Developing Strategies to Target Glioblastoma Stemness and Immunosuppression

Brain tumor-initiating cells (BTICs) drive tumor progression, immunosuppression, and resistance to treatments, posing formidable challenges to advancing effective treatments against glioblastoma (GBM). In this dissertation, we demonstrate that clemastine, an over-the-counter drug for treating hay fever and allergy symptoms, effectively attenuated the stemness and suppressed the propagation of primary BTIC cultures bearing PDGFRA amplification. These effects on BTICs were accompanied by altered gene expression profiling indicative of their more differentiated states, resonating with the activity of clemastine in promoting the differentiation of normal oligodendrocyte progenitor cells (OPCs) into mature oligodendrocytes. Functional assays for pharmacological targets of clemastine revealed that the Emopamil Binding Protein (EBP), an enzyme in the cholesterol biosynthesis pathway, is a target that mediates the suppressive effects of clemastine. Consistently, we showed that a neural stem cell-derived mouse glioma model displaying predominantly proneural features was similarly susceptible to clemastine treatment in vitro and in vivo. Surprisingly, we discovered that EBP protein is essential for BTIC propagation and stemness properties, and revealed a potential lipid-independent function of EBP in regulating epigenetic programming. Collectively, this original work identifies pathways indispensible for maintaining the stemness and progenitor features of GBMs, uncovers BTIC dependency on EBP, and suggests that non-oncology, low-toxicity drugs with OPC differentiation-promoting activity can be repurposed to target GBM stemness and aid in their treatment.Another key strategy extensively pursued for treating GBMs focuses on targeting endolysosomes, mainly on the basis that the intact function of these subcellular organelles is crucial for tumor cell autophagy and survival. Through gene expression analyses and cell type abundance estimation in GBMs, we showed that genes associated with the endolysosomal machinery are more prominently featured in non-tumor cells in GBMs than in the tumor cells themselves, and that tumor-associated macrophages represent the primary immune cell type that contributes to this phenomenon. Further analyses uncovered an enrichment of endolysosomal pathway genes in immunosuppressive and pro-tumorigenic macrophages, such as M2-like macrophages or those associated with worse prognosis in glioma patients, but not in those linked to inflammation and anti-tumorigenic properties. Specifically, genes critical to the hydrolysis function of endolysosomes, including progranulin and cathepsins, were among the most positively correlated with immunosuppressive macrophages, and elevated expression of these genes is associated with worse patient survival in GBMs. Together, these results implicate the hydrolysis function of endolysosomes in shaping the immunosuppressive microenvironment of GBM. We propose that targeting endolysosomes, in addition to its detrimental effects on tumor cells, can be leveraged for modulating immunosuppression to render GBMs more amendable to immunotherapies.