Methotrexate has a distinct mechanism of action regarding its use in chemotherapy and immunosuppression in autoimmune diseases.
In cancer, methotrexate acts as an antifolate antimetabolite. Methotrexate is taken up into the cell by carriers called the human reduced folate carriers (SLC19A1), and it forms methotrexate-polyglutamate. Both the methotrexate and the methotrexate-polyglutamate inhibit the enzyme dihydrofolate reductase, which catalyzes the conversion of dihydrofolate into tetrahydrofolate, the active form of folic acid. Tetrahydrofolate is necessary for the synthesis of the nucleotides of both DNA and RNA. Methotrexate-polyglutamate further inhibits the de novo purine synthesis of both purine and thymidylate synthase, thereby inhibiting DNA synthesis. This mechanism is utilized in the treatment of cancer because of its cytotoxic effect.