title: Antifolate resistance associated with loss of MRP1 expression and function in Chinese hamster ovary cells with markedly impaired export of folate and cholate.
abstract: Export of folates from a Chinese hamster ovary PyrR100 cell line is markedly impaired, resulting in expansion of cellular folate pools and high-level antifolate resistance. We now report that MRP1 expression is absent in PyrR100 cells along with a marked decrease in MRP5 expression with 3-fold cross-resistance to thiopurines. PyrR100 and wild-type cells had comparable low levels of MRP2 expression; both lacked the breast cancer resistance protein. PyrR100 cells showed a 4-fold decrease in cholate (an MRP substrate) efflux with a 6-fold increase in cellular cholate accumulation compared with wild-type cells. Prostaglandin A1 increased cholate accumulation in wild-type cells to levels comparable with PyrR100 cells. Calcein (an MRP1 substrate) fluorescence increased 5-fold in PyrR100 cells; probenecid increased the intracellular calcein level in wild-type cells to that of PyrR100 cells. Consistent with the loss of MRP1 expression, PyrR100 cells showed modest collateral sensitivity to cholate, etoposide, doxorubicin, and vincristine. Transfection of MRP5 into PyrR100 cells did not alter sensitivity to pyrimethamine or MTX but restored sensitivity to mercaptopurines, indicating that decreased MRP5 expression did not play a role in antifolate resistance. Hence, although MRP-mediated anticancer drug resistance has been associated with gain of function (i.e., overexpression), this is the first report that loss of MRP1 efflux function can expand intracellular folate pools to result in acquired antifolate resistance. The data also suggest that MRP1, and possibly other MRPs that transport folates, can play a role in the maintenance of cellular folate homeostasis.
authors.affiliation: Department of Biology, The Technion-Israel Institute of Technology, Haifa 32000, Israel.
authors.forename: Michal
authors.initials: M
authors.lastname: Stark
authors.forename: Lilah
authors.initials: L
authors.lastname: Rothem
authors.forename: Gerrit
authors.initials: G
authors.lastname: Jansen
authors.forename: George L
authors.initials: GL
authors.lastname: Scheffer
authors.forename: I David
authors.initials: ID
authors.lastname: Goldman
authors.forename: Yehuda G
authors.initials: YG
authors.lastname: Assaraf
mesh_terms: D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D000818:Animals; D001692:Biological Transport; D016466:CHO Cells; D002455:Cell Division / Q000187:drug effects; D002478:Cells, Cultured; D020355:Cholates / Q000378:metabolism*; D006224:Cricetinae; D004351:Drug Resistance / Q000502:physiology*; D005434:Flow Cytometry; D005452:Fluoresceins / Q000378:metabolism; D005492:Folic Acid / Q000378:metabolism*; D006801:Humans; D008727:Methotrexate / Q000494:pharmacology; D027425:ATP-Binding Cassette, Sub-Family C Proteins / Q000235:genetics / Q000378:metabolism* / Q000502:physiology; D011454:Prostaglandins A / Q000494:pharmacology; D011739:Pyrimethamine / Q000494:pharmacology*; D014316:Tritium
chemical_list: D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D020355:Cholates; D005452:Fluoresceins; D005492:Folic Acid; D008727:Methotrexate; D027425:ATP-Binding Cassette, Sub-Family C Proteins; D011454:Prostaglandins A; D011739:Pyrimethamine; D014316:Tritium; C120809:ABCC5 protein, human; C085925:calcein AM; C100573:prostaglandin A1; C421109:multidrug resistance-associated protein 1