This Article Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Pizzorno, G. Articles by Beardsley, G. P. Search for Related Content PubMed PubMed Citation Articles by Pizzorno, G. Articles by Beardsley, G. P.

Multifactorial Resistance to 5,10-Dideazatetrahydrofolic Acid in Cell Lines Derived from Human Lymphoblastic Leukemia CCRF-CEM¹

Departments of Pediatrics, Pharmacology, and Internal Medicine, Yale University School of Medicine, New Haven, Connecticut 06510 [G. P., B. A. M., A. R. C., O. R., J. R. M., G. P. B.]; Wadsworth Center for Laboratories and Research, New York State Department of Health, Albany, New York 12201 [J. G.]; and Department of Biochemistry, Medical University of South Carolina, Charleston, South Carolina 29425 [M. A. B., D. G. P.]

5,10-dideaza-5,6,7,8-terrahydrofolic acid (DDATHF) is a potent anti-proliferative agent in cell culture systems and in vivo in a number of murine and human xenograft tumors. In contrast to classical antifolates, which are dihydrofolate reductase inhibitors, DDATHF primarily inhibits GAR transformylase, the first folate-dependent enzyme along the pathway of de novo purine biosynthesis. The (6R) diastereomer of DDATHF (Lometrexol), currently undergoing clinical investigation, was used to develop CCRF-CEM human leukemia sublines resistant to increasing concentrations of the drug. Three cell lines were selected for ability to grow in medium containing 0.1 µM, 1.0 µM, and 10 µM of (6R)DDATHF, respectively. Impaired polyglutamylation was identified as a common mechanism of resistance in all three cell lines. A progressive decrease in the level of polyglutamylation was associated with diminished folylpolyglutamate synthetase activity and paralleled increasing levels of resistance to the drug. However, the expression of folylpolyglutamate synthetase RNA was not altered in the resistant cell lines compared to the parent cells. The most resistant cell subline also displayed an increased activity of -glutamyl hydrolase.

The sublines were scrutinized for other possible mechanisms of resistance. No alterations in drug transport or in purine economy were found. Modest increases were found in the activity of methylene tetrahydrofolate dehydrogenase but no alterations of other folate-dependent enzymes were observed. Increases in accumulation and conversion of folic acid to reduced forms, particularly 10-formyltetrahydrofolate, was also seen. The resistant cell lines were sensitive to dihydrofolate reductase inhibitors, methotrexate and trimetrexate, for a 72-h exposure period but showed cross-resistance to methotrexate for 4 and 24 h exposures. Cross-resistance was also shown toward other deazafolate analogues for both short- and long-term exposures.

¹ Supported by NIH Research Grants CA42367, CA57320, CA 25933, and RR01237.

² Special Fellow of the Leukemia Society of America.

³ Recipient of an American Cancer Society Faculty Research Award.

⁴ To whom requests for reprints should be addressed, at Yale University School of Medicine, Department of Pediatrics, 333 Cedar Street, P.O. Box 208064, New Haven, CT 06520-8064.

Received 7/ 8/94. Accepted 12/ 1/94.

This article has been cited by other articles:

				M. Stark, C. Wichman, I. Avivi, and Y. G. Assaraf Aberrant splicing of folylpolyglutamate synthetase as a novel mechanism of antifolate resistance in leukemia Blood, April 30, 2009; 113(18): 4362 - 4369. [Abstract] [Full Text] [PDF]

				S. Chattopadhyay, R. Zhao, S. A. Krupenko, N. Krupenko, and I. D. Goldman The inverse relationship between reduced folate carrier function and Pemetrexed activity in a human colon cancer cell line. Mol. Cancer Ther., February 1, 2006; 5(2): 438 - 449. [Abstract] [Full Text] [PDF]

