Estimation of the future numbers of patients with circulatory diseases in Japan based on the results of national patient surveys]

Virological response to treatment of chronic hepatitis B is defined as the loss of serum hepatitis B virus DNA (HBV DNA) and hepatitis B e antigen (HBeAg). The quantitative measurement of HBV DNA is useful for monitoring and predicting the response to therapy with interferon-alpha (IFN-alpha). In this study, we evaluated whether quantitative measurement of serum HBeAg and IgM antibody to hepatitis B core antigen (HBcAb) could also be used in this manner. Using a microparticle-capture enzyme immunoassay (IMx), a standard curve of fluorescence rate vs HBeAg concentration was constructed to provide quantitative results. The IgM HBcAb index was also measured using a microparticle enzyme immunoassay and serum HBV DNA was measured by a solution hybridization assay. We studied 48 patients who were initially positive for HBeAg and HBV DNA and who were treated with IFN-alpha2b. Their sera were serially evaluated for HBeAg concentration, and results were compared with HBV DNA levels. In the 14 patients who responded to IFN, similar disappearance curves were observed with good intraindividual correlation between the levels of the two markers. In the 34 non-responders, HBeAg levels decreased during treatment but never became negative; HBV DNA levels also decreased during treatment and became transiently undetectable in six patients, falsely suggesting treatment success. The IgM HBcAb index paralleled changes in alanine aminotransferase (ALT) concentration and did not provide additional information. Multiple logistic regression indicated that baseline ALT and HBeAg concentrations were independent predictors of the response to treatment and the addition of neither HBV DNA nor IgM HBcAb improved the model. We conclude that quantitative measurement of HBeAg provides information similar to that of HBV DNA in monitoring and predicting the response to treatment; this technique could be readily adaptable to clinical laboratories.     

Use of quantitative assays for hepatitis B e antigen and IgM antibody to hepatitis B core antigen to monitor therapy in chronic hepatitis B

OBJECTIVES: We evaluated the clinical utility of IgM antibody to the hepatitis B (HB) core antigen (anti-HBc) and HB e antigen (HBeAg) serum levels in patients with chronic HB receiving interferon alfa. METHODS: Stored serum from 47 patients with chronic HB participating in a controlled trial of interferon alfa therapy (10 million U three times a week for 16 wk) were analyzed. All were seropositive for HB surface Ag, HBeAg, and HB virus (HBV) DNA before entry. IgM anti-HBc index values and HBeAg standard values were determined by automated microparticle enzyme immunoassay on samples drawn just before therapy and 6 months later. Ten normal subjects were tested as controls. IgM anti-HBc and HBeAg levels were compared to initial serum HBV DNA, DNA polymerase, serum aminotransferase levels, and demographic features. Serial IgM anti-HBc levels were also obtained during and after therapy in 10 responders and five nonresponders, and serial HBeAg levels were also obtained during and after therapy in four responders and four nonresponders. RESULTS: Neither IgM anti-HBc nor HBeAg levels correlated significantly with values for serum HBV DNA, DNA polymerase, aminotransferases, or demographic features. The initial mean IgM anti-HBc level among the 15 responders to therapy (loss of HBeAg and HBV DNA from serum) was no different from that in nonresponders (mean 1.15 vs 1.27, p = not significant). However, the initial mean HBeAg level was significantly lower in responders than in nonresponders (749.4 vs 1356.4, p = 0.019). Among 10 responders, IgM anti-HBc levels decreased progressively over time, so that at latest follow-up (1.5-4 yr later, mean 2.6 yr), the mean had decreased from 1.325 to 0.312 (p = < 0.001). Among five nonresponders, the mean did not change significantly over 1.5-3 yr (mean 2.2 yr) (1.26 vs 1.08, p = not significant). HBeAg values fell in parallel with HBV DNA and DNA polymerase values in four responders tested but remained elevated in four nonresponders. CONCLUSIONS: HBeAg levels, but not IgM anti-HBc levels, are useful in predicting response to interferon alfa, with responders tending to have lower pretreatment HBeAg levels than nonresponders. HBeAg levels may be used to monitor response to interferon alfa in patients with chronic HB.     

