Endothelins influence the proliferation-migration balance of IDH1-mutant glioma cells and promote proneural to mesenchymal transition

Adult diffuse gliomas are the deadliest brain tumours including IDH-wildtype glioblastomas of worst prognosis and diffuse low grade IDH-mutant astrocytomas and oligodendrogliomas. These glial tumours display distinct tumoral cell population defeating current therapies. Our group has unveiled the role of NOTCH signalling in glioblastoma cell plasticity and in the conversion of oligodendrocytic-like to astrocytic-like tumoral cells in IDH-mutant low-grade gliomas which escalate inevitably to higher grade malignant gliomas. To gain insight into signalling pathways regulating glioma cell plasticity and malignancy, we focused our work on endothelin signalling including endothelin peptide ligands (ET-1, ET-2, ET-3) binding to G-protein coupled endothelin receptors A and B (EDNRA, EDNRB). Here, using glioma patient samples and glioma patient-derived cell lines, we showed that endothelin reduces glioma cell proliferation while increasing migration initiating a proneural to mesenchymal transition. Mechanistically, EDNRB activation led to IP3-dependent calcium mobilization, apamin-sensitive KCNN2/KCNN3 potassium currents and phosphorylation of ERK1/2 and STAT3 in glioma cells. Finally, we studied endothelin receptor regulation by tumoral microenvironment stimuli highlighting a role for EDNRA induced by NOTCH and hypoxia in perivascular hypoxic area in glioblastoma. Altogether, this study demonstrates endothelin signalling as a key player in mesenchymal transformation of diffuse IDH-mutant gliomas and glioblastomas. For RNA sequencing experiments upon endothelin, LGG275, LGG336, BT237, Gb4, Gb7, Gb21 were seeded at 260,000 cells/wells in 3.5 mL of +GFs medium in 6-well plates (#140685, ThermoFisher Nunclon). Cells were treated either with endothelin dilution buffer (PBS with BSA 0.1%) for control condition or with ET-1 at 10 nM final once a day for 5 days before proceeding to RNA extraction 8 hours after the last treatment.