Development of an anti-invasive therapeutic strategy for paediatric high grade glioma

Paediatric high grade glioma (pHGG) and diffuse intrinsic pontine glioma (DIPG) are highly aggressive brain tumours. Their invasive phenotype contributes to their limited therapeutic response and novel treatments that block brain tumour invasion are needed. Here, two novel anti-invasive therapeutic strategies are evaluated: glycogen synthase kinase-3 (GSK-3) inhibitors and oncolytic viruses (OVs). Small molecule GSK-3 inhibitors, lithium chloride (LiCl) and the indirubin derivative 6-bromoindirubin-oxime (BIO), reduced migration and invasion of pHGG cell lines (SF188 and KNS42) and one patient-derived DIPG cell line (HSJD-DIPG-007) in 2D and 3D in vitro assays. Following drug treatment, pHGG cells demonstrated loss of polarity and altered morphology as seen by live cell imaging and cytoskeletal rearrangement of actin fibres and focal adhesions as seen by immunofluorescence. OVs (herpes simplex virus (HSV), reovirus and vaccinia virus (VV)) were able to inhibit the migration and invasion of pHGG and DIPG cell lines. Oncolytic HSV was the most interesting candidate, as anti-migratory and anti-invasive effects did not appear to be a consequence of cytotoxicity or altered proliferation. Oncolytic HSV altered pHGG cytoskeletal dynamics, stabilising microtubules through the accumulation of post-translational tubulin modifications, as evaluated by Western blotting and immunofluorescent labelling. Furthermore, oncolytic HSV treatment of pHGG cell lines inhibited GSK-3β activity and prevented the localised clustering of adenomatous polyposis coli to the leading edge of the cell. These observations are highly novel and begin to document the molecular mechanisms by which oncolytic HSV may inhibit pHGG migration and invasion. In conclusion, this study is the first to demonstrate that it is possible to target migration and invasion of pHGG and DIPG in vitro using either small molecule GSK-3 inhibitors or OVs, such as HSV. These agents warrant further in vivo pre-clinical investigation as potential anti-invasive therapeutics and have the potential to improve outcomes of these devastating childhood diseases