IL28B polymorphism and cytomegalovirus predict response to treatment in Egyptian HCV type 4 patients

AIM:To test whether the status of positive cytomegalovirus(CMV) DNA detection adds to the predictive value of IL28B and to further categorize C/T allele carriers.METHODS:This study included 166 chronic hepatitis C(CHC) patients who received combined interferon and ribavirin therapy for 48 wk,84 spontaneous hepatitis C virus(HCV) resolvers who were positive for IgG anti-HCV antibody and negative for HCV RNA,and 100 healthy subjects who were negative for both HCV antibodies and RNA as controls.Genomic DNA from peripheral blood was used for IL28B rs.12979860 single nucleotide polymorphism(SNP) and CMV DNA detection.A 139 bp fragment containing IL28B SNP was amplified in all subjects by polymerase chain reaction using a specifically designed primer.Then the IL28B rs.12979860 SNP was detected by restriction fragment length polymorphism(RFLP) genotyping.The presence of CMV DNA was tested by amplification of the gB1 gene using nested polymerase chain reaction.The role of CMV and IL28B rs.12979860 SNP genotypes in determining the response rate to combined interferon therapy and clinical status of patients were statistically analyzed.RESULTS:Current data showed that 67% of patients carrying the IL28B 12979860 C/C allele had a sustained viral response(SVR) while the genotypes C/T and TT were associated with lower SVR rates,50% and 48%,respectively.SVR rates for the C/C allele were lower than other HCV genotypes and/or other populations.Genotype CC was associated with the response to interferon(P = 0.025).Genotype C/C was reduced from 48% in controls to 14% in CHC patients suggesting its protective role against progression to chronicity.The majority of spontaneously cleared subjects(86%) were C/C,confirming its protective role.The C/T allele was present in 71% of CHC patients compared with 38% of controls,so the use of IL28B SNP genotyping only in these patients may be of little value as a predictor of response.CMV reactivation occurred in 40% of CHC patients.Co-infection with CMV seriously diminished the response to interferon(IFN) therapy,with SVR rates in C/C genotypes 87.5% in CMV-negative patients and 12.5% in CMV-positive patients(P < 0.0001).SVR rates among C/T carriers were reduced to < 50% in patients with positive CMV DNA while the non-response rate doubled.These data indicate that a supplemental assay for CMV viremia adds to the prognostic value of IL28B genotyping.CONCLUSION:The results suggest that both genetic(i.e.,spontaneous) and therapeutic(IFN-based therapy) arms are complementary in the battle against HCV.CMV DNA testing may be of value to better predict the response to IFN,particularly in IL28B C/T carriers.

Tumor necrosis factor-α-G308A polymorphism is associated with liver pathological changes in hepatitis C virus patients

AIM To investigate the association of tumor necrosis factor alpha(TNFα)-G308 A polymorphism with different liver pathological changes in treatment-na?ve Egyptian patients infected with hepatitis C virus(HCV) genotype 4.METHODS This study included 180 subjects,composed of 120 treatment-na?ve chronic HCV patients with different fibrosis grades(F0-F4) and 60 healthy controls. The TNFα-G308 A region was amplified by PCR and the different genotypes were detected by restriction fragment length polymorphism analysis. The TNFα protein was detected by enzyme-linked immunosorbent assay. The influence of different TNFα-G308 A genotypes on TNFα expression and liver disease progression were statistically analyzed. The OR and 95%CI were calculated to assess the relative risk confidence.RESULTS Current data showed that the TNFα-G308 A SNP frequency was significantly different between controls and HCV infected patients(P = 0.001). Both the AA genotype and A allele were significantly higher in late fibrosis patients(F2-F4,n = 60) than in early fibrosis patients(F0-F1,n = 60)(P = 0.05,0.04 respectively). Moreover,the GA or AA genotypes increased the TNFα serum level greater than the GG genotype(P = 0.002). The results showed a clear association between severe liver pathological conditions(inflammation,steatosis and fibrosis) and(GA + AA) genotypes(P = 0.035,0.03,0.04 respectively). The stepwise logistic regression analysis showed that the TNFα genotypes(GA + AA) were significantly associated with liver inflammation(OR = 3.776,95%CI: 1.399-10.194,P = 0.009),severe steatosis(OR = 4.49,95%CI: 1.441-14.0,P = 0.010) and fibrosis progression(OR = 2.84,95%CI: 1.080-7.472,P = 0.034). Also,the A allele was an independent risk factor for liver inflammation(P = 0.003),steatosis(P = 0.003) and fibrosis(P = 0.014). CONCLUSION TNFα SNP at nucleotide-308 represents an important genetic marker that can be used for the prognosis of different liver pathological changes in HCV infected

Positional effect of mutations in 5'UTR of hepatitis C virus 4a on patients' response to therapy

AIM:To investigate the effects of mutations in domain Ⅲ of the hepatitis C virus(HCV)internal ribosome entry sequences(IRES)on the response of chronic HCV genotype 4a patients to interferon therapy.METHODS:HCV RNA was extracted from 19 chronic HCV 4a patients receiving interferon/ribavirin therapy who showed dramatic differences in their response to combination therapy after initial viral clearance.IRES domainⅢ was cloned and 15 clones for each patient were sequenced.The obtained sequences were aligned with genotype 4a prototype using the ClustalW program and mutations scored.Prediction of stem-loop secondary structure and thermodynamic stability of the major quasispecies in each patient was performed using the MFOLD 3.2 program with Turner energies and selected constraints on base pairing.RESULTS:Analysis of RNA secondary structure revealed that insertions in domainⅢ altered WatsonCrick base pairing of stems and reduced molecular stability of RNA,which may ultimately reduce binding affinity to ribosomal proteins.Insertion mutations in domainⅢwere statistically more prevalent in sustained viral response patients(SVR,n=14)as compared to breakthrough(BT,n=5)patients.CONCLUSION:The influence of mutations within domainⅢ on the response of HCV patients to combination therapy depends primarily on the position,but not the frequency,of these mutations within IRES domain Ⅲ.