Transplantation of human iPSC-derived photoreceptor precursors isolated by targeting of the surface antigen CD73

Abstract

Purpose : For cell therapy involving transplantation of new photoreceptors derived from human induced pluripotent stem (hiPSCs), it is crucial to obtain a homogenous population of transplantation-competent cells and achieve adequate maturation and synaptic connections in the recipient retina. We previously showed that the cell surface antigen CD73 is specifically expressed in photoreceptors in hiPSC-derived retinal organoids. Here, we investigated the integration competence of transplanted CD73+ photoreceptors and their ability to mediate a restoration of visual function.

Methods : Retinal organoids were generated from hiPSCs and cultured for 120 days. CD73+ photoreceptor precursors were isolated by Magnetic Activated Cell Sorting (MACS). Sorting efficacy was verified by flow cytometry. CD73+ cells were injected as cell suspension in the subretinal space of hemizygous P23H rats aged of 1.5 months. In each animal only one eye was transplanted. Retinas were collected at 6 and 10 weeks after and analyzed by immunostaining with photoreceptor-specific and human-specific antibodies. Additionally, fluorescence in situ hybridization (FISH) against human Y chromosome was performed. Visual function was evaluated in both eyes by recording full field electroretinogram (ERG) at 4 and 8 weeks following transplantation.

Results : Targeting of CD73 by MACS led to enrichment to 90% of CD73+ cells in the sorted fraction. CD73+ photoreceptors survived following transplantation and matured in close proximity to host inner retina during several weeks. Transplanted cells corresponded mostly to cones, expressing the cone marker CONE-ARRESTIN. Human cells expressed human-specific cytoplasmic and nuclear markers and displayed distinct morphological features from host rat photoreceptors. FISH signal was also restricted to donor cells, excluding the occurrence of cytoplasmic material exchange between human and rat photoreceptors. ERG recording failed to detect any significant improvement of the visual function in transplanted eyes compared to contralateral non-injected eyes at the time points tested.

Conclusions : Although the ability of donor cells to establish functional synaptic connections and mediate a significant rescue of the visual function remains to be assessed, these data demonstrate that CD73+ photoreceptor precursors hold great promise for the development of a future clinical translation.

This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.