Overexpression of stably transfected human glutathione S-transferase P1-1 protects against DNA damage by benzo[a]pyrene diol-epoxide in human T47D cells.
The (+)-anti enantiomer of benzo[a]pyrene-7,8-dihydrodiol-9, 10-epoxide (BPDE) is a potent mutagenic and carcinogenic metabolite of benzo[a]pyrene (BP), and a major fraction is conjugated with glutathione in vivo. The chemopreventive role of glutathione S-transferases (GSTs) in protecting against covalent modification of DNA and other cellular macromolecules by BPDE was modeled in human T47D and MCF-7 cell lines previously stably transfected with human GSTpi1 (hGSTP1). Cells were exposed to [3H]BPDE (30-600 nM). Dose-response experiments indicated that the high level of expression of hGSTP1-1 in the T47Dpi cell line (4411 +/- 183 milliunits/mg of cytosolic protein, using 1-Cl-2,4-dinitrobenzene as substrate), resulted in 70-90% reduction in the covalent 3H-adduct formation in DNA or RNA isolated from the GSTP1-transfected T47Dpi cell line. The lower level of hGSTP1-1 expression in the transfected MCF-7 cell line (91 milliunits/mg) provided only marginal protection against [3H]BPDE adduct formation and did not affect sensitivity to BPDE-induced cytotoxicity. Protection against BPDE-induced cytotoxicity was observed only in the T47Dpi cell line, which had an IC50 value 5.8-fold greater than that of the T47Dneo control cell line. Measurement of glutathione conjugates of BPDE indicated that the total conjugation was 5-fold higher in the GSTpi-transfected T47D line, most of which was exported into the culture medium over the 20-min exposure period. These results indicate that hGSTP1-1 protects effectively against DNA and RNA modification by BPDE, but moderate to high level expression may be required for strong protection against BPDE-induced genotoxicity and cytotoxicity.
Fields
W R
WR
Biochemistry Department and Wake Forest University Comprehensive Cancer Center, Wake Forest University School of Medicine, Winston-Salem, North Carolina 27157, USA.
Morrow
C S
CS
Doss
A J
AJ
Sundberg
K
K
Jernström
B
B
Townsend
A J
AJ
D015123:7,8-Dihydro-7,8-dihydroxybenzo(a)pyrene 9,10-oxide / Q000494:pharmacology*; D016587:Antimutagenic Agents / Q000494:pharmacology; D002460:Cell Line; D018736:DNA Adducts; D004249:DNA Damage; D051549:Glutathione S-Transferase pi; D005982:Glutathione Transferase / Q000502:physiology*; D006801:Humans; D007527:Isoenzymes / Q000502:physiology*; D009153:Mutagens / Q000494:pharmacology*; D014162:Transfection
D016587:Antimutagenic Agents; D018736:DNA Adducts; D007527:Isoenzymes; D009153:Mutagens; D015123:7,8-Dihydro-7,8-dihydroxybenzo(a)pyrene 9,10-oxide; C496556:GSTP1 protein, human; D051549:Glutathione S-Transferase pi; D005982:Glutathione Transferase