Program Director/Principal Investigator (Last, First, Middle): Kwiatkowski, David J. et. al., Project 3 Abstract In this project, we will characterize transcriptional and epigenetic events in TSC hamartomas that are required to enable tumor growth in the presence of two hit loss of TSC1/TSC2, and consequent downstream activation of mTORC1. Because we have recently shown that TSC hamartomas display bi-allelic loss of TSC1/TSC2 alone, without accompanying oncogene activation, we hypothesize that the core transcriptional regulatory circuitry causally contributes to TSC tumor growth and drives particular therapeutic vulnerabilities. Using H3K27ac ChIP-Seq and RNA-Seq data from AML, we have identified several transcription factors including MITF as relatively highly expressed and highly marked with H3K27ac, suggesting that these transcription factors are part of the core transcriptional regulatory circuitry of AML. Furthermore, the CDK7-targeted transcriptional inhibitor THZ1 causes growth inhibition and apoptosis in a TSC2-null AML cell line, as well as in TSC1-null human bladder cancer cell lines, both in vitro and in vivo, suggesting that transcription overall is a key dependency in TSC null tumors. Our Specific Aims are: 1) To characterize MITF as a transcriptional driver and therapeutic target in AML, identifying downstream targets of MITF that may be targeted; 2) To analyze the epigenetic landscape and core transcriptional regulatory circuitry in TSC-related hamartomas, thereby enabling identification of the master transcription factors that drive development of these tumors; and 3) To target transcription in TSC tumors and cancers with TSC1/TSC2 loss using THZ1 and other CDK7 inhibitors.