Pilot study of combination therapy with ribavirin and interferon alfa for the retreatment of chronic hepatitis B e antibody-positive patients.

Twenty-four patients with chronic hepatitis B virus (HBV), antibody to hepatitis B e antigen (anti-HBe), HBV DNA positivity, and alanine transaminase (ALT) elevation who failed previous interferon alfa (IFN-alpha) therapy were included in a pilot study of combination therapy with ribavirin and IFN-alpha. The patients received daily oral ribavirin (1,000-1,200 mg according to body weight) plus 5 million units (MU) IFN-alpha2b three times a week for 12 months and were followed-up for 12 months. The median viremia level decreased significantly at the end of treatment (1.2 x 10(3) copies/mL) and follow-up (4.0 x 10(2) copies/mL) compared with the baseline (3.0 x 10(6) copies/mL; P <.05). After 12 months, 8 of 24 (33%) patients had cleared HBV DNA and 12 (50%) had normal ALT levels. At the end of the study virological and biochemical response was 50% and 21%, respectively. Thus, virological and biochemical response sustained in 5 of 24 (21%) patients retreated with ribavirin and IFN-alpha; none of them lost hepatitis B surface antigen (HBsAg). Liver histology improved in 2 of 4 sustained responders but in none of the 12 nonresponders with paired biopsies (P =.05). The response was independent of dose and duration of previous treatment, viral load, or the distribution of HBV precore wild-type/mutant variants. However, sustained responders had significantly higher necroinflammation (P =.036) and fibrosis (P =.007) scores. IFN-alpha-related side effects were mild and reversible on discontinuation. In 4 (17%) patients who suffered nausea and diarrhea the ribavirin dosage was reduced by 50% after 1 month of therapy and finally discontinued in all of them. No patient had liver disease decompensation. In summary, combination therapy with ribavirin and IFN-alpha may be efficacious to treat viremic anti-HBe-positive patients with chronic hepatitis B who have failed previous IFN therapy.

Analysis of the hepatitis B virus precore and ORF-X sequences in patients with antibody to hepatitis B e antigen with and without normal ALT levels.

Serum samples from 20 anti-hepatitis B e antigen-positive patients with and without normal alanine aminotransferase (ALT) levels who had serum hepatitis B virus (HBV) DNA detectable only by polymerase chain reaction (PCR) were examined. Viral DNA was amplified by PCR, using primers that encompassed precore and ORF-X regions and sequenced directly, to investigate whether mutations in the nucleotide sequences of X and precore gene regions of HBV-DNA might be responsible for the difference in the activity of disease and in the levels of viral replication. The HBV-DNA concentration in patients with abnormal ALT levels was higher than in those with normal ALT. The amount of HBV-DNA correlated with the ALT levels (P < 0.05). Seventy-two percent of patients had HBV-DNA harboring the 1896 precore stop mutation, and there was a negative correlation between the percentage of precore mutant genotype and the HBV-DNA concentration (P < 0.05). Thirty percent of patients had mutations in ORF-X. Patients with ORF-X mutations had lower levels of HBV-DNA than those who had wild-type virus. The presence of mutations in precore and X regions may be related to a low HBV-DNA concentration and reduced biochemical activity in patients with anti-HBe.

GB virus C RNA in serum, liver, and peripheral blood mononuclear cells from patients with chronic hepatitis B, C, and D.

BACKGROUND & AIMS: No conclusive data about GB virus C (GBV-C) tropism are available. We have studied the presence of genomic and antigenomic GBV-C RNA in serum, liver, and peripheral blood cells of 56 patients with chronic hepatitis B, C, or D virus infection. METHODS: Genomic and antigenomic GBV-C RNA were detected by reverse-transcription nested polymerase chain reaction. Specificity was confirmed by sequencing, by chemical modification of the RNA, and by using tagged primers. RESULTS: Genomic GBV-C RNA was found in 10 of 56 (18%) of the sera. In contrast, antigenomic strand was not detected. The sequence of the amplified GBV-C RNA from 3 patients showed a 96% homology among them and from 83% to 88% with previously described isolates. Genomic GBV-C RNA was found in 7 of 7 liver samples of the patients with serum GBV-C RNA. In 6 of these 7 patients (85%), antigenomic strand was found. Genomic RNA was found in 7 of 7 of the peripheral blood cell samples of the same 7 patients. Antigenomic GBV-C RNA was not found in these cells. CONCLUSIONS: These results suggest that GBV-C is a hepatotropic virus that replicates in the human liver. The data do not support a role for GBV-C in chronic liver disease.

Decreased nitric oxide production in chronic viral hepatitis B and C.

