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

Brian R. Lcyland-Jones, Alan J. Townsend, Kenneth H. Cowan, Merrill E. Goldsmith

Research output: Contribution to journalArticle

65 Citations (Scopus)

Abstract

Studies haâ»csuggested that the a class glutathione S-transferase (GST) may protect cells from the chemotherapeutic drugs chlorambucil and melphalan. In order to further define the function of humana class GST, a complementary DNA which encodes it was (¡gatedinto 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 eumene hydroperoxide as the substrate (1.9-to 3.8-fold) which is consistent with the intrinsic peroxidase activity of a class GSTs. Southern blot analysis indicated that genomic DNA from these cells contained a fragment indistinguishable from the transfected a class GST complementary DNA (850 base pairs); Northern blot analysis of total cellular RN'Aindicated that these cells contained appropriately sized a class GST RNA (980 nucleotides); and Western blot analysis indicated that, while MCF-7 cells contained no detectable a class GST protein, the transfected cells contained markedly elevated levels of a class GST but no detectable in or ir class GST. These a 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 resist ance measured at the drug dosage which inhibited 50% of the colony formation to the chemotherapeutic drugs chlorambucil, melphalan, Adri amycin, 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-fâ«r/iã-7,8-dihydrodâ¡ol-9,10-epoxâ¡de unni ).or l-chloro-2.4-dinitrobenzene. These studies indicate that expres sion of this humana 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.

Original languageEnglish (US)
Pages (from-to)587-594
Number of pages8
JournalCancer Research
Volume51
Issue number2
StatePublished - Jan 1991

Fingerprint

Glutathione Transferase
Antineoplastic Agents
Breast Neoplasms
Genes
Chlorambucil
Melphalan
Pharmaceutical Preparations
Benzo(a)pyrene
Peroxidase
Complementary DNA
Dinitrobenzenes
Ethacrynic Acid
Cell Line
Metallothionein
MCF-7 Cells
Cytotoxins
Southern Blotting
Base Pairing
Northern Blotting
Hydrogen Peroxide

ASJC Scopus subject areas

  • Oncology
  • Cancer Research

Cite this

Antineoplastic Drug Sensitivity of Human MCF-7 Breast Cancer Cells Stably Transfected with a Human α Class Glutathione S-Transferase Gene. / Lcyland-Jones, Brian R.; Townsend, Alan J.; Cowan, Kenneth H.; Goldsmith, Merrill E.

In: Cancer Research, Vol. 51, No. 2, 01.1991, p. 587-594.

Research output: Contribution to journalArticle

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abstract = "Studies ha{\^a}»csuggested that the a class glutathione S-transferase (GST) may protect cells from the chemotherapeutic drugs chlorambucil and melphalan. In order to further define the function of humana class GST, a complementary DNA which encodes it was ({\^A}¡gatedinto 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 eumene hydroperoxide as the substrate (1.9-to 3.8-fold) which is consistent with the intrinsic peroxidase activity of a class GSTs. Southern blot analysis indicated that genomic DNA from these cells contained a fragment indistinguishable from the transfected a class GST complementary DNA (850 base pairs); Northern blot analysis of total cellular RN'Aindicated that these cells contained appropriately sized a class GST RNA (980 nucleotides); and Western blot analysis indicated that, while MCF-7 cells contained no detectable a class GST protein, the transfected cells contained markedly elevated levels of a class GST but no detectable in or ir class GST. These a 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 resist ance measured at the drug dosage which inhibited 50{\%} of the colony formation to the chemotherapeutic drugs chlorambucil, melphalan, Adri amycin, 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-f{\^a}«r/i{\~a}-7,8-dihydrod{\^a}¡ol-9,10-epox{\^a}¡de unni ).or l-chloro-2.4-dinitrobenzene. These studies indicate that expres sion of this humana 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.",
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