853. Persistent Expression of Single Chain Antibodies Mediated by AAV5 and AAVrh.10 Vectors

Abstract

Top of pageAbstract Passive transfer of immunity with monoclonal antibodies is an effective strategy to augment host defenses, but it is a challenge to maintain effective antibody levels due to the short half-life of the antibodies and the consequent requirement for repetitive administration once every 1 to 4 wk. We have recently demonstrated (Kasuya et al. Molecular Therapy 2005; 11:237-244) the efficiency of an adenovirus (Ad)-based gene transfer vector in delivering single chain antibodies, but while effective, the protective immunity lasted only 2 wk secondary to the immune responses against the Ad vector. To refine the technology to provide a longer duration of antibody expression, we tested the hypothesis that a gene encoding a single chain antibody delivered by adeno-associated virus (AAV) vectors would exhibit sustained antibody expression in the serum of experimental animals. To test this hypothesis, the gene encoding a cobratoxin-specific single chain antibody was cloned into two vectors, one based on an AAV serotype 5 vector (AAV5|[alpha]|CTX) and one based on the non-human primate AAV serotype rh.10 (AAVrh.10|[alpha]|CTX). The expression cassette in these vectors consists of (5' to 3'): the chicken |[beta]|-actin promoter; an Ig|[kappa]| secretion signal to direct expression into the extracellular space; the cDNA sequence corresponding to the anti-cobratoxin single chain antibody, including a human constant |[kappa]| (C|[kappa]|) domain; and the SV40 poly A signal. The expression of the antibody from both vectors was confirmed by Western analysis of infected cell supernatants. Groups of 5 BALB/c mice received a dose of 1011 particle units (pu) by the intrapleural route. Mice that received AAV5|[alpha]|CTX had serum Ig|[kappa]| titers that reached a peak of 102 at 4 wk. These levels remained stable through an 8 wk observation period. For mice that received 1011 pu of the AAVrh.10|[alpha]|CTX, the anti-human Ig|[kappa]| titer rose from undetectable levels at wk 0 to 9|[times]|102 at 4 wk, a level 9-fold higher levels than that for the AAV5-based vector. These levels also remained stable through the 8 wk observation period. These data demonstrate that it is possible to achieve sustained expression of antibodies using AAV-based vectors, and that AAVrh.10-based vectors provide a higher efficiency of antibody gene expression compared to AAV5-based vectors when administered by the intrapleural route. This strategy should prove useful in applications requiring persistent augmentation of host defenses with monoclonal antibodies.