Screening and in vitro testing of antifolate inhibitors of human cytosolic serine hydroxymethyltransferase

Metabolic reprogramming of tumor cells toward serine catabolism is now recognized as a hallmark of cancer. Serine hydroxymethyltransferase (SHMT), the enzyme providing one-carbon units by converting serine and tetrahydrofolate (H4 PteGlu) to glycine and 5,10-CH2 -H4 PteGlu, therefore represents a target of interest in developing new chemotherapeutic drugs. In this study, 13 folate analogues under clinical evaluation or in therapeutic use were in silico screened against SHMT, ultimately identifying four antifolate agents worthy of closer evaluation. The interaction mode of SHMT with these four antifolate drugs (lometrexol, nolatrexed, raltitrexed, and methotrexate) was assessed. The mechanism of SHMT inhibition by the selected antifolate agents was investigated in vitro using the human cytosolic isozyme. The results of this study showed that lometrexol competitively inhibits SHMT with inhibition constant (Ki ) values in the low micromolar. The binding mode of lometrexol to SHMT was further investigated by molecular docking. These results thus provide insights into the mechanism of action of antifolate drugs and constitute the basis for the rational design of novel and more potent inhibitors of SHMT.