Decreased melphalan accumulation in a human breast cancer cell line selected for resistance to melphalan.
An in vitro model of acquired melphalan resistance was developed by serial incubation of an MCF-7 human breast cancer cell line in increasing concentrations of melphalan. The resulting derivative cell line, Me1R MCF-7, was 30-fold resistant to melphalan. Uptake studies demonstrated decreased initial melphalan accumulation in Me1R MCF-7 cells. Inverse-reciprocal plots of initial melphalan uptake revealed a 4-fold decrease in the apparent Vmax of Me1R MCF-7 compared with WT MCF-7 (516 amol cell-1 min-1 vs 2110 amol cell-1 min-1 respectively) as well as a decrease in the apparent Kt (36 microM vs 70 microM respectively). Two amino acid transporters have previously been identified as melphalan transporters: system L, which is sodium-independent and inhibited by 2-amino-bicyclo[2,2,1]heptane-2-carboxylic acid (BCH), and system ASC which is sodium dependent and unaffected by BCH. At low concentrations of melphalan (3-30 microM), 1mM BCH competition eliminated the differences between the two cell lines, thus implicating an alteration of the system L transporter in the transport defect in the resistant cells. Me1R MCF-7 cells were also evaluated for glutathione-mediated detoxification mechanisms associated with melphalan resistance. There was no difference between Me1R MCF-7 and WT MCF-7 in glutathione content, glutathione-S-transferase activity and expression of pi class glutathione S-transferase RNA. In addition, buthionine sulfoximine did not reverse melphalan resistance in Me1R MCF-7 cells. Therefore, Me1R MCF-7 cells provide an in vitro model of transport-mediated melphalan resistance in human breast cancer cells.
Medical Breast Cancer Section, National Cancer Institute.
J A
JA
Moscow
C A
CA
Swanson
K H
KH
Cowan
D026905:Amino Acid Transport Systems; D000596:Amino Acids / Q000494:pharmacology; D000598:Amino Acids, Cyclic; D001943:Breast Neoplasms / Q000378:metabolism*; D002352:Carrier Proteins / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004351:Drug Resistance; D005978:Glutathione / Q000378:metabolism; D005982:Glutathione Transferase / Q000378:metabolism; D006801:Humans; D008558:Melphalan / Q000493:pharmacokinetics*; D013997:Time Factors; D014407:Tumor Cells, Cultured
D026905:Amino Acid Transport Systems; D000596:Amino Acids; D000598:Amino Acids, Cyclic; D002352:Carrier Proteins; C006504:2-aminobicyclo(2,2,1)heptane-2-carboxylic acid; D005982:Glutathione Transferase; D005978:Glutathione; D008558:Melphalan