Identification and Characterization of Transcriptional Regulators as Therapeutic Vulnerabilities in Hematologic Malignancies
CitationErb, Michael A. 2017. Identification and Characterization of Transcriptional Regulators as Therapeutic Vulnerabilities in Hematologic Malignancies. Doctoral dissertation, Harvard University, Graduate School of Arts & Sciences.
AbstractTranscription regulation is orchestrated on a chromatin template via the complex interactions between cis-regulatory DNA elements, trans-acting co-regulators, and RNA polymerase (Pol) II. These factors combine to regulate cell-type specific gene expression programs that enable the derivation of diverse cell types from a static genome, or when dysregulated, cause and support oncogenesis. Here, I present a collaborative body of work aimed to identify and delineate the mechanistic role of transcriptional regulators as cancer- specific dependencies.
Our group previously reported dBET1, a proof-of-principle small molecule capable of directing BET family (BRD2, BRD3, and BRD4) proteins for proteasome-dependent degradation. BRD4, which utilizes a pair of tandem bromodomains to bind acetyl-lysine residues in active regions of chromatin, promotes pathogenic transcription in cancer. Using dBET1, our group previously reported BET degradation as having improved anti-cancer activity over BET bromodomain inhibition, but through unclear mechanisms. Here, we use a chemically-optimized degrader molecule to demonstrate that BET degradation results in global shutdown of transcription elongation. While BRD4 has previously been understood to recruit the master regulator of Pol II pause release, P-TEFb, we find persistence of P-TEFb on chromatin following BET degradation. Unexpectedly, recruitment of the transcription elongation factor, SPT5, is globally diminished, repositioning BRD4 as a master regulator of transcription elongation independent of P-TEFb recruitment.
While the function of BRD4 as a principal mediator of oncogenic transcription is well established, the YEATS domain, also an acyl-lysine reader, has remained unexplored in human malignancies. In a genome-scale CRISPR/Cas9 knockout screen of a human acute leukemia cell line, we identified ENL as particularly indispensable for proliferation, an observation extended to diverse acute leukemia cell lines and a xenotransplantation model of disseminated leukemia. Importantly, ENL-dependent leukemic growth was contingent upon an intact YEATS chromatin reader domain. To explain the mechanistic role for ENL in leukemia pathogenesis and dynamic transcription control, we pursued a chemical genetic strategy utilizing targeted protein degradation. Acute ENL loss suppresses transcription initiation and elongation genome- wide, with pronounced effects at genes featuring disproportionate ENL load. These findings reveal a novel dependency in acute leukemia and a first mechanistic rationale for disrupting the YEATS domain in disease.
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