IL28B gene are associated with the clinical outcome of pegylated interferon α (peg-IFN-α) plus ribavirin therapy for patients with chronic hepatitis C virus (HCV). However, it is unclear whether genetic variations near the IL28B gene influence hepatic interferon (IFN)-stimulated gene (ISG) induction or cellular immune responses, lead to the viral reduction during IFN treatment. DESIGN: Changes in HCV-RNA levels before therapy, at day 1 and weeks 1, 2, 4, 8 and 12 after administering peg-IFN-α plus ribavirin were measured in 54 patients infected with HCV genotype 1. Furthermore, we prepared four lines of chimeric mice having four different lots of human hepatocytes containing various single nucleotide polymorphisms (SNP) around the IL28B gene. HCV infecting chimeric mice were subcutaneously administered with peg-IFN-α for 2 weeks. RESULTS: There were significant differences in the reduction of HCV-RNA levels after peg-IFN-α plus ribavirin therapy based on the IL28B SNP rs8099917 between TT (favourable) and TG/GG (unfavourable) genotypes in patients; the first-phase viral decline slope per day and second-phase slope per week in TT genotype were significantly higher than in TG/GG genotype. On peg-IFN-α administration to chimeric mice, however, no significant difference in the median reduction of HCV-RNA levels and the induction of antiviral ISG was observed between favourable and unfavourable human hepatocyte genotypes. CONCLUSIONS: As chimeric mice have the characteristic of immunodeficiency, the response to peg-IFN-α associated with the variation in IL28B alleles in chronic HCV patients would be composed of the intact immune system.

Age and total ribavirin dose are independent predictors of relapse after interferon therapy in chronic hepatitis C revealed by data mining analysis.

BACKGROUND: This study aimed to define factors associated with relapse among responders to pegylated interferon (PEG-IFN) plus ribavirin (RBV) therapy in chronic hepatitis C. METHODS: A cohort of genotype 1b chronic hepatitis C patients treated with PEG-IFN plus RBV and who had an undetectable HCV RNA by week 12 (n=951) were randomly assigned to model derivation (n=636) or internal validation (n=315) groups. An independent cohort (n=598) were used for an external validation. A decision tree model for relapse was explored using data mining analysis. RESULTS: The data mining analysis defined five subgroups of patients with variable rates of relapse ranging from 13% to 52%. The reproducibility of the model was confirmed by internal and external validations (r(2)=0.79 and 0.83, respectively). Patients with undetectable HCV RNA at week 4 had the lowest risk of relapse (13%), followed by patients <60 years with undetectable HCV RNA at week 5-12 who received ≥3.0 g/kg of body weight of RBV (16%). Older patients with a total RBV dose <3.0 g/kg had the highest risk of relapse (52%). Higher RBV dose beyond 3.0 g/kg was associated with further decrease of relapse rate among patients <60 years (up to 11%) but not among older patients whose relapse rate remained stable around 30%. CONCLUSIONS: Data mining analysis revealed that time to HCV RNA negativity, age and total RBV dose was associated with relapse. To prevent relapse, ≥3.0 g/kg of RBV should be administered. Higher dose of RBV may be beneficial in patients <60 years.

Data mining model using simple and readily available factors could identify patients at high risk for hepatocellular carcinoma in chronic hepatitis C.

BACKGROUND & AIMS: Assessment of the risk of hepatocellular carcinoma (HCC) development is essential for formulating personalized surveillance or antiviral treatment plan for chronic hepatitis C. We aimed to build a simple model for the identification of patients at high risk of developing HCC. METHODS: Chronic hepatitis C patients followed for at least 5 years (n=1003) were analyzed by data mining to build a predictive model for HCC development. The model was externally validated using a cohort of 1072 patients (472 with sustained virological response (SVR) and 600 with nonSVR to PEG-interferon plus ribavirin therapy). RESULTS: On the basis of factors such as age, platelet, albumin, and aspartate aminotransferase, the HCC risk prediction model identified subgroups with high-, intermediate-, and low-risk of HCC with a 5-year HCC development rate of 20.9%, 6.3-7.3%, and 0-1.5%, respectively. The reproducibility of the model was confirmed through external validation (r(2)=0.981). The 10-year HCC development rate was also significantly higher in the high-and intermediate-risk group than in the low-risk group (24.5% vs. 4.8%; p<0.0001). In the high-and intermediate-risk group, the incidence of HCC development was significantly reduced in patients with SVR compared to those with nonSVR (5-year rate, 9.5% vs. 4.5%; p=0.040). CONCLUSIONS: The HCC risk prediction model uses simple and readily available factors and identifies patients at a high risk of HCC development. The model allows physicians to identify patients requiring HCC surveillance and those who benefit from IFN therapy to prevent HCC.