				H. Li, T. J. Ryan, K. J. Chave, and P. Van Roey Three-dimensional Structure of Human gamma -Glutamyl Hydrolase. A CLASS I GLUTAMINE AMIDOTRANSFERASE ADAPTED FOR A COMPLEX SUBSTRATE J. Biol. Chem., June 28, 2002; 277(27): 24522 - 24529. [Abstract] [Full Text] [PDF]

				F. M. Sirotnak Enhancing Cytotoxic Sensitivity of Tumor Cells to Antifolates: Another Opportunity for Gene Therapy? J Natl Cancer Inst, July 21, 1999; 91(14): 1178 - 1179. [Full Text] [PDF]

				M. G. Rots, R. Pieters, G. J. Peters, P. Noordhuis, C. H. van Zantwijk, G. J.L. Kaspers, K. Hahlen, U. Creutzig, A. J.P. Veerman, and G. Jansen Role of Folylpolyglutamate Synthetase and Folylpolyglutamate Hydrolase in Methotrexate Accumulation and Polyglutamylation in Childhood Leukemia Blood, March 1, 1999; 93(5): 1677 - 1683. [Abstract] [Full Text] [PDF]

				A. Tse and R. G. Moran Cellular Folates Prevent Polyglutamation of 5,10-Dideazatetrahydrofolate. A NOVEL MECHANISM OF RESISTANCE TO FOLATE ANTIMETABOLITES J. Biol. Chem., October 2, 1998; 273(40): 25944 - 25952. [Abstract] [Full Text] [PDF]

				M. S. Rhee, B. Lindau-Shepard, K. J. Chave, J. Galivan, and T. J. Ryan Characterization of Human Cellular gamma -Glutamyl Hydrolase Mol. Pharmacol., June 1, 1998; 53(6): 1040 - 1046. [Abstract] [Full Text]

				M. C. Waltham, W.-W. Li, H. Gritsman, W. P. Tong, and J. R. Bertino gamma -Glutamyl Hydrolase from Human Sarcoma HT-1080 Cells: Characterization and Inhibition by Glutamine Antagonists Mol. Pharmacol., May 1, 1997; 51(5): 825 - 832. [Abstract] [Full Text]

				K. Roy, M. G. Egan, S. Sirlin, and F. M. Sirotnak Posttranscriptionally Mediated Decreases in Folylpolyglutamate Synthetase Gene Expression in Some Folate Analogue-resistant Variants of the L1210 Cell. EVIDENCE FOR AN ALTERED COGNATE mRNA IN THE VARIANTS AFFECTING THE RATE OF DE NOVO SYNTHESIS OF THE ENZYME J. Biol. Chem., March 14, 1997; 272(11): 6903 - 6908. [Abstract] [Full Text] [PDF]

				L. Chen, H. Qi, J. Korenberg, T. A. Garrow, Y.-J. Choi, and B. Shane Purification and Properties of Human Cytosolic Folylpoly-gamma -glutamate Synthetase and Organization, Localization, and Differential Splicing of Its Gene J. Biol. Chem., May 31, 1996; 271(22): 13077 - 13087. [Abstract] [Full Text] [PDF]

				R. Yao, Z. Nimec, T. J. Ryan, and J. Galivan Identification, Cloning, and Sequencing of a cDNA Coding for Rat [IMAGE]-Glutamyl Hydrolase J. Biol. Chem., April 12, 1996; 271(15): 8525 - 8528. [Abstract] [Full Text] [PDF]

				C. H. Takimoto New Antifolates: Pharmacology and Clinical Applications Oncologist, February 1, 1996; 1(1): 68 - 81. [Abstract] [Full Text] [PDF]

				R. Zhao, S. Titus, F. Gao, R. G. Moran, and I. D. Goldman Molecular Analysis of Murine Leukemia Cell Lines Resistant to 5,10-Dideazatetrahydrofolate Identifies Several Amino Acids Critical to the Function of Folylpolyglutamate Synthetase J. Biol. Chem., August 18, 2000; 275(34): 26599 - 26606. [Abstract] [Full Text] [PDF]