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 domain III 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 domain III altered Watson-Crick base pairing of stems and reduced molecular stability of RNA, which may ultimately reduce binding affinity to ribosomal proteins. Insertion mutations in domain III were 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 domain III 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.