Role of glutathione depletion in the mechanism of action of N-methylformamide and N,N-dimethylformamide in a cultured human colon carcinoma cell line.

The mechanism of the antitumor action of N-methylformamide (NMF), an agent currently undergoing clinical trials, and its congener, N,N-dimethylformamide (DMF), was examined in the DLD-1 Clone A human colon carcinoma cell line in vitro. The primary action of NMF and DMF on these cells is a depletion of cellular reduced glutathione levels which results in cytostasis. Evidence to support this hypothesis include (a) the extent of growth inhibition produced by NMF and DMF is directly proportional to the extent of depletion they effect on cellular reduced glutathione levels; (b) removal of NMF (170 mM) or DMF (103 mM) from the culture medium results in a parallel restoration of cell growth and cellular glutathione levels; (c) coaddition of the glutathione precursor, L-cysteine (0.5 mM), or certain precursors for this amino acid, significantly reverses NMF- and DMF-induced glutathione depletion and cytostasis; and (d) the specific glutathione-depleting agent, buthionine sulfoximine (7.5 mM) mimics the ability of NMF and DMF to induce cytostasis. In addition, NMF- and DMF-mediated reduced glutathione depletion accounts for the previously reported ability of these agents at high concentrations (less than 100 mM) to induce a more benign phenotype in DLD-1 Clone A human colon carcinoma cells. This is evidenced by the findings that (a) L-cysteine (0.5 mM) reverses NMF- and DMF-mediated (170 and 103 mM, respectively) increases in doubling time, decreases in saturation density, and decreases in clonogenicity; (b) buthionine sulfoximine (7.5 mM) mimics these actions of NMF and DMF; and (c) the tumorigenicity of DLD-1 Clone A cells in nude mice, which is completely eliminated by the in vitro treatment of these cells for 4 passages with 170 mM NMF prior to inoculation, is fully restored if the cells are passaged in the presence of 170 mM NMF and 0.5 mM L-cysteine. Thus, NMF- and DMF-induced depletion of cellular reduced glutathione is responsible for not only the cytostatic effect of these agents on human colon carcinoma cells but also for their ability to induce more benign characteristics in these cells.