Development of Novel Recombinant AAV Vectors and Strategies for the Potential Gene Therapy of Hemophilia.

Recombinant vectors based on a non-pathogenic human parvovirus, the adeno-associated virus (AAV), have gained attention as a potentially safe and useful alternative to the more commonly used retroviral and adenoviral vectors. AAV vectors are currently in use in Phase I/II clinical trials for gene therapy of a number of diseases such as cystic fibrosis, α-1 antitrypsin deficiency, muscular dystrophy, Batten's disease, and Parkinson's disease, and have shown efficacy in patients with Leber's congenital amaurosis, and hemophilia B. For patients with hemophilia B, however, relatively large vector doses are needed to achieve therapeutic benefits. Large vector doses also trigger an immune response as significant fraction of the vectors fails to traffic efficiently to the nucleus, and is targeted for degradation by the host cell proteasome machinery. With a better understanding of the various steps in the life cycle of AAV vectors, strategies leading to the development of novel AAV vectors that are capable of high-efficiency transduction at lower doses are needed. In this review, we summarize our strategies to develop novel AAV vectors for the potential gene therapy of both hemophilia B and hemophilia A, based on our recent studies on the basic molecular biology of AAV. These strategies, including the development of novel AAV vectors by site-directed mutagenesis of critical surface-exposed tyrosine residues on AAV2 capsids to circumvent the ubiquitination step and the use of different AAV serotypes and self-complementary (sc) AAV2 vectors, and their use as helper vectors to circumvent the obstacles of second-strand DNA synthesis of single-stranded (ss) AAV, should dramatically accelerate the progress towards the potential gene therapy of both hemophilia A and hemophilia B.