Lack of an effect of breast cancer resistance protein (BCRP/ABCG2) overexpression on methotrexate polyglutamate export and folate accumulation in a human breast cancer cell line.
Accumulation of methotrexate (MTX) and its polyglutamates (PGs) has been recognized as an important factor in MTX efficacy. We have previously described a multidrug-resistant human breast cancer cell line, MCF7/MX, that exhibits reduced accumulation of total MTX as well as MTX-PGs, and that is resistant to continuous MTX exposure [Volk EL, Rohde K, Rhee M, McGuire JJ, Doyle LA, Ross DD, et al. Methotrexate cross-resistance in a mitoxantrone selected multidrug-resistant MCF7 breast cancer cell line is due to enhanced energy-dependent drug efflux. Cancer Res 2000;60:3514-21]. These cells express high levels of the breast cancer resistance protein (BCRP/ABCG2) that has been shown to actively transport MTX and short-chain MTX-PGs in vitro. However, the effect of BCRP on MTX-PG accumulation in intact cells was unclear. Here, we show that MTX transport by BCRP is required for the observed lower levels of MTX-PGs in the resistant cells. When BCRP was inhibited with fumitremorgin C, or in cells expressing a mutated form of BCRP that is unable to transport MTX, MTX-PG accumulation was similar or even higher than that in the parental cells that do not express BCRP. Concomitantly, there was increased inhibition of thymidylate synthase. It had previously been suggested that BCRP-mediated efflux of MTX-PGs contributed to the reduced MTX-PG accumulation. However, we found no evidence of BCRP-mediated efflux of MTX-PGs from intact cells, suggesting that direct efflux of MTX-PGs does not play a major role in MTX resistance. Together, these data show that BCRP overexpression can cause a reduction in total MTX accumulation as well as a reduction in the proportion of long-chain MTX-PGs. In contrast, BCRP overexpression did not affect natural folate accumulation or the relative distribution of folylpolyglutamates in the resistant, as compared to the parental, cells. Thus, it appears that BCRP overexpression affects the metabolism of the antifolate MTX, but not that of natural folates, although indirect effects cannot be excluded.
Rhee
Myung S
MS
Wadsworth Center, New York State Department of Health, Empire State Plaza, Albany, NY 12201, USA.
Schneider
Erasmus
E
D000070997:ATP Binding Cassette Transporter, Subfamily G, Member 2; D018528:ATP-Binding Cassette Transporters / Q000096:biosynthesis*; D001943:Breast Neoplasms / Q000378:metabolism*; D045744:Cell Line, Tumor; D004305:Dose-Response Relationship, Drug; D005492:Folic Acid / Q000378:metabolism*; D015972:Gene Expression Regulation, Neoplastic / Q000502:physiology*; D006801:Humans; D008727:Methotrexate / Q000031:analogs & derivatives* / Q000378:metabolism*; D009363:Neoplasm Proteins / Q000096:biosynthesis*; D011099:Polyglutamic Acid / Q000031:analogs & derivatives* / Q000378:metabolism*
C116614:ABCG2 protein, human; D000070997:ATP Binding Cassette Transporter, Subfamily G, Member 2; D018528:ATP-Binding Cassette Transporters; D009363:Neoplasm Proteins; D011099:Polyglutamic Acid; C014085:methotrexate polyglutamate; D005492:Folic Acid; D008727:Methotrexate