Abstract 257: The E3 ligase HECTD4 regulates COX-2 dependent tumor progression and metastasis

Abstract E3 ubiquitin ligases mediating turnover of proteins engaged in cancer progression point to key regulatory nodes. To uncover modifiers of metastatic competency, we conducted an in vivo genome-wide CRISPR-inactivation screen using cultured breast circulating tumor cells, following intravascular seeding and lung colonization. We identified HECTD4, a previously uncharacterized gene encoding a conserved potential HECT domain-containing ubiquitin transferase, as a potent tumor suppressor. We show that purified HECTD4 mediates ubiquitin conjugation in vitro, and proteomic studies combined with ubiquitin remnant profiling identify a major degradation target as the prostaglandin synthetic enzyme cyclooxygenase-2 (COX-2; PTGS2). In addition to COX-2 itself, HECTD4 targets its regulatory kinase MKK7. In breast cancer models, HECTD4 expression is induced as cells lose adherence to the matrix, and its depletion massively increases COX-2 expression, enhancing anchorage-independent proliferation and tumorigenesis. Genetic or pharmacologic suppression of COX-2 reverses the pro-tumorigenic phenotype of HECTD4-depleted cells. Thus, HECTD4 encodes an E3 ubiquitin ligase that downregulates COX-2 suppressing anchorage-independence in epithelial cancer cells. Citation Format: Joanna Vuille, Cem Tanriover, Douglas S. Micalizzi, Richard Y. Ebright, Sambhavi Animesh, Robert Morris, Soroush Hajizadeh, Zachary J. Nicholson, Hunter C. Russell, Eric F. Zaniewski, Ben S. Wittner, Ben K. Wesley, Julian Grünewald, Regan N. Szalay, Ezgi Antmen, Douglas B. Fox, Min Yang, Jae Keith Joung, Doga C. Gulhan, Andrew E. Elia, Wilhelm Haas, Eugene Oh, Shyamala Maheswaran, Daniel A. Haber. The E3 ligase HECTD4 regulates COX-2 dependent tumor progression and metastasis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 257.