Project Grant R01CA282785

INVESTIGATING MECHANISMS OF TUMOR PLASTICITY IN HUMAN BLADDER CANCER - PROJECT SUMMARY/ABSTRACT THE PROGRESSION OF BLADDER CANCER FROM NON-INVASIVE TO MUSCLE INVASIVE DISEASE REPRESENTS A CRITICAL STEP THAT IS ASSOCIATED WITH ADVERSE CLINICAL OUTCOMES, YET IS POORLY UNDERSTOOD AT THE MOLECULAR LEVEL. HOWEVER, THERE IS INCREASING EVIDENCE FROM US AND OTHERS THAT PROGRESSION IS OFTEN ASSOCIATED WITH LINEAGE PLASTICITY, IN WHICH THE TUMOR PHENOTYPE UNDERGOES A SWITCH FROM A LUMINAL TO A BASAL SUBTYPE. THEREFORE, WE WILL PURSUE A COMPREHENSIVE MOLECULAR ANALYSIS OF LUMINAL-BASAL LINEAGE PLASTICITY TO ELUCIDATE THE MECHANISTIC BASIS FOR THIS PHENOTYPIC SWITCH. IN PRELIMINARY STUDIES, WE HAVE SHOWN THAT PATIENT-DERIVED BLADDER TUMOR ORGANOIDS REPRESENT A MODEL SYSTEM FOR STUDYING TUMOR PLASTICITY, AND THAT THIS PLASTICITY IS REVERSIBLE. WE HAVE IDENTIFIED THE KMT2D HISTONE METHYLTRANSFERASE AND THE KDM1A HISTONE DEMETHYLASE AS HAVING OPPOSING ACTIVITIES IN MODULATING LUMINAL-BASAL PLASTICITY IN ORGANOIDS, CONSISTENT WITH THEIR EFFECTS ON HISTONE H3 LYSINE 4 METHYLATION. MODULATION OF LUMINAL-BASAL PLASTICITY AFFECTS OUTGROWTH OF TUMOR ORGANOIDS IN CULTURE, CONSISTENT WITH A ROLE IN PROGRESSION TO INVASIVE DISEASE, AND CAN RESULT IN IMPROVED RESPONSE TO ENFORTUMAB VEDOTIN, AN ANTIBODY-DRUG CONJUGATE APPROVED FOR TREATMENT OF LOCALLY ADVANCED AND METASTATIC BLADDER CANCER. FINALLY, USING A CELLULAR BARCODING APPROACH FOR LINEAGE TRACING, WE FIND THAT KMT2D MUTATIONS PROMOTE CLONAL EXPANSION IN ORGANOIDS, SUGGESTING THAT CLONAL COMPETITION CAN DRIVE THE SPREAD OF PHENOTYPIC ALTERATIONS DUE TO PLASTICITY DURING TUMOR PROGRESSION. BASED ON OUR PRELIMINARY DATA, WE HYPOTHESIZE THAT LUMINAL-BASAL PLASTICITY AND CLONAL COMPETITION IN HUMAN BLADDER CANCER IS DRIVEN BY EPIGENOMIC REPROGRAMMING. TO INVESTIGATE THIS HYPOTHESIS, WE WILL PURSUE THREE COMPLEMENTARY AIMS THAT INTEGRATE INNOVATIVE IN VITRO, IN VIVO, MOLECULAR, AND BIOINFORMATIC APPROACHES AS FOLLOWS: (1) ANALYSIS OF KMT2D FUNCTION IN MODULATING LUMINAL-BASAL PLASTICITY AND DRUG RESPONSE TO ESTABLISH THE PRECISE ROLES OF KMT2D IN PLASTICITY, INVASIVENESS, AND RESPONSE TO ENFORTUMAB VEDOTIN; (2) ANALYSIS OF CHROMATIN LANDSCAPES IN BLADDER CANCER PLASTICITY AND PROGRESSION TO IDENTIFY CHROMATIN STATES ASSOCIATED WITH LUMINAL- BASAL PHENOTYPIC TRANSITIONS; AND (3) INVESTIGATION OF KMT2D FUNCTION IN CLONAL EVOLUTION TO EXAMINE MECHANISMS BY WHICH KMT2D LOSS PROMOTES CLONAL EXPANSION IN BLADDER CANCER. OVERALL, OUR FINDINGS WILL HAVE A STRONG TRANSLATIONAL IMPACT THROUGH THE IDENTIFICATION OF EPIGENETIC MECHANISMS THAT PROMOTE PLASTICITY AND TUMOR PROGRESSION IN BLADDER CANCER, AND MAY LEAD TO IMPROVED THERAPIES FOR AGGRESSIVE DISEASE.