Evaluation of pedicle screw insertion monitored by intraoperative evoked electromyography

The clinical importance of hepatitis B virus (HBV) genome variability has been reported recently. One example is the occurrence of hepatitis B virus pre-core mutants, which arise during spontaneous or interferon-induced seroconversion from HBeAg to anti-HBe and are thought to be selected by immune pressure. A survey of HBV pre-core mutants and viral genotypes in 35 HBeAg negative patients during interferon therapy was carried out to understand viral pathogenesis in this form of chronic hepatitis B. Seventeen patients responded to interferon therapy as assessed by the sustained normalization of serum ALT levels and the significant decrease of viremia levels. The response rate to interferon was independent of both initial serum viral DNA level and interferon doses. During interferon therapy, a significant decrease of M0 (wild-type pre-core sequence at pos. 1887-1908), M1 (TGG to TAG at pos. 1896) or M2 (TGG to TAG at pos. 1896, and GGC to GAC at pos. 1899) positive viral genomes was found in 48%, 42%, and 33% of patients, respectively. A higher response rate to interferon therapy was observed in patients infected with HBV genotype A (70%) or M0 positive strains (75%) as compared to patients infected with genotype D/E (40%) or M1/M2 positive strains (44%). The data support the hypothesis that pre-core defective HBV represent viral mutants with an increased capacity to resist exogenous alpha interferon. These findings emphasize that characterization of HBV genome variability prior to interferon therapy may help to predict antiviral response in HBeAg negative patients.     

Treatment of chronic viral hepatitis

Recent advances have been made in the treatment of chronic viral hepatitis, mainly with recombinant interferon (IFN) alpha. However, the present treatment of chronic viral hepatitis is not entirely satisfactory because the efficacy is inconstant and/or incomplete. In chronic hepatitis B IFN-alpha induces a sustained interruption of hepatitis B virus (HBV) replication, with a HBeAg to anti-HBe seroconversion in about 30% of patients. Patients most likely to respond are those with no immunosuppression, HBV infection acquired during adulthood or active liver disease with low HBV replication. Responders usually show a significant decrease in serum HBV DNA levels during the first 2 months of therapy, followed by a significant increase in the level of aminotransferases. New nucleoside analogues might be useful in combination with IFN-alpha in the treatment of those who do not respond to IFN therapy. In chronic hepatitis B-D, the rate of sustained response to IFN-alpha therapy is low. To be effective, IFN-alpha must be used at a high dosage (9-10 mega units) with a long duration (1 year). In chronic hepatitis C, IFN-alpha at a dosage of 3 mega units over 6 months, induces a sustained response in about 20% of patients. A higher dosage of IFN (5-10 mega units) and a longer duration of treatment increases the rate of sustained response but is associated with poor tolerance. Non-responders to a first course of IFN do not respond to a second course of treatment. In patients who respond but relapse after treatment, the rate of sustained response after a second course of IFN needs to be assessed. Ribavirin, which has a significant antiviral effect on hepatitis C virus, might be useful in combination with IFN-alpha. At the dosage (3-6 mega units) usually used, IFN-alpha is relatively well tolerated. In about 10% of the patients therapy is interrupted, mainly because of severe fatigue, thyroid dysfunction or depression.     

High degree of conservation in the hepatitis B virus core gene during the immune tolerant phase in perinatally acquired chronic hepatitis B virus infection

BACKGROUND: Mutations in the hepatitis B virus genome have been implicated in the persistence of hepatitis B virus infection and the pathogenesis of hepatitis B virus related liver disease. In view of the heterogeneity in published sequences, data from cross-sectional studies of unrelated subjects cannot differentiate true mutations from infections with variant sequences. AIMS/METHODS: We compared the hepatitis B virus core gene sequences of 42 HBsAg positive subjects from 11 Chinese families with those of the index patients (maternal carriers) to determine the frequency and rate of true hepatitis B virus core gene mutations in patients with chronic hepatitis B virus infection. RESULTS: Completely identical nucleotide sequences were present in all the family members and index patients in two families, suggesting that the hepatitis B virus core gene can be conserved for more than 20 years. The high degree of sequence conservation in these families is related to the young age of the subjects (mean 19.2+/-8.9 years), the fact that they were all HBeAg positive and that 75% of them had persistently normal aminotransferase levels. Longitudinal studies confirmed that mutations were rare in those who remained HBeAg positive with normal aminotransferase levels (immune tolerant phase), but significantly more common in HBeAg positive subjects who had elevated aminotransferase levels and in those who cleared HBeAg (immune clearance phase), the rates of nucleotide and amino acid changes were respectively: 0.28+/-0.12 vs 1.30+/-0.26/10(3) nt position/yr and 0.04+/-0.01 vs 0.18+/-0.5/10(2) codon/yr. CONCLUSIONS: Identical nucleotide differences could be found in the sequences of all the subjects in some families. These differences were more likely to be due to intra-familial transmission of stable variants. Sequence analysis based on comparisons with published sequences would have led to over-reporting of mutations. The hepatitis B virus core gene can remain highly conserved for more than two decades during the immune tolerant phase of perinatally acquired chronic hepatitis B virus infection. However, significant changes can occur within 2-3 years during the immune clearance phase.     

