Function and Targeting of ETV6 in Ewing Sarcoma - PROJECT SUMMARY/ABSTRACT Ewing sarcoma is a highly aggressive bone and soft tissue tumor mainly affecting children and young adults, which still lacks effective therapy. Due to its low mutational burden, there are very few acquired vulnerabilities in Ewing sarcoma cells. During my postdoctoral work, I discovered that transcription factor ETV6 is among the very few top Ewing sarcoma-biased dependencies. Remarkably, although ubiquitously expressed, ETV6 is not essential for cancer cells outside Ewing sarcoma, based on CRISPR screening efforts from us and others. Genetic studies in mice also revealed that ETV6 is dispensable in the majority of tissues. Hence, perturbing ETV6 function in Ewing sarcoma will have a wide therapeutic index. In the proposed project, I aim to further study the detailed mechanisms of how ETV6 maintains the cancer cell state, and leverage this new knowledge to develop therapies for Ewing sarcoma patients with exceptional potency and specificity. To do so, I will characterize the mesenchymal differentiation phenotype of ETV6-deficient Ewing sarcoma cells using high- throughput functional genomics and single cell transcriptomics. Integration of these `-omic' approaches allow me to obtain a deep biochemical understanding of the cancer maintenance function of ETV6 (Aim 1). Moreover, I showed that expression of the Sterile Alpha Motif (SAM) domain of ETV6, which is responsible for its self- oligomerization, has a dominant-negative effect to endogenous ETV6, and inhibits sarcomagenesis in vivo. Therefore, I propose to optimize this SAM peptide to increase its potency and further engineer it for exogenous delivery (Aim 2). Successful generation of a potent, and tumor penetrating ETV6 blocker will benefit therapy development. Finally, as transcription factors, like ETV6, have proven to be challenging targets for drug development, I profiled for endogenous metabolites that can bind ETV6 to unveil druggable pockets, and identified its association with phosphatidic acid. I will further explore the regulatory effects of phosphatidic acid binding to ETV6 in Ewing sarcoma (Aim 3). Results from these studies will guide small molecule development, and more importantly, will also reflect a novel mechanism for metabolic control of gene expression through direct allosteric regulation of transcription factors. During the mentored K99 phase, I will work closely with my mentor Dr. Christopher Vakoc and co-mentor Dr. Carolyn Fein Levy, and collaborators Drs. Stegmaier, Kentsis, Shi and Furukawa, recognized experts in pediatric oncology, peptide therapy, screening methodology and structural biology respectively. I have also established an exceptional advisory committee at CSHL, constituted by Drs. Joshua-Tor and Beyaz, who will monitor and support my transition to independence. In addition, CSHL will provide me an outstanding scientific environment for my research and training, being a conference hub for world-renowned meetings and courses. My objective is to obtain a faculty position to develop an impactful research program, where the K99/R00 funding mechanism will serve as an essential step in my transition to independence.
