An engineered glioblastoma model yields macrophage-secreted drivers of invasion

Glioblastomas (GBMs) are highly invasive brain tumors replete with brain- and blood-derived macrophages, collectively known as tumor-associated macrophages (TAMs). Targeting TAMs has been proposed as a therapeutic strategy but has thus far yielded limited clinical success in slowing GBM progression, due in part to an incomplete understanding of TAM function in GBM. Here, by using an engineered hyaluronic acid-based 3D invasion platform, patient-derived GBM cells, and multi-omics analysis of GBM tumor microenvironments, we show that M2-polarized macrophages stimulate GBM stem cell (GSC) mesenchymal transition and invasion. We identify TAM-derived transforming growth factor beta induced (TGFßI/BIGH3) as a pro-tumorigenic factor in the GBM microenvironment. In GBM patients, BIGH3 mRNA expression correlates with poor patient prognosis and is highest in the most aggressive GBM molecular subtype. Inhibiting TAM-derived BIGH3 signaling with a blocking antibody or small molecule inhibitor suppresses GSC invasion. Our work highlights the utility of 3D in vitro tumor microenvironment platforms to investigate TAM-cancer cell crosstalk and offers new insights into TAM function to guide novel TAM-targeting therapies. Overall design: To investigate how M2-polarized THP-1-derived macrophage conditioned media (CM) alters the transcriptional profile of GSCs, we performed bulk RNA-sequencing (RNA-seq) on GSCs in hydrogels cultured in GSC maintenance (control) media or M2 CM. We utilized a previously developed microchannel invasion platform, which consists of a cylindrical cell reservoir channel embedded within a 3D hydrogel and enables the physical separation of highly invasive and non-invasive (core) cell fractions from a single device. We then performed Differentiaion Gene Expression Analysis between three samples (M2 CM invasive, M2 CM core, Control) collected at the same timepoint (28 day after start of invasion assay). Comparative Gene Expression profiling analysis of RNA-seq data for GSC-20 cells exposed to M2-polarized macrophage CM or Control media. Cells treated with M2 CM were further separated based on distance migrated in 3D invasion device (core = no invasion, invasive = invasive fraction).