Project Grant R01CA288596

DECIPHERING MECHANISMS OF TRANSCRIPTIONAL REPRESSION WHICH DRIVE MIDLINE GLIOMA - PROJECT ABSTRACT: DIFFUSE MIDLINE GLIOMA (DMG) IS A UNIVERSALLY LETHAL FORM OF BRAIN CANCER THAT AFFECTS BOTH CHILDREN AND ADULTS, WITH MOST PATIENTS PASSING AWAY WITHIN 1 YEAR OF DIAGNOSIS FOLLOWING STANDARD OF CARE RADIATION THERAPY. NOT A SINGLE DRUG HAS BEEN FDA APPROVED DESPITE A MULTITUDE OF CLINICAL TRIALS, WITH MOST AGENTS TESTED NOT DESIGNED SPECIFICALLY FOR THIS DISEASE. THIS HIGHLIGHTS THE NEED FOR IDENTIFICATION OF THERAPEUTIC TARGETS DIRECTLY LINKED TO THE PATHOGENIC MECHANISM IN DMGS. THE HALLMARK OF DMGS ARE DOMINANT NEGATIVE MUTATIONS IN HISTONE H3, A KEY STRUCTURAL AND REGULATORY CONSTITUENT OF THE CHROMATIN STRUCTURE IN WHICH DNA IS PACKAGED. MUTANT H3 DRIVES TUMORIGENESIS BY PROMOTING TRANSCRIPTIONALLY SILENT CHROMATIN AT SPECIFIC TUMOR SUPPRESSOR GENES INCLUDING CDKN2A. HOW TRANSCRIPTION IS SILENCED AT THESE GENOMIC SITES IS POORLY UNDERSTOOD AND IS A KEY KNOWLEDGE GAP IN THE FIELD. OUR PRELIMINARY DATA IMPLICATES A MECHANISM IN WHICH MUTANT H3 INDUCES SILENT CHROMATIN AT TUMOR SUPPRESSOR GENES BY HIJACKING THE ACTIVITY OF A SPECIFIC CONFIGURATION OF THE POLYCOMB REPRESSIVE COMPLEX 1 (PRC1). PRC1 IS AN EPIGENETIC SILENCING COMPLEX WITH NUMEROUS PROTEIN SUBUNITS WHICH CAN ADOPT HIGHLY HETEROGENEOUS CONFIGURATIONS (>150 COMBINATIONS), MAKING IT PARTICULARLY CHALLENGING TO STUDY. LEVERAGING POOLED CRISPR SCREENS INTEGRATED WITH QUANTITATIVE PROTEOMICS, WE HAVE IMPLICATED A PRC1 COMPLEX CONTAINING CHROMOBOX 4 (CBX4-PRC1) AS THE MEDIATOR OF INAPPROPRIATE TRANSCRIPTIONAL SILENCING IN DMGS. OUR CENTRAL HYPOTHESIS IS THAT CBX4-PRC1 IS CO-OPTED IN DMGS TO DRIVE TUMOR GROWTH BY TRANSCRIPTIONALLY REPRESSING TUMOR SUPPRESSOR GENES INCLUDING CDKN2A. THE GOAL OF THIS PROPOSAL IS TO DEFINE CBX4-PRC1 AS THE FUNCTIONAL CONFIGURATION WHICH MEDIATES INAPPROPRIATE TRANSCRIPTIONAL SILENCING IN DMGS AND DETERMINE WHETHER TARGETING CBX4 CAN BE COMBINED WITH STANDARD OF CARE RADIATION TREATMENT IN DMG MODELS. WE WILL LEVERAGE OUR STRONG PRELIMINARY DATA, MULTI-DISCIPLINARY TEAM, AND NOVEL PATIENT-DERIVED DMG MODELS TO TEST OUR HYPOTHESIS THROUGH THE FOLLOWING AIMS: 1) DEFINE THE FUNCTIONALLY ESSENTIAL PRC1 COMPLEX CONFIGURATION IN DMGS. 2) ELUCIDATE THE ROLE OF CBX4 IN PRC1 CHROMATIN RECRUITMENT AND SILENCING FUNCTION IN DMGS. 3) EVALUATE THE THERAPEUTIC POTENTIAL OF TARGETING CBX4 IN CONJUNCTION WITH STANDARD OF CARE RADIATION THERAPY IN DMGS. THIS PROPOSAL WILL LAY THE MECHANISTIC FRAMEWORK FOR CBX4 AS A NOVEL THERAPEUTIC TARGET IN DMGS, A CURRENTLY INCURABLE BRAIN CANCER. WE WILL REVEAL THE SPECIFIC MECHANISM BY WHICH MUTANT H3 PROMOTES INAPPROPRIATE TRANSCRIPTIONAL SILENCING TO DRIVE DMG GROWTH AND DETERMINE THE RELEVANCE OF THIS MECHANISM ACROSS DMG SUBTYPES TO GUIDE PATIENT SELECTION IN THE FUTURE.