Adeno-Associated Virus Mediated Gene Therapy for Retinal Degenerative Diseases

The rhodopsin crystal structure provides a structural basis for understanding the function of this and other G protein-coupled receptors (GPCRs). The major structural motifs observed for rhodopsin are expected to carry over to other GPCRs, and the mechanism of transformation of the receptor from inactive to active forms is thus likely conserved. Moreover, the high expression level of rhodopsin in the retina, its specific localization in the internal disks of the photoreceptor structures [termed rod outer segments (ROS)], and the lack of other highly abundant membrane proteins allow rhodopsin to be examined in the native disk membranes by a number of methods. The results of these investigations provide evidence of the propensity of rhodopsin and, most likely, other GPCRs to dimerize, a property that may be pertinent to their function.

The safety and efficacy of gene therapy for inherited retinal diseases is being tested in humans affected with Leber's congenital amaurosis (LCA), an autosomal recessive blinding disease. Three independent studies have provided evidence that the subretinal administration of adeno-associated viral (AAV) vectors encoding RPE65 in patients affected with LCA2 due to mutations in the RPE65 gene, is safe and, in some cases, results in efficacy. We evaluated the long-term safety and efficacy (global effects on retinal/visual function) resulting from subretinal administration of AAV2-hRPE65v2. Both the safety and the efficacy noted at early timepoints persist through at least 1.5 years after injection in the three LCA2 patients enrolled in the low dose cohort of our trial. A transient rise in neutralizing antibodies to AAV capsid was observed but there was no humoral response to RPE65 protein. The persistence of functional amelioration suggests that AAV-mediated gene transfer to the human retina does not elicit immunological responses which cause significant loss of transduced cells. The persistence of physiologic effect supports the possibility that gene therapy may influence LCA2 disease progression. The safety of the intervention and the stability of the improvement in visual and retinal function in these subjects support the use of AAV-mediated gene augmentation therapy for treatment of inherited retinal diseases.

Genes associated with inherited retinal degeneration have been found to encode proteins required for phototransduction, metabolism, or structural support of photoreceptors. Here we show that mutations in a novel photoreceptor-specific homeodomain transcription factor gene (CRX) cause an autosomal dominant form of cone-rod dystrophy (adCRD) at the CORD2 locus on chromosome 19q13. In affected members of a CORD2-linked family, the highly conserved glutamic acid at the first position of the recognition helix is replaced by alanine (E80A). In another CRD family, a 1 bp deletion (E168 [delta1 bp]) within a novel sequence, the WSP motif, predicts truncation of the C-terminal 132 residues of CRX. Mutations in the CRX gene cause adCRD either by haploinsufficiency or by a dominant negative effect and demonstrate that CRX is essential for the maintenance of mammalian photoreceptors.