title: Up-regulation of glutathione-related genes, enzyme activities and transport proteins in human cervical cancer cells treated with doxorubicin.
abstract: Doxorubicin (DOX), one of the most effective anticancer drugs, acts in a variety of ways including DNA damage, enzyme inhibition and generation of reactive oxygen species. Glutathione (GSH) and glutathione-related enzymes including: glutathione peroxidase (GPX), glutathione reductase (GSR) and glutathione S-transferases (GST) may play a role in adaptive detoxification processes in response to the oxidative stress, thus contributing to drug resistance phenotype. In this study, we investigated effects of DOX treatment on expression and activity of GSH-related enzymes and multidrug resistance-associated proteins in cultured human cervical cancer cells displaying different resistance against this drug (HeLa and KB-V1). Determination of expression level of genes encoding GST isoforms and MRP proteins (GCS, GPX, GSR, GSTA1-3, GSTM1, GSTP1, ABCC1-3, MGST1-3) was performed using StellARray™ Technology. Enzymatic activities of GPX and GSR were measured using biochemical methods. Expression of MRP1 was examined by immunofluorescence microscopy. This study showed that native expression levels of GSTM1 and GSTA3 were markedly higher in KB-V1 cells (2000-fold and 200-fold) compared to HeLa cells. Resistant cells have also shown significantly elevated expression of GSTA1 and GSTA2 genes (200-fold and 50-fold) as a result of DOX treatment. In HeLa cells, exposure to DOX increased expression of all genes: GSTM1 (7-fold) and GSTA1-3 (550-fold, 150-fold and 300-fold). Exposure to DOX led to the slight increase of GCS expression as well as GPX activity in KB-V1 cells, while in HeLa cells it did not. Expression of ABCC1 (MRP1) was not increased in any of the tested cell lines. Our results indicate that expression of GSTM1 and GSTA1-3 genes is up-regulated by DOX treatment and suggest that activity of these genes may be associated with drug resistance of the tested cells. At the same time, involvement of MRP1 in DOX resistance in the given experimental conditions is unlikely.
authors.affiliation: National Medicines Institute, Department of Biochemistry and Biopharmaceuticals, Chełmska Str. 30/34, 00-725 Warsaw, Poland. Electronic address: e.drozd@nil.gov.pl.
authors.forename: Ewa
authors.initials: E
authors.lastname: Drozd
authors.affiliation: National Medicines Institute, Department of Biochemistry and Biopharmaceuticals, Chełmska Str. 30/34, 00-725 Warsaw, Poland.
authors.forename: Jolanta
authors.initials: J
authors.lastname: Krzysztoń-Russjan
authors.affiliation: National Medicines Institute, Department of Biochemistry and Biopharmaceuticals, Chełmska Str. 30/34, 00-725 Warsaw, Poland.
authors.forename: Jadwiga
authors.initials: J
authors.lastname: Marczewska
authors.affiliation: National Medicines Institute, Department of Biochemistry and Biopharmaceuticals, Chełmska Str. 30/34, 00-725 Warsaw, Poland.
authors.forename: Janina
authors.initials: J
authors.lastname: Drozd
authors.affiliation: National Medicines Institute, Department of Biochemistry and Biopharmaceuticals, Chełmska Str. 30/34, 00-725 Warsaw, Poland.
authors.forename: Irena
authors.initials: I
authors.lastname: Bubko
authors.affiliation: National Medicines Institute, Department of Biochemistry and Biopharmaceuticals, Chełmska Str. 30/34, 00-725 Warsaw, Poland.
authors.forename: Magda
authors.initials: M
authors.lastname: Bielak
authors.affiliation: National Medicines Institute, Department of Cell Biology, Chełmska Str. 30/34, 00-725 Warsaw, Poland.
authors.forename: Katarzyna
authors.initials: K
authors.lastname: Lubelska
authors.affiliation: National Medicines Institute, Department of Cell Biology, Chełmska Str. 30/34, 00-725 Warsaw, Poland.
authors.forename: Katarzyna
authors.initials: K
authors.lastname: Wiktorska
authors.affiliation: National Medicines Institute, Department of Cell Biology, Chełmska Str. 30/34, 00-725 Warsaw, Poland.
authors.forename: Zdzisław
authors.initials: Z
authors.lastname: Chilmonczyk
authors.affiliation: National Medicines Institute, Department of Biochemistry and Biopharmaceuticals, Chełmska Str. 30/34, 00-725 Warsaw, Poland.
authors.forename: Elżbieta
authors.initials: E
authors.lastname: Anuszewska
authors.affiliation: National Medicines Institute, Department of Biochemistry and Biopharmaceuticals, Chełmska Str. 30/34, 00-725 Warsaw, Poland.
authors.forename: Beata
authors.initials: B
authors.lastname: Gruber-Bzura
mesh_terms: D002352:Carrier Proteins / Q000235:genetics* / Q000378:metabolism; D045744:Cell Line, Tumor; D004317:Doxorubicin / Q000494:pharmacology*; D005260:Female; D005455:Fluorescent Antibody Technique; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D005978:Glutathione / Q000378:metabolism*; D005979:Glutathione Peroxidase / Q000235:genetics / Q000378:metabolism; D005980:Glutathione Reductase / Q000235:genetics / Q000378:metabolism; D006801:Humans; D027425:ATP-Binding Cassette, Sub-Family C Proteins / Q000235:genetics / Q000378:metabolism; D015854:Up-Regulation / Q000187:drug effects*; D002583:Uterine Cervical Neoplasms / Q000201:enzymology* / Q000235:genetics*
chemical_list: D002352:Carrier Proteins; D004317:Doxorubicin; D005978:Glutathione; D005979:Glutathione Peroxidase; D005980:Glutathione Reductase; D027425:ATP-Binding Cassette, Sub-Family C Proteins; C421109:multidrug resistance-associated protein 1
keywords: Cervical cancer; Doxorubicin; GST; Glutathione; Resistance