[Quantitative analysis of hepatitis B surface antigen as a clinical marker].

There are 1.5 million hepatitis B virus (HBV) infected patients in Japan. Anti-viral therapy is important for chronic hepatitis B patients to prevent hepatocellular carcinoma. Recently, HBs antigen (HBsAg) quantification has been reported to be useful for not only HBV screening but also for monitoring of anti-viral treatment. In this paper, we evaluated the clinical utility of quantitative assay of HBsAg by HISCL HBsAg kit. Although there can be a significant difference in age, HBeAg positive/negative and viral genotype, there is not in the disease stage. Moreover, the weak correlation was confirmed between HBsAg and HBV-DNA levels with or without anti-virus treatment. In the clinical practice, as HBV-DNA becomes undetectable immediately by anti-viral therapy such as entecavir, it may be difficult to evaluate the efficacy. The monitoring of the HBsAg concentration in addition to HBV-DNA would be useful for the evaluation. Hence, the clinical role of HBsAg concentration could spread widely in Japan.

Relationship between polymorphisms of the inosine triphosphatase gene and anaemia or outcome after treatment with pegylated interferon and ribavirin.

BACKGROUND: A genome-wide association study revealed an association between variants of the inosine triphosphatase (ITPA) gene and ribavirin (RBV)-induced anaemia. The aim of this study was to replicate this finding in an independent Japanese cohort and to define a method to allow pretreatment prediction of anaemia in combination with other factors. METHODS: Genotype 1b chronic hepatitis C patients (n=132) treated with pegylated interferon (PEG-IFN)-α and RBV for 48 weeks were genotyped for ITPA rs1127354 and examined for anaemia and treatment outcome. RESULTS: Variants of the ITPA gene protected against severe anaemia throughout the 48-week treatment period and were associated with lower incidence of anaemia-related RBV dose reduction. A combination of the ITPA genotype with baseline haemoglobin (Hb) and creatinine clearance (CLcr) levels predicted severe anaemia with high accuracy (90% sensitivity and 62% specificity). Among a subset of patients with the IL28B genotype of TT at rs8099917, patients with variants of the ITPA gene were associated with a higher rate of receiving >80% of the expected RBV dose, a higher rate of sustained virological response (SVR), and a lower rate of relapse. CONCLUSIONS: The variants of the ITPA gene, which could protect against haemolytic anaemia and RBV dose reduction, were associated with a high rate of SVR by standard PEG-IFN and RBV therapy in a subset of Japanese patients with the favourable TT genotype at rs8099917 of IL28B. A combination of ITPA genetic polymorphisms with baseline Hb and CLcr levels further improves the predictive accuracy of severe anaemia.

Pretreatment prediction of anemia progression by pegylated interferon alpha-2b plus ribavirin combination therapy in chronic hepatitis C infection: decision-tree analysis.

BACKGROUND: This study aimed to develop a model to predict the development of severe anemia during pegylated interferon alpha-2b plus ribavirin combination therapy. METHODS: Data were collected from 1081 genotype 1b chronic hepatitis C patients who were treated at 6 hospitals in Japan. These patients were randomly assigned to a model-building group (n = 691) or an internal validation group (n = 390). Factors predictive of severe anemia (hemoglobin, Hb < 8.5 g/dl) were explored using data-mining analysis. RESULTS: Hb values at baseline, creatinine clearance (Ccr), and an Hb concentration decline by 2 g/dl at week 2 were used to build a decision-tree model, in which the patients were divided into 5 subgroups based on variable rates of severe anemia ranging from 0.4 to 11.8%. The reproducibility of the model was confirmed by the internal validation group (r² = 0.96). The probability of severe anemia was high in patients whose Hb value was <14 g/dl before treatment (6.5%), especially (a) in those whose Ccr was <80 ml/min (11.8%) and (b) those whose Ccr was ≥ 80 ml/min but whose Hb concentration decline at week 2 was ≥ 2 g/dl (11.5%). The probability of severe anemia was low in the other patients (0.4-2.5%). CONCLUSIONS: The decision-tree model that included Hb values at baseline, Ccr, and an Hb concentration decline by 2 g/dl at week 2 was useful for predicting the probability of severe anemia, and has the potential to support clinical decisions regarding early dose reduction of ribavirin.

