STEM-06. ROLE OF CARBOHYDRATE-BINDING PROTEINS IN CONTROLLING GLIOBLASTOMA STEM CELL FATE AND TUMORIGENESIS

Abstract Galectins (GALs, LGALS) are a family of proteins recognized for their β-galactoside-binding activity, which underlies their diverse intra- and extracellular functions. Some members of the GAL family are associated with tumorigenesis, but their precise role in glioblastoma (GB) development remains insufficiently explored. To elucidate the role of GALs in modulating GB progression, we first conducted an in silico analysis of LGALS1, -3, -8, and -9 expression in GB using TCGA and CCGA datasets. We found that the combination of high expression of these GALs is associated with the poorest prognosis compared to other combinations. The following scRNASeq analysis has revealed that GALs are distributed heterogeneously in GB, where GAL-3 is highly expressed in tumor cells, macrophages, and T-cells. To assess the impact of GAL3 on glioblastoma stem cells (GSCs), we subsequently conducted in vitro experiments using multiple patient-derived lines. Our findings demonstrated that GAL-3 suppression, achieved through siRNA silencing and CRISPR-Cas9, led to a decrease in GSC numbers, which is related to an effect on proliferation but not apoptosis. Pharmacological inhibition with the selective GAL-3 inhibitor GB1107 confirmed these findings. The PI-based cell cycle analysis demonstrated that CRISPR knockout cells may exhibit an accumulation in the G1 phase. Notably, growth factor deficiency resulted in the opposite phenotype of Gal-3 knockout cells, characterized by increased proliferation and a decreased G1-phase cell fraction. We suggest that these effects may be linked to a less effective G1 checkpoint and enhanced DNA damage, as indicated by elevated levels of γH2AX in knockout cells. Finally, temozolomide (TMZ) treatment in GAL-3 knockout cells led to a significantly greater reduction in cell viability compared to treatment in control cells. Our findings indicate that GAL-3 may contribute to GB development by regulating GSC proliferation, cell cycle progression, response to deprivation of growth factors and TMZ treatment.