460. Very Low Prevalence of Circulating Antibodies Against Common AAV Serotypes in a Population of Cats

Abstract

An important goal in applied reproductive biology is to devise a reliable method to induce permanent sterility in cats and dogs following a single administration of a bioactive molecule. Although the reproductive system can be manipulated at different levels, it appears desirable to directly target the brain neurons that govern reproductive function. AAV vectors are promising candidates for this role, but their usefulness is hampered by the lack of tissue accessibility and the need of administering high doses. To address this issue and develop novel AAV vectors that effectively target neurons in the cat brain following a single intravenous injection of vector, we are using AAV Barcode-Seq, which allows us to study the biological properties of hundreds of different AAV capsids in an unprecedented, high-throughput manner using next-generation sequencing (NGS) technology. However, success of this approach depends on low prevalence of circulating antibodies against common AAV serotypes such as AAV2 and AAV9 because these serotypes serve as internal reference controls in this system. Therefore, it is essential to investigate the prevalence of neutralizing antibodies against various serotypes in cats for applying the AAV Barcode-Seq to preclinical studies and ultimately for successful translation of AAV vector-mediated gene therapy approaches into the veterinary clinic. Here we report the results of the initial screening of 30 cat serum samples obtained from the Liberty Research Inc. cat colony for the presence of antibodies against a total of 11 different AAV serotypes (AAV1 through 11). Such a study is imperative because a vaccine against feline panleukopenia virus (FPV), a feline parvovirus, is widely used to immunize cats and all the cats to be used for our study will have received that vaccine. In the study, serum samples were incubated in ELISA plates coated with serotype-specific particles followed by addition of an HRP-conjugated anti-cat IgG antibody. Anti-FPV antibody titers were determined by a hemagglutination inhibition test (Cornell University, Animal Health Diagnostic Center). The results showed that the majority of the 30 cats did not harbor any detectable antibodies against AAV1, 2, 3, 4, 5, 7, 8, 9, 10 or 11, while modest levels of anti-AAV6 antibodies were detected in two-thirds of the animals. Spearman's rank correlation coefficient between anti-AAV6 and anti-FPV antibody levels was 0.40, showing a positive correlation. In addition, the anti-FPV and anti-AAV6 antibody levels were significantly higher in the cats that had been vaccinated within 99 days prior to the blood sample collection (p=0.005 and p=0.008, respectively). These observations indicate that the prevalence of antibodies against the common AAV serotypes is very low in the population of cats we investigated even though the cats had developed high titers of anti-FPV antibodies, which appear to only cross-react with AAV6. We are currently investigating whether anti-AAV6 antibodies found in cat sera exhibit neutralizing activity.