Genome-wide association study identified ITPA/DDRGK1 variants reflecting thrombocytopenia in pegylated interferon and ribavirin therapy for chronic hepatitis C.

Hematologic abnormalities during current therapy with pegylated interferon and ribavirin (PEG-IFN/RBV) for chronic hepatitis C (CHC) often necessitate dose reduction and premature withdrawal from therapy. The aim of this study was to identify host factors associated with IFN-induced thrombocytopenia by genome-wide association study (GWAS). In the GWAS stage using 900K single-nucleotide polymorphism (SNP) microarrays, 303 Japanese CHC patients treated with PEG-IFN/RBV therapy were genotyped. One SNP (rs11697186) located on DDRGK1 gene on chromosome 20 showed strong associations in the minor-allele-dominant model with the decrease of platelet counts in response to PEG-IFN/RBV therapy [P = 8.17 × 10(-9); odds ratio (OR) = 4.6]. These associations were replicated in another sample set (n = 391) and the combined P-values reached 5.29 × 10(-17) (OR = 4.5). Fine mapping with 22 SNPs around DDRGK1 and ITPA genes showed that rs11697186 at the GWAS stage had a strong linkage disequilibrium with rs1127354, known as a functional variant in the ITPA gene. The ITPA-AA/CA genotype was independently associated with a higher degree of reduction in platelet counts at week 4 (P < 0.0001), as well as protection against the reduction in hemoglobin, whereas the CC genotype had significantly less reduction in the mean platelet counts compared with the AA/CA genotype (P < 0.0001 for weeks 2, 4, 8, 12), due to a reactive increase of the platelet count through weeks 1-4. Our present results may provide a valuable pharmacogenetic diagnostic tool for tailoring PEG-IFN/RBV dosing to minimize drug-induced adverse events.

Recommendation of lamivudine-to-entecavir switching treatment in chronic hepatitis B responders: Randomized controlled trial.

AIM: In the 2007-2008 guidelines of the study group (Ministry of Health, Labor and Welfare of Japan), lamivudine (LAM)-continuous treatment was recommended in patients treated with LAM for more than 3 years who maintained hepatitis B virus (HBV) DNA less than 2.6 log copies/mL, because in these patients LAM resistance might exist and switching treatment to entecavir (ETV) might cause ETV resistance. However, there was no evidence on whether switching treatment to ETV- or LAM-continuous treatment was better in those patients. In the present study, we performed a randomized controlled trial of LAM-to-ETV switching treatment. METHODS: Twenty-seven patients treated with LAM for more than 3 years whose HBV DNA levels were less than 2.6 log copies/mL were enrolled and randomly divided into two groups, LAM-continued group or switching to ETV group. Then, we examined incidence of virological breakthrough (VBT) and breakthrough hepatitis (BTH) in each group. RESULTS: There was no BTH in any of the patients. VBT was observed in six patients of the LAM group (6/15, 40%), and no patient of the ETV group (0/11, 0%) (P = 0.02). The differences of the proportion of cumulated VBT using a log-rank test with Kaplan-Meier analysis were significant between the LAM and ETV groups (P = 0.025). CONCLUSION: In patients treated with LAM for more than 3 years maintaining HBV DNA less than 2.6 log copies/mL, switching treatment to ETV is recommended at least during the 2 years' follow-up period.

Transmissibility of hepatitis B virus (HBV) infection through blood transfusion from blood donors with occult HBV infection.

