Although the development of anti-interferon (IFN)-alpha neutralizing antibodies (NAbs) is likely to be a common clinical problem for patients with various diseases treated with IFN, anti-IFN-alpha NAb has been exceptionally considered to have no clinical significance in the treatment of chronic hepatitis C with pegylated IFN-alpha (Peg-IFN-alpha). However, we recently clarified that the presence of NAb was associated with a non-response to the Peg-IFN plus ribavirin (RBV) therapy. In this study, we used the HCV-replicon system with genotype 1 b, and investigated the role of anti-IFN-alpha NAb in the response to telaprevir (TVR)-containing new antiviral therapy for hepatitis C virus (HCV). Anti-IFN-alpha NAb-positive sera specifically inhibited the anti-HCV effects of IFN-alpha, without any effect on the activity of IFN-beta in vitro. The NAb-positive sera also inhibited theIFN-alpha-dependent induction of interferon-stimulated genes, MxA and OAS-1, in a dosedependent manner. Although TVR monotherapy decreased the HCV-RNA in vitro, the HCV-RNA was increased again with the development of TVR-resistant mutations. When IFN-alpha was administrated with TVR, the replication of HCV was continuously suppressed for more than a month. However, in the presence of anti-IFN-alpha NAb-positive sera, even when IFN-alpha was combined with TVR, the levels of HCV-RNA exhibited a time-course similar to that with TVR monotherapy, and HCV with TVR-resistant mutations emerged. In conclusion, our findings suggest that the presence of IFN-alpha NAb decreases the antiviral effects of IFN-alpha and may be related to the development of TVR-resistant mutated viruses. (C) 2014 Elsevier Inc. All rights reserved.