A randomized, controlled trial of a 24-month course of interferon alfa 2b in patients with chronic hepatitis B who had hepatitis B virus DNA without hepatitis B e antigen in serum

Short-term interferon treatment of serum hepatitis B e antigen (HBeAg)-negative carriers with serum hepatitis B virus (HBV) DNA and histological features of chronic hepatitis B has been largely unsuccessful. In a pilot study of long-term treatment, 42 such patients were randomly assigned to 6 million units of interferon alfa 2b (IFN-alpha2b) three times per week for 24 consecutive months (n = 21, 4 with cirrhosis) or to no therapy (n = 21, 3 with cirrhosis). Five patients (24%) discontinued therapy because of treatment-related adverse reactions. Serum levels of alanine transaminase (ALT) became persistently normal and HBV DNA undetectable by dot-blot assay in 8 patients receiving interferon and in 2 untreated controls (38% vs. 10%; P = .03). Hepatitis flare-ups disappeared in 17 patients during therapy compared with 6 controls (81% vs. 29%; P < .001). During a median period of 22 months after interferon was stopped, 2 treated patients (10%) lost serum hepatitis B surface antigen (HBsAg) and seroconverted to antibodies to hepatitis B surface antigen (anti-HBs). Serum ALT remained persistently normal and HBV DNA undetectable by dot-blot assay in 6 initial responders and 1 initial nonresponder, compared with none of the 21 untreated controls (sustained response: 33% vs. 0; P < .001). Comparative analysis of pre- and posttreatment liver biopsies showed that mean Knodell scores dropped in the treated group (10.3 to 5.3; P = .01), but not in the untreated group (9.3 to 9.8; not significant). In conclusion, a 24-month course of treatment with 6 MU IFN-alpha2b was well tolerated by most patients, led to sustained suppression of HBV in one third, and attenuated hepatitis in 81% of patients.     

Serum hepatitis G virus RNA in patients with chronic viral hepatitis

OBJECTIVES: The hepatitis G virus (HGV) is a newly described flavivirus that affects a high proportion of patients with chronic viral hepatitis: our objective was to determine what role HGV might play in the course of disease. METHODS: We evaluated stored serum samples from 108 patients with chronic hepatitis B and 99 patients with chronic hepatitis C who participated in trials of alpha-interferon or ribavirin for the presence of hepatitis B virus (HBV) DNA and hepatitis C virus (HCV) RNA by branched DNA and for the presence of HGV RNA by polymerase chain reaction (PCR), using primers from the NS5 region of the genome. RESULTS: Initially, 20 (19%) patients with hepatitis B and 11 (11%) with hepatitis C had HGV RNA in their serum. Patients with and without HGV infection were similar with regard to clinical features, laboratory tests, and hepatic histology. HGV RNA levels fell during interferon therapy and became undetectable in those receiving the highest doses; however, HGV RNA levels returned to pretreatment values when therapy was stopped. With ribavirin therapy, HGV RNA levels did not change. Two- to 12-yr follow-up serum samples were available from 17 initially HGV RNA-positive patients, of whom only 10 (59%) were still positive. CONCLUSIONS: HGV infection is common among patients with chronic hepatitis B and C but has little effect on the short-term course of disease or response to therapy. HGV RNA levels are suppressed but not eradicated by alpha-interferon and are unaffected by ribavirin treatment. Spontaneous loss of HGV RNA occurs over time in a proportion of patients.     

