Abstract 3089: Evaluation of highly potent and selective single-molecule dual EGFR/PI3K inhibitors in preclinical models of adult and pediatric high-grade glioma
Authors: Sun Yusha Y. , Bailey Cavan P. , Carter Trever R. , Whitehead Christopher E. , Sebolt-Leopold Judith S. , Chandra Joya 
Published: August 2019

High grade gliomas (HGG), including glioblastoma (GBM) and diffuse intrinsic pontine glioma (DIPG), carry poor prognosis with median survival rates under 15 months post diagnosis. Due to dysregulation of the RTK/PI3K pathways within these tumors, including RTK pathway amplifications, targeted kinase inhibition has been considered a promising strategy to improve patient outcomes. However, many single-agent inhibitors of EGFR or PI3K have shown limited response in gliomas due to poor blood-brain barrier (BBB) penetrance and activation of compensatory signaling. The design of BBB-penetrant novel therapeutics that target multiple kinases is crucial for overcoming these obstacles. Single-molecule multikinase inhibitors may decrease resistance, present a single pharmacokinetic dosage profile, and reduce risks of multi-agent toxicities, supporting this strategy over dual drug approaches. A panel of inhibitors exploiting known binding modes of structurally-related ATP binding site inhibitors of EGFR/PI3K were synthesized and characterized, of which six promising candidates were chosen for in vitro study. The interaction of the inhibitors with common drug efflux proteins were probed to predict BBB penetrance. Of the compounds, MTX-241 was least likely to act as a substrate for efflux proteins such as P-glycoprotein. Treatment of MTX-241 in three human U87 GBM lines, some of which overexpress wild-type or vIII EGFR, three patient-derived DIPG lines, and two glioma stem cell (GSC) lines, one of which is radioresistant, showed strong cytotoxic potency measured by growth inhibitory activity, with IC50s in the &lt;10 µM range. MTX-241 was significantly more potent than clinically relevant inhibitors targeting EGFR/RTKs (gefitinib, lapatinib, dasatinib, imatinib) or PI3K (alpelisib, idelalisib). Notably, selectivity of MTX-241 towards these HGG cell lines in comparison to normal human astrocytes (NHA) was observed, with NHA nearly insensitive to MTX-241 even at &gt;100 µM. We found significant inhibition of p-EGFR and p-Akt in these cell lines compared to DMSO controls. Further, we identified a unique glycolytic-suppressing activity of MTX-241 in the U87/DIPG lines, to a greater extent than treatment with gefitinib or alpelisib. Additionally, as the clinically relevant HDAC inhibitor vorinostat upregulated EGFR activity, we examined the combination of MTX-241 and vorinostat, which significantly reduced proliferative abilities, achieving synergistic combination indices. MTX-241 and vorinostat treatment in the NHA model was not synergistic, highlighting the selectivity for HGG tumor models. Our data suggests that a dual inhibitor of EGFR and PI3K, alone or in combination with an HDAC inhibitor, represents a viable therapeutic strategy in adult/pediatric HGG. Future studies will focus on evaluation of in vivo efficacy in tumor-bearing mouse models.

Citation Format: Yusha Y. Sun, Cavan P. Bailey, Trever R. Carter, Christopher E. Whitehead, Judith S. Sebolt-Leopold, Joya Chandra. Evaluation of highly potent and selective single-molecule dual EGFR/PI3K inhibitors in preclinical models of adult and pediatric high-grade glioma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 3089.