Effect of metformin on glucose-starved and 2-deoxyglucose exposed triple negative MDA-MB 231 breast cancer cells

C. R. Triggle, Y. Majeed, H. Ding, S. M. Samuel. Pharmacology, Weill Cornell Medicine - Qatar, Doha, Qatar.

Introduction: Metformin exhibits anti-cancer properties and we have reported that treatment with metformin (2mM) in glucose-starved or 2-deoxyglucose (2DG;5mM) exposed tumour endothelial cells reversed pro-survival autophagy, inhibited the mTOR pathway and reduced cell viability ¹. Triple-negative breast cancers (TNBC) lack estrogen and progesterone receptors as well as HER2 and do not benefit from hormonal or HER2 targeted therapies ^{2, 3}. Although chemotherapy is the primary established systemic treatment for patients with TNBC, the lack of targeted therapies, its aggressiveness, early recurrence, rapid metastasis and poor prognosis ^{2, 3} calls for alternative therapeutic approaches. It was reported that the anti-proliferative effect of metformin in triple-negative MDA-MB-231 breast cancer cells is highly dependent on glucose concentration ⁴. Thus, we investigated the effects of metformin in glucose-starved and 2DG exposed MDA-MB 231 TNBC cells.

Method: MDA-MB 231 cells were glucose-starved/treated with 2DG (10mM) for 48h in the absence/presence of metformin (2mM). Western blot analysis (n=3-4; to assess the status of the mTOR and autophagy pathway), cell proliferation (MTT assay; n=5), migration (Radius Cell Migration Assay; n=4) and viability (propidium iodide assay; n=5) were performed. Statistical significance was determined by one-way analysis of variance (ANOVA).

Results: Cell proliferation rate significantly decreased in 1) glucose-starved cells treated with metformin (by 95%) when compared to non-treated glucose-starved cells (by 88%) and 2) 2DG exposed cells treated with metformin (by 46%) when compared to cells that were treated with either metformin (by 20%) or 2DG (by 28%) alone. Cell viability significantly (P<0.05) decreased in glucose-starved cells treated with metformin (by 34%) when compared to non-treated glucose-starved cells (by 18%). We observed a marked reduction in cell migration in 2DG exposed cells treated with metformin. Western blot analysis revealed that treatment with metformin significantly decreased (fold change) the levels of LC3A-II (2.4&1.8) and LC3B-II (2.3&1.6), pmTOR(S2448;1.6&1.4) and downstream p4E-BP1(T36/47;1.5&1.3), pS6(S235/236;4.7&2.8) and pS6(S240/244;3.2&1.8) in glucose-starved and 2DG-exposed cells treated with metformin.

Conclusion: Results show that using metformin in glucose-starved and 2DG exposed TNBC cells could prove to be a potential anti-cancer therapy as an adjunct in the treatment of triple-negative breast cancers.

References:

1. Samuel SM, et al. (2017). Biochem Pharmacol. 132: 118-132.

2. Bianchini G, et al. (2016). Nat Rev Clin Oncol. 13: 674-690.

3. Hudis CA, et al. (2011). Oncologist. 16 Suppl 1: 1-11.

4. Zordoky BN, et al. (2014). Biochim Biophys Acta. 1840: 1943-1957.

*Supported by Qatar National Research Fund grants: NPRP-04-910-3-244 & JSREP-03-016-3-009