Recreating pathophysiology of CLN2 disease and demonstrating reversion by TPP1 gene therapy in hiPSCs-derived retinal organoid and retina-on-chip

Mutations in tripeptidyl peptidase 1 (TPP1) gene lead to late infantile neuronal ceroid lipofuscinosis CLN2, characterized by lysosomal accumulation of lipofuscins predominantly found in brain and retina. The ocular phenotype is characterized by bilateral outer retinal degeneration that leads to complete vision loss. CLN2 animal models struggle in recapitulating the retinal phenotype observed in patients. Here, we leveraged human induced pluripotent stem cell (hiPSC)-derived retinal organoids (ROs), retinal pigmented epithelial (RPE) cells, and retina-on-chip (RoC) technologies to model CLN2 disease in vitro in patient-specific microphysiological models of the human retina. Using these models, we were able to recreate the major clinical histological hallmark of CLN2 patients, namely the accumulation of lipofuscins containing subunit C of mitochondrial ATP synthase (SCMAS) and lipids mainly in the outer retina. Furthermore, single-cell RNA-sequencing of CLN2 ROs revealed a dysregulation of translational and mitochondrial function in cones. Finally, we demonstrate that an adeno-associated virus (AAV)-mediated TPP1 gene therapy was able to restore TPP1 expression and could decrease and even prevent SCMAS accumulations. In summary, our study produced novel human-relevant microphysiological retinal disease models, uncovered new mechanisms of CLN2 pathophysiology in the human retina, and demonstrated the immense potential of AAV9.hCLN2 gene therapy for CLN2 disease potentially treating and even curing blindness of affected individuals.