Divergence from the human astrocyte developmental trajectory in glioblastoma
Glioblastoma (GBM) is defined by heterogenous and resilient cell populations that closely reflect neurodevelopmental cell types. While it is clear that GBM echoes early and immature cell states, identifying the specific developmental programs disrupted in these tumors has been hindered by a lack of high-resolution trajectories of glial and neuronal lineages. Here, we delineate the course of human astrocyte maturation to uncover discrete developmental stages and attributes mirrored by GBM. We generated a transcriptomic and epigenomic map of human astrocyte maturation using cortical organoids maintained in culture for nearly two years. Through this approach, we chronicled a multi-phase developmental process orchestrated by a series of transcription factor and gene regulatory networks. Our time course of human astrocyte maturation includes a novel and molecularly distinct intermediate period that serves as a lineage commitment checkpoint upstream of mature quiescence. This intermediate stage acts as a site of developmental deviation separating IDH-wildtype neoplastic astrocyte lineage cells from quiescent astrocyte populations. Interestingly, IDH1-mutant tumor astrocyte lineage cells are the exception to this developmental perturbation, where immature properties are suppressed as a result of D2HG oncometabolite exposure. We propose that this defiance is likely a consequence of IDH1mt-associated epigenetic dysregulation and we identified biased DNA hydroxymethylation (5hmC) in maturation genes as a possible mechanism. Together, this study illustrates a novel cellular state aberration in GBM astrocyte lineage cells and presents new developmental targets for experimental and therapeutic exploration. Overall design: This study consists of multiple sequencing datasets. (1) We purified astrocytes from fresh human cortical organoids and GBM tumor and margin tissue samples for bulk RNA-seq and ATAC-seq. (2) Nuclei were isolated from frozen GBM tumor and margin tissue samples for single-nucleus multiome (ATAC- and RNA-seq) sequencing. (3) We performed MeDIP and hMe-Seal with flash-frozen tissue from IDH1mt and IDHwt tumors (with corresponding margin samples) for 5mC and 5hmC analyses. (4) Bulk RNA-seq data for ASCL1 overexpression experiments and D2HG functional experiments with human fetal astrocytes.