Transcriptome-wide analysis of CRISPR-edited iPSC-derived glioma models

CRISPR-edited human induced pluripotent stem cell (iPSC)-derived glioma models provide a robust platform to investigate the biology of these aggressive tumors in an isogenic background or to test possible treatments in preclinical settings. We created iPSC12 lines that carry different combinations of glioma driver mutations using CRISPR/Cas9 genome editing technology, including TP53-/-, ATRX-/-, IDH1-R132H/WT, PTEN-/-, CDKN2A/B-/-, TERT promoter(TERTp) C228T/WT, MTAP-/-, and overexpression (OE) of EGFRvIII oncogenic isoform. Edited iPSCs were then differentiated into neural progenitor cells (NPC) using a small molecule protocol. We cultured the edited NPCs in 3D organoid with an FBS-containing differentiation medium. After 14 days, different genotypes of NPCs formed organoids were harvest and RNAs were isolated and subjected to RNA-seq analysis. To create an in vivo model system, NPCs with different genotypes were intracranially transplanted into athymic mice. Mice were sacrificed when pathologic symptoms developed resulting from tumor burden or 120 days post brain transplantation. RNAs were extracted from the tumor region of brain tissue sections and subjected to RNA-seq analysis. Our study showed that the iPSC-based model of gliomas displayed distinct mutation-dependent variation in transcriptome, which recapitulated the gene expression signatures of human gliomas. Overall design: Gene expression and alternative splicing profiling analysis of CRISPR-edited iPSC-derived glioma models in vitro and in vivo. Samples include in vitro organoids of NPCs with genotypes of T (TP53-/-), TA (TP53-/-, ATRX-/-), TI (TP53-/-, IDH1-R132H/WT), TIA (TP53-/-, IDH1-R132H/WT, ATRX-/-), PCT (PTEN-/-, CDKN2A/B-/-, TERTp-C228T/WT), PCTM (PTEN-/-, CDKN2A/B-/-, TERTp-C228T/WT, MTAP-/-), PCTE (PTEN-/-, CDKN2A/B-/-, TERTp-C228T/WT, EGFRvIII-OE), and PCTME (PTEN-/-, CDKN2A/B-/-, TERTp-C228T/WT, MTAP-/-, EGFRvIII), as well as in vivo xenografts (triplicates of each genotypes) from NPC-TI, TIA, PCTE, and PCTME.