Alternative splicing of GTBP in normal human tissues

The relationship between serum hepatitis Be antigen (HBeAg) and serum hepatitis B virus (HBV) DNA determined by two commercially available assays was examined in 345 Chinese patients with chronic HBV infection. HBV DNA was detected by these commercial assays in 85% of the HBeAg-positive patients. Discrepancies between test results were found to occur when serum HBV DNA levels were low (< 5 pg ml-1 for the Abbott Genostics and < 100 MEq ml-1 for Chiron Quantiplex assays). An equation for the conversion between results generated by these two assays was derived, which was found to be very similar to the equation recently described by Kapke et al.     

Nested polymerase chain reaction (PCR) and multiple cloned antibody capture PCR for the examination of serum hepatitis B virus DNA in negative hepatitis B surface antigen patients suffering from liver diseases

In order to find out rapidly the causes of the liver diseases suffered by patients with negative hepatitis B surface antigen (HBsAg), nested polymerase chain reaction (PCR) and multiple cloned antibody capture PCR techniques were established to examine serum hepatitis B virus (HBV) DNA. By using both techniques along with the examination of hepatitis C virus (HCV) infection, the causes of chronic liver diseases with negative HBsAg were studied. It is found that nested-PCR can increase the sensitivity of single PCR more than 1,000 fold and multiple cloned antibody capture-PCR can detect concentration of HBV DNA as low as 0.1-0.01 pg/L. HBV DNA positive patients were found in 45.5%, 30.8%, 13.3% and 100% respectively of the patients suffering from liver cirhosis with negative HBsAg (group A, 22 cases), chronic hepatitis with negative HBsAg (group B, 13 cases), normal subjects with negative HBsAg and positive hepatitis B core antibody (HBcAb, group C, 30 cases) and liver cirhosis with positive HBsAg and negative HBeAg (group D, 12 cases). HBV DNA can be also found in the serum of HBsAb positive patients and subjects supposed to be healthy, 81.8% and 53.8% of the patients were infected with HBV and/or HCV in group A and group B respectively. All these results suggest that nested-PCR and multiple cloned antibody capture-PCR are rapid and highly sensitive methods for detection of serum HBV DNA. HBV infection is an important cause of chronic liver diseases in patients with negative HBsAg. The causes of most of the HBsAg-negative chronic liver diseases are related with infection of viruses. The clinical significance of serum HBsAb in naturally infected patients should be reconsidered.     

Post-hemorrhagic shock mesenteric lymph is cytotoxic to endothelial cells and activates neutrophils

The aim of this study was to evaluate the Chiron branched DNA (bDNA) assay for detection of serum hepatitis B virus (HBV) DNA in patients with chronic hepatitis B lacking hepatitis B e antigen (HBeAg) and undergoing interferon (IFN) therapy. Results obtained with the bDNA assay were compared with those obtained using the Abbott liquid hybridization (LH) assay and the polymerase chain reaction (PCR). Serial samples (274) from 34 patients were analysed. Analysis of variance results indicated that bDNA values were more significantly correlated than LH values with both PCR positive/negative results (probability of artifact (Prob > F) = 0.7 and 0.09 for LH and bDNA assays, respectively) and presence/absence of precore mutations (Prob > F = 0.21 and 0.001 for LH and bDNA assays, respectively). Both bDNA and LH results correlated highly with alanine aminotransferase (ALT) values (both had Prob > F values of 0.0) while PCR was not correlated with ALT (Prob > F = 0.05). In 26 evaluable patients, a model based on a generalized Knodell score was used to predict response to IFN therapy, as defined by normalization of ALT values during therapy. This model discriminated well between non-responders and responders. The bDNA results correlated well with the generalized Knodell score, while the LH results did not (Prob > F = 0.04 and 0.19 for the bDNA and LH assays, respectively). In conclusion, the bDNA assay appears to be useful for quantification of HBV DNA levels in HBeAg-negative chronic hepatitis as it correlates with biochemical and histological indications of disease severity as well as with response to IFN therapy.     

