Abstract
Overexpression of ATP-binding cassette (ABC) transporters causing multidrug resistance (MDR) in cancer cells is one of the major obstacles in cancer chemotherapy. The 5-FU resistant subclone (HL-60/5FU) of the human HL-60 promyelocytic leukemia cell line was selected by the conventional method of continuous exposure of the cells to the drug up to 0.08 mmol/L concentration. HL-60/5FU cells exhibited six-fold enhanced resistance to 5-FU than HL-60 cells. RT-PCR and ELISA assay showed significant overexpression of MDR-related ABC transporters, ABCB1, ABCG2 but especially ABCC1 in the HL-60/5FU as compared with the parental cell line. Three novel synthetic 5-methylidenedihydrouracil analogs, U-236, U-332 and U-359, selected as highly cytotoxic for HL-60 cells in MTT test, showed similar cytotoxicity in the resistant cell line. When co-incubated with 5-FU, these analogs were found to down-regulate the expression of all three transporters. However, the most pronounced effect was caused by U-332 which almost completely abolished ABCC1 expression in the resistant HL-60/5FU cells. Additionally, U-332 inhibited the activity of ATPase, an enzyme which catalyzes hydrolysis of ATP, providing energy to efflux drugs from the cells through the cellular membranes. Taken together, the obtained data suggest that acquired 5-FU resistance in HL-60/5FU cells results from overexpression of ABCC1 and that targeting ABCC1 expression could be a potential approach to re-sensitize resistant leukemia cells to 5-FU. The synthetic uracil analog U-332, which can potently down-regulate ABC transporter expression and therefore disturb drug efflux, can be considered an efficient ABCC1 regulator in cancer cells.
Highlights
Acute myeloid leukemia (AML), characterized as a heterogeneous clonal disorder of hematopoietic progenitor cells, is known to be a frequent cause of cancer-related deaths [1]
Among them P-glycoprotein (ABCB1), multidrug resistance-associated protein 1 (ABCC1) and breast cancer resistance protein (ABCG2) are three best known transmembrane proteins from the ATP-binding cassette (ABC) family that in many cases reduce the cellular uptake of drugs into cancer cells, defending them from medical interventions [7, 8]
ABC transporters mediate the transport of substrates against a concentration gradient using energy derived from ATP hydrolysis, which is proportional to the transporter activity and could be detected by a luminescence method
Summary
Acute myeloid leukemia (AML), characterized as a heterogeneous clonal disorder of hematopoietic progenitor cells, is known to be a frequent cause of cancer-related deaths [1]. The ABC transporter family consists of transmembrane proteins that use the energy from ATP hydrolysis to efflux various potentially dangerous compounds of diverse structure across a cell membrane [3,4,5]. While such efflux is a normal physiological process, it is a known mechanism of drug resistance in cancer cells. Among them P-glycoprotein (ABCB1), multidrug resistance-associated protein 1 (ABCC1) and breast cancer resistance protein (ABCG2) are three best known transmembrane proteins from the ABC family that in many cases reduce the cellular uptake of drugs into cancer cells, defending them from medical interventions [7, 8]
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