Detection of Intact rAAV Particles up to 6 Years After Successful Gene Transfer in the Retina of Dogs and Primates

Abstract

Gene transfer to the retina using recombinant adeno-associated viral (rAAV) vectors has proven to be an effective option for the treatment of retinal degenerative diseases in several animal models and has recently advanced into clinical trials in humans. To date, intracellular trafficking of AAV vectors and subsequent capsid degradation has been studied only in vitro, but the fate of AAV particles in transduced cells following subretinal injection has yet to be elucidated. Using electron microscopy and western blot, we analyzed retinas of one primate and four dogs that had been subretinally injected with AAV2/4, -2/5, or -2/2 serotypes and that displayed efficient gene transfer over several years. We show that intact AAV particles are still present in retinal cells, for up to 6 years after successful gene transfer in these large animals. The persistence of intact vector particles in the target organ, several years postadministration, is totally unexpected and, therefore, represents a new and unanticipated safety issue to consider at a time when gene therapy clinical trials raise new immunological concerns.

Human Treg responses allow sustained recombinant adeno-associated virus–mediated transgene expression

“…This finding is consistent with an earlier study that documented the detection of rAAV particles up to 6 years after administration in the retina of dogs and nonhuman primates (25) and may explain why Tregs would continue to reside within the injected muscle months after administration.…”

Long-term VEGF-A expression promotes aberrant angiogenesis and fibrosis in skeletal muscle

“…[7][8][9] There are studies following the transgene expression and effects in the long-term. [10][11][12] However, not much is known about the expression of angiogenic growth factors delivered by AAV in large animal models. Previously, the effects of AAV vascular endothelial growth factor A (AAV-VEGF-A) on angiogenesis in ischemic skeletal muscle have been primarily evaluated in small animal models for time points no longer than 6 months.…”

Gene therapy for retinal ganglion cell neuroprotection in glaucoma

“…13 Tetracycline-regulatable promoter systems are also being investigated as a means to regulate AAVmediated transgene expression in RGCs. 14 Other advantages of the AAV vector system for in vivo gene delivery include that it is not pathogenic and has not been implicated in the etiology of any known human disease, it mediates long-term transgene expression that can last for several years in the retina 15 and it has low immunogenicity. In the absence of helper virus, wild-type AAV can integrate at a specific site on the 19q to establish latent infection.…”