Nitric oxide is a free radical gas molecule which may be implicated in antiviral defense. However, there is no information about its possible role in chronic viral hepatitis B and C. In this study we have analyzed the serum levels of NO2- (as an index of nitric oxide generation) from patients with chronic viral hepatitis B and C and relationship of same with the response to interferon therapy. Serum samples were analysed from 61 patients with chronic hepatitis B, 60 patients with chronic hepatitis C, 11 with chronic liver disease of nonviral origin, and 23 healthy controls. Levels of NO2- were statistically higher in healthy controls (P < 0.001) than in patients with chronic liver disease. No relation was found between NO2- and viremia or response to interferon therapy in patients with chronic hepatitis B. In contrast in chronic hepatitis C, responder patients had significantly higher NO2- than nonresponders (P < 0.01). With respect to the relation between NO2- levels and liver damage, patients with cirrhosis had lower NO2- levels than the rest of the patients (P < 0.001). In conclusion, patients with chronic viral hepatitis have low serum NO2- levels.

Impaired interferon induction of human MxA protein in chronic hepatitis B virus infection.

MxA protein is interferon inducible, and its role as an antiviral mediator is being studied in various viral diseases. Several cytokines, including type 1 interferons (alpha and beta), interleukins 2 and 12, and granulocyte, macrophage, and granulocyte-macrophage colony-stimulating factors, were tested for their ability to induce human MxA protein synthesis in peripheral blood mononuclear cells from 15 chronic hepatitis B virus-infected patients and 6 healthy subjects as controls. Constitutive MxA expression was scarce in patients and controls but increased significantly in response to type I interferons. MxA responsiveness to interferon alpha was diminished significantly in chronic hepatitis B patients, compared with healthy donors (P < 0.05); this effect was more marked in patients with high viremia levels. Interleukins 2 and 12, and none of the colony-stimulating factors tested, induced low, but detectable, MxA protein levels. These results indicate that chronic infection by hepatitis B virus may impair activation of the immune cells and their capacity to respond to type 1 interferons.

Efficacy of prolonged interferon-alpha treatment in chronic hepatitis B patients with HBeAb: comparison between 6 and 12 months of therapy.

We compared the response and the relapse rates of HBeAb-positive patients treated with interferon-alpha for 6 or 12 months. Thirty-eight HBeAb-positive patients which chronic hepatitis B were randomly allocated into two groups: Group I (19 patients receiving 10 MU of recombinant interferon-alpha 2b three times a week for 2 months, followed by 5 MU three times a week for 2 months and then 3 MU three times a week for 2 months); Group II (19 patients receiving 10 MU of recombinant interferon-alpha 2b three times a week for 2 months, followed by 5 MU three times a week for 2 months and then 3 MU three times a week for 8 months). At the end of treatment, alanine aminotransferase normalization was higher but not more significant in Group I than in II (53% vs 26%), while hepatitis B virus DNA clearance was similar in both groups (21% in Group I vs 26% in Group II). However, at 12 months of follow-up, biochemical relapses occurred only in Group I (60% vs 0% in Groups I and II, respectively). Five complete responders cleared hepatitis B surface antigen at that time. In conclusion, prolonged treatment of HBeAb patients is efficient in reducing the biochemical relapse.

Hepatitis E virus seroprevalence in acute viral hepatitis in a developed country confirmed by a supplemental assay.

Hepatitis E virus (HEV) infection is prevalent among cases of acute viral hepatitis in young adults in developing countries. HEV infection is not restricted to endemic areas, but would appear to be worldwide in distribution. In order to document the incidence of HEV infection in acute hepatitis cases in a developed country, IgG and IgM anti-HEV antibodies and HEV RNA were tested in 101 Caucasian patients with acute viral hepatitis; 92 of these cases had markers of acute viral hepatitis other than HEV. Forty-seven (46.5%) cases had IgG anti-HEV; IgM anti-HEV and HEV viremia were not detected. As the incidence of anti-HEV was higher than would be expected, the possibility of the occurrence of false positive results was subsequently investigated. Supplemental antibody testing, using a broadly reactive epitope region, reduced the frequency of anti-HEV to 17%. Therefore, supplemental antibody testing confirms the hepatitis E virus seroprevalence in a developed country. Since IgM anti-HEV and HEV viremia were not detected, persons with IgG anti-HEV may be "subclinical HEV cases," or have long-lived antibodies in their circulation.

Analysis of hepatitis B virus precore variants in hepatitis B e antibody-positive patients treated with prednisone plus interferon.

