Retinoblastoma from Human Stem Cell-Derived Retinal Organoids (RNA-Seq)

Retinoblastoma is a childhood cancer of the developing retina that initiates with biallelic inactivation of the RB1 gene. To develop a laboratory model of human retinoblastoma formation, we made induced pluripotent stem cells (iPSCs) from 15 participants with germline RB1 mutations. After 45 days in culture, the retinal organoids were dissociated and injected into the vitreous of eyes of immunocompromised mice to support retinoblastoma tumor growth. Retinoblastomas formed from retinal organoids made from patient-derived iPSCs had molecular, cellular and genomic features indistinguishable from human retinoblastomas. Overall design: We made induced pluripotent stem cells (iPSCs) from 15 participants with germline RB1 mutations. After 45 days in culture, the retinal organoids were dissociated and injected into the vitreous of eyes of immunocompromised mice to support retinoblastoma tumor growth. We validated tumors by comparing whole genome sequencing, RNA-sequencing, methylation analysis, and single cell sequencing of organoid derived tumors to patient tumors and patient derived xenografts.

视网膜母细胞瘤（Retinoblastoma）是一种发生于发育中视网膜的儿童恶性肿瘤，其起始事件为RB1基因的双等位基因失活。为构建人类视网膜母细胞瘤发生的实验室模型，我们从15名携带生殖系RB1突变的受试者中诱导生成了诱导多能干细胞（induced pluripotent stem cells, iPSCs）。培养45天后，将视网膜类器官解离并注射至免疫缺陷小鼠（immunocompromised mice）的玻璃体腔内，以支持视网膜母细胞瘤的生长。由患者来源iPSCs构建的视网膜类器官所形成的视网膜母细胞瘤，其分子、细胞及基因组特征与人类视网膜母细胞瘤无显著差异。整体实验设计：我们从15名携带生殖系RB1突变的受试者中诱导生成了诱导多能干细胞（iPSCs）。培养45天后，将视网膜类器官解离并注射至免疫缺陷小鼠的玻璃体腔内以支持肿瘤生长。我们通过全基因组测序（whole genome sequencing）、RNA测序（RNA-sequencing）、甲基化分析（methylation analysis）以及类器官来源肿瘤的单细胞测序（single cell sequencing），并将结果与患者肿瘤及患者来源异种移植瘤（patient derived xenografts）进行比对，完成了对肿瘤的验证。