BACKGROUND: Studies of the transmissibility of hepatitis B virus (HBV) in occult hepatitis B (OHB) through blood transfusion are scarce. We aimed to determine the transmissibility of HBV in blood donors with OHB through transfusion in animal and human studies. METHODS: Among 217,595 blood donors, 67 donors with OHB were identified. Four chimeric mice populated with human hepatocytes were inoculated with 2 donor serum samples. Serial serum and liver HBV DNA levels were measured. Forty-nine recipients of blood transfusions traced from 10 donors with OHB (9 of whom were positive for antibody to hepatitis B surface antigen [anti-HBs]) were tested for HBV infection. Homology and phylogenetic analyses between the HBV genomic sequences of donors and recipients were performed. RESULTS: Serum HBV DNA was detectable (10(4) copies/mL) in 1 mouse at weeks 5 and 7 after inoculation. Total HBV DNA and HBV replication template (covalently closed circular DNA) and hepatitis B core antigen were detected in the mouse liver. After transfusion, 45 recipients (91.8%) had no HBV infection (ie, they tested negative for hepatitis B surface antigen and HBV DNA). Four tested positive for HBV DNA. In 3 recipients, 83%-86% homology and distant phylogenetic relatedness with their donor HBV excluded transmission through transfusion. The remaining recipient HBV had 95% sequence homology with her donor HBV, compatible with acquisition of HBV infection from the transfusion. High anti-HBs levels in 7 other recipients suggested recent transfusion-related HBV immune response. CONCLUSIONS: OHB donor blood infectivity was shown in our animal and human studies. However, the risk of HBV transmission in humans was low, especially from blood products obtained from donors with OHB who were anti-HBs positive.

Virological and clinical characteristics on reactivation of occult hepatitis B in patients with hematological malignancy.

The virological characteristics of hepatitis B virus (HBV) implicated in the reactivation of occult hepatitis B in patients who have received hematopoietic stem-cell transplantation or chemotherapy for the hematological malignancy are not well defined. Twenty-eight HBsAg-negative patients who received hematopoietic stem-cell transplantation and 138 HBsAg-negative patients treated for malignant lymphoma with chemotherapy including rituximab were enrolled. Three of the 28 patients (10.7%) received hematopoietic stem-cell transplantation and one of the 138 (0.72%) patients treated for malignant lymphoma with chemotherapy developed de novo HBV hepatitis. Anti-HBc was detected in four and anti-HBs in two patients. Genotype Bj was detected in two and C in two of they all possessed wild-type sequences in the core promoter region. A precore stop mutation (A1896) was detected in a patient with genotype Bj who developed fulminant hepatic failure. HBV DNA was detected in pretreatment HBsAg-negative samples in two of four patients, and the HBV genome sequence identified from sera before chemotherapy and at the time of de novo HBV hepatitis showed 100% homology. In an in vitro replication model, genotype Bj with the A1896 clone obtained from a fulminant case had a replication level much higher than clones obtained from de novo hepatitis B patients with genotype Bj or C with G1896. In conclusion, this is the first report demonstrating de novo hepatitis B from the reactivation of occult HBV infection confirmed by molecular evolutional analysis. The fulminant outcome of HBV reactivation can be associated with genotype Bj exhibiting high replication due to the A1896 mutation.

Sequences in the interferon sensitivity-determining region and core region of hepatitis C virus impact pretreatment prediction of response to PEG-interferon plus ribavirin: data mining analysis.

The aim of the present study was to clarify the significance of viral factors for pretreatment prediction of sustained virological response to pegylated-interferon (PEG-IFN) plus ribavirin (RBV) therapy for chronic hepatitis C using data mining analysis. Substitutions in the IFN sensitivity-determining region (ISDR) and at position 70 of the HCV core region (Core70) were determined in 505 patients with genotype 1b chronic hepatitis C treated with PEG-IFN plus RBV. Data mining analysis was used to build a predictive model of sustained virological response in patients selected randomly (n = 304). The reproducibility of the model was validated in the remaining 201 patients. Substitutions in ISDR (odds ratio = 9.92, P < 0.0001) and Core70 (odds ratio = 1.92, P = 0.01) predicted sustained virological response independent of other covariates. The decision-tree model revealed that the rate of sustained virological response was highest (83%) in patients with two or more substitutions in ISDR. The overall rate of sustained virological response was 44% in patients with a low number of substitutions in ISDR (0-1) but was 83% in selected subgroups of younger patients (<60 years), wild-type sequence at Core70, and higher level of low-density lipoprotein cholesterol (LDL-C) (≥ 120 mg/dl). Reproducibility of the model was validated (r(2) = 0.94, P < 0.001). In conclusion, substitutions in ISDR and Core70 of HCV are significant predictors of response to PEG-IFN plus RBV therapy. A decision-tree model that includes these viral factors as predictors could identify patients with a high probability of sustained virological response.

