GSK-3β

Despite continuous advances in the knowledge of breast cancer pathophysiology, this type of neoplasia remains a leading cause of cancer-related death in women, with more than 450 000 deaths every year worldwide.¹ Surgery, coupled with radiation and/or chemotherapy, remains the main approach for breast cancer treatment. However, another common therapeutic option is based on the use of the estrogen receptor (ER) antagonist, tamoxifen.² Indeed, approximately 60–70% of early-stage breast cancers overexpress the ER, making their growth dependent on estrogens. Tamoxifen blocks estrogen signaling by competitively binding the ER, thus antagonizing its proliferative and pro-survival effects.

Breast cancers can be inherently drug-resistant or develop resistance after exposure to chemotherapeutic drugs, such as the anthracyline, doxorubicin.¹ Resistance can also develop in patients receiving tamoxifen.² Therefore, it is very important to understand how breast cancers become drug- and hormone-resistant, and whether or not their drug-resistance can be reversed. There also is a need for novel targeted therapies for breast cancer patients who develop resistance to traditional therapies.

GSK-3β is a serine/threonine kinase involved in several signal transduction cascades, including the PI3K/Akt/mTOR, Wnt/β-catenin, and MEK/ERK pathways.³ In particular, Akt and other kinases phosphorylate GSK-3β at Ser9. This phosphorylative event inactivates GSK-3β³. GSK-3β regulates cell cycle progression, differentiation, survival, embryogenesis, migration, and metabolism. However, aberrant GSK-3β activity has been linked with an increasing number of pathologies, including cancer, heart disease, immune system disorders, diabetes, atherosclerosis, and different neurological diseases.³

Although its role in cancer remains controversial, GSK-3β has also been implicated in breast cancer development and drug resistance.^4,5 Using ER-positive MCF-7 breast cancer cells, Sokolosky and coworkers have performed a detailed analysis of the roles played by GSK-3β in causing resistance to doxorubicin or tamoxifen.⁶ They found that MCF-7 cells overexpressing a kinase-dead (KD) form of GSK-3β were more resistant to doxorubicin and tamoxifen compared with MCF-7 cells overexpressing either wild-type (WT) or constitutively active (CA) GSK-3β. Ectopic expression of GSK-3β(KD) also resulted in increased clonogenic activity of MCF-7 cells in comparison with either GSK-3β(WT) or GSK-3β(CA) overexpression.

Moreover, treatment of parental MCF-7 and MCF-7/GSK-3β(WT) cells with doxorubicin abrogated the phosphorylation of GSK-3β at Ser9. In contrast, Ser9 p-GSK-3β was still detectable in MCF-7/GSK-3β(KD) and MCF-7/GSK-3β(CA) cells. This indicated that one of the effects of doxorubicin on MCF-7 cells was suppression of Ser9 p-GSK-3β, which could then result in increased GSK-3β activity. Downregulation of Ser9 p-GSK-3β levels was detected despite increased Akt activity induced in these cell types by doxorubicin. This finding documented that the control of GSK-3β activity is complex and multi-factorial in MCF-7 cells, as there may be several alternate routes of GSK-3β inactivation that are independent of Akt,³ including downregulation of protein phosphatase activity or upregulation of other kinases (Fig. 1).

Figure 1. Schematic of the effects of GSK-3β inactivation on MCF-7 cell sensitivity to doxorubicin or tamoxifen. Arrows indicate activating events, whereas perpendicular lines indicate inhibitory events. At present, it is not clear how doxorubicin could dephosphorylate GSK-3β and activate Akt at the same time. Doxorubicin could either inhibit other protein kinases, targeting Ser9 p-GSK-3β, or stimulate protein phosphatases.

What is even more interesting, MCF-7/GSK-3β(KD) cells displayed an elevated sensitivity to the mTORC1 inhibitor, rapamycin, compared with MCF-7/GSK-3β(WT) or MCF-7/GSK-3β(CA) cells, while concurrent MEK/ERK inhibition alleviated doxorubicin and tamoxifen resistance in MCF-7/GSK-3β(KD) cells (Fig. 1). Overall, these results demonstrated that GSK-3β is a key regulatory molecule in sensitivity of breast cancer cells to chemo-, hormonal, and targeted therapy (Fig. 1). Moreover, they fit well with the conclusions of an immunohistochemical study performed on paraffin-embedded samples from 72 consecutive invasive mammary carcinomas, which documented how higher Ser9 p-GSK-3β levels correlated with a worse clinical outcome in ER-positive cases.⁷ GSK-3β-controlled drug resistance may be mediated by the BH₃-only member of the Bcl-2 family, Mcl-1. Indeed, the expression levels of this anti-apoptotic protein inversely correlated with GSK-3β activity (assessed by immunostochemical staining for p-GSK-3β) in breast cancer specimens, and active GSK-3β was found to be required for proteasome-mediated Mcl-1 degradation.⁸

In conclusion, the findings by Sokolosky et al.⁶ unequivocally demonstrated that loss of GSK-3β kinase activity could dramatically increase the drug and hormonal resistance of breast cancer cells. However, this may confer an Achilles’ heel by sensitizing cancer cells to targeted therapy with small-molecule kinase inhibitors. Although further studies are needed to determine the clinical relevance of the complex interactions between GSK-3β, mTORC1, and MEK/ERK, it could be envisaged that mTORC1 and MEK/ERK inhibitors should be able to potentiate the effects of chemo- and hormonal therapy, thereby presenting an attractive treatment route for overcoming GSK-3β-mediated drug-resistance in breast cancer.

Sokolosky M, et al. Cell Cycle. 2014;13:820–33. doi: 10.4161/cc.27728.