Method Profile
Track live/dead fluorescence over time to separate compound-induced death from growth arrest in patient-derived glioma spheroids.
Screen extracellular-ligand modulation of glioma drug-induced death responses and test mechanistic perturbations.
Low-passage patient-derived glioblastoma (GBM) gliomaspheres and diffuse intrinsic pontine glioma (DIPG) spheroids with nuclear mKate2 expression.
Ultra-low-attachment 384-well spheroids; ligand pretreatment followed by compound dose series; imaging through five days in the screen.
Live/dead reporters
Nuclear mKate2 and SYTOX Green with spheroid imaging and a combined death score.
Comparative assays
ATP-based CellTiter-Glo 3D, trypan-blue death measurements, pathway and genetic perturbations.
Death responses are integrated across doses and time against matched ligand-free BSA controls. ATP-based viability is assessed separately.
Compare HADES with ATP/trypan-blue assays, then profile 50 ligands with 13 compounds and validate selected responses in additional models.
Main ligand screen in GS025 patient-derived GBM spheroids; additional GBM lines and three named DIPG models used for selected validation.
Ligand-free BSA controls, BMP3, pharmacological/genetic pathway controls, and distinct assay readouts.
HADES resolved compound-death differences missed by ATP readouts. BMP2/4 reduced selected drug lethality while increasing lipid-dependent-necrosis sensitivity.
An imaging assay separates growth arrest from death in glioma spheroids. BMP signaling protected against selected alkylating agents and kinase inhibitors while increasing sensitivity to lipid-dependent necrosis, including tegavivint-induced death.
