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- Title
492. Biodistribution and Integration Assessment of AAV1 gag-PR-RT(ΔRNaseH) after Intramuscular Administration in Rabbits.
- Authors
Schnepp, Bruce C.; Soult, Michael C.; Allen, Tara; Anklesaria, Pervin; Johnson, Philip R.; Munson, Keith
- Abstract
Clade C HIV-1 is responsible for approximately half of all HIV infections worldwide. An initial AAV2 vaccine containing the Clade C HIV-1 genes for the gag, protease and part of the reverse transcriptase proteins (gag-PR-ΔRT) is currently being tested in clinical trials. A vaccine based on pseudotyped AAV 1 vectors is being developed to be used either on its own on or in combination with other vectors to improve the immune response. Similar to the AAV2-based HIV vaccine, this vaccine also contains the gag, protease and part of the reverse transcriptase proteins (gag-PR-RTΔRNaseH) flanked by the AAV2 ITR but is pseudotyped with the AAV1 capsids.In the current study we determined the biodistribution, persistence and potential chromosomal integration of the rAAV1-HIV vaccine after a single intramuscular injection in New Zealand White rabbits. At sacrifice on days 5, 91 and 364, blood and tissues were collected for biodistribution analysis. Tissues positive for vaccine DNA on days 91 and 364 were further evaluated for the potential of the vaccine DNA to integrate into host genomic DNA. The assay utilized was similar to the previously described PCR-based assay that detects the presence of integrated rAAV vectors present in the mouse genome (Schnepp et al., 2003).Biodistribution and persistence of the vaccine DNA was most common at the injection site and highly perfused tissues. The amount of vaccine DNA in tissues was dose dependent and decreased over time. By day 364 in the low dose (4e9 DRP) and mid dose (4e10 DRP) animals, vaccine DNA was confined to the injection site and liver. In the high dose (4e11 DRP) animals, vaccine DNA was detectable in the injection site, liver, iliac lymph nodes, spleen and the one testis sample. In the testis sample, the quantity of vaccine DNA was <10 copies/μg and the epididymal sperm samples were negative from all dose groups on day 364 suggesting that germline cells in the testes were not transduced. Tissue samples (∼n= 111) with quantifiable vaccine DNA at days 91 and 364 were analyzed for integration. One liver sample from the day 91 time-point injected with the highest dose scored as positive in the integration assay at the limit of detection. When this sample was re-tested in the C-repeat PCR assay at a lower level of DNA input, the sample scored as negative. Based upon the input level, it was determined that this sample had <624 potential integrated copies/μg and that the associated potential mutagenic rate induced by the rAAV integration was at least 200 times lower than the spontaneous mutation rate.Thus, biodistribution of vaccine DNA was limited to the injection site and highly perfused tissues and decreased with dose administered and time. The data are similar to those observed with the AAV2-based HIV vaccine and provide further evidence that the vaccine DNA persists primarily as unintegrated episomal concatamers.Molecular Therapy (2006) 13, S190–S191; doi: 10.1016/j.ymthe.2006.08.562
- Subjects
PROTEOLYTIC enzymes; REVERSE transcriptase; PROTEINS; TISSUES; RABBITS
- Publication
Molecular Therapy, 2006, Vol 13, pS190
- ISSN
1525-0016
- Publication type
Article
- DOI
10.1016/j.ymthe.2006.08.562