TMIC-75. Cancer-associated fibroblasts mediate the invasion of glioblastoma stem cells and promote therapeutic resistance

Cancer-associated fibroblasts (CAFs), which regulate the microenvironment of solid tumors, have recently been identified in glioblastoma (GBM), but their impact on the GBM microenvironment and therapeutic response remains largely unexplored. We investigated the impact of CAFs on the pro-tumorigenic niche driving therapeutic resistance. Due to the lack of a single defining surface marker for GBM CAFs, we developed a novel protocol and established the first biobank of six patient-derived GBM-associated CAFs, achieving >90% purity by sorting out PTPRZ1+ tumor cells, CD45+ immune cells and MBP+ oligodendrocytes from serially trypisinized patient GBM samples. Intracranial implantation of GBM43 neurospheres and GBM patient-derived CAFs into athymic mice led to poorer survival compared to mice injected with neurospheres alone. In the presence of CAFs, the response to radiation decreases, with a significant reduction in median survival. In a similar experimental setting, the presence of CAFs also attenuated the efficacy of temozolomide therapy, implicating CAFs in conferring therapeutic resistance. Mechanistic insights have emerged by studying the impact of GBM CAFs on GBM cells growing in biomimetic 3D HA- hydrogels, where CAF conditioned media was found to significantly increase the invasion of several GBM stem cell lines. In parallel, guided by our bioinformatic analyses, we have identified Thy1 as a robust cell surface marker of CAFs in an SB28 murine GBM model, facilitating positive selection of CAFs via flow cytometry and the development of antibody-based depletion strategies. While Thy1 is minimally expressed in a small fraction of T cells, it exhibits a ~6-fold upregulation in GBM CAFs compared to other cellular compartments within the SB28 tumor microenvironment. Intranasal delivery of anti-Thy1 antibody results in ECM depletion enabling us to study the role of CAF driven processes in an immunocompetent mouse model, towards the goal of developing the targeting of GBM CAFs as a novel therapeutic strategy.