Glioblastoma (GBM) is a lethal disease for which there is no known cure. Following ~2 years of initial treatment, which includes surgical resection, radiation, and chemotherapy, more than 90% of GBM patients succumb to disease progression. Inhibitors of phosphatidylinositol-4,5-bisphosphate 3-kinase (PI3K) have been used clinically to treat original and recurrent GBMs with modest benefits. Our recent research finds that PIK3CB (PI3K catalytic subunit ?, encoding p110?), but not other PI3K catalytic subunits, shows a strong association with GBM progression. Moreover, depletion or pharmaceutical inhibition of p110? induces growth inhibition/cell death in GBM cells highly expressing this subunit. In contrast, blocking other PI3K catalytic subunits fails to do so. Hence, PIK3CB/p110? is a selective survival factor for GBM. Our results strongly support that targeting one PI3K isoform that is dominant in GBM may be a more effective approach to treat GBM. While past research on PI3K isoforms has identified PI3K isoform-selective inhibitors, the clinical benefits of these chemical compounds are limited. The lack of molecular details pertaining to p110? selective activation and structural information of native p110 complexes likely contributes to the poor outcomes of current therapies. Understanding the molecular/structural details of p110? will permit a better design of more selective and effective p110?-based therapies for GBM. To this end, we will complete the following two specific aims.
In aim 1, we will acquire high-resolution 3D conformations of p110? native protein complexes using immuno-capture cryoEM. Resolution of cryo-EM images we acquired previously was not high enough to provide 3D conformations of p110?/p85 complexes at a greater detail. Access to a FEI Titan Krios G2 electron microscope has rendered high resolution 3D conformations of native p110?/p85 complexes possible. To acquire more clinically relevant protein structures, primary GBM xenograts derived from patient specimens will be used.
In aim 2, we will test the hypothesis that p110?C2in changes 3D conformations of p110? native protein complexes, thus inactivating p110?. Primary GBM cells will be treated with p110?C2in or a control scramble peptide. 3D conformations of p110? native complexes will be revealed by immuno-capture cryo-EM. Structural differences between p110? native complexes with or without p110?C2in will then be determined. Results from this R21 application will be highly impactful, particularly to our future research in PI3K signaling and on the therapeutic intervention for GBM. The molecular details of p110? native complexes revealed by cryo-EM will encourage us to further our understanding of molecular mechanisms underlying selective activation of p110? in GBM and to identify novel vulnerabilities of p110? at atomic levels. Visualization of p110? native complexes with or without p110?C2in will not only help us understand how this peptide acts but also will facilitate a further development of p110?C2in into a GBM treatment.
Patients with deadly glioblastoma typically live for an average of only one year after diagnosis before another tumor inevitably occurs. We have discovered that one isoform of PI3K (phosphatidylinositol-4,5-bisphosphate 3- kinase) gene family, but not other PI3K isoforms, plays a critical role in glioblastoma disease progression. Therefore, elucidating the molecular/structural basis of this specific PI3K isoform in cellular signaling will not only gain in-depth understanding of PI3K in glioblastoma, but it will also help us design new and effective PI3K isoform-selective therapies to halt progression of this deadly disease.
