Impact of MTHFD2 Expression in Bladder/Breast Cancer\nand Screening of Its Potential Inhibitor

Genes\nof folate-mediated 1 carbon metabolism are found to be highly\nupregulated in tumor cells and promote cancer cell proliferation.\nThe current study aimed to determine the expression of the MTHFD2\ngene in bladder and breast cancers. Furthermore, the determination\nof potential ligand-based inhibitors against MTHFD2 was performed\nin comparison with those of chemotherapeutic drugs and natural plant-based\ncompounds. Semiquantitative expression analysis along with structure-based\nvirtual ligand library screening was done to find plausible inhibitors.\nMTHFD2 expression was significantly increased with tumor stage progression\nboth in low- and high-grade bladder cancer and especially in triple-negative\nbreast cancer. Virtual ligand-based library screening against the\nthree-dimensional MTHFD2 protein structure led to the identification\nof plausible inhibitors like MCULE-8109969891-0 and MCULE-9715677418-0-1\nthat displayed lower binding free energy as compared to that of already\ndocumented LY345899. Similar scaffold commercial drugs leucal (LEU),\nepirubicin (EPI), and lometrexol also displayed strong binding to\nthe active site of MTHFD2. EPI and LEU in combinatorial therapy were\nalso tested in vitro on MDA-MB-231 cells. The high doses of LEU in\ncombination with EPI showed a significant reduction in cell viability\nat 2 and 3 μM concentrations. The interaction of breast cancer\nserum with high expression of MTHFD2 also showed an increase in binding\nof epirubicin in the presence of leucovorin. The decrease in the absorbance\nspectra of epirubicin at 37 and 53 °C displayed the stability\ninduced by LEU on the interaction of EPI with the MTHFD2 binding pocket.\nLeucovorin tends to stabilize the interaction as the binding affinity\nis high even at 53 °C. Thus, MTHFD2 might be used as a cancer\nbiomarker since its expression level changes drastically with tumor\nprogression. Further experimental studies are required to establish\nthe potential mode of inhibition of the novel small ligands. Future\nin vivo trials may validate the effectiveness of the combinatorial\ntherapy.