Abstract 6675: Exploiting rewired metabolism in cancer: A biochemical focus on malic enzyme inhibitors using structure-based drug design

Abstract Background: Cancer continues to be a formidable adversary, claiming countless lives globally and fueling the pursuit of more effective therapies to enhance treatment outcomes and quality of life. A hallmark of cancer is the reprogramming of cellular metabolism, including the aberrant biosynthesis of essential metabolites that support survival and rapid cellular proliferation. This metabolic shift often involves increased reliance on glutamine as a fuel source and elevated expression of malic enzyme 2 (ME2) within the mitochondria. ME2 catalyzes the generation of pyruvate and NAD(P)H and is upregulated in pancreatic, melanoma, and lung cancer, making it a promising yet underexplored therapeutic target. We have solved the structure of the active site small molecule inhibitor NPD-389 in complex with ME2 and used virtual screening to identify novel scaffolds to develop into new drug leads. Methods: We expressed recombinant human ME1-3 isoforms and detailed their enzymatic kinetics with and without inhibitors. Molecular snapshots of the potent malic enzyme inhibitor were trapped within a complete set of X-ray crystal structures for the three isoforms ME1-3, and this structural data was used to conduct the virtual screening of up to 12 million drug-like and fragment molecules to identify novel, potent malic enzyme inhibitors. To complement the multidisciplinary characterization of malic enzyme inhibition, the efficacy of NPD-389 in treating triple-negative breast cancer was assessed. Results: Our kinetic analyses reveal unique inhibition patterns, particularly for the least studied isoform, ME3. We detailed its substrate and cofactor binding affinities and inhibitory constants for EA, MDSA, and NPD-389. The revelation of X-ray crystallographic visualization of the potent but isoform non-selective inhibitor NPD-389 in complex is a metal-binding mode and the adoption of an unexpected malic enzyme conformation. Virtual screening based on the unique protein conformation identified several new scaffolds with promising inhibitory activity against ME’s. These include molecules confirmed by X-ray crystallography to bind in an allosteric site and that, critically, display distinct isoform selectivity. In triple-negative breast cancer cells, NPD-389 inhibited proliferation in most cell lines, suggesting its potential as a lead compound for further development. Conclusions: Our studies underscore the potential of ME2 as a therapeutic target in cancer treatment. By detailing the binding mode of NPD-389 and those of newly identified selective inhibitors, we provide a robust platform for the future clinical development of potent inhibitors of metabolic pathways driving cancers. We envisage that isoform selective ME drugs could serve as adjunct therapies alongside existing checkpoint inhibitors or chemotherapy, enhancing treatment efficacy and improving patient outcomes. Citation Format: Ben Krinkel, Mark Slayton, Jin Heon Jeon, Jack Copping, Jack Flanagan, Sofia Merajver, Chris Squire, Kerry Loomes. Exploiting rewired metabolism in cancer: A biochemical focus on malic enzyme inhibitors using structure-based drug design [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 6675.