THE PACKAGING CAPACITY OF ADENOASSOCIATED VIRUS (AAV) AND THE POTENTIAL FOR WILD-TYPE-PLUS AAV GENE-THERAPY VECTORS

Because of its ability to integrate chromosomally and its non-pathogen ic nature, adeno-associated virus (AAV) has significant potential as a human gene therapy vector. Here we investigate the maximum amount of DNA which can be inserted into the AAV genome and still allow efficien t packaging into an infectious virus particle. Altered wild-type AAV g enomes were constructed with inserts, which increased in size by 100 b p, ligated at map unit 96. These large wild-type-plus genomes were abl e to replicate and produce infectious virus, at levels slightly reduce d but comparable to normal sized wild type, until the insert size reac hed 1 kb. These data indicate that the maximum effective packaging cap acity of AAV is approximately 900 bp larger than wild type, or 119%, F urthermore, it is demonstrated that these large AAV genomes are able t o latently infect cells by chromosomal integration as does wild-type A AV. These data suggest that therapy vectors carrying a foreign gene of 900 bp or less can be generated from AAV, by ligation into non-essent ial locations, and result in a recombinant AAV virus with a fully wild -type phenotype, Such wild-type-plus AAV vectors will have both advant ages and disadvantages over defective recombinant AAV virus - the most important advantages being the ease in which high titers of infectiou s virus can be generated and the ability to specifically integrate wit hin chromosome 19, Once the concern subsides over the presence of wild -type AAV in clinical applications, wild-type AAV vectors may find spe cific application niches for use in human gene therapy. (C) 1997 Feder ation of European Biochemical Societies.