Targeting the chromatin binding domains of BPTF for neuroblastoma epigenetic therapy

ABSTRACT The goal of this proposal is to develop high-quality chemical probes to functionally annotate the epigenetic effector “reader” domains within Bromodomain PHD Finger Transcription Factor (BPTF) in neuroblastoma (NB) and to serve as new therapeutic leads. BPTF is an essential component of the nucleosome remodeling complex NURF. Nucleosome remodeling complexes are emerging as important anticancer drug targets due to their role in regulating chromatin accessibility and gene expression. We hypothesize that interfering with the chromatin binding of BPTF will disrupt oncogenic regulatory networks present in cancer cells. Our preliminary data supports a new tumorigenic role of BPTF in NB, an aggressive pediatric cancer which accounts for ~15% of cancer deaths in children. Intriguingly, BPTF copy number gain is frequent in NB and high BPTF expression at initial diagnosis is associated with increased risk of disease progression and recurrence. Reduction of BPTF protein levels using genetic depletion resulted in robust loss of either MYC or MYCN and a concomitant loss of cell viability. Our results are consistent with the reported role of BPTF in many adult cancers including melanoma, glioma, breast, lung, and pancreatic cancer. However, high-quality chemical probes for BPTF are lacking, and the individual roles of the BPTF chromatin binding domains for regulating these mechanisms in cancer have yet to be fully elucidated. To address these shortcomings, we will develop chemical probes to assess the function of the BPTF C-terminal plant homeodomain (PHD) and bromodomain (BRD) with respect to chromatin binding and accessibility, transcriptional regulation of MYC and MYCN and their downstream targets, and anti-cancer activity. In Aim 1, we will address liabilities of current BRD chemical probes and optimize ADME pharmacokinetic properties; in Aim 2, we will develop PHD inhibitors using biochemical, structural, and biophysical assays; in Aim 3, we will establish an epigenetic mechanism linking BPTF, NURF, MYC, and MYCN in NB and evaluate the roles of the individual BPTF chromatin binding domains. Optimized, drug-like BRD and PHD inhibitors will enable in vivo evaluation of these compounds as therapeutic leads and offer starting points for PROTACs. The innovation in our approach is the use of new structural biology methods based on room temperature crystallography, protein NMR, and molecular dynamics for capturing dynamic protein states to facilitate structure- based-design of selective inhibitors and virtual screening of over a billion small molecules. Our proposal benefits from extensive preliminary data based on our second generation nanomolar-affinity BRD inhibitors BZ1 and BZ2, the first room temperature crystal structures guiding our design, the first reported inhibitors developed from a screen targeting the PHD finger, and BPTF domain-edited cancer cell lines. High-quality chemical tools emerging from this project will enable the elucidation of the oncogenic roles of the BPTF chromatin binding domains in NB and provide a potential therapeutic option for an aggressive cancer that lacks effective treatments. Furthermore, our work will empower the research community to probe BPTF biology in other adult and pediatric cancers.