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152. Retinal Tropism of Exosome-Associated AAV Vector Via Intravitreal Delivery

2015; Elsevier BV; Volume: 23; Linguagem: Inglês

10.1016/s1525-0016(16)33757-1

1525-0024

Lívia S. Carvalho, Bence György, Dakai Mu, Samiksha Shah, Luk H. Vandenberghe, Casey A. Maguire,

Extracellular vesicles in disease

Recent clinical studies have validated the use of adeno-associated viral vectors (AAV) as a safe and efficient gene delivery vehicle for the treatment of retinal disorders. Different studies targeting the retina using AAV vectors have been achieved transduction of retinal cells via one of two delivery routes: subretinal or intravitreal injections, with subretinal delivery by far the most efficient method. Current treatments for age-related macular degeneration (AMD) heavily rely on protein drug delivery via regular and repeated intravitreal injections making the current standard of care clearly not optimal. One alternative for AMD treatment would be to use an AAV-based gene delivery and indeed preclinical studies in both mouse and non-human primates have shown that intravitreal delivery of AAV2 can mediate sustained expression of transgenes in the vitreous cavity following a single injection. Since the DNA is encapsulated in the AAV capsid, it is the capsid that determines the majority of the pharmacological interactions with the host, including cellular specificity, binding and uptake, immune activation and stability. However intravitreal injections using AAV vectors are inefficient regarding expression levels and poses higher inflammatory risk to vector or transgene antigens. In the current study we evaluated the retinal tropism of exosome-associated AAV9 vector (vexosomes). Exosomes are naturally secreted membrane nanovesicles involved in intercellular communication. We have previously shown that vexosomes highly outperforms regular AAV in efficacy in vitro and in vivo, and are resistant to neutralizing antibodies due to shielding of the AAV by the exosomal membrane. AAV9-vexosomes were isolated from AAV-producer cell (293T) media by ultracentrifugation. AAV vexosomes were injected into C57bl/6 wild type mice and transgene expression was evaluated at 2 and 4 weeks post injections via in vivo fundus imaging. Eyes were collected at 4 weeks post injections and retinal cell tropism was analyzed. Comparison of transgene expression levels between the different vexosomes and traditionally formulated AAV vectors was done using real time PCR. Intravitreal delivery of AAV9-vexosomes showed increased level of retinal cells transduction compared to regular AAV9 vectors which show very poor transduction. Apparently, vexosomes were able to penetrate through the ILM and transduce the retina in patches, as assessed by fundus imaging. Due to the immune issues surrounding gene delivery in the eye via an intravitreal route, the results presented here indicate a promising alternative gene delivery platform using vexosomes that could circumvent the inflammatory risks expected by intravitreal injection. Further studies in assessing the extent vexosomes evade neutralization in the vitreous will be extremely important to complement the results presented here.