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Antineoplastic Drug Sensitivity of Human MCF-7 Breast Cancer Cells Stably Transfected with a Human Class Glutathione S-Transferase Gene

Medicine Branch, National Cancer Institute, National Institutes of Health, Bethesda, Maryland 20892 [B. R. L.-J., A. J. T., K. H. C., M. E. G.], and Department of Molecular and Cell Biology, The Pennsylvania State University, University Park, Pennsylvania 16802 [C.-P. D. T.]

Studies have suggested that the class glutathione S-transferase (GST) may protect cells from the chemotherapeutic drugs chlorambucil and melphalan. In order to further define the function of human class GST, a complementary DNA which encodes it was ligated into an expression vector under the direction of the human metallothionein-II_A promoter and stably transfected into human MCF-7 breast cancer cells in conjunction with the G418-selectable plasmid pSV2neo. Clonal cell lines were identified which expressed increased levels of GST enzyme activity (2.2- to 5.6-fold). The transfected cell lines also had increased peroxidase activity using cumene hydroperoxide as the substrate (1.9- to 3.8-fold) which is consistent with the intrinsic peroxidase activity of class GSTs. Southern blot analysis indicated that genomic DNA from these cells contained a fragment indistinguishable from the transfected class GST complementary DNA (850 base pairs); Northern blot analysis of total cellular RNA indicated that these cells contained appropriately sized class GST RNA (980 nucleotides); and Western blot analysis indicated that, while MCF-7 cells contained no detectable class GST protein, the transfected cells contained markedly elevated levels of class GST but no detectable µ or class GST. These class GST transfected cells had increased resistance to ethacrynic acid (2.1- to 3.0-fold). However, the transfected cells failed to show any increased resistance measured at the drug dosage which inhibited 50% of the colony formation to the chemotherapeutic drugs chlorambucil, melphalan, Adriamycin, or cisplatin under conditions of either continuous or 1-h drug exposure. Neither was there any change in sensitivity to the cytotoxins benzo(a)pyrene, benzo(a)pyrene-trans-7,8-dihydrodiol-9,10-epoxide (anti), or 1-chloro-2,4-dinitrobenzene. These studies indicate that expression of this human class GST by itself in MCF-7 human breast cancer cells does not confer resistance to the chemotherapeutic drugs tested under the conditions used in these studies.

¹ Present address: Department of Oncology, McGill University, Montreal, PQ, Canada H3G1Y6.

² Present address: Department of Biochemistry, Bowman-Gray School of Medicine, Winston-Salem, NC 27127.

³ To whom requests for reprints should be addressed, at Medicine Branch, National Cancer Institute, National Institutes of Health, Bldg. 10, Rm. 12N226, Bethesda, MD 20892.

Received 8/ 3/90. Accepted 11/ 1/90.

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