Pretreatment prediction of response to peginterferon plus ribavirin therapy in genotype 1 chronic hepatitis C using data mining analysis.

BACKGROUND: This study aimed to develop a model for the pre-treatment prediction of sustained virological response (SVR) to peg-interferon plus ribavirin therapy in chronic hepatitis C. METHODS: Data from 800 genotype 1b chronic hepatitis C patients with high viral load (>100,000 IU/ml) treated by peg-interferon plus ribavirin at 6 hospitals in Japan were randomly assigned to a model building (n = 506) or an internal validation (n = 294). Data from 524 patients treated at 29 hospitals in Japan were used for an external validation. Factors predictive of SVR were explored using data mining analysis. RESULTS: Age (<50 years), alpha-fetoprotein (AFP) (<8 ng/mL), platelet count (≥ 120 × 10(9)/l), gamma-glutamyltransferase (GGT) (<40 IU/l), and male gender were used to build the decision tree model, which divided patients into 7 subgroups with variable rates of SVR ranging from 22 to 77%. The reproducibility of the model was confirmed by the internal and external validation (r (2) = 0.92 and 0.93, respectively). When reconstructed into 3 groups, the rate of SVR was 75% for the high probability group, 44% for the intermediate probability group and 23% for the low probability group. Poor adherence to drugs lowered the rate of SVR in the low probability group, but not in the high probability group. CONCLUSIONS: A decision tree model that includes age, gender, AFP, platelet counts, and GGT is useful for predicting the probability of response to therapy with peg-interferon plus ribavirin and has the potential to support clinical decisions regarding the selection of patients for therapy.

Pre-treatment prediction of response to pegylated-interferon plus ribavirin for chronic hepatitis C using genetic polymorphism in IL28B and viral factors.

BACKGROUND & AIMS: Pegylated interferon and ribavirin (PEG-IFN/RBV) therapy for chronic hepatitis C virus (HCV) genotype 1 infection is effective in 50% of patients. Recent studies revealed an association between the IL28B genotype and treatment response. We aimed to develop a model for the pre-treatment prediction of response using host and viral factors. METHODS: Data were collected from 496 patients with HCV genotype 1 treated with PEG-IFN/RBV at five hospitals and universities in Japan. IL28B genotype and mutations in the core and IFN sensitivity determining region (ISDR) of HCV were analyzed to predict response to therapy. The decision model was generated by data mining analysis. RESULTS: The IL28B polymorphism correlated with early virological response and predicted null virological response (NVR) (odds ratio=20.83, p<0.0001) and sustained virological response (SVR) (odds ratio=7.41, p<0.0001) independent of other covariates. Mutations in the ISDR predicted relapse and SVR independent of IL28B. The decision model revealed that patients with the minor IL28B allele and low platelet counts had the highest NVR (84%) and lowest SVR (7%), whereas those with the major IL28B allele and mutations in the ISDR or high platelet counts had the lowest NVR (0-17%) and highest SVR (61-90%). The model had high reproducibility and predicted SVR with 78% specificity and 70% sensitivity. CONCLUSIONS: The IL28B polymorphism and mutations in the ISDR of HCV were significant pre-treatment predictors of response to PEG-IFN/RBV. The decision model, including these host and viral factors may support selection of optimum treatment strategy for individual patients.

Performance of two Real-Time RT-PCR assays for quantitation of hepatitis C virus RNA: evaluation on HCV genotypes 1-4.

