PCR-based in vitro synthesis of hepatitis C virus NS3 protease for rapid phenotypic resistance testing of protease inhibitors.
J Clin Microbiol
; 52(4): 1139-45, 2014 Apr.
Article
em En
| MEDLINE
| ID: mdl-24452171
ABSTRACT
Protease inhibitors (PIs) targeting the hepatitis C virus (HCV) NS3 protease, such as telaprevir, have significantly improved the sustained virologic response (SVR) rates of HCV genotype 1 antiviral therapy. Given the expanding antiviral therapy regimen, fast HCV PI resistance assays are urgently needed. In this view, we have developed a novel phenotypic resistance test for HCV PIs based on in vitro synthesis of patient-derived HCV NS3 protease and subsequent enzymatic testing in a fluorescent readout. The enzymatically active HCV NS3 proteases were synthesized from PCR-derived templates by an Escherichia coli S30 extract system. Tests of the protease genes with known mutations for telaprevir resistance showed that the phenotypic resistance test was fast, with a total turnaround time of <10 h, and was fully in agreement with the previous resistance results. The initial tests with 38 treatment-naive serum samples showed that the method was significantly less laborious and faster than currently available phenotypic resistance assays of HCV NS3 PIs.
Texto completo:
1
Base de dados:
MEDLINE
Assunto principal:
Antivirais
/
Inibidores de Proteases
/
Reação em Cadeia da Polimerase
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Proteínas não Estruturais Virais
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Hepacivirus
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Farmacorresistência Viral
Idioma:
En
Ano de publicação:
2014
Tipo de documento:
Article