Metformin Enhances TKI-Afatinib Cytotoxic Effect, Causing Downregulation of Glycolysis, Epithelial–Mesenchymal Transition, and EGFR-Signaling Pathway Activation in Lung Cancer Cells

Abstract

The combination of metformin and TKIs for non-small cell lung cancer has been proposed as a strategy to overcome resistance of neoplastic cells induced by several molecular mechanisms. This study sought to investigate the effects of a second generation TKI afatinib, metformin, or their combination on three adenocarcinoma lung cancer cell lines with different EGFRmutation status. A549, H1975, and HCC827 cell lines were treated with afatinib, metformin, and their combination for 72 h. Afterwards, several parameters were assessed including cytotoxicity, interactions, apoptosis, and EGFR protein levels at the cell membrane and several glycolytic, oxidative phosphorylation (OXPHOS), and EMT expression markers. All cell lines showed additive to synergic interactions for the induction of cytotoxicity caused by the tested combination, as well as an improved pro-apoptotic effect. This effect was accompanied by downregulation of glycolytic, EMT markers, a significant decrease in glucose uptake, extracellular lactate, and a tendency towards increased OXPHOS subunits expression. Interestingly, we observed a better response to the combined therapy in lung cancer cell lines A549 and H1975, which normally have low affinity for TKI treatment. Findings from this study suggest a sensitization to afatinib therapy by metformin in TKI-resistant lung cancer cells, as well as a reduction in cellular glycolytic phenotype.

Will We Unlock the Benefit of Metformin for Patients with Lung Cancer? Lessons from Current Evidence and New Hypotheses

“…Metformin–TKI treatment was tested in transformed TKI-resistant LC cell lines with epithelial–mesenchymal transition (EMT) patterns, and the treatment showed sensitization of TKI-resistant cell lines promoting EMT regression, increasing adhesion cadherins and inhibiting IL-6 signaling [ 120 ]. Recently, our group reported a synergic effect between metformin and afatinib, and this effect was associated with a reduction in the EGFR pathway, glycolytic, and EMT markers [ 9 ]. In contrast, another study described an increase in EGFR protein expression with metformin monotherapy, but when combined with erlotinib, metformin showed synergism and growth inhibition of EGFR wild-type LC cells [ 121 ].…”

“…Metformin is an appealing candidate for drug repurposing in oncology for several reasons; as one of the World Health Organization´s essential medicines, it has been massively prescribed and has a well-established safety profile [ 5 , 6 ]. Further, it is one of the most available and low-cost drugs in the world today; last, several basic studies have provided information which put metformin centerfold as an important metabolic regulator, inhibiting proliferative pathways [ 7 , 8 , 9 ], making this a justifiable pharmacological intervention for a disease highly characterized by a metabolic dysregulation which supports uncontrolled proliferation [ 10 , 11 , 12 , 13 ]. Nonetheless, use of metformin in this indication is currently not advised outside a clinical trial setting, and though some studies have shown remarkable benefits by adding metformin to standard anticancer drugs, other trials have shown a lack of effect or even deleterious outcomes, particularly in trials among patients with lung cancer [ 14 , 15 , 16 , 17 , 18 , 19 , 20 ] ( Table 1 ).…”

Advances in Non-Small Cell Lung Cancer (NSCLC) Treatment—A Paradigm Shift in Oncology

“…Novel approaches such as inhalation therapy [ 20 ], improving drug solubility for delivery [ 21 ] and drug repurposing [ 20 , 22 ] are discussed in Review articles in this Special Issue. The promise of drug repurposing and combination therapy was demonstrated by Barrious-Bernal et al [ 23 ], in which Metformin, an anti-diabetic drug, when combined with EGFR TKI-Afatinib, demonstrated additive to synergic anti-tumour effects in TKI-resistant lung cancer cells. Alternative paradigms proposed for NSCLC therapy in this Special Issue include targeting nuclear receptors [ 24 ], DNA damage responses [ 25 ] and inflammation [ 20 ].…”

Obesity paradox and lung cancer, metformin-based therapeutic opportunity?

“…In cancer cells, metformin has been shown to modify tumor metastatic properties by cell migration inhibition and suppressing epithelial-mesenchymal transition (EMT). Similarly, positive immunogenic properties have been associated with metformin through the activation of cytotoxic T- cells and enhancement of oxidative metabolism, which promotes an antitumor immune response [ 3 , 5 ].…”