Glioblastoma: multifaceted immunosuppression mediated through galectin family members

Abstract

Glioblastoma (GBM) is an aggressive brain cancer with a near-uniformly lethal prognosis. Anti-tumoural immunity remains low due to a highly suppressive tumour microenvironment. This project set out to identify immunomodulatory proteins secreted by glioblastoma, describe the distribution of intratumoural immune cells, and characterise the effects of secreted immunomodulatory on adaptive immune effector cells. Mass spectrometry data identified galectin family members in the glioblastoma secretome. ELISA assays confirmed galectin presence in GBM-derived liquid samples; immunofluorescence confirmed their presence in tumour sections, as well as the presence of T-cells, B-cells and macrophages. Changes in cell surface marker expression and phagocytosis following exposure to recombinant proteins were assayed using flow cytometry and pHrodo-conjugated Escherichia coli respectively. B-cells and macrophages were predominantly found in tumour bulk (p=0.0053 and 0.0087) with T-cells located in the perivascular niche (p=0.0932). Additionally, 22% of tumours contained T-cell aggregations. Galectins 1, 3, 4 and 7 were also found in the majority of GBM assayed; immune checkpoints galectin-9 and PD-L1 were found in 100% and 50% of assayed tumours, respectively. The perivascular location of galectin-1 approached significance (p=0.0844), whereas other galectins showed a more equitable distribution. Consistent, though insignificant, downregulation of MHCII and phagocytosis was observed in pro-inflammatory adult macrophages exposed to GBM-relevant galectin-1 concentrations. These results suggested a T-cell-specific mechanism of suppression resulting in reduced infiltration, whereas B-cells and macrophages were able to effectively infiltrate GBM. Antigen presentation by macrophages may be inhibited by perivascular galectin-1, thus limiting T-cell retention; encounter with known immunosuppressive proteins such as galectins 1, 3, 9 and PD-L1 following effector extravasation may be a key mechanism through which GBM immunosuppression is mediated. These results posit the perivascular region as a uniquely immunosuppressive environment, and the presence of galectins as a potential key mediator of immunosuppression within glioblastoma.