LINEAGE REPROGRAMMING OF HUMAN GLIOBLASTOMA CELLS INTO NEURONS USING PATIENT-DERIVED TUMOR ORGANOIDS

Glioblastoma (GBM) is the most common and lethal primary brain tumor in adults. Treatment failures are explained by a high degree of interpatient and intratumoral cellular and transcriptional heterogeneity. Malignant cells exist in multiple cell states showing dynamic transitions between states implying substantial plasticity involved in tumor initiation and progression, including therapy resistance and relapse. Since neurons are post‑mitotic cells, redirecting glioblastoma cells towards a neuronal identity could durably alter proliferative capacity and phenotypic plasticity of tumour cells. Here, we show that virally-driven expression of the neurogenic transcription factors (TF) Neurog2 or Ascl1/Dlx2 in 2D cultures derived from patient tumours (n=3 patients) and kept at low passages, induces their efficient conversion into induced neurons (iNs) characterized by rapid acquisition of neuronal morphology and expression of the immature neuronal marker DCX (80% of transduced cells). Over time, most iNs mature, as illustrated by expression of the mature marker NeuN. Importantly, EdU proliferation assays reveal that most iNs exit the cell cycle between one and two weeks after TF induction. In parallel, using 3D patient-derived tumor organoids, that faithfully recapitulate cell heterogeneity, DNA mutations and copy number variations of parental tumors, we show that a significant number of malignant cells can be converted into iNs, albeit with reduced reprogramming efficiency compared to 2D cultures. Together, our results indicate that neuronal reprogramming can divert primary GBM cells from their oncogenic trajectory and reduce their proliferation, thereby opening a promising therapeutic avenue for GBM patients.