A Tetra-Primer Amplification Refractory Mutation System–Polymerase Chain Reaction (T-ARMS-PCR) for Genotyping of rs8099917 & rs12979860 IL28B Polymorphisms and Its Correlation of Various Variables in Iranian HCV Patients
Ali Bahari
Department of Gastroenterology and Liver Disease, Emam Reza Hospital, Mashhad University of Medical Sciences, Mashhad, Iran.
Mohammad Hashemi
Cellular and Molecular Research Center, Zahedan University of Medical Sciences, Zahedan, Iran.
Gholam Reza Bahari
Cellular and Molecular Research Center, Zahedan University of Medical Sciences, Zahedan, Iran.
Tahereh Fakharian *
Department of Internal Medicine, Vali Asr Hospital, Birjand University of Medical Sciences, Birjand, Iran.
Sina Gerayli
Department of Biology, Faculty of Sciences, Ferdowsi University of Mashhad, Mashhad, Iran.
Abbas Esmail Zadeh
Department of Gastroenterology and Liver Disease, Emam Reza Hospital, Mashhad University of Medical Sciences, Mashhad, Iran.
Hamid Reza Sima
Department of Gastroenterology and Liver Disease, Emam Reza Hospital, Mashhad University of Medical Sciences, Mashhad, Iran.
Hooman Mosannan Mozaffari
Department of Gastroenterology and Liver Disease, Emam Reza Hospital, Mashhad University of Medical Sciences, Mashhad, Iran.
Ali Reza Bakhshipour
Department of Gastroenterology and Liver Disease, Ali-Ebne-Abitaleb Hospital, Zahedan University of Medical Sciences, Zahedan, Iran.
Zohreh Bari
Department of Gasteroenterology, Mazandaran University of Medical Science, Sari, Iran.
*Author to whom correspondence should be addressed.
Abstract
Background: Selecting patients for new direct acting antiviral treatment of HCV has prompted a conflicting matter worldwide because of its high cost and limited availability. Genotyping of IL28B polymorphisms will aid clinical decision making for identifying priorities of urgent treatment in resource-limited countries.
Objectives: The aim of the present study was to design a simple tetra-primer amplification refractory mutation system–polymerase chain reaction (T-ARMS-PCR) for genotyping of the rs8099917 and rs12979860 IL28B gene polymorphisms. Furthermore, we identify the correlation of variables such as gender, serum ALT level, histology of liver and baseline viral load with these polymorphisms.
Patients and Methods: We efficiently designed a T-ARMS-PCR for detection of rs12979860 and rs8099917 IL28B gene polymorphisms. Using this method, we genotyped 148 hepatitis C patients. To ensure T-ARMS genotyping quality, we, regenotyped samples with the PCR- sequencing method.
Results: Results of genotyping of rs12979860 and rs8099917 by T-ARMS PCR method were 100% concordant with the sequencing results. Among these 148 patients with chronic hepatitis C, the frequency of the rs12979860 CT, TT and CC genotypes was 72.3%, 14.2% and 13.5%, respectively and the frequency of the rs8099917 TT, GT and GG genotypes was 58.1%, 38.5% and 3.4%. Low frequency (2.7%) of association of two unfavourable homozygous genotypes (TT rs12979860 / GG rs809917) as well as 56.7% of association of 3 or 4 favorable alleles could explain good response of Iranians to HCV treatment with interferon-based regimens. About correlation of polymorphisms with different variables, only high viral load showed a statistically significant correlation to unfavorable genotype of TT rs12979860 ( p value = 0/05 ) and there was no correlation of serum ALT level, gender and histology of liver to IL28B genotypes.
Conclusions: We report that rs8099917 polymorphisms could predict outcomes better than rs12979860 in Iranian HCV patients.
Keywords: Hepatitis C, IL28B, single nucleotide polymorphism, Tetra-ARMS-PCR, Iranian.