Oncogenic potential of PIK3CD in glioblastoma is exerted through cytoskeletal proteins PAK3 and PLEK2

Abstract

The Class I_A phosphoinositide-3-kinase catalytic isoforms p110α, p110β, and p110δ have been implicated to play vital but overlapping roles in various cancers, including glioblastoma (GBM). We have previously shown that PIK3CD, encoding p110δ, is highly expressed in multiple glioma cell lines and involved in glioma cell migration and invasion. Based on the RNA sequencing data from The Cancer Genome Atlas (TCGA) database, we found the level of PIK3CD expression is significantly higher in GBM than WHO grade II and III gliomas and is closely related to poor survival. To further dissect the oncogenic roles of PIK3CD in glioma progression, we employed CRISPR/Cas9 to completely abrogate its expression in the GBM cell line U87-MG and have successfully isolated two knockout clones with different gene modifications. As expected, the knockout clones exhibited significantly lower migration and invasion capabilities when compared with their parental cells. Interestingly, knockout of PIK3CD also dramatically reduced the colony formation ability of the knockout cells. Further study revealed that PIK3CD deficiency could negate tumorigenesis in nude mice. To determine the downstream effect of PIK3CD depletion, we performed RT² profiler PCR array of selected gene sets and found that knockout of PIK3CD impaired the activity of p-21 activated kinase 3 (PAK3) and pleckstrin 2 (PLEK2), molecules involved in cancer cell migration and proliferation. This explains why the glioma cells without the PIK3CD expression exhibited weaker oncogenic features. Further, RNAseq analysis of parent and knockout clones revealed that this interaction might happen through axonogenesis signaling pathway. Taken together, we demonstrated that PIK3CD could be a potential prognostic factor and therapeutic target for GBM patients.