Triple therapy of initial high-dose interferon with ribavirin and amantadine for patients with chronic hepatitis C.

AIMS: We previously reported the potential effect of combination therapy of an initial high-dose interferon (IFN) and amantadine on the eradication of HCV-RNA in patients with chronic hepatitis C. The additive effects of amantadine on interferon and ribavirin combination therapy remain controversial. In this study we investigated the efficacy of initial high-dose IFN with ribavirin and amantadine on the virological response in patients with chronic hepatitis C with a high viral load of genotype 1b. METHODS: Twenty-two patients with high viral loads of genotype 1b hepatitis C virus were enrolled in this pilot study. Patients were administered IFN-beta for four weeks and then IFN-alpha2b for 22 weeks with daily oral administration of ribavirin and amantadine. RESULTS: A sustained virological response (SVR) was shown in 31.8% (seven of 22 patients). With the naïve patients, the SVR rate was 21.4% (three of 14 patients). In patients who could not eradicate HCV-RNA by previous IFN monotherapy, the SVR rate was 50% (four of eight patients). CONCLUSION: Triple therapy with an initial high dose of IFN with ribavirin and amantadine may be effective, especially for chronic hepatitis C IFN-retreatment patients with a high viral load of genotype 1b.

A pilot study of combination therapy with initial high-dose interferon and amantadine hydrochloride for patients with chronic hepatitis C with the genotype 1b virus.

BACKGROUND/AIMS: Interferon monotherapy for patients with chronic hepatitis C has been suboptimal. We studied the effect of the combination therapy of an initial high-dose of interferon and amantadine. METHODOLOGY: We investigated the virological response of 20 patients with naive chronic hepatitis C with a high viral load of the genotype 1b virus. Seven patients were administered 6MU of interferon-beta once daily for 6 weeks and then thrice weekly for 20 weeks, and 13 were administered 6 MU of interferon-beta daily for 4 or 6 weeks and then 10 MU of natural interferon-alpha thrice weekly for 22 or 20 weeks. All patients were treated with amantadine hydrochloride (100 mg/day) for 26 weeks during interferon administration. RESULTS: The complete response, transient response and no response rate were 15.0%, 60.0%, and 25%, respectively. After daily administration of interferon-beta intravenously, 19 patients (95.0%) showed negative tests for serum HCV-RNA by the polymerase chain reaction method. At the end of treatment, the serum HCV-RNA was not detected in any patients treated with daily interferon-beta and intermittent interferon-alpha with amantadine. At 6-month follow-up, three patients had eradicated HCV-RNA, who were in the group of daily interferon-beta and intermittent interferon-alpha with amantadine. In the patients treated with daily interferon-beta and intermittent interferon-alpha with amantadine, the complete response, transient response and no response rates were 23.1%,-76.9% and 0%, respectively. CONCLUSIONS: These findings suggest that the combination of an initial high-dose interferon and amantadine shows promising effects on the eradication of HCV-RNA in the chronic hepatitis C patients with a high viral load of the genotype 1b virus.

The combination therapy of interferon and amantadine hydrochloride for patients with chronic hepatitis C.

BACKGROUND/AIMS: The effect of interferon treatment for chronic hepatitis C patients with genotype 1b virus has been suboptimal. We studied the effect of the combination therapy of interferon and amantadine on patients with a high serum viral load of genotype 1b virus. METHODOLOGY: We studied the virological response of naive chronic hepatitis C patients with a high viral load of genotype 1b virus (4.5 log copies/50 microL or 100 kcopies/mL and higher) during interferon and amantadine administration for 6 months and 6 months after the end of treatment. Twenty patients were treated with interferon alone (natural interferon-beta 6 MU daily for 6 weeks and thrice-a-week for 20 weeks) for 26 weeks. Eleven patients were treated with the combination therapy of interferon and amantadine hydrochloride (100 mg orally daily) for 26 weeks. RESULTS: After daily administration of interferon-beta intravenously once a day for 6 weeks, all patients showed the negative tests of serum HCV-RNA by polymerase-chain-reaction methods by the combination therapy, while 13 patients (65.0%) showed the negative tests by interferon alone (p = 0.0257). At the end of treatment, serum HCV-RNA were not detected in 54.5% of patients treated with interferon and amantadine, while it was detected in 50.0% of patients treated with interferon alone. At 6 months follow-up, only one patient (9.1%) could eradicate HCV-RNA in patients with interferon and amantadine, while no patient could with interferon monotherapy (not significantly). CONCLUSIONS: Amantadine hydrochloride has the additive effects to interferon treatment on the virological responses of serum HCV-RNA during a co-administration, although the combination therapy has not shown a significantly promising effect on the eradication of HCV-RNA in the patients with chronic hepatitis C with a high viral load of genotype 1b virus.

