The small packaging capacity of adeno-associated virus (AAV) vectors limits the utility of this promising vector system for transfer of large genes. We explored the possibility that larger genes could be reconstituted following homologous recombination between AAV vectors carrying overlapping gene fragments. An alkaline phosphatase (AP) gene was split between two such AAV vectors (rec vectors) and packaged using AAV2 or AAV6 capsid proteins. Rec vectors having either capsid protein recombined to express AP in cultured cells at about 1–2% of the rate observed for an intact vector. Surprisingly, the AAV6 rec vectors transduced lung cells in mice almost as efficiently as did an intact vector, with 10% of airway epithelial cells, the target for treatment of cystic fibrosis (CF), being positive. Thus AAV rec vectors may be useful for diseases such as CF that require transfer of large genes.
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We thank John M. Alfano and Stephanie R. Moe for excellent technical assistance. This work was supported by grants DK47754 and HL66947 from the National Institutes of Health.
The authors own small amounts of stock in several gene therapy companies and may share in royalties from patents covering inventions related to vector development which are owned by the Fred Hutchinson Cancer Research Center.
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Halbert, C., Allen, J. & Miller, A. Efficient mouse airway transduction following recombination between AAV vectors carrying parts of a larger gene. Nat Biotechnol 20, 697–701 (2002). https://doi.org/10.1038/nbt0702-697
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