AAV-Mediated Gene Delivery to Foveal Cones.

Abstract

Adeno-associated virus (AAV) has emerged as the vector of choice for delivering genes to the mammalian retina. From the first gene therapy to receive FDA approval for the inherited retinal disease (Luxturna™) to more recent clinical trials using microbial opsins to regain light sensitivity, therapeutic transgenes rely on AAV vectors for safe and efficient gene delivery to retinal cells. Such vectors are administered to the retina via subretinal (SR) injection or intravitreal (IVT) injection routes depending on the targeted retinal cell type. An attractive target for gene therapy is the fovea, bearing the highest concentration of cone cells responsible for our high acuity daylight vision. However, previous clinical trials and large animal studies reported that SR administration of vector under the cone-exclusive fovea disrupts its fine structure and might impair visual acuity. Due to its technical difficulty and potential risks, alternatives to vector injection under this delicate region have been investigated by using novel AAV capsid variants identified via rational design or directed evolution. We recently established new vector-promoter combinations to overcome the limitations associated with AAV-mediated cone transduction in the fovea. Our methods provide efficient foveal cone transduction without detaching this delicate region and rely on the use of engineered AAVs and optimal promoters compatible with optogenetic vision restoration. Here we describe in detail our AAV vectors, methods for intravitreal and subretinal injections as well as pre- and postoperative procedures as performed in cynomolgus macaques.