Clinical and virological evaluation of the detection of pre-S1 and pre-S2 antigens in serum from patients with chronic hepatitis B

OBJECTIVES AND METHODS: We performed a prospective study to determine the clinical and virological significance of pre-S antigen detection in serum samples from patients with chronic hepatitis B virus infection. Four hundred thirty seven consecutive serum samples from 116 patients were tested for the presence of both pre-S1 and pre-S2 antigens by radioimmunoassay using specific monoclonal antibodies. RESULTS: The pre-S1 antigen/HBs antigen ratio, gave an estimation of the number of pre-S1 epitopes expressed on the surface of circulating viral particles, and was positively correlated with the intensity of viral replication intensity (P < 0.05). Moreover, the pre-S1 antigen/HBs antigen ratio was significantly higher in patients suffering from chronic hepatitis associated with viral replication (24% +/- 13); in anti-HBe positive patients, the pre-S1 antigen/HBs antigen ratio was higher in patients replicating a HBe antigen minus variant of the hepatitis B virus and suffering from chronic hepatitis (17% +/- 9) than in asymptomatic HBs antigen carriers (5% +/- 6) (P < 0.05). The pre-S2 antigen/HBs antigen ratio was not correlated with the level of viral replication or with the patient's clinical status. CONCLUSION: This study confirms that pre-S1 antigen detection is a reliable marker of hepatitis B virus replication which can be easily performed in chronically infected patients. This assay is especially useful in identifying anti-HBe positive carriers who replicate a minus pre-core mutant and could benefit from antiviral therapy.     

The effect of interferon and desferrioxamine on serum ferritin and hepatic iron concentrations in chronic hepatitis B

BACKGROUND/AIMS: Recent reports indicate that an individual's iron status might affect the response rate achieved with Interferon therapy for the treatment of chronic viral hepatitis. METHODOLOGY: Forty individuals, 29 men and 11 women, with chronic viral hepatitis B, who had elevated serum ferritin levels, were randomized to receive either Interferon (IFN) 5 MU TIW SQ for 6 months alone (n=21) or Interferon in combination with repetitive cycles of desferrioxamine infused at a dose of 80 mg/kg per cycle (n=19) over 3 consecutive days in an effort to reduce their metabolically active iron pool during the course of IFN treatment. These cycles were continued until a serum ferritin level of less than 250 ng/ml (normal values <220 ng/ml) was achieved. Additionally, all desferrioxamine treated subjects were placed on a low iron containing diet. An interferon response was defined as normalization of the serum ALT and seroconversion from eAg positive to eAb positive. All other responses were defined as failures. RESULTS: The mean ages of the subjects in the 2 groups were 39+/-6 and 38+/-5 years. The initial serum ALT levels were 150+/-27 and 151+/-13 IU/l. The hepatic iron concentrations were 916+/-29 and 896+/-15 microg/g/dry liver weight. The serum ferritin levels were 386+/-12 and 393+/-18 ng/ml. None of these values differed significantly between the 2 treatment groups. The desferrioxamine treated group consisted of 14 men and 5 women. This group experienced a reduction in their serum ferritin to a level of 237+/-13 ng/ml as a result of the desferrioxamine treatment (p<0.05). Additionally, a reduction in their hepatic iron concentration, to a level 766+/-29 microg/g/dry liver weight, occurred with treatment (p<0.05). Twelve of the 19 (63%) desferrioxamine-treated subjects and 8 of the 21 (38%) control subjects experienced a normalization of their serum ALT levels with treatment (p<0.05). Thirteen of 19 (68%) of the desferrioxamine-treated subjects but only 8 of 21 (38%) of the IFN alone treated group seroconverted to anti-e positive (p<0.05). Moreover, a greater improvement in the hepatic histologic score and rate of HBV-DNA loss occurred in the desferrioxamine-treated group. CONCLUSIONS: Based upon these data, it can be concluded that desferrioxamine infusion to achieve a normal serum ferritin level enhances the likelihood of an individual with chronic hepatitis B responding to IFN therapy. The precise mechanism responsible for this phenomenon is not clear, but would appear to be due to a reduction in the hepatic free iron pool as reflected by sequential changes in the serum ferritin and hepatic iron concentrations.     

