Csig-29. Investigating the Role of Cdkn2b in Glioblastoma

Despite extensive genomic characterization of glioblastoma (GBM), current treatments are ineffective. Therefore, we treated a panel of 78 glioma lines with ~400 different compounds to identify potential biomarkers for effective treatments. We found that both TP53 mutation and loss of the CDKN2A/B locus predicted response to CHK1/2 inhibitors. Deletion of both CDKN2A and CDKN2B occur together in ~58% of GBMs due to their close genomic locations. And all of these factors are known regulators of the cell cycle, with CDKN2A and CDKN2B regulating the G1/S cell cycle checkpoint, CHK1 and CHK2 regulating the G2/M checkpoint, and TP53 regulating both checkpoints. We hypothesized that TP53 mutation and CDKN2A/B loss sensitized cells to CHK1/2 inhibition through complete loss of cell cycle control. Supporting this hypothesis, CDKN2A/B-null U87 cells were more sensitive to G2/M inhibition after TP53 knockout. However, only depleting CDKN2A did not sensitize TP53-null M059K GBM cells to G2/M checkpoint inhibition. Therefore, we wondered whether CDKN2B provided residual cell cycle control, reducing dependence on the G2/M checkpoint. Indeed, parental M059K cells with TP53 mutations expressed CDKN2A with only a negligible amount of CDKN2B, and CDKN2B protein levels increased substantially only upon CDKN2A knockout. We generated double knockouts of CDKN2A and CDKN2B in M059K cells and obtained modest but significant increases in sensitivity to both the CHK1/2 inhibitor Prexasertib and the WEE1 inhibitor Adavosertib, another inhibitor of the G2/M checkpoint. This demonstrated that the additional loss of CDKN2B along with CDKN2A loss and TP53 mutation was necessary to sensitize these cells to G2/M checkpoint inhibition. This work suggests that inactivation of both CDKN2A and CDKN2B is necessary to accurately model this homozygous deletion.