EP190

Immune responses to Adeno-associated virus (AAV) capsids for retinal gene therapy

GEHRKE Miranda, DIEDRICHS-MÖHRING Maria, BOGEDEIN Jacqueline, BÜNING Hildegard, MICHALAKIS Stylianos, WILDNER Gerhild

Purpose

Ocular gene therapy using adeno-associated viral (AAV) vectors has emerged as an approved approach for treating inherited retinal diseases. Despite the ocular immune privilege, the viral vectors often induce uveitis by eliciting granulomatous inflammation. Two AAV2 capsid variants containing small peptides insert in their capsid were designed for improved tropism for photoreceptors even after intravitreal injection, allowing for a dose reduction of therapeutic vectors.

Methods

Here we have investigated the pre-existing immune responses of blood donors without prior gene therapy to AAV2 wildtype and AAV2 capsid variants with respect of antibody binding by ELISA, neutralization by inhibiting transduction of HeLa cells and cellular immune responses like proliferation, evaluated by immunofluorescence staining and FACS analysis and cytokine secretion determined by multiplex bead analysis.

Results

Comparing the three different vectors we found no differences in total antibody binding, but a decreased neutralization of the two variants AAV2.GL and AAV2.NN compared to AAV2 WT. There was no adaptive cellular immune response detected to any of the vectors after in vitro-stimulation of the donor’s PBMS, but secretion of inflammatory cytokines and chemokines mainly produced by innate immune cells like CXCL1, IL-6, CCL3 and VEGF. CD11c+ dendritic cells (DC) and CD14+ monocytes producing IFN-b and CD11c+/CD14+ monocyte-derived DC (moDC) coproducing IL-1b and IFN-b were detected after in vitro stimulation, pointing to a primarily innate response elicited by the viral vectors.

Conclusions

MoDCs recognize bacteria and viruses via their pattern-recognition receptors and are found in chronic granulomas of infectious diseases and thus might be the drivers of intraocular inflammation observed after ocular gene therapy.

Previous
Next