Malic enzyme inhibition as a strategy to exploit metabolic reprogramming in cancer: A structure-based approach

Background: Cancer remains a global health challenge and urgently requires therapies that improve clinical outcomes and patient quality of life. Tumor cells commonly reprogram metabolism to sustain proliferation and survival, notably by increasing glutamine dependence and upregulating mitochondrial malic enzyme 2 (ME2), which produces pyruvate and NAD(P)H. ME2 is overexpressed in pancreatic, melanoma, and lung cancers, making it a compelling therapeutic target that remains underexplored. Methods: Recombinant human ME1, ME2, and ME3 were expressed and kinetically characterized in the presence and absence of inhibitors. We solved a complete set of X-ray crystal structures for the three isoforms with a potent malic enzyme inhibitor bound, revealing ligand and metal coordination. These structures guided virtual screening of a curated library of up to 14 million drug-like and fragment molecules to identify novel malic enzyme inhibitors. Biochemical hits were advanced to cellular assays to evaluate efficacy; lead compounds NPD-389 and FLA were tested for antiproliferative activity in melanoma and triple-negative breast cancer cell lines. Results: Kinetic analyses show distinct activity and inhibition patterns across ME isoforms. X-ray crystallography visualized the potent, isoform-nonselective inhibitor NPD-389 bound in a metal-coordinating mode and captured an unexpected malic enzyme conformation. Structure-based virtual screening against this conformation identified multiple novel scaffolds with biochemical inhibitory activity. One scaffold, FLA, exhibited a unique selectivity profile and was found bound to ME2 in a previously cryptic pocket that rearranges upon ligand engagement and sits adjacent to the NAD + cofactor and malate-binding site. In a panel of melanoma and triple-negative breast cancer cell lines, NPD-389 and FLA reduced proliferation in most models, supporting their potential as chemical leads for further development. Conclusions: These results establish mitochondrial malic enzymes, particularly ME2/ME3, as tractable targets for cancer therapy. Structural elucidation of NPD-389's metal-binding mode and of FLA's cryptic/allosteric binding pocket provides a robust foundation for rational optimization of potency and isoform selectivity. Selective ME inhibitors-either isoform-specific or mitochondrially targeted-have potential as adjuncts to checkpoint inhibitors or chemotherapy, enabling more effective, metabolism-directed cancer treatment. References: 1.Heon, J., Slayton, M., Krinkel, B., et al., Interplay Between Malic Enzyme 2, de novo Serine Synthesis, and the Malate-Aspartate Shuttle Drives Metabolic Adaptation in Triple-Negative Breast Cancer. Cancer & Metabolism , 2025.