Spontaneous Retinoblastoma (RB) in Patient Induced Pluripotent Stem Cell (iPSC) Derived Retinal Organoid

Abstract

Purpose : The study of RB is limited to cell culture, genetics knock out models, and patient derived orthotopic xenografts. Each, though useful, has limitations. RBs derived from patient derived iPSC retinal organoids could further our understanding of tumor biology and provide patient specific tumors for preclinical testing. RETCELL (NCT02193724) studied the feasibility, validation and differentiation of iPSCs from patients with heritable RB.

Methods : After IRB approval, iPSCs were created from 15 individuals with germline RB1. Multiple clones form each iPSC line were validated for retention of germline mutations, subjected to whole genome sequencing (WGS) and characterized for neurogenic potential. Representative clones from each patient were then differentiated into retinal organoids, dissociated and injected into eye of immunocompromised mice to monitor for tumor formation.

Results : Of 15 participants, 11 were in active treatment and 4 were parents, of which 2 were unaffected carriers of the RB1 mutation. 4 individuals with no family history were diagnosed < 12 months, 2 had 13q deletions and 9 represented 4 family cohorts. 71 iPSC clones were isolated, and the DNA mutation was verified in each clone. WGS of representative clones and patient germline DNA confirmed no secondary mutations were accumulated during reprogramming. At day 45, retinal organoids from each line were dissociated and injected into the vitreous immunocompromised mice. 3 lines produced spontaneous tumors and 7 produced tumors with CRISPR-Cas9 inactivation of RB1. Tumors showed biallelic inactivation of RB1 with no secondary mutations, upregulation of SYK and were indistinguishable from patient tumors.

Conclusions : Taken together, these data suggest that spontaneous retinoblastoma can form from iPSCs differentiated into retinal organoids. This modeling provides a new methodology to study retinoblastoma development and to develop patient specific target therapies. </font></p>

This is a 2020 ARVO Annual Meeting abstract.