Antineoplastic drug sensitivity of human MCF-7 breast cancer cells stably transfected with a human alpha class glutathione S-transferase gene.
Studies have suggested that the alpha class glutathione S-transferase (GST) may protect cells from the chemotherapeutic drugs chlorambucil and melphalan. In order to further define the function of human alpha class GST, a complementary DNA which encodes it was ligated into an expression vector under the direction of the human metallothionein-IIA 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 alpha class GSTs. Southern blot analysis indicated that genomic DNA from these cells contained a fragment indistinguishable from the transfected alpha class GST complementary DNA (850 base pairs); Northern blot analysis of total cellular RNA indicated that these cells contained appropriately sized alpha class GST RNA (980 nucleotides); and Western blot analysis indicated that, while MCF-7 cells contained no detectable alpha class GST protein, the transfected cells contained markedly elevated levels of alpha class GST but no detectable mu or pi class GST. These alpha 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 alpha 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.
Leyland-Jones
B R
BR
Medicine Branch, National Cancer Institute, National Institutes of Health, Bethesda, Maryland 20892.
Townsend
A J
AJ
Tu
C P
CP
Cowan
K H
KH
Goldsmith
M E
ME
D000970:Antineoplastic Agents / Q000494:pharmacology*; D001483:Base Sequence; D001943:Breast Neoplasms; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects*; D002999:Clone Cells; D005260:Female; D015723:Gene Library; D005822:Genetic Vectors; D005978:Glutathione / Q000378:metabolism; D005979:Glutathione Peroxidase / Q000378:metabolism; D005982:Glutathione Transferase / Q000235:genetics* / Q000378:metabolism; D006801:Humans; D007700:Kinetics; D008099:Liver / Q000201:enzymology; D015183:Restriction Mapping; D014162:Transfection
D000970:Antineoplastic Agents; D005979:Glutathione Peroxidase; D005982:Glutathione Transferase; D005978:Glutathione