title: Inhibition of GSK-3β activity can result in drug and hormonal resistance and alter sensitivity to targeted therapy in MCF-7 breast cancer cells.
abstract: The PI3K/Akt/mTORC1 pathway plays prominent roles in malignant transformation, prevention of apoptosis, drug resistance, and metastasis. One molecule regulated by this pathway is GSK-3β. GSK-3β is phosphorylated by Akt on S9, which leads to its inactivation; however, GSK-3β also can regulate the activity of the PI3K/Akt/mTORC1 pathway by phosphorylating molecules such as PTEN, TSC2, p70S6K, and 4E-BP1. To further elucidate the roles of GSK-3β in chemotherapeutic drug and hormonal resistance of MCF-7 breast cancer cells, we transfected MCF-7 breast cancer cells with wild-type (WT), kinase-dead (KD), and constitutively activated (A9) forms of GSK-3β. MCF-7/GSK-3β(KD) cells were more resistant to doxorubicin and tamoxifen compared with either MCF-7/GSK-3β(WT) or MCF-7/GSK-3β(A9) cells. In the presence and absence of doxorubicin, the MCF-7/GSK-3β(KD) cells formed more colonies in soft agar compared with MCF-7/GSK-3β(WT) or MCF-7/GSK-3β(A9) cells. In contrast, MCF-7/GSK-3β(KD) cells displayed an elevated sensitivity to the mTORC1 blocker rapamycin compared with MCF-7/GSK-3β(WT) or MCF-7/GSK-3β(A9) cells, while no differences between the 3 cell types were observed upon treatment with a MEK inhibitor by itself. However, resistance to doxorubicin and tamoxifen were alleviated in MCF-7/GSK-3β(KD) cells upon co-treatment with an MEK inhibitor, indicating regulation of this resistance by the Raf/MEK/ERK pathway. Treatment of MCF-7 and MCF-7/GSK-3β(WT) cells with doxorubicin eliminated the detection of S9-phosphorylated GSK-3β, while total GSK-3β was still detected. In contrast, S9-phosphorylated GSK-3β was still detected in MCF-7/GSK-3β(KD) and MCF-7/GSK-3β(A9) cells, indicating that one of the effects of doxorubicin on MCF-7 cells was suppression of S9-phosphorylated GSK-3β, which could result in increased GSK-3β activity. Taken together, these results demonstrate that introduction of GSK-3β(KD) into MCF-7 breast cancer cells promotes resistance to doxorubicin and tamoxifen, but sensitizes the cells to mTORC1 blockade by rapamycin. Therefore GSK-3β is a key regulatory molecule in sensitivity of breast cancer cells to chemo-, hormonal, and targeted therapy.
authors.affiliation: Department of Microbiology and Immunology; Brody School of Medicine at East Carolina University; Greenville, NC USA.
authors.forename: Melissa
authors.initials: M
authors.lastname: Sokolosky
authors.affiliation: Department of Microbiology and Immunology; Brody School of Medicine at East Carolina University; Greenville, NC USA.
authors.forename: William H
authors.initials: WH
authors.lastname: Chappell
authors.affiliation: Department of Microbiology and Immunology; Brody School of Medicine at East Carolina University; Greenville, NC USA.
authors.forename: Kristin
authors.initials: K
authors.lastname: Stadelman
authors.affiliation: Department of Microbiology and Immunology; Brody School of Medicine at East Carolina University; Greenville, NC USA.
authors.forename: Stephen L
authors.initials: SL
authors.lastname: Abrams
authors.affiliation: Department of Microbiology and Immunology; Brody School of Medicine at East Carolina University; Greenville, NC USA.
authors.forename: Nicole M
authors.initials: NM
authors.lastname: Davis
authors.affiliation: Department of Microbiology and Immunology; Brody School of Medicine at East Carolina University; Greenville, NC USA.
authors.forename: Linda S
authors.initials: LS
authors.lastname: Steelman
authors.affiliation: Department of Microbiology and Immunology; Brody School of Medicine at East Carolina University; Greenville, NC USA.
authors.forename: James A
authors.initials: JA
authors.lastname: McCubrey
mesh_terms: D000903:Antibiotics, Antineoplastic / Q000494:pharmacology*; D018931:Antineoplastic Agents, Hormonal / Q000494:pharmacology*; D004317:Doxorubicin / Q000494:pharmacology*; D019008:Drug Resistance, Neoplasm; D005260:Female; D038362:Glycogen Synthase Kinase 3 / Q000037:antagonists & inhibitors* / Q000378:metabolism; D000071679:Glycogen Synthase Kinase 3 beta; D006801:Humans; D061986:MCF-7 Cells; D000076222:Mechanistic Target of Rapamycin Complex 1; D058990:Molecular Targeted Therapy; D046912:Multiprotein Complexes / Q000037:antagonists & inhibitors / Q000378:metabolism; D010766:Phosphorylation; D020123:Sirolimus / Q000494:pharmacology; D058570:TOR Serine-Threonine Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D013629:Tamoxifen / Q000494:pharmacology*
chemical_list: D000903:Antibiotics, Antineoplastic; D018931:Antineoplastic Agents, Hormonal; D046912:Multiprotein Complexes; D013629:Tamoxifen; D004317:Doxorubicin; C000605506:GSK3B protein, human; D000071679:Glycogen Synthase Kinase 3 beta; D000076222:Mechanistic Target of Rapamycin Complex 1; D058570:TOR Serine-Threonine Kinases; D038362:Glycogen Synthase Kinase 3; D020123:Sirolimus
keywords: GSK-3β; MEK; PI3K; mTOR; rapamycin; sorafenib; targeted therapy; therapy resistance; β-catenin