Novel pyrrolo[2,3-d]pyrimidine antifolate TNP-351: cytotoxic effect on methotrexate-resistant CCRF-CEM cells and inhibition of transformylases of de novo purine biosynthesis.
N-[4-[3-(2,4-Diamino-7H-pyrrolo[2,3-d]pyrimidin-5- yl)propyl]benzoyl]-L-glutamic acid (TNP-351), characterized by a pyrrolo[2,3-d]pyrimidine ring, is a novel antifolate that exhibits potent antitumor activities against mammalian solid tumors. The mechanism of action of TNP-351 was evaluated using some methotrexate-resistant CCRF-CEM human lymphoblastic leukemia cell lines as well as partially purified enzymes folylpolyglutamate synthetase (FPGS), aminoimidazolecarboxamide ribonucleotide transformylase (AICARTFase), and glycinamide ribonucleotide transformylase (GARTFase) from parent CCRF-CEM cells. TNP-351 was found to inhibit the growth of L1210 and CCRF-CEM cells in culture, with the doses effective against 50% of the cells (ED50 values) being 0.79 and 2.7 nM, respectively. The growth inhibition caused by TNF-351 was reversed by leucovorin or a combination of hypoxanthine and thymidine. The methotrexate-resistant CCRF-CEM cell line, which has an impaired methotrexate transport, showed less resistance to TNP-351 than to methotrexate. TNP-351 was also found to be an excellent substrate for FPGS with a Michaelis constant (Km) of 1.45 microM and a maximum of velocity (Vmax) of 1,925 pmol h-1 mg-1. Inhibitory activities of TNF-351-Gn (n = 1-6) for AICARTFase were found to be significantly enhanced with increasing glutamyl chain length [inhibition constants (Ki): G1, 52 microM; G6, 0.07 microM]. Neither TNP-351 nor its polyglutamates were very strong inhibitors of GARTFase. These findings have significant implications regarding the mechanism of action of TNP-351.
Pharmaceutical Research Laboratories III, Takeda Chemical Industries, Ltd., Osaka, Japan.
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Itoh
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Russello
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Akimoto
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Beardsley
D000217:Acyltransferases / Q000037:antagonists & inhibitors; D000818:Animals; D002455:Cell Division / Q000187:drug effects; D004351:Drug Resistance; D005493:Folic Acid Antagonists / Q000378:metabolism / Q000494:pharmacology*; D006801:Humans; D019877:Hydroxymethyl and Formyl Transferases; D007945:Leukemia, Lymphoid / Q000188:drug therapy* / Q000201:enzymology; D008727:Methotrexate / Q000031:analogs & derivatives* / Q000378:metabolism / Q000494:pharmacology / Q000627:therapeutic use*; D051379:Mice; D010453:Peptide Synthases / Q000378:metabolism; D051027:Phosphoribosylaminoimidazolecarboxamide Formyltransferase; D051026:Phosphoribosylglycinamide Formyltransferase; D014407:Tumor Cells, Cultured
D005493:Folic Acid Antagonists; C078101:TNP 351; D019877:Hydroxymethyl and Formyl Transferases; D051026:Phosphoribosylglycinamide Formyltransferase; D051027:Phosphoribosylaminoimidazolecarboxamide Formyltransferase; D000217:Acyltransferases; D010453:Peptide Synthases; C023103:folylpolyglutamate synthetase; D008727:Methotrexate