To assess the effects of prednisone and interferon on the distribution of hepatitis B virus (HBV) precore mutants, nine hepatitis B e antibody (HBeAb)-positive patients with HBV chronic infection were studied. Patients were treated with prednisone (30 mg day-1 for 4 weeks, followed by 20 mg day-1 for 2 weeks and by 10 mg day-1 for 1 week), followed by recombinant interferon-alpha (15 MU thrice per week) for 6 months, without a clearance period. The HBV precore region was amplified by polymerase chain reaction (PCR) and distribution of the precore mutants was determined by hybridization of PCR products. Moreover, the glucocorticoid-responsive element (GRE) was sequenced to determine whether changes in the sequence were produced at the end of prednisone treatment. During prednisone treatment, changes in alanine transaminase (ALT) were observed in only two patients, in who ALT decreased to nearly normal values. In three patients ALT normalized at the end of interferon treatment. At baseline, wild-type HBV alone was detected in one patient, while seven patients were infected by a mixture of wild-type and precore mutants, predominantly wild type. At the end of prednisone treatment, two patients were infected by only wild-type HBV. The proportion of precore mutants decreased in three cases, while no changes were observed in three. At the end of interferon treatment, the precore mutant proportion decreased in the three responders, while tending to increase or remain unchanged in the rest. No significant changes in GRE sequence were found as a result of prednisone treatment. Our results would appear to confirm the role of the immune system in the selection of precore mutants.

Analysis of hepatitis B precore region in serum and liver of chronic hepatitis B virus carriers.

Using an oligonucleotide hybridization assay we studied the prevalence of wild-type and the predominant pre-core mutant hepatitis B virus in serum and liver of 49 antibody to hepatitis B e antigen carriers and three hepatitis B e antigen positive patients. Of the 45 serum samples from the anti-HBe carriers analyzed (no serum sample was available in four patients), 36 (80%) had hepatitis B virus DNA. In 26 of these 36 patients (72%) a mixed population was detected, wild-type genome alone was found in six patients (16%), the single mutant (nucleotide position 1896), in three cases (8%) and in one patient (2%) the viral DNA had the two nucleotide mutation (1896 and 1899). Of the liver biopsies from the 36 anti-HBe patients studied (no liver biopsy was available in 13 patients), 33 (92%) had hepatitis B virus DNA. A mixed viral population was detected in 23 patients (69%), only wild-type virus or a single mutation was found in eight (34%) and two patients (8%), respectively. In all cases, wild-type was the predominant genome. In serum and liver samples from the same patient, we found a concordance of the presence of wild-type HBV and the pre-core mutants studied in 23/26 (88%) of the patients. Alanine aminotransferase levels were higher (p < 0.01) and the duration of hepatitis B surface antigen carrier lower (p < 0.02) in patients with a predominance of precore mutant in comparison to wild-type.(ABSTRACT TRUNCATED AT 250 WORDS)

Treatment of chronic hepatitis D virus infection with low and high doses of interferon-alpha 2a: utility of polymerase chain reaction in monitoring antiviral response.

We examined the efficacy of decreasing high doses (beginning at 18 MU/day) of interferon-alpha 2a vs. that of daily low doses (3 MU) in the treatment of chronic hepatitis delta virus infection. Patients treated with 18 MU had a somewhat higher frequency of normalization of serum ALT levels than patients treated with low doses (31% and 12%, respectively, on an intention-to-treat basis). A decrease in the percentage of hepatitis D virus RNA positivity was observed in both groups at the end of treatment. Thus, whereas in baseline samples 10 (62%) of the patients in each group were positive for hepatitis D virus RNA in serum on slot-blot hybridization, these numbers decreased to 5 (31%) and 4 (25%) patients in groups 1 and 2, respectively, at the end of therapy. However, hepatitis D virus RNA, detected by means of nested polymerase chain reaction, remained in all but two (one in each group) patients who completed the treatment. Finally, during posttreatment follow-up, hepatitis D virus RNA levels returned to baseline values, and only one patient remained negative for this marker. The beneficial effect of interferon-alpha was only transient. Only two patients (one from each treatment group) had persistently normal serum ALT levels after 18 mo of follow-up. Finally, the presence of serum hepatitis D virus RNA at the end of therapy, detected with nested polymerase chain reaction, might be a good marker for the prediction of viral replication relapse.

Significance of low HDV replication levels during the natural history of HDV infection. Long-term follow-up.

The use of genetic amplification of the hepatitis delta virus (HDV) genome reveals the existence of different HDV replicative behaviours during the natural history of chronic HDV infection. While some of the patients (8/19, 42%) presented high and long-term maintained levels of HDV replication, as detected by slot-blot hybridization, others showed fluctuations from positive to negative, and in 5/7 (71%) polymerase chain reaction (PCR) demonstrated the presence of the HDV genome. Finally, 4 patients were persistently slot-blot-negative and in 3 of them HDV-RNA was detected by PCR in all samples tested. The correlation observed between the low levels of HDV replication and the ALT values, as well as the reactivation observed in one of the patients, suggests that PCR is useful in the virological surveillance of HDV infection, and indicates its usefulness in evaluating the effectiveness of antiviral therapy for chronic hepatitis D.