Loss of reduced folate carrier function and folate depletion result in enhanced pemetrexed inhibition of purine synthesis.
Pemetrexed is a novel antifolate with polyglutamate derivatives that are potent inhibitors of thymidylate synthase (TS) and to a lesser extent glycinamide ribonucleotide formyltransferase (GARFT). Conditions that might modulate relative suppression of these sites were assessed by the pattern of hypoxanthine and thymidine protection. When grown with 25 nmol/L racemic 5-formyltetrahydrofolate, thymidine alone fully protected wild-type HeLa cells to at least 1 micromol/L pemetrexed, but protection of a reduced folate carrier (RFC)-null subline required both thymidine and hypoxanthine above a concentration of 30 nmol/L pemetrexed. As medium 5-formyltetrahydrofolate was decreased, protection by thymidine alone decreased, and was further diminished when HeLa cells were grown in dialyzed serum. There was little protection by thymidine of RFC-null HeLa cells under the latter conditions. Thymidine alone was not protective, and hypoxanthine alone produced only a small (2-fold) increase in IC(50), in a HeLa-derived line 8-fold resistant to pemetrexed due to a modest increase in TS. Finally, in MCF-7 breast cancer cells there was greater protection with thymidine alone than in HeLa cells when cells were grown in medium containing a low concentration of 5-formyltetrahydrofolate. These observations indicate that as intracellular folates decrease in HeLa cells, due to decreased extracellular reduced folate, or loss of RFC function, pemetrexed inhibition of GARFT increases. These data support the concept that the contribution to pemetrexed activity by inhibition of GARFT, particularly at low folate levels, is a contributing factor to drug activity but relative inhibition of TS and GARFT may vary among human tumors and cell lines.
Zhao
Rongbao
R
Departments of Medicine and Molecular Pharmacology, the Albert Einstein College of Medicine and Cancer Research Center, 1300 Morris Park Avenue, Bronx, NY 10461, USA.
Zhang
Shubing
S
Hanscom
Marie
M
Chattopadhyay
Shrikanta
S
Goldman
I David
ID
D000818:Animals; D015153:Blotting, Western; D002417:Cattle; D002455:Cell Division / Q000187:drug effects; D045744:Cell Line, Tumor; D003470:Culture Media / Q000737:chemistry / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D005492:Folic Acid / Q000378:metabolism*; D005493:Folic Acid Antagonists / Q000494:pharmacology*; D005971:Glutamates / Q000494:pharmacology*; D006147:Guanine / Q000031:analogs & derivatives* / Q000494:pharmacology*; D006367:HeLa Cells; D006801:Humans; D019271:Hypoxanthine / Q000494:pharmacology; D002955:Leucovorin / Q000494:pharmacology; D026901:Membrane Transport Proteins / Q000502:physiology; D000068437:Pemetrexed; D011687:Purines / Q000096:biosynthesis*; D058978:Reduced Folate Carrier Protein; D044967:Serum / Q000737:chemistry; D013940:Thymidylate Synthase / Q000378:metabolism; D013941:Thymine / Q000494:pharmacology
D003470:Culture Media; D005493:Folic Acid Antagonists; D005971:Glutamates; D026901:Membrane Transport Proteins; D011687:Purines; D058978:Reduced Folate Carrier Protein; C548699:SLC19A1 protein, human; D000068437:Pemetrexed; D019271:Hypoxanthine; D006147:Guanine; D005492:Folic Acid; D013940:Thymidylate Synthase; D002955:Leucovorin; D013941:Thymine; C030985:purine