Hepatoma-derived growth factor is highly expressed in developing liver and promotes fetal hepatocyte proliferation.

Hepatoma-derived growth factor (HDGF) is a heparin-binding protein, which has been purified from the conditioned media of HuH-7 hepatoma cells. Recent studies have suggested the involvement of HDGF in development of the kidney and cardiovascular systems. In the present study, we investigated the possibility that HDGF was also involved in liver development. Northern blot and immunostaining revealed unique expression patterns of HDGF in liver development. HDGF expression was strongly detected in the fetal liver of the midgestation stage and was markedly decreased near birth. Its expression was mainly detected in stromal cells, including immature hepatocytes. Expression in hepatocytes decreased with differentiation. Administration of recombinant HDGF enhanced the growth of primary cultured fetal hepatocytes significantly, although the effect was small. The effect of exogenous HDGF on the proliferation of neonatal hepatocytes was also small and significant only at one point, despite the lower expression of endogenous HDGF, suggesting that the differences exist between fetal and neonatal hepatocytes. However, adenoviral introduction of HDGF antisense cDNA into the fetal hepatocytes significantly suppressed their proliferation, and the inhibitory effect of HDGF antisense virus was reversed by exogenous HDGF. In conclusion, HDGF helps regulate the hepatocyte proliferation in liver development.

Antisense oligonucleotides of hepatoma-derived growth factor (HDGF) suppress the proliferation of hepatoma cells.

BACKGROUND/AIMS: Human hepatoma-derived growth factor, purified from the conditioned medium of hepatoma-derived cell line, HuH-7, stimulates the growth of Swiss 3T3 fibroblasts and HuH-7 cells. To evaluate the role of hepatoma-derived growth factor on the growth of hepatoma cells, we investigated the effects of recombinant hepatoma-derived growth factor protein and hepatoma-derived growth factor antisense oligonucleotides on the proliferation of several hepatoma cell lines. METHODOLOGY: We examined the effects of hepatoma-derived growth factor antisense oligonucleotides on the growth of hepatoma cells by cell growth assay. RESULTS: Hepatoma-derived growth factor stimulated the proliferation of some hepatoma cells (HuH-7, HLF, HepG2, AH66tc cells) about 15-70% over than the control. Hepatoma-derived growth factor antisense oligonucleotides, phosphorothioate-linked or encapsulated in liposome, can inhibit the growth of hepatoma cells. The ID50 of hepatoma-derived growth factor antisense phosphorothioate oligonucleotides for HuH-7 cells, in which hepatoma-derived growth factor expression was abundant, was 3 microM by the assay of cell proliferation and [3H]-thymidine incorporation. Their ID50 for AH66tc cells, on which the effects of exogenous hepatoma-derived growth factor were weak, was higher than 10 microM. To omit the toxic effects due to phosphorothioate modification of oligonucleotides and keep the cellular uptake more without their destruction in the culture medium, we used oligonucleotides encapsulated in cationic liposome. Hepatoma-derived growth factor antisense oligonucleotides encapsulated in liposome suppressed the growth of hepatoma cells effectively (ID50:2.0 microM). CONCLUSIONS: These findings suggest that hepatoma-derived growth factor is one of important autocrine, and/or intracrine factors for hepatoma cells, and that hepatoma-derived growth factor anti-sense oligonucleotides may be useful for human hepatocellular carcinoma as an anti-cancer agent.

Interferon treatment for patients with chronic hepatitis C infected with high viral load of genotype 2 virus.

BACKGROUND/AIMS: Interferon treatment is more effective in patients with chronic hepatitis C infected with genotype 2a virus than those with genotype 1b virus. We analyzed patients with chronic hepatitis C treated by interferon in our clinics to develop a more effective regimen of interferon treatment for patients with genotype 2 virus infection. METHODOLOGY: We retrospectively analyzed the virological response of 36 patients with chronic hepatitis C with a high viral load, including 28 cases infected with the genotype 2a virus and 8 cases with the genotype 2b virus. The serum viral load of these patients were 6.0 log copies/mL and higher by the competitive polymerase chain reaction assay method. All patients could be treated with interferon-alpha or -beta for 6 months. Eleven patients were administered 6 million units of interferon-beta once daily for 6 weeks and then thrice weekly (group A). Twelve patients were administered 6 million units of interferon-alpha daily initially for 2 weeks and then thrice weekly (group B), and 10 patients were treated with the same dose of interferon-alpha thrice weekly from the first administration (group C). We decided the criteria of complete remission as the absence of serum HCV-RNA at both points of the end of interferon treatment and 6 months later. RESULTS: For all patients with genotype 2a virus infection, the complete remission, transient response and no response rates were 46.4%, 39.3% and 14.3%, respectively. The complete remission rates in group A, B and C were 100%, 41.7% and 20%, respectively. The transient remission rates in group B and C were 41.7% and 60%, respectively. The no response rates in group B and C were 16.7% and 20%, respectively. All patients with a high viral load of genotype 2a virus showed eradicated serum HCV-RNA virus in group A. The eradication rate of serum HCV-RNA in patients infected with the genotype 2a virus in group A was significantly higher than that of group B (p < 0.02) or group C (p < 0.01). For all patients with genotype 2b virus infection, complete remission, transient remission and no response rates were 12.5%, 50.0% and 37.5%, respectively. The complete remission rate of patients with the genotype 2b virus in group A and group B plus C was 0% and 25.0%, respectively. The eradication rate of patients with the genotype 2a virus in group A was significantly higher than that of patients with the genotype 2b virus (p < 0.01). CONCLUSIONS: These findings suggest that the initial sufficient dose of interferon administration is effective to eradicate serum HCV-RNA in patients with a high viral load of genotype 2a virus in chronic hepatitis C.

