A molecular standard for circulating HBV RNA detection and quantification assays in patients with chronic hepatitis B.

Background & Aims: Circulating HBV RNAs have been proposed as a biomarker that reflects the transcriptional activity of covalently closed circular DNA (cccDNA) and may help to evaluate HBV treatment activity. Different research assays have been proposed and, although two PCR-based research use only investigational assays have been developed, the lack of standardized protocols represents an important limitation. Here we have designed and generated a stable clonal cell line producing an RNA-based standard for the calibration of PCR-based circulating HBV RNA assays.

Quantification of circulating HBV RNA expressed from intrahepatic cccDNA in untreated and NUC treated patients with chronic hepatitis B.

Objective: A convenient, reproducible biomarker of hepatitis B virus (HBV) covalently closed circular DNA (cccDNA) transcriptional activity is lacking. We measured circulating HBV RNA (cirB-RNA) in untreated and nucleos(t)ide analogues (NUC) treated chronic hepatitis B (CHB) patients to define its correlation with intrahepatic viral markers and HBV core-related antigen (HBcrAg).

Design: Paired liver biopsy and serum samples were collected from 122 untreated and 30 NUC-treated CHB patients.

Predictors of hepatitis B surface antigen loss, relapse and retreatment after discontinuation of effective oral antiviral therapy in noncirrhotic HBeAg-negative chronic hepatitis B.

Reliable predictors of outcomes after treatment discontinuation in HBeAg-negative chronic hepatitis B (CHB) patients have not been established. We investigated the role of hepatitis B surface antigen (HBsAg), interferon-inducible protein-10 (IP10) and hepatitis B core-related antigen (HBcrAg) serum levels as predictors of HBsAg loss, relapse and retreatment in noncirrhotic HBeAg-negative CHB patients who discontinued long-term antiviral therapy. All HBsAg-positive (n = 57) patients of the prospective DARING-B study were included and followed monthly for 3 months, every 2/3 months until month-12 and every 3/6 months thereafter.

Serum hepatitis B core-related antigen (HBcrAg) correlates with covalently closed circular DNA transcriptional activity in chronic hepatitis B patients.

Background & Aims: It has been proposed that serum hepatitis B core-related antigen (HBcrAg) reflects intrahepatic covalently closed circular (ccc)DNA levels. However, the correlation of HBcrAg with serum and intrahepatic viral markers and liver histology has not been comprehensively investigated in a large sample. We aimed to determine if HBcrAg could be a useful therapeutic marker in patients with chronic hepatitis B.

Inhibition of hepatitis B viral entry by nucleic acid polymers in HepaRG cells and primary human hepatocytes.

Hepatitis B virus (HBV) infection remains a major public health concern worldwide with 240 million individuals chronically infected and at risk of developing cirrhosis and hepatocellular carcinoma. Current treatments rarely cure chronic hepatitis B infection, highlighting the need for new anti-HBV drugs. Nucleic acid polymers (NAPs) are phosphorothioated oligonucleotides that have demonstrated a great potential to inhibit infection with several viruses.

Intrahepatic innate immune response pathways are downregulated in untreated chronic hepatitis B.

Background & Aims: Hepatitis B virus (HBV) persistence and the pathobiology of chronic HBV (CHB) infections result from the interplay between viral replication and host immune responses. We aimed to comprehensively analyse the expression of intrahepatic host genes as well as serum and liver HBV markers in a large cohort of untreated CHB patients.

Methods: One-hundred and five CHB patients untreated at the time of liver biopsy (34 HBeAg[+] and 71 HBeAg[-]) were analysed for the intrahepatic expression profile of 67 genes belonging to multiple innate immunity pathways.

Association of anti-E1E2 antibodies with spontaneous recovery or sustained viral response to therapy in patients infected with hepatitis C virus.

Unlabelled: The monoclonal antibody (mAb) D32.10 recognizes a discontinuous epitope encompassing three regions E1 (amino acids 297-306), E2A (amino acids 480-494), and E2B (amino acids 613-621) juxtaposed on the surface of serum-derived hepatitis C virus (HCV) particles (HCVsp). The mAb D32.

HCV infection in northeastern Brazil: unexpected high prevalence of genotype 3a and absence of African genotypes.

The genomic diversity of HCV embraces 6 genotypes and at least 52 subtypes with clinical and epidemiological correlations. There is a paucity of studies assessing HCV genotypes and biomolecular epidemiology in Brazil. We studied genotype distribution and epidemiological aspects in 232 HCV carriers, 133 (57.

Early detection of viral resistance by determination of hepatitis B virus polymerase mutations in patients treated by lamivudine for chronic hepatitis B.

We have analyzed the molecular dynamics of emergence of drug-resistant strains in patients receiving lamivudine therapy for chronic hepatitis B. Twenty consecutive patients with lamivudine resistance were studied (13 hepatitis B e antigen [HBeAg]-positive patients and 7 HBe antibody [anti-HBe]-positive patients). Determination of viral genotype, precore mutants, and polymerase gene mutants (L528M, M552V, M552I) was performed using the research version of Lipa-HBV.

Hepatitis C virus genotypes in a northeastern area of Brazil.

