Abstract C013: Discovery of novel functional allosteric pockets on p53 using the AMPSTM screening platform

Abstract Allosteric modulation represents a promising strategy for addressing intractable targets by improving selectivity and leveraging novel mechanisms of action. One such target is the tumor suppressor p53, a transcription factor with high clinical relevance that has proven difficult to drug due to challenges in identifying binding pockets. Mutated in over 50% of human cancers, p53’s central role in oncology—along with emerging evidence that stabilization of mutant p53 can restore function—make it a compelling focus for therapeutic intervention, particularly through pan-mutant allosteric modulators. To overcome the challenge in identifying novel allosteric sites for undrugged targets, Atavistik Bio has established the AMPSTM (Atavistik Metabolite Proprietary Screening) platform, a systematic and scalable discovery engine that enables high-throughput screening of metabolites against macromolecular targets. AMPS combines equilibrium dialysis with highly sensitive mass spectrometry to screen a proprietary, metabolite-enriched library for identification of protein–small molecule interactions and discovery of allosteric functional binding sites. These sites serve as targets for small-molecule drug candidates, which are developed into potent and highly selective leads using structure-based drug design. This approach, which combines empirical data with state-of-the-art computational and structural enablement, rapidly discovers and validates functional allosteric modulators. We applied this platform to p53, where we screened multiple p53 variants, including wildtype and clinically relevant mutants, both with and without DNA. From this comprehensive AMPS dataset, we identified small molecule hits with distinct binding profiles: some were selective for multiple -mutants, others for single mutants; some bound to the apo protein, while others engaged with DNA-bound states. Follow-up biophysical studies confirmed thermal stabilization for some mutants, supporting functional relevance. Moreover, NMR analysis of a top hit compound showed perturbations localized to a novel allosteric pocket on p53. While we continue to characterize and optimize our lead hits, these results validate this platform as a robust means to identify new allosteric binding sites for challenging targets. Citation Format: Jonathan J. Ipsaro, Kaitlyn Selmer, Joseph H. LaPointe, Gordon Murray, Benjamin Brigham, Dave Church, Aaron Coffin, Rajiv G. Govindaraj, Bingsong Han, Trisha Moro, Joann Prescott-Roy, Shomit Sengupta, María-Jesus Blanco, Marion Dorsch, Anil K. Padyana, Thomas P. Roddy. Discovery of novel functional allosteric pockets on p53 using the AMPSTM screening platform [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference on Molecular Targets and Cancer Therapeutics; 2025 Oct 22-26; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2025;24(10 Suppl):Abstract nr C013.