Methotrexate and the need for continued research.

With the current national commitment to cancer research, new and challenging areas of investigation are being planned. Hopefully, this inevitably expanding search into the nature and treatment of malignant tumors will include a continued examination of current chemotherapeutic agents. These standard tumoricidal drugs have proved to be effective in the therapy of numerous cancers despite an incomplete understanding of their mode of action in many cases (1-4). A further unraveling of the pharmacologic basis of these drugs should undoubtedly lead to a greater therapeutic effect, perhaps producing longer remissions and increased cures. A recent example of how continued basic research on available antineoplastic agents may reveal findings pertinent to clinical chemotherapy concerns metho-trexate, a drug frequently employed in the treatment of cancer (1-5, 66). Traditionally , the mode of action of this drug has assumed to be as an analog of the vitamin folic acid. In humans, folic acid is biologically inactive, requiring reduction by an enzyme termed "dihydrofolate reductase" to dehydrofolate, and then to tet-rahydrofolate. This latter compound accepts 1-carbon fragments from various sources to produce the folate coenzymes. These compounds act as 1-carbon donors in numerous biochemical reactions, including the synthesis of the B-carbon of serines, the formation of the C-2 and C-8 units of the purine skeleton, and the then, by irreversibly binding to and inhibiting the enzyme dihydrofolate reductase thereby interfering with the maintenance of intracellular pools of reduced folates, particularly N5' '0-methylenetetrahydrofolate (1, 5, 11-13, 66). This latter compound is necessary for the conversion of deoxyuridylate to thymidylate which is an essential component of DNA. As is evident, methotrexate, as well as other antifo-late agents, essentially act by inhibiting DNA synthesis For many years it was accepted that the biological effects of methotrexate were due entirely to its inhibition of the enzyme dihydrofolate reductase. Since free or un-bound intracellular methotrexate accumulates only after all the dihydrofolate reductase binding sites are saturated, little biological activity or significance has been ascribed to this free intracellular drug component. However, evidence has been accumulating that methotrexate may have a second site of action of equal or greater importance than the inhibition of dihydrofolate reductase and furthermore, that it is the unbound intracellular drug that is involved with this alternative site. Roberts et al. (35-37), several years ago, demonstrated a lack of correlation between the inhibition of dihydrofolate reductase activity in human leukemia cells and the response of leukemia patients …