Metformin impairs growth of endometrial cancer cells via cell cycle arrest and concomitant autophagy and apoptosis

Takahashi, Akimasa; Kimura, Fuminori; Yamanaka, Akiyoshi; Takebayashi, Akie; Kita, Nobuyuki; Takahashi, Kentaro; Murakami, Takashi

doi:10.1186/1475-2867-14-53

Cited by 115 publications

(102 citation statements)

References 37 publications

(49 reference statements)

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“…These adaptor protein-ubiquitin complexes consequently depend on autophagy machinery for macromolecule clearance (37). Consistently, selective autophagy via p62 is enhanced via the autophagy agonist metformin in a dose-dependent manner, whereas silencing of autophagy with the modulator BECLIN1 attenuates the process (24). A recent study showed that HACE1 proteins play an intricate role in mediating autophagy flux by promoting optineurin-ubiquitin-p62 conjugation (38).…”

Section: Discussionmentioning

confidence: 87%

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Metformin increases degradation of phospholamban via autophagy in cardiomyocytes

Teng

Miyake

Yokoe

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

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Phospholamban (PLN) is an effective inhibitor of the sarco(endo) plasmic reticulum Ca 2+ ATPase (SERCA). Here, we examined PLN stability and degradation in primary cultured mouse neonatal cardiomyocytes (CMNCs) and mouse hearts using immunoblotting, molecular imaging, and [ 35 S]methionine pulse-chase experiments, together with lysosome (chloroquine and bafilomycin A1) and autophagic (3-methyladenine and Atg5 siRNA) antagonists. Inhibiting lysosomal and autophagic activities promoted endogenous PLN accumulation, whereas accelerating autophagy with metformin enhanced PLN degradation in CMNCs. This reduction in PLN levels was functionally correlated with an increased rate of SERCA2a activity, accounting for an inotropic effect of metformin. Metabolic labeling reaffirmed that metformin promoted wild-type and R9C PLN degradation. Immunofluorescence showed that PLN and the autophagy marker, microtubule light chain 3, became increasingly colocalized in response to chloroquine and bafilomycin treatments. Mechanistically, pentameric PLN was polyubiquitinylated at the K3 residue and this modification was required for p62-mediated selective autophagy trafficking. Consistently, attenuated autophagic flux in HECT domain and ankyrin repeat-containing E3 ubiquitin protein ligase 1-null mouse hearts was associated with increased PLN levels determined by immunoblots and immunofluorescence. Our study identifies a biological mechanism that traffics PLN to the lysosomes for degradation in mouse hearts.selective autophagy | ubiquitinylation | protein degradation P hospholamban (PLN) is a 52-amino acid peptide located in the sarcoplasmic reticulum (SR) membrane in cardiac, slowtwitch skeletal, and smooth muscle, where it exists as a monomer or pentamer. Whereas monomeric PLN physically interacts with sarco(endo)plasmic reticulum Ca 2+ ATPase type 2a (SERCA2a) to antagonize its function, pentameric PLN complexes are thought to be a reservoir of inactive PLN (1-3). The physical interaction between SERCA2a and PLN reduces the apparent affinity of SERCA2a for Ca 2+ , thereby making SERCA2a less active in transporting Ca 2+ from the cytoplasm to the lumen of the SR at the same concentration of cytoplasmic Ca 2+. The physical interaction between the two proteins is regulated by phosphorylation of PLN at Ser16 by protein kinase A or at Thr17 by Ca 2+ /calmodulin-dependent protein kinase II (2). Phosphorylation of PLN reduces its affinity for SERCA2a, thereby increasing SERCA2a activity (2). Evidence from transgenic mice also supports the inhibitory function of PLN. Although targeted PLN deletion enhances baseline cardiac performance, cardiac-specific overexpression of superinhibