The challenge for successful CF gene therapy for CF is to express the normal CFTR coding sequences in sufficient numbers of airway epithelial cells on a persistent basis without significant toxicity. In the past, our program has focused on achieving these goals using adenovirus (Ad) vectors. Although we have achieved considerable success, despite intense efforts to develop Ad vectors to circumvent host responses that limit Ad-mediated gene transfer, host defense arrayed against Ad vectors are so robust that strategies to circumvent anti-Ad host responses and improve efficiency of Ad entry and trafficking are insufficient to achieve the goal of gene therapy for CF. In this context, over the past 18 months, we have developed strategies to achieve persistent CFTR expression using adenoassociated virus (AAV) and lentivirus vectors. To be useful, both vector systems have to improve entry efficiency into the airway epithelium, and for AAV, size constraints make it difficult to transfer the full length normal CFTR coding sequences controlled by a robust constitutive promoter or, better yet, the 5' flanking region of the normal CFTR gene. This refocused P01, representing a collaboration of the Weill Cornell CF Gene Therapy program with the Wilson laboratory at U. Pennsylvania, is designed to overcome these challenges. Together with the Crystal laboratory development of """"""""segmental trans-splicing"""""""", which functions at the mRNA level to double the genetic cargo space of any vector system, and the development in the Wilson laboratory of novel AAV serotypes and pseudotyped lentiviruses with high efficiency for gene transfer to the respiratory epithelium, we have combined our efforts and refocused our CF gene therapy program to novel AAV and pseudotyped lentivirus gene transfer vectors to achieve the goals of transfer and persistent expression of normal human CFTR coding sequences in airway epithelium. The proposal includes: Project 1, G. Gao, AAV Vectors Based on New Serotypes; Project 2, P. Leopold, Intracellular Trafficking of Novel AAV Serotypes; Project 3, R. Crystal, AAV-mediated Gene Therapy for CF Using Segmental Trans-splicing; Project 4, J. Wilson, Novel Lentiviral Pseudotypes; Core A, S. Kaminsky, Vector; Core B, N. Wivel, Animal Models; and Core C, R. Crystal, Administrative.
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