Ebselen augments its peroxidase activity by inducing nrf-2-dependent transcription.
Ebselen is an organoselenium compound that acts as a glutathione peroxidase mimic. Since ebselen is a hydrophobic, thio-reactive compound capable of interacting with Keap-1, we tested its ability to activate nrf-2-dependent responses in the human hepatocarcinoma derived cell line, HepG2. Ebselen (25 microM) increased expression of an nrf-2 response element reporter in transient transfection experiments by 4-fold. Although, the induction was lower than that observed with classic nrf-2 inducer, sulforaphane (10 microL; 7-fold), ebselen also induced expression of native NAD(P)H:quinone oxidoreductase (1.6-fold) activity; induction of this protein is known to be dependent on nrf-2 action. Treatment of HepG2 cells with ebselen increased glutathione levels after 12 (1.5-fold) or 24 (1.9-fold)h of treatment. Treatment of the cells with either sulforaphane or ebselen 24 h prior to treatment with varying concentrations of t-butyl hydroperoxide increased the half maximal lethal dose from 28 to 42 microM and 58 microM for sulforaphane and ebselen, respectively. The protective effects of ebselen treatment were greater with pretreatment (IC50=58 microM) than simultaneous addition (IC50=45 microM). The protein synthesis inhibitor cycloheximide blocked increases in intracellular glutathione synthesis and partially blocked the protective effects of this regimen on increasing cell survival following t-butyl hydroperoxide treatment. Likewise co-treatment with the MEK 1 inhibitor, PD98059, which has been shown to inhibit nrf-2-dependent gene activation, partially inhibited the ebselen-dependent increases in IC50 while not affecting the control cells. We conclude that nrf-2 activation augments the role of ebselen as an antioxidant or by indirect induction of cellular antioxidant defences.
Department of Biochemistry and Molecular Biology, University of Louisville School of Medicine, Louisville, KY 40292, USA.
Viola
V
Tamasi
Julianne M
JM
Jeffries
Gavin E
GE
Arteel
K Cameron
KC
Falkner
D000975:Antioxidants / Q000494:pharmacology*; D001393:Azoles / Q000494:pharmacology*; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D004268:DNA-Binding Proteins / Q000187:drug effects* / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation; D051268:GA-Binding Protein Transcription Factor; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D017930:Genes, Reporter; D005978:Glutathione / Q000032:analysis / Q000187:drug effects / Q000378:metabolism; D005979:Glutathione Peroxidase / Q000378:metabolism*; D018197:Hepatoblastoma / Q000378:metabolism; D006801:Humans; D054833:Isoindoles; D007700:Kinetics; D008156:Luciferases / Q000378:metabolism; D016566:Organoselenium Compounds / Q000494:pharmacology*; D020218:Response Elements / Q000187:drug effects; D013880:Thioredoxin-Disulfide Reductase / Q000378:metabolism; D014157:Transcription Factors / Q000187:drug effects* / Q000378:metabolism; D014158:Transcription, Genetic; D015533:Transcriptional Activation
D000975:Antioxidants; D001393:Azoles; D004268:DNA-Binding Proteins; D051268:GA-Binding Protein Transcription Factor; D054833:Isoindoles; D016566:Organoselenium Compounds; D014157:Transcription Factors; C042986:ebselen; D005979:Glutathione Peroxidase; D008156:Luciferases; D013880:Thioredoxin-Disulfide Reductase; D005978:Glutathione