Isolated and combined action of tamoxifen and metformin in wild-type, tamoxifen-resistant, and estrogen-deprived MCF-7 cells

Abstract

Resistance to tamoxifen (TAM) and aromatase inhibitors represents a major drawback to the treatment of hormone-dependent breast cancer, and strategies to overcome this problem are urgently needed. The anti-diabetic biguanide metformin (MF) exerts pleiotropic effects which could enhance the effectiveness of available hormonal therapies. This study modeled several aspects of hormonal therapy in women and examined the effectiveness of MF under those conditions. For cell growth evaluation, wild-type (wt), TAM-resistant (TAM-R), and long-term estradiol-deprived (LTED) MCF-7 cells, as a model of aromatase inhibitor resistance, were grown in the presence or absence of TAM or MF for 5 days. For immunoblot analysis and aromatase activity measurements, these cells were grown for 48 h. Wild-type and LTED cells were equally sensitive to the growth inhibitory effects of TAM and MF, while TAM-R cells were less sensitive to TAM than to MF. Partial additive effects on cell number of TAM combined with MF were greatest (if compared with isolated TAM action) in TAM-R and LTED cells. In contrast to the decrease in PCNA values in TAM-resistant cells treated with the TAM and MF combination, no other changes were found in the levels of this proliferation marker. These findings suggested a major component of apoptosis in the growth inhibitory effect. This was confirmed with Western blot analysis of PARP and caspase 7 as well as with apoptosis ELISA assay. MF also altered signaling pathways. AMP-kinase was stimulated by MF approximately equally in MCF-7, TAM-R, and LTED cells, while inhibition by biguanide of p-S6K as a downstream target of mTOR was strongest in TAM-R cells. Under the influence of MF, expression of ER-α was decreased in wt MCF-7 cells suggesting possible involvement of this compound in estrogen signaling. Metformin interacts additively with TAM to reduce neoplastic cells growth. The cellular context (including loss of sensitivity to TAM and estrogen deprivation) is of importance in influencing breast cancer responses to MF and to a combination of MF and TAM.

Metformin in cancer: translational challenges

“…In primary human breast adipose stromal cells, metformin treatment led to an increase in the phosphorylation of AMPK, which was associated with an inhibition of nuclear translocation of CRTC2, a CREB co-activator known to increase aromatase expression (Brown et al 2010). Furthermore, metformin interacted additively with tamoxifen to reduce breast cancer cell proliferation (Berstein et al 2010b). These results indicate that metformin may represent a potentially effective therapy in women with ER-positive breast tumours.…”

Phosphorylated S6K1 is a possible marker for endocrine therapy resistance in hormone receptor-positive breast cancer

“…The main downstream effect of AMPK activation is the inhibition of mTOR [26]. Recent study showed that the effect of p-S6K1 (a target of mTOR) inhibition by metformin is stronger in tamoxifen-resistant MCF-7 cells than in wild-type MCF-7 cells and the additive effect of metformin combined with tamoxifen is also greater in tamoxifen-resistant MCF-7 cells [27]. In addition, the mTOR inhibition by rapamycin derivatives may activate compensatory pathways such as insulin-like growth factor-1 receptor (IGF-1R)/PI3 K signaling to Akt [28], and this collateral effect of mTOR blockade is another potential explanation for the limited activity of mTOR inhibitors.…”

Molecular Pathways: Preclinical Models and Clinical Trials with Metformin in Breast Cancer

“…Given that aromatase inhibitors are now the mainstay of neoadjuvant, adjuvant, and extended treatment in postmenopausal women with breast cancer, the potential for synergism between an aromatase inhibitor and metformin is under exploration in a Korean neoadjuvant trial and is planned for an international extended adjuvant trial. For premenopausal women, where tamoxifen remains the principal adjuvant therapy for ER-positive breast cancer, the additive effects of metformin to tamoxifen, at least in terms of reducing cell proliferation (46), merits subgroup analysis within the ongoing MA32 adjuvant metformin trial (47).…”