Novel platform for optimizing AAV transgene expression to improve efficacy of ocular gene therapies

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Abstract Gene therapy applications based on adeno-associated virus (AAV) have demonstrated promise in clinical applications for the treatment of diverse genetic diseases highlighted by the recent FDA approval of AAV vector formulations as new drugs for ocular and neurological diseases. In these instances, along with all applications of clinical AAV gene therapy to date, constitutive transcription of the therapeutic cassette is employed without any safeguards in place to modulate transgene production in the targeted tissue. At the mechanistic level, the AAV vector transduction pathway is not well understood. The episomal AAV vector genomes form circular monomers and concatemers and a limited number of studies have demonstrated their associations with histones, transcriptional activators and repressors implying an untapped level of control related to overall transgene production. These observations suggest that AAV episomes may be restricted for maximal expression and allude to the ability for targeted approaches for modulating their epigenetic composition to enhance and/or repress AAV vector transduction. We recently developed a technology that couples a dCas9-based protein targeting system at a promoter with a small bifunctional molecule to control gene expression. Spefically, the dCas9-FKBP system was used to successfully activate epigenetically-silenced endogenous loci (e.g., MyoD1 and CXCR4) in HEK293 cells. Epigenos Biosciences proposes adding a new functionality, referred as CEMtrol, to AAV modules that will be designed to mitigate epigenetic dampening of transgene expression to ensure the efficacy of gene therapies will not be subject to uncontrollable epigenetic regulation. In collaboration with Dr. Matthew Hirsch, who is developing therapies for ocular diseases, Phase I will focus on developing an AAV that maximizes transgene expression in cell culture and explant cornea models as a proof-of-concept feasibility study.