Supplementary Figures S1-S26 from Screening for Tumor Microtube–Targeting Drugs Identifies PKC Modulators as Multipotent Inhibitors of Glioblastoma Progression

Supplementary Figures, Supplementary Figure Legends, and Supplementary Video Legends. Supplementary Figure S1: Stemness markers and in-vivo-like multicellular networks in the 2DTM growth assay. Supplementary Figure S2: Tumor cells do not form TMs and do not functionally interconnect under classical neurosphere conditions. Supplementary Figure S3: Development of a 2D morphometry pipeline for TM detection and quantification using ilastik software. Supplementary Figure S4: Ground truth TM annotations of S24, BG5 and T269 GBSCs cultured in 2DTM assay conditions. Supplementary Figure S5: Ground truth and Ilastik-based quantification of tumor cells and cell death via live-cell imaging. Supplementary Figure S6: Lead identification strategy for anti-TM drug classes. Supplementary Figure S7: Morphological disruption of BG5 and T269 GBSC TM-networks by PKC activators. Supplementary Figure S8: Basal and phosphorylation levels of PKC. Supplementary Figure S9: PKC activators disrupt TM network functionality. Supplementary Figure S10: AIVIA-based 3D Morphometry Pipeline. Supplementary Figure S11: Cell networks in a human glioblastoma-inbrain-spheroid (hGliCS) model. Supplementary Figure S12: Consequences of TPPB and PMA exposition in a human brain spheroid model. Supplementary Figure S13: Radioresistance of TM-connected vs TM-unconnected glioblastoma cells in an orthotopic mouse model. Supplementary Figure S14: In vivo drug testing pipeline. Supplementary Figure S15: In vivo evaluation of dynamic antitumor and anti-TM effects of short-term TPPB plus radiotherapy in the tumor cores. Supplementary Figure S16: In vivo antitumor and anti-TM effects in the tumor cores of individual mice having received combined radiotherapy and short-term TPPB administration. Supplementary Figure S17: Lunaphore COMETTM-based Ku80, MKI67 and cleaved Caspase-3 signal recognition and quantification in mice brains. Supplementary Figure S18: In vivo antitumor and anti-TM effects in the mouse cohort having received combined radiotherapy and long-term TPPB administration. Supplementary Figure S19: In vivo antitumor and anti-TM effects in individual mice having received combined radiotherapy and long-term TPPB administration. Supplementary Figure S20: Basal and phosphorylation levels of the known TM network drivers Cx43 and GAP43. Supplementary Figure S21: Global changes in gene expression after PKC activators treatments. Supplementary Figure S22: Gene expression changes per type of PKC modulating drug and GBSC. Supplementary Figure S23: Effects of the PKC activators TPPB and PMA on TTYH1, CHI3L1 and GAP43 RNA expression in GBSCs. Supplementary Figure S24: Anti-TM effects of TPPB on patient tumoroids. Supplementary Figure S25: Lunaphore COMETTM-based TTYH1 RNA recognition and quantification in mice brains. Supplementary Figure S26: Graphical summary. Multiple anti-tumor effects of TPPB in Glioblastoma.