Use of Induced Pluripotent Stem Cell Models to Elucidate Retinal Disease Pathogenesis and to Develop Gene-based Therapies

Choroideremia (CHM) is a rare monogenic, X-linked recessive inherited retinal degenerative disease caused by mutations in the Rab Escort Protein-1 (REP1) encoding CHM gene. CHM is characterized by childhood-onset night blindness (nyctalopia), progressive peripheral vision loss due to the degeneration of neural retina, RPE and choroid in a peripheral-to-central fashion. Most of CHM mutations are loss-of-function mutations leading to the complete lacking of REP1 protein. However, the primary retinal cell type leading to CHM and molecular mechanism remains unknown in addition to the fact of lacking proper disease models. In this study, we explored the utility of induced pluripotent stem cell-derived models of retinal pigment epithelium (iPSC-RPE) to study disease pathogenesis and a potential gene-based intervention in four different genetically distinct forms of CHM. A number of abnormal cell biologic, biochemical, and physiologic functions were identified in the CHM patient cells. Transduction efficiency testing using 11 recombinant adeno-associated virus (AAV) serotype 1-9, 7m8 and 8b showed a differential cell tropism on iPSC and iPSC-derived RPE. We identified AAV7m8 to be optimal for both delivering transgenes to iPSC-RPEs as well as to appropriate target cells (RPE cells and rod photoreceptors) in the primate retina. To establish the proof of concept of AAV7m8 mediated CHM gene therapy, we developed a AAV7m8.hCHM viral vector, which delivers the human CHM cDNA under control of CMV-enhanced chicken β-actin promoter (CβA). Delivery of AAV7m8.CMV.CβA.hCHM to CHM iPSC-RPEs restored protein prenylation, trafficking and phagocytosis defects. The results confirm that AAV-mediated delivery of the REP1-encoding gene can rescue defects in CHM iPSC-RPE regardless of the type of disease-causing mutation. The results also extend our understanding of mechanisms involved in the pathophysiology of choroideremia