Accuracy for monitoring of the concentration of hepatitis C virus (HCV) RNA represents a major challenge throughout the management of patients with chronic hepatitis C. To investigate the genotype-independent efficiency and the accuracy of two real-time detection reverse transcription-polymerase chain reaction (RT-PCR) assays; the Cobas Ampliprep/Cobas TaqMan (CAP/CTM); and the Abbott RealTime HCV (ART), a total of 184 samples with different HCV subtypes were examined; 1b (n=58), 2a (n=39), 2b (n=26), 3a (n=20), and 4 (n=41). A robust linear correlation was observed between the two assays applied to genotypes 1b, 2a, 2b, and 3a [the correlation coefficient (R) ranged from 0.99 to 0.98], but not to genotype 4 specimens (R=0.78). A significant difference in measurements of HCV RNA using CAP/CTM and ART in serum samples with genotypes 1b and 4 was observed (0.72, -0.53 log IU/ml, P<0.0001, 0.01, respectively). A robust correlation was observed between the HCV core antigen and HCV RNA values by either of the HCV RNA quantitation assays applied to all genotypes with exception of genotype 4, for which R was higher with ART (R=0.95) than with CAP/CTM (R=0.80). The lower limit of detection of CAP/CTM and ART were 41.4 and 28.5 IU/ml using the WHO standards, respectively. In conclusion, two RT-PCR assays had a high efficiency and accuracy for quantitation of HCV RNA of genotypes 2a, 2b, and 3a, but the mean values of HCV RNA differed for genotype 1b and 4.

Reactivation of hepatitis B virus in HBsAg-negative patients with multiple myeloma: two case reports.

It was recently reported that hepatitis B virus (HBV) reactivation had occurred in HBsAg-negative lymphoma patients who received rituximab plus steroid combination chemotherapy. HBV reactivation in myeloma patients have not been reported extensively. We describe here two cases of HBV reactivation in HBsAg-negative myeloma patients receiving systemic chemotherapy: one from the medical records of 40 patients and another from 61 patients with prospective HBV-DNA monitoring. In the first case positive for anti-HBs, HBV reactivation was diagnosed when hepatitis developed during conventional chemotherapy such as MP and MCP regimen in a relapsed patient after autologous stem cell transplantation (APBSCT); in the second case positive for anti-HBc and anti-HBs, elevation of HBV-DNA was recognized by serial HBV-DNA monitoring performed prospectively following APBSCT. Interestingly, these two cases had the reduction of the titer of anti-HBs during the treatment, followed by HBV reactivation. These clinical data suggest that the HBV-DNA monitoring is necessary for not only HBsAg-positive but also HBsAg-negative myeloma patients with anti-HBc-positive and/or anti-HBs-positive following transplantation and after conventional chemotherapy in the salvage setting. Establishment of a standard strategy to prevent HBV reactivation is important for myeloma patients receiving systemic chemotherapy.

Positive selection of core 70Q variant genotype 1b hepatitis C virus strains induced by pegylated interferon and ribavirin.

BACKGROUND: Approximately 20% of patients with hepatitis C virus (HCV) genotype 1b infection have nonresponse to the most current treatment, pegylated interferon with ribavirin. Mutations in the HCV core region were recently proposed to be associated with nonresponse. Our aim was to evaluate the viral factors associated with treatment failure. METHODS: HCV variants were determined directly and after cloning in 66 HCV-1b-infected Japanese patients and in 5 urokinase-type plasminogen activator transgenic severe combined immunodeficiency mice with human hepatocytes (chimeric mice), at baseline, during treatment, and after treatment. RESULTS: At baseline, glutamine at position 70 of the HCV core protein (70Q) was detected by direct sequencing in 20% of patients with virologic response and in 43.8% of patients with nonresponse. Among patients with nonresponse, who were examined during and after treatment, the prevalence of the 70Q substitution increased to 56.3%, which indicates that treatment-induced selection occurred in all patients with nonresponse who had 70Q quasispecies detectable by cloning. This observation was reinforced by the results from experimentally infected chimeric mice. Logistic regression analysis indicated that detection of 70Q quasispecies was associated with a statistically significantly increased risk of nonresponse (odds ratio, 15.1; P = .004) in the studied cohort. CONCLUSION: Presence of the 70Q quasispecies at baseline was associated with an increased risk of treatment failure, as indicated by the positive selection of the 70Q clones induced by treatment with pegylated interferon with ribavirin. These results urge further investigation of the mechanisms of this association.

Genome-wide association of IL28B with response to pegylated interferon-alpha and ribavirin therapy for chronic hepatitis C.