Functional analysis of HBV genomes from patients with fulminant hepatitis

Two previous case reports suggest that hepatitis B virus (HBV) core promoter variants with a high replication competence contribute to the pathogenesis of fulminant hepatitis B (FHB). We recently found in HBV genomes from patients with FHB an accumulation of mutations within the core promoter region. Therefore, the aim of this study was to investigate the phenotype of these HBV variants. Replication competence and expression of hepatitis B surface antigen (HBsAg) and hepatitis B e antigen (HBeAg) of viral genomes from seven patients with FHB and one patient with fulminant recurrent hepatitis after liver transplantation were analyzed by transfection experiments in human hepatoma cells. Compared with wild-type virus, the HBV variants from the seven patients with FHB produced similar or slightly lower levels of intracellular replicative intermediates and extracellular viral particles. In contrast, the HBV genomes from the patient with fulminant recurrent hepatitis synthesized and secreted significantly more HBV DNA. All genomes tested expressed similar or even higher levels of HBeAg compared with wild-type virus, except for those from four patients with a precore stop codon mutation in the respective dominant viral populations. The level of HBsAg produced by all variant genomes was similar or reduced compared with wild-type virus. These data indicate that in some cases HBV variants with enhanced replication competence and/or a defect in HBeAg expression may contribute to the development of FHB. However, neither phenotype is an essential prerequisite; thus, an additional role of other viral or host factors in the pathogenesis of FHB is suggested.     

Response of patients with dual hepatitis B virus and C virus infection to interferon therapy

Patients with dual infection with hepatitis B virus (HBV) and delta virus (HDV) responded poorly to interferon (IFN) therapy. Little is known about the effect of IFN therapy in patients with HBV and hepatitis C virus (HCV) dual infection. The patients in two randomized controlled trials with chronic HBV infection were retrospectively assayed for HCV markers. The HBV responses to IFN therapy in patients with and without HCV markers were compared. An open trial was conducted in 4 patients who had lost their serum HBV surface antigen (HBsAg) but had continuing HCV viremia and hepatitis. Of the 15 patients seropositive for HCV marker(s), only 1 (6.7%) responded with seroclearance of HBV DNA and HBV e antigen, as compared with 46 (28%) of 164 HCV-negative patients (p = 0.058). Icteric hepatitis developed in 1 patient on emergence of serum HCV RNA in association with seroclearance of HBV DNA. In contrast, good response was demonstrated in 3 of the 4 patients who had lost serum HBsAg before therapy. The results suggest that IFN therapy is not only of limited value in patients with dual infection with HBV and HCV but also has a potential risk of severe hepatitis if the clearance of one virus removes its suppressive effect on and facilitates the emergence of the other. However, patients with continuing HCV hepatitis after termination of the chronic HBsAg carrier state responded well to IFN therapy.     

Fibrosing cholestatic hepatitis in a transplant recipient with hepatitis B virus precore mutant

A patient with hepatitis B virus (HBV) precore mutant (seropositive for hepatitis B surface antigen [HBsAg], anti-hepatitis B e antigen [HBeAg], and HBV DNA) who underwent orthotopic liver transplantation for end-stage liver disease is described. Sequencing of the HBV precore region of the pretransplant serum sample confirmed the presence of the precore stop-codon mutant (G-->A mutation in codon 1896) only. The patient received HBV immunoglobulin prophylaxis for 6 months but HBV recurred thereafter with a mild hepatitic flare, and he remained seropositive for HBsAg, anti-HBe, and HBV DNA. The initial hepatitic illness resolved in 3 months. The patient remained well for another 16 months before presenting with fibrosing cholestatic hepatitis (FCH). During his entire initial hepatitic flare, quiescent period, and final FCH phase, he remained seropositive for HBsAg, anti-HBe, and HBV DNA. Moreover, sequencing of the serum HBV DNA in final FCH phase showed the presence of the identical HBV precore mutant. Immunohistochemical staining showed extensive expression of HBsAg/pre-S1, pre-S2, and hepatitis B core antigen, but HBeAg was scarcely detectable. This case illustrates that (1) recurrence of HBV precore mutant infection can occur in liver; (2) it can give rise to FCH; and (3) hepatic accumulation of HBeAg is not essential for the development of FCH.     

