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Many viruses, including hepatitis C virus (HCV), are major threats to public health, but few treatment options are available. Systemic administration of a combination of interferon-a and ribavirin is the only approved treatment for HCV. However, half of all patients are not cured by such treatment and a wide spectrum of systemic side-effects limits its effectiveness. I developed a gene therapy approach with three goals: 1) the restoration of local interferon secretion in cells infected with HCV, 2) no secretion of interferon in normal cells not infected with the virus, and 3) suppression of the emergence of resistant viral strains. A recombinant transcription factor was constructed, the intracellular localization of which is controlled by a viral protease to stimulate focal interferon secretion at sites of infection. A recombinant adenovirus associated virus expressing the transcription factor based on the described strategy inhibited HCV replication effectively in a HCV in vitro culture system

ABSTRACTThe hepatitis C virus (HCV) is a continuing threat to public health. The systemic administration of interferon alpha with ribavirin is the only currently approved treatment. However, this treatment is associated with a wide spectrum of systemic side effects that limits its effectiveness; thus, there is an urgent need for new treatment modalities. In this study, we describe a novel anti-HCV strategy employing a recombinant transcription factor that we have engineered in such a way that NS3/4a viral protease controls its intracellular localization, thereby restoring interferon secretion specifically in cells infected with HCV. Proof-of-concept experiments validated the strategy, showing that the recombinant transcription factor was triggered to stimulate interferon promoter by NS3/4A and remained inactive in cells without NS3/4a. Using an adenovirus-associated viral vector delivery system, we found that the recombinant transcription factor inhibited HCV replication effectively in vitroin cultured cells.

Coxsackievirus B3 (CVB3) has been identified as a major causative agent of acute and chronic myocarditis, but the involvement of other CVB serotypes in myocarditis has not been investigated. To dissect the pathological properties of different CVB serotypes toward primary cardiomyocytes, we tested their effects on primary cardiomyocyte cultures from neonatal rats. Morphological abnormalities were examined by both light and fluorescence microscopy after Hoechst 33342 staining, and loss of cell viability was estimated by MTT assay. All six CVB serotypes showed a similar degree of severe toxicity toward primary cardiomyocytes. CVB clinical isolates had cytopathic effects (CPEs) similar to those of their respective CVB reference strains. Within 1–2 days of infection with multiplicities of infection MOI 50, the cells began to experience morphological changes including cell shrinkage, rounding‐up, and slight nuclear condensation. The irreversible loss of cell viability was readily observed within 3–5 days following virus infection. These results suggest that all six CVB serotypes induce direct, irreversible toxicity towards cardiomyocytes, which eventually leads to the death of infected cells. These findings indicate that the variations in CVB serotype are not the limiting factor determining the susceptibility of cardiomyocytes to CVB infection. J. Med. Virol. 75:290–294, 2005. © 2004 Wiley‐Liss, Inc.