Detailed knowledge about the life cycle of hepatitis C virus has been hampered by the lack of a robust cell culture system. The recent development of replicon and infectious cell culture system are major breakthroughs. The mechanisms of interferon action and resistance in the treatment of hepatitis C remain largely unknown.? ? About half of hepatitis C virus (HCV)-infected patients do not respond to current interferon (IFN)-alfa combination therapy. To understand IFN resistance in vivo, we examined the dynamic responses to human IFN (hIFN)-alfa, -gamma and consensus IFN in the chimpanzee model. In vitro study with peripheral blood mononuclear cells (PBMCs) of naive chimpanzees and healthy human donors showed that chimpanzee responded less well than human to hIFNs. Naive and HCV-infected chimpanzees were treated with hIFNs using higher doses to compensate for the lower efficacy of hIFNs in chimpanzees. The in vivo responses of PBMCs to all three IFNs were much lower in the HCV-infected chimpanzees than those in the naive chimpanzees. The difference was more dramatic in the liver as the hepatic IFN-induced gene inductions were barely detectable in the infected animals. Assessment of various IFN signaling inhibitors showed that following IFN administration, the expression of suppressor of cytokine signaling 3 (SOCS3) was significantly upregulated, possibly through the induction of IL-6, in the liver of HCV-infected chimpanzees. In conclusion, these data indicate a defective response, particularly in the liver, to IFNs in HCV-infected chimpanzees, and this defect is possibly mediated through the activation of SOCS3. Further study on the mechanism of IFN pathway blockage by HCV infection in chimpanzee may provide novel insights into the clinical issue of nonresponse to IFN therapy.? ? To further explore the mechanisms of IFN action and resistance in HCV patients, we compared hepatic gene expression in patients prior to and during peginterferon and ribavirin therapy. Patients were divided into 2 groups. Group A (treatment group) received treatment prior to liver biopsy: 5 patients received peginterferon alone 24 hours prior to biopsy (Peg) and 6 patients received ribavirin for 72 hours plus peginterferon 24 hours prior to biopsy (Peg+Rbv). Treated patients were divided by response: rapid responders (RR) >2 log drop in HCV RNA by week 2 (n=6) and slow responders (SR) <2 log drop in HCV RNA by week 2 (n=5). In Group B (control group) patients had a liver biopsy performed before starting antiviral therapy and were grouped to RR and SR based on their subsequent treatment response with Peg and Rbv. Biopsy tissue was snap-frozen and preserved in liquid nitrogen. After RNA extraction, gene expression profiling was performed using the full human genome with standard Affymetrix microarray technology (54,000 gene platform). Gene expression was compared between the groups and by treatment outcome. Expression differences of at least 1.5 fold with a p-value <0.01 were considered significant. Eleven patients in the treatment group were compared to 19 in the control group. The groups were matched for gender, race, viral load, genotype and severity of histological disease. A total of 1274 genes differed between the treatment and control groups. Known ISGs as well as genes involved in the immune response were induced in the treated patients. 563 genes differed between patients that received Peg+Rbv and those receiving Peg alone at the time of liver biopsy. Peg+Rbv patients had heightened induction of many IFN-related genes, including the IFN-? receptor, and also showed significant down-regulation of genes involved in IFN inhibition, apoptosis and stellate cell activation as compared to the Peg patients. Examining treatment response, 1064 genes differed between RR and SR patients. Similar to the pattern seen in the RBV-treated patients, IFN inhibitory pathways and genes involved in stellate cell activation and apoptosis were down-regulated in the RR group. As previously reported, NR baseline liver biopsies had higher expression of numerous ISGs than those of SVR in the control group. Induction of interferon inhibitory pathways may account for reduced treatment efficacy as was seen in the slow responder group. Patients achieving a rapid decline in viral load and those treated with peginterferon and ribavirin showed similar expression patterns with down-regulation of genes involved in apoptosis, hepatic stellate cell activation and interferon inhibition. Ribavirin may also provide synergy to interferon through up-regulation of the interferon receptor. Collectively these mechanisms may provide a molecular basis for treatment response and account for the improved efficacy of combination therapy.? ? Polymorphisms of cytokine genes are associated with disease outcome and treatment response in hepatitis C. We performed a comprehensive analysis of interferon (IFN)-gamma gene polymorphisms for association with IFN-gamma-induced and spontaneous recovery from HCV infection. We genotyped eight single nucleotide polymorphisms (SNPs) spanning the entire IFN-gamma gene in two cohorts and assessed the association between those polymorphisms and treatment response or spontaneous viral clearance. The first cohort was composed of 284 chronic HCV-infected patients who had received IFN-alfa-based therapy and the second was 251 intravenous drug users who had either spontaneously cleared HCV or become chronically infected. A SNP variant located in the proximal IFN-gamma promoter region, -764G, was significantly associated with sustained virological response (SVR) [P=0.01, OR=2.7 (1.3-5.6)]. The association was independently significant in multiple logistic regression (P=0.04) along with race, viral titer and genotype. This variant was also significantly associated with spontaneous recovery [P=0.04, OR=3.51(1.0-12.5)] in the second cohort. In the promoter assay, the -764G IFN-gamma promoter variant had a two- to three-fold higher activity than the common -764C. Electrophoretic mobility shift assay demonstrated that this SNP is located within a heat shock transcription faction binding site and the binding affinity was significantly higher with the G than the C allele. Our study suggests that the SNP at position -764 of the IFN-gamma promoter is functionally important in determining viral clearance and treatment response and may be used as a genetic marker to predict SVR in HCV-infected patients.
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