Metabolic effects and kill of human T-cell leukemia by 5-deazaacyclotetrahydrofolate, a specific inhibitor of glycineamide ribonucleotide transformylase.
Metabolic effects and mode of cytotoxicity of 5-deazaacyclotetrahydrofolate (5-DACTHF, BW543U76), a glycineamide ribonucleotide transformylase inhibitor, were studied in MOLT-4 cells, a human T-cell leukemia line. 5-DACTHF inhibits purine synthesis with 50% inhibitory concentration values of 0.5 microM and 0.08 microM following 6- or 24-h exposure to drug, respectively. At 6 h, adenine nucleotide synthesis is preferentially inhibited over guanine nucleotide synthesis. A similar effect was observed with another glycineamide ribonucleotide transformylase inhibitor, 5,10-dideazatetrahydrofolate. GTP was depleted to 40% of control and ATP to 10% of control by 5 microM 5-DACTHF. After a transitory increase, UTP and CTP were depleted to 30% of control. Deoxynucleotides were also depleted by the drug; dCTP was depleted to the greatest extent, followed by dATP, dTTP, and dGTP, respectively. MOLT-4 cell growth was inhibited by 5-DACTHF with a 50% inhibitory concentration of 0.066 microM. Complete reversal was effected by hypoxanthine, and there was no reversal by thymidine. The drug was cytotoxic to MOLT-4 cells in the range 0.25 to 5.0 microM, but a minimum of 48 h was required for trypan blue-staining dead cells to appear. The rate and extent of kill with the thymidylate synthase inhibitor 2-methyl-10-propargyl-5,8-dideazafolate was greater than with 5-DACTHF, which indicates that kill by inhibition of thymidylate synthase is more effective than that by inhibition of purine synthesis. Electron microscopy of MOLT-4 cells exposed to 5-DACTHF showed electron-dense mitochondria and nuclear changes reminiscent of apoptosis. These morphological changes were accompanied by the appearance of DNA strand breaks at approximately 180-base pair intervals (internucleosomal breaks). Concomitant proteolysis of nuclear proteins poly(ADP-ribose) polymerase and lamin B was observed.
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Burroughs-Wellcome Co., Wellcome Research Labs, Research Triangle Park, North Carolina 27709.
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Kaufmann
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D000217:Acyltransferases / Q000037:antagonists & inhibitors*; D016923:Cell Death / Q000187:drug effects; D002455:Cell Division / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D003854:Deoxyribonucleotides / Q000378:metabolism; D006801:Humans; D019877:Hydroxymethyl and Formyl Transferases; D066298:In Vitro Techniques; D015458:Leukemia, T-Cell; D009711:Nucleotides / Q000378:metabolism; D051026:Phosphoribosylglycinamide Formyltransferase; D011687:Purines / Q000378:metabolism; D013763:Tetrahydrofolates / Q000494:pharmacology*; D014407:Tumor Cells, Cultured / Q000187:drug effects* / Q000378:metabolism
D003854:Deoxyribonucleotides; D009711:Nucleotides; D011687:Purines; D013763:Tetrahydrofolates; C062810:N-(4-((3-(2,4-diamino-1,6-dihydro-6-oxo-5-pyrimidinyl)propyl)amino)benzoyl)glutamic acid; D019877:Hydroxymethyl and Formyl Transferases; D051026:Phosphoribosylglycinamide Formyltransferase; D000217:Acyltransferases