rAAV8 biodistribution and shedding after subretinal injection in non-human primates

Abstract

Purpose : The aim of this study was to analyse virus distribution and shedding after a single subretinal administration of clinical grade recombinant adeno-associated virus (rAAV8) in non-human primates. This is important for environmental risk assessment in retinal gene therapy trials.

Methods : 18 non-human primates (Macacca fascicularis) underwent 23G pars plana vitrectomy and subretinal injection in three cohorts (high dose: 1x10¹² vector genomes [vg], low dose: 1x10¹¹ vg, or vehicle only). Four additional animals received intravitreal injections to mimic via falsa biodistribution. Tissues samples were harvested at necropsy (day 91) from the treated eye, draining lymph nodes, salivary gland and spleen, optic nerve, brain and spinal cord, heart, lung, liver, adrenal glands and gonads. Blood, urine, lacrimal and nasal swabs were harvested from each animal prior to dosing and 1, 2 and 3 days and 1, 4 and 13 weeks after application of the vector for DNA extraction and quantification of vector genomes by qPCR.

Results : Dose dependent rAAV8 DNA was found in the treated retina. Persistence of rAAV8 in other tissues was most notable after intravitreal injection (via falsa control). Shedding was found in all bio fluids. The highest concentrations were found in lacrimal fluid of the high dose group. DNA was not detected in the germ line tissues and apart from sporadic signals detected in a small number of animals in the liver and lung, all remaining tissues were negative. Concerning the biofluids at day 31 only blood samples showed remaining virus DNA.

Conclusions : These data are relevant for clinical retinal gene therapy trials, where trial subjects, investigators and regulators alike are interested to identify environmental risks associated with application of genetically modified organisms. While shedding into biofluids seems to occur in a dose dependent manner, transduction of off-target organs seems minimal.

This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.