Anti-IL-6 receptor monoclonal antibody inhibits leukocyte recruitment and promotes T-cell apoptosis in a murine model of Crohn's disease.

BACKGROUND: To assess the contribution of IL-6 signaling to the physiopathology of Crohn's disease, we introduced anti-IL-6 receptor monoclonal antibody to a murine colitis model. METHODS: Colitis was induced in C.B-17-scid mice to which were transferred CD45RBhigh CD4+ T cells from Balb/c mice. Anti-IL-6 receptor monoclonal antibody or rat IgG was given intraperitoneally after T-cell transfer, followed by weekly injection. Vascular adhesion molecules and inducible nitric oxide synthase were visualized by immunostaining. Cytokine expression was determined by RT-PCR, and apoptotic cells were determined by the TUNEL method. RESULTS: Mice treated with anti-IL-6 receptor monoclonal antibody showed normal growth while controls lost weight. Colitis was improved histologically with reduced infiltration of LFA-1+ monocytes/macrophages and VLA-4+ T cells. ICAM-1 and VCAM-1 expression in the colonic vascular endothelium was markedly suppressed by the treatment, whereas no significant difference was seen in MAdCAM-1. IFN-gamma, TNF-alpha, and IL-1beta mRNAs were markedly reduced, but no difference was observed in the expression of IL-4, IL-10, and TGF-beta. Inducible nitric oxide synthase was upregulated in the mucosa of colitic mice and downregulated in the treated mice. Apoptotic cells were very sparse despite massive CD4+ T-cell infiltration in colitic mice, whereas increased apoptosis was seen in the treated mice with an apparently reduced number of T cells. CONCLUSIONS: Anti-IL-6 receptor monoclonal antibody abrogated murine colitis. It effectively blocked the expression of adhesion molecules, thereby blocking leukocyte recruitment, and increased T-cell apoptosis. These results strongly suggest the therapeutic potential of anti-IL-6 receptor monoclonal antibody for human Crohn's disease.

Hepatoma-derived growth factor stimulates cell growth after translocation to the nucleus by nuclear localization signals.

Hepatoma-derived growth factor (HDGF) is the original member of the HDGF family of proteins, which contains a well-conserved N-terminal amino acid sequence (homologous to the amino terminus of HDGF; hath) and nuclear localization signals (NLSs) in gene-specific regions other than the hath region. In addition to a bipartite NLS in a gene-specific region, an NLS-like sequence is also found in the hath region. In cells expressing green fluorescence protein (GFP)-HDGF, green fluorescence was observed in the nucleus, whereas it was detected in the cytoplasm of cells expressing GFP-HDGF with both NLSs mutated or deleted. GFP-hath protein (GFP-HATH) was distributed mainly in the nucleus, although some was present in the cytoplasm, whereas GFP-HDGF with a deleted hath region (HDGFnonHATH) was found only in the nucleus. Exogenously supplied GFP-HDGF was internalized and translocated to the nucleus. GFP-HATH was internalized, whereas GFP-HDGFnonHATH was not. Overexpression of HDGF stimulated DNA synthesis and cellular proliferation, although HDGF with both NLSs deleted did not. Overexpression of HDGFnonHATH caused a significant stimulation of DNA synthesis, whereas that of hath protein did not. HDGF containing the NLS sequence of p53 instead of the bipartite NLS did not stimulate DNA synthesis, and truncated forms without the C- or N-terminal side of NLS2 did not. These findings suggest that the gene-specific region, at least the bipartite NLS sequence and the N- and C-terminal neighboring portions, is essential for the mitogenic activity of HDGF after nuclear translocation.