A newly identified pattern of K-ras mutations at codons 12 and 13 is associated with long-term survival in colorectal cancer

BACKGROUND/AIMS: Hepatitis B virus (HBV) core gene heterogeneity may influence the outcome of liver disease and the response to interferon (IFN) therapy in adult HBV carriers. The aim of this study was to evaluate the possible association between HBV core gene variability and evolution of chronic hepatitis in children. METHODS: We examined serum samples from 25 children with HBV chronic hepatitis and HBe antigen (HBeAg) positivity who were followed-up for a mean of 7.4 years. Seven cases spontaneously seroconverted to anti-HBe, becoming HBV healthy carriers; nine cases were successfully treated with IFN; nine cases were non-responders to IFN therapy. HBV-DNA was extracted from one serum sample ("I") collected during the HBeAg positive phase, and from a second sample ("II") collected after the anti-HBe seroconversion or, in non-responders, after stopping therapy. The entire core gene of the HBV isolates was amplified and sequenced. RESULTS: Each isolate showed single or no missense mutation independently of the clinical behavior of the patients. HBeAg-defective viruses were detected in one case in both samples and in two cases only in sample "II". CONCLUSIONS: Core gene variability does not seem to be involved either in the outcome of infection or in the response to IFN treatment in children with HBV chronic hepatitis. Considering that most of the HBV carriers in our area acquire the infection in childhood, our data suggest that core gene heterogeneity is not a major cause of progression to chronicity.     

Efficacy of the carbocyclic 2'-deoxyguanosine nucleoside BMS-200475 in the woodchuck model of hepatitis B virus infection

Daily oral treatment with the cyclopentyl 2'-deoxyguanosine nucleoside BMS-200475 at doses ranging from 0.02 to 0.5 mg/kg of body weight for 1 to 3 months effectively reduced the level of woodchuck hepatitis virus (WHV) viremia in chronically infected woodchucks as measured by reductions in serum WHV DNA levels and endogenous hepadnaviral polymerase activity. Within 4 weeks of daily therapy with 0.5 or 0.1 mg of BMS-200475 per kg, endogenous viral polymerase levels in serum were reduced about 1,000-fold compared to pretreatment levels. Serum WHV DNA levels determined by a dot blot hybridization technique were comparably decreased in these treated animals. In the 3-month study, the sera of animals that had undetectable levels of WHV DNA by the dot blot technique were further analyzed by a highly sensitive semiquantitative PCR assay. The results indicate that BMS-200475 therapy reduced mean WHV titers by 10(7)- to 10(8)-fold, down to levels as low as 10(2) to 10(3) virions/ml of serum. Southern blot hybridization analysis of liver biopsy samples taken from animals during and after BMS-200475 treatment showed remarkable reductions in the levels of WHV DNA replicative intermediates and in the levels of covalently closed circular viral DNA. WHV viremia in BMS-200475-treated WHV carriers eventually returned to pretreatment levels after therapy was stopped. These results indicate that BMS-200475 should be evaluated in clinical trials for the therapy of chronic human hepatitis B virus infections.     

Strategy to design peptide inhibitors: structure of a complex of proteinase K with a designed octapeptide inhibitor N-Ac-Pro-Ala-Pro-Phe-DAla-Ala-Ala-Ala-NH2 at 2.5 A resolution

To assess the relationship between hepatitis C virus infection and Fas antigen expression on hepatocytes, we examined changes in hepatic Fas antigen expression in the presence or absence of active hepatitis C virus infection. Twenty patients with chronic hepatitis C infection were treated with interferon and underwent pre- and posttreatment liver biopsies. Patients were classified according to the absence (group A; n = 9) or the presence (group B; n = 11) of hepatitis C virus RNA (HCV-RNA) in the liver after interferon therapy. An immunohistochemical assay showed Fas antigen staining in hepatocytes membranes and cytoplasm with expression concentrated mainly in periportal areas. The percentage of Fas-positive cells in the liver before treatment was not different between group A (39.5+/-19.1%) and group B (32.5+/-15.6%). Hepatic Fas expression was reduced significantly after treatment (24.3+/-10.6%) compared with the pretreatment values in group A (p < 0.05) but not in group B (25.9+/-16.9%). There was no significant difference between the two groups in the degree of histologic improvement. These results suggest that hepatic Fas expression is associated with persistent infection of hepatitis C virus.