| Co-Investigator(Kenkyū-buntansha) |
KUROSAKI Masayuki Tokyo Medical and Dental University, Faculty of Medicine, Assistant, 医学部, 助手 (10280976)
ENOMOTO Nobuyuki Tokyo Medical and Dental University, Faculty of Medicine, Assistant, 医学部, 助手 (20251530)
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| Budget Amount *help |
¥7,700,000 (Direct Cost: ¥7,700,000)
Fiscal Year 1997: ¥3,600,000 (Direct Cost: ¥3,600,000)
Fiscal Year 1996: ¥4,100,000 (Direct Cost: ¥4,100,000)
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| Research Abstract |
To predict IFN efficacy, it is important to detect IFN-resistant HCV with the wild-type NS5A rather than to detect IFN-sensitive HCV with the mutant-type NS5A in individuals. The purpose of the present study was to develop a simple method to detect IFN-resistant HCV in the serum. The RFLP method, the hybridization method using type-specific probes, the PCR method with type-specific primers were considered as candidates. However, these methods were turned out to be inappropriate because it was not nucleotide mutations but amino acid mutations that determine the type of the NS5A,and there were many silent mutations in this region. The Multipin methods, which can detect many peptide at one time, did not work well enough. At presnt, therefore, the direct sequencing method appeared to be the most cost-effective method to detect HCV with wild-type NS5A.
Then, using the direct sequencing method and the cloning method, it was studied whether wild-type, intermediate-type, and mutant type HCV co-existed in same individuals. In patients in whom the mutant-type was replaced by the intermediate-type after interferon therapy, it was found that these change were not caused by the mutation of the virus but the selection of the quasispecies as shown by the cloning method.
These cases were rare, however, among a certain number of patients. It was concluded that the direct sequencing method could be a useful method to detect interferon-resistant HCV after improving the reading of nucleotide sequences.
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