We used a reverse transcription-polymerase chain reaction (RT-PCR) to obtain the genotypes of circulating hepatitis C virus (HCV) in patients from a Gastro-Hepatology Unit in the city of Salvador (Bahia State) in northeastern Brazil. Viral RNA was detected in 83 (65.4%) of 127 anti-HCV seropositive serum samples.

Persistence of viral replication after anti-HBe seroconversion during antiviral therapy for chronic hepatitis B.

Background/aims: Hepatitis B virus genome mutants may be selected during the immune-mediated clearance of infection or during long-term nucleoside analog administration and may escape both antiviral pressures. The pattern of anti-HBe seroconversion was analyzed in patients receiving new nucleoside analogs, lamivudine or famciclovir, in comparison with patients treated with interferon alpha.

Methods: Eighteen consecutive patients who seroconverted to anti-HBe were included in the study.

In vivo tropism of hepatitis C virus genomic sequences in hematopoietic cells: influence of viral load, viral genotype, and cell phenotype.

Extrahepatic sites capable of supporting hepatitis C virus (HCV) replication have been suggested. We analyzed the influence of virological factors such as viral genotype and viral load, and cellular factors such as cell phenotype, on the detection rate of HCV sequences in hematopoietic cells of infected patients. Thirty-eight chronically infected patients were included in the study: 19 infected by genotype 1 isolates (1a and 1b), 13 by nongenotype 1 isolates (including genotypes 2 a/c, 3a, and 4), and 6 coinfected by genotype 1 and 6 isolates.

Specific detection of hepatitis C virus minus strand RNA in hematopoietic cells.

The presence of hepatitis C virus (HCV) negative strand RNA in extrahepatic compartments based on PCR detection assays has been suggested in many reports with a very heterologous detection rate (from 0 to 100%). In this study, we have analyzed the presence of HCV negative strand in hepatic (liver biopsies, n = 20) and extrahepatic (sera, n = 32; PBMC, n = 26 and fresh bone marrow cells, n = 8) compartments from infected patients with three different reverse transcriptase (RT)-PCR-based assays using primers located in the 5' noncoding region, with or without a tag selected to display different viral loads (10(5)-3 x 10(7) genomic equivalent/ml or gram) and viral genotypes (n = 5). Using synthetic as well as biological templates, we could document extensive artifactual detection of negative strand RNA, due to self priming and mispriming events, even either 5' noncoding region primer pair was used, whereas both artifacts were dramatically reduced (mispriming) or eliminated (selfpriming) using CAP-based RT-PCR assay.

[Treatment of hepatitis C].

The treatment of hepatitis C virus (HCV) infections is essentially known for chronic hepatitis C and is mainly restricted to interferon alpha. Initial trials have indicated that around 50% of the patients with chronic hepatitis C respond to alpha interferon (administered at 3 MU, thrice weekly, during 6 months) by normalizing alanine aminotransferase at the end of therapy, although 25% were found to relapse after therapy. Normalization of biochemical tests is associated with an improvement in liver histological features and with decrease or loss of HCV from serum and liver.

Interferon therapy for hepatitis C.

Initial trials indicated that around 50% of patients respond to recombinant alpha interferon by normalizing alanine aminotransferase (ALT) at the end of therapy and that half of these relapsed within 6 months following cessation of treatment. Both dose and duration of treatment are critical in the response to therapy. Higher doses and longer duration have been suggested to be more effective than the current recommendations of 3 MUI thrice weekly for 6 months based on results of these initial studies which used ALT and histological scores to evaluate the efficacy of interferon therapy.

Hepatitis C virus genotypes in France: comparison of clinical features of patients infected with HCV type I and type II.

Two major hepatitis C virus genotypes, F1 and F2, corresponding to hepatitis C virus type I and type II respectively, were found in France. To investigate the correlation between infection with these genotypes (F1 and F2) and clinical features of patients, serum samples proven to be hepatitis C virus positive by polymerase chain reaction amplification on 5' non-coding region were further amplified in the NS3 region with nested polymerase chain reaction. The NS3-polymerase chain reaction products were Southern blotted and hybridized with specific probes to identify the genotype of hepatitis C virus.

Heterogeneity of hepatitis C virus genotypes in France.

The genotypes of French hepatitis C virus (HCV) isolates were investigated by amplification of a domain from the non-structural region 3 (NS3) using nested PCR, followed by hybridization with two genotype-specific probes, F1 (HCV type I-specific) and F2 (HCV type II-specific). Among 119 HCV RNA-positive sera, 91% of samples were NS3 PCR positive. Most samples (83.

Serological responses to different genotypes of hepatitis C virus in France.

The relationship between hepatitis C virus (HCV) genotypes and antibody status was studied in 104 chronic non-A, non-B hepatitis patients and asymptomatic HCV-infected blood donors. On the basis of amplification of the nonstructural protein 3 (NS3) coding region by PCR and hybridization with specific probes, 55 and 42 patients were identified as being infected with type I and type II, respectively, according to the classification by H. Okamoto, K.

[Importance of PCR in the diagnosis of hepatitis C].

The identification of hepatitis C virus, based on DNA amplification, gives a precise estimation of the prevalence of the most frequent agent of NANB hepatitis. The first ELISA allowing the detection of anti-HCV antibodies, had too many false positive results and required the development of more sensitive and specific assays to confirm its results. PCR, allowing the hepatitis C virus diagnosis by showing directly HCV RNA sequences, offers a complementary approach to immunoserological tests.