Therapeutic targeting mesenchymal transition in newly diagnosed and recurrent GBM

PROJECT SUMMARY IDHwt glioblastoma (GBM) patients still live an average of ~15-months, despite advances in multimodal therapy. A central issue confounding treatment is the heterogeneous nature of this aggressive tumor. Transcriptomics has defined three GBM molecular subtypes - proneural (PN), classical (CL), and mesenchymal

(MES).7 Individual tumors typically harbor mixtures of all three subtypes in spatially distinct subpopulations with different mutation profiles, making mutation- or pathway-specific therapies less effective. While cell-intrinsic mechanisms of therapeutic resistance have garnered considerable scientific attention, much less is known about

cellular interactions in the tumor microenvironment that contribute to therapeutic recalcitrance. A hallmark mutation in 60% of GBM is amplification and mutation of the epidermal growth factor receptor (EGFR). The most common EGFR alteration, EGFRvIII, results from deletion within its extracellular domain, yielding a constitutively

active receptor that conveys tumor-enhancing and therapy-resisting functions.8 Genetic and pharmacological data from our lab show that EGFRvIII activity, specifically in the context of PI3K pathway activation, results in nuclear localization of the NF-kB subunit RelA (p65), its association with members of the BET (bromodomain

and extra terminal domain) family of acetylated lysine-binding proteins, and transcriptional activation of inflammatory genes.6,9 The requirement of RelA acetylation at lysine 310 (acK310-RelA) for BET bromodomain interactions10 and the central role of NF-kB in driving a PN/CL to MES phenotype transition (collectively MES

transition – MESt),6,11 immune evasion,3 and therapy resistance12 leads us to postulate that the acK310- RelA:BET protein complex is a druggable regulatory switch mediating MESt and treatment resistance. The goal of this project is to dissect and target mechanisms whereby tumors undergo MESt and acquire

therapeutic resistance through the acK310-RelA:BET switch. We have identified a druggable node of specific bromodomains, through precise drug targeting, that controls the pro-inflammatory activity of the acK310- RelA:BET complex.6 The following lines of experimentation will be employed in the newly diagnosed and

recurrent GBM settings. SA1 will use genetic and pharmacological approaches to determine upstream effector mechanisms mediating RelA K310 acetylation and associated MESt, including abundance of tumor-associated microglia and macrophages (TAM), and resistance to standard-of-care (SOC) therapy. SA2 will functionally

analyze BET family members BRD2-4 through gene knockout, gene editing, inducible protein degradation, selective bromodomain (BD1, BD2) drug targeting, and assessment of acK310-RelA:BET induced transcription programs and associated epigenomic rewiring. SA3 will specifically focus on the recurrent GBM setting, where

we will investigate direct drug targeting of the acK310-RelA:BET interaction to mitigate resistance to salvage radiation and promote phagocytotic recognition by TAM. These studies will be central to identifying a therapeutically tractable node promoting MESt and nominate a specific drug for further development.