HGG-20. NEURONAL ACTIVITY PROMOTES PEDIATRIC GLIOMA INVASION

The diffusely infiltrating nature of pediatric high-grade glioma (pHGG) is a characteristic hallmark of this disease and represents a major contributor to the dire prognosis seen for pHGG of all neuroanatomical locations, most notably in H3K27M-mutant diffuse midline gliomas such as DIPG. Despite the fact that widespread infiltration is a defining aspect of this group of diseases, the molecular and cellular determinants of pHGG migration and invasion are not well understood. We have previously shown that neuronal activity regulates pHGG growth. Here, we investigated the role of neuronal activity on glioma invasion using conditioned media (CM) generated from optogenetically stimulated, spontaneously active or TTX-silenced acute cortical slices. RNA-seq data revealed gene expression changes in cortical pHGG cells exposed to active CM, with an increased expression of several known glioma migratory/invasive factors including AQP4, TTYH1 and CHI3L1. We observed a marked increase in the invasive capacity of pHGG and DIPG patient-derived cultures upon the addition of active CM in both transwell and three-dimensional (3D) in vitro invasion assays. Analysis of several candidate proteins identified via proteomic screens highlighted two secreted factors, GRP78 and CRTAC1, which alone were sufficient to increase pHGG invasion. Abrogation of this pro-infiltrative effect was seen upon immunodepletion of GRP78 from the CM. Conversely CRISPR knockout of the Nogo receptor (NgR), on which CRTAC1 acts as an exogenous antagonist, markedly increased in vitro DIPG cell invasion. Both CRTAC1 and GRP78 have been shown to modulate RhoA activity in putatively complimentary ways, and we observed a significant reduction in RhoA activation upon exposure to active CM. As Rho-ROCK signalling modulates actin reorganisation and cell motility, this highlights a novel means by which the neural microenvironment drives glioma progression and potentially identifies a new set of therapeutic targets to limit glioma spread.