Abstract 259: Germline p53 R181 mutants and DNA binding cooperativity in tumorigenesis

Abstract TP53 is the most frequently mutated gene in cancer, and its encoded protein p53 has many tumor-suppressive functions. There are several different classes of mutant p53 acquired in human cancer and inherited in cancer-prone families with Li-Fraumeni Syndrome, where individuals have an 80-90% increased risk of cancer. These include structural mutations that are generally misfolded (R175H), DNA contact mutations (R273H); and oligomerization mutations (A347D). A new class of variants called “cooperativity” mutations affect the binding of the p53 tetramer to DNA via disruption of a salt bridge formed by the negatively charged glutamic acid (E) 180 residue of one p53 monomer and the positively charged arginine (R) 181 residue of another. p53 cooperativity mutations at the R181 residue (R181H and R181C) have been increasingly identified in cancer-prone families undergoing genetic testing; however, the mechanism by which these variants disrupt p53 tumor suppression in humans is not understood. We show that R181H and R181C have significantly diminished ability to transactivate a curated set of ∼300 known p53 target genes in CRISPR knock-in colorectal and breast cancer cell lines. This loss of transactivation ability is not explained by defects in p53 structure or oligomerization, but through reduced cooperative binding to p53 target sites on DNA as determined by fluorescence polarization and chromatin-immunoprecipitation experiments. Interestingly, we observe residual apoptotic activity in R181H and R181C mutant cells upon DNA damage, despite the poor transcriptional activation of p53’s proapoptotic targets. We observe increased p53 at the mitochondria for transcription-independent apoptosis in R181-mutant cells. Understanding the tumor suppressive activities that are lost versus retained will guide the definition of the pathogenicity of R181 variants, which account for 5000 cancer cases per year in the United States. Citation Format: Renyta Moses, Alexandra Indeglia, Alison S. Levine, Ryan Hausler, Gregory Kelly, Sven A. Miller, Melissa Heller, Anh N. Le, Wendy Kohlmann, Anne Naumer, Jennie Vagher, Sophie H. Cahill, Luke D. Maese, John Karanicolas, Judy E. Garber, Maureen E. Murphy, Kara N. Maxwell. Germline p53 R181 mutants and DNA binding cooperativity in tumorigenesis [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 259.