311. AAV8 Vexosome Vectors Enhance Cell Transduction In Vitro and Outperform Conventional AAV8 Vectors in Liver-Transduction In Vivo in the Presence of Anti-AAV Neutralizing Antibodies
Authors: Amine Meliani, Zachary Fitzpatrick, Severine Charles, Dakai Mu, Florence Boisgerault, Casey A. Maguire, Federico Mingozzi
Journal: Molecular Therapy (2015)

Adeno-associated virus vectors based on serotype 8 (AAV8) have demonstrated superior efficiency of transduction of hepatocytes in vivo in animal models and in humans. Recently, we described extracellular vesicle (exosomes) -associated AAV vectors termed vexosomes (vAAV). We tested vAAV1, 2, and 9 and found them to enhance transduction and antibody evasion capabilities compared to conventional AAV vectors. With the goal of developing an enhanced vector for liver gene therapy, we sought to characterize the in vitro and in vivo transduction, biodistribution, and immunogenicity profile of vAAV8 vectors compared to conventional AAV8 vectors purified from the same preparation. Several cell lines were transduced with conventional AAV8 or vAAV8 vectors expressing luciferase, at multiplicity of infection (MOI) ranging from 250 to 25000, and luciferase expression was measured after an overnight incubation. In HeLa and HEK293 cells, levels of luciferase up to 1 log higher were observed with vAAV8 compared with AAV8 vectors, with detectable levels of luciferase at MOIs as low as 250. In hepatocyte cell lines, the magnitude of enhancement was about 3-fold, with lower max relative light unit signal measured compared to the other cell lines. Notably, although at low levels, transduction with vAAV8 vectors was also observed in a Jurkat T cell line, in which no transduction was observed with conventional AAV8 vectors. We then evaluated the efficiency of liver targeting in vivo with this vector system using the secreted factor IX (F.IX) transgene. Conventional or vAAV8 vectors expressing human F.IX under the control of a liver-specific promoter were administered i.v. to C57BL/6J male mice at 5×1010 vg/mouse. In naïve animals, no difference in F.IX transgene expression levels was observed between the two vector types; similarly, vector genome were found at similar levels in liver, spleen, muscle, lungs, heart, and kidneys. In these animals, i.v. vector administration also resulted in identical levels of anti-AAV antibody formation. We next evaluated the effect of vAAV8 on liver transduction in the presence of antibodies. Animals were passively immunized with IVIg intraperitoneally (0.5, 5.0, and 15 mg/mouse) followed by administration of vector (5×1010 vg) intravenously 24 hours later. At a neutralizing antibody titer of~1:10 (0.5mg IVIg), vAAV8 vector-injected mice had levels of expression identical to those in naïve animals. At higher titers (>1:100, 5.0 mg), low levels of residual expression of F.IX were measure with vAAV8 vectors and no expression was measured with conventional AAV. No expression was detected with either AAV8 or vAAV8 vectors in animals immunized with the highest dose of IVIg. In conclusion, vAAV8 vectors have unique transduction characteristics that may be exploited in vitro to transduce various cell types. In vivo, these vectors showed identical levels of liver transduction in naïve male mice and provided some level of protection from antibody-mediated vector neutralization.