The recommended treatment for patients with chronic hepatitis C, pegylated interferon-alpha (PEG-IFN-alpha) plus ribavirin (RBV), does not provide sustained virologic response (SVR) in all patients. We report a genome-wide association study (GWAS) to null virological response (NVR) in the treatment of patients with hepatitis C virus (HCV) genotype 1 within a Japanese population. We found two SNPs near the gene IL28B on chromosome 19 to be strongly associated with NVR (rs12980275, P = 1.93 x 10(-13), and rs8099917, 3.11 x 10(-15)). We replicated these associations in an independent cohort (combined P values, 2.84 x 10(-27) (OR = 17.7; 95% CI = 10.0-31.3) and 2.68 x 10(-32) (OR = 27.1; 95% CI = 14.6-50.3), respectively). Compared to NVR, these SNPs were also associated with SVR (rs12980275, P = 3.99 x 10(-24), and rs8099917, P = 1.11 x 10(-27)). In further fine mapping of the region, seven SNPs (rs8105790, rs11881222, rs8103142, rs28416813, rs4803219, rs8099917 and rs7248668) located in the IL28B region showed the most significant associations (P = 5.52 x 10(-28)-2.68 x 10(-32); OR = 22.3-27.1). Real-time quantitative PCR assays in peripheral blood mononuclear cells showed lower IL28B expression levels in individuals carrying the minor alleles (P = 0.015).

A genetic variant of hepatitis B virus divergent from known human and ape genotypes isolated from a Japanese patient and provisionally assigned to new genotype J.

Hepatitis B virus (HBV) of a novel genotype (J) was recovered from an 88-year-old Japanese patient with hepatocellular carcinoma who had a history of residing in Borneo during the World War II. It was divergent from eight human (A to H) and four ape (chimpanzee, gorilla, gibbon, and orangutan) HBV genotypes, as well as from a recently proposed ninth human genotype I, by 9.9 to 16.5% of the entire genomic sequence and did not have evidence of recombination with any of the nine human genotypes and four nonhuman genotypes. Based on a comparison of the entire nucleotide sequence against 1,440 HBV isolates reported, HBV/J was nearest to the gibbon and orangutan genotypes (mean divergences of 10.9 and 10.7%, respectively). Based on a comparison of four open reading frames, HBV/J was closer to gibbon/orangutan genotypes than to human genotypes in the P and large S genes and closest to Australian aboriginal strains (HBV/C4) and orangutan-derived strains in the S gene, whereas it was closer to human than ape genotypes in the C gene. HBV/J shared a deletion of 33 nucleotides at the start of preS1 region with C4 and gibbon genotypes, had an S-gene sequence similar to that of C4, and expressed the ayw subtype. Efficient infection, replication, and antigen expression by HBV/J were experimentally established in two chimeric mice with the liver repopulated for human hepatocytes. The HBV DNA sequence recovered from infected mice was identical to that in the inoculum. Since HBV/J is positioned phylogenetically in between human and ape genotypes, it may help to trace the origin of HBV and merits further epidemiological surveys.

Case-control study for the identification of virological factors associated with fulminant hepatitis B.

BACKGROUND: Host and viral factors can promote the development of fulminant hepatitis B (FHB), but there have been no case-control studies for figuring out virological parameters that can distinguish FHB. METHODS: In a case-control study, virological factors associated with the development of FHB were sought in 50 patients with FH developed by transient hepatitis B virus (HBV) infection (FH-T) and 50 with acute self-limited hepatitis B (AHB) who were matched for sex and age. In addition, 12 patients with FH developed by acute exacerbation (AE) of asymptomatic HBV carrier (ASC) (FH-C) were also compared with 12 patients without FH by AE of chronic hepatitis B (AE-C). RESULTS: Higher HBV DNA levels, subgenotype B1/Bj, A1762T/G1764A, G1896A, G1899A and A2339G mutation were significantly more frequent (P < 0.05), while hepatitis B e-antigen was less frequent in the FH-T patients than AHB. In multivariate analysis, G1896A mutation (odds ratio [OR], 13.53; 95% confidence interval [CI], 2.75-66.64), serum HBV DNA more than 5.23 log copies/mL (OR, 5.14; 95% CI, 1.10-24.15) and total bilirubin more than 10.35 mg/mL (OR, 7.81; 95% CI, 1.77-34.51) were independently associated with a fulminant outcome by transient HBV infection. On the other hand, in comparison with the patients between FH-C and AE-C groups, there was no significant difference of virological factors associated with the development of FHB. CONCLUSION: A number of virological factors have been defined that may distinguish FH-T from AHB in a case-control study. The pathogenic mechanism of FHB between transient HBV infection and AE of ASC would be different.