Abrogation of PIK3CA or PIK3R1 reduces proliferation, migration, and invasion in glioblastoma multiforme cells

Glioblastoma multiforme (GBM) is the most common primary brain malignancy, with a current median survival of approximately 15 months in patients with newly diagnosed disease following treatment with surgery, chemotherapy and radiotherapy. The invasive nature of GBM has been frequently implicated as a key feature of the tumor's resistance to therapy. Targeting pathways involved in the invasive phenotype of the tumor could be an important component of successful treatment in combination with strategies that more directly target the cancer cell.;The phosphatidylinositol 3-kinase (PI3K) signaling cascade is an important pathway known to be involved in proliferation, invasion, and migration in GBM and other cancers. Class IA PI3Ks are heterodimers of a p85 regulatory subunit and a p110 catalytic subunit. Three catalytic subunits exist, and are designated p110alpha, p110beta, and p110delta. Five class IA PI3K regulatory subunits include p85alpha, p85beta, p50alpha, p55alpha, and p55gamma. Class IA PI3Ks localize to the plasma membrane via binding of Src-homology 2 (SH2) domains on the regulatory subunit to activated tyrosine kinase receptors. At the plasma membrane, the catalytic subunit phosphorylates phosphatidylinositol 4,5-bisphosphate (PIP2), converting it to phosphatidylinositol 3,4,5-triphosphate (PIP3). PIP3 can activate several downstream signaling cascades, including the Akt and mTOR pathways, which are involved in proliferation and cell survival. Both the class IA catalytic PIK3CA gene encoding p110alpha and the regulatory PIK3R1 gene encoding p85alpha, p50alpha, and p55alpha have been found to be mutated in GBM. While less is known about the role of PIK3R1 in GBM, mutations in PIK3CA have been shown to constitutively activate p110alpha, and generally result in increased Akt pathway activation and tumor progression.;Our hypothesis for this research was that there are common mutations that activate a defined set of pathways that are responsible for the invasive properties in GBM cells. We performed in-depth informatics analyses of invasion and migration-related pathways that are enriched with mutations in GBM. These analyses made use of the Ingenuity Pathway Analysis, Partek Genomics Suite, and Spotfire DecisionSite software. With the recent availability of a well defined set of mutations responsible for GBM, we were able to determine the likely mutational basis responsible for at least part of GBM's invasive phenotype. We mapped many GBM mutations to pathways that may influence the invasive or migratory phenotype of this tumor, including focal adhesion signaling, integrin signaling, and extracellular component and receptor signaling. We showed that the PI3K signaling cascade is involved in each of these pathways and is likely to play an important role in both the proliferative and migratory/invasive phenotypes observed in GBM cells. shRNA-mediated knockdown of the catalytic PIK3CA or regulatory PIK3R1 gene resulted in reduced proliferation, migration, and invasion in GBM cell lines, and in the case of the PIK3R1 gene these effects are independent of the Akt pathway. Both PIK3CA and PIK3R1 knockdown decreased focal adhesion kinase (FAK) activity. PIK3R1 knockdown reduced MMP2 activity while PIK3CA knockdown did not. Our studies suggest for the first time that PIK3R1 may play an important role in the invasive and migratory capabilities of GBM cells, and that the p85alpha product of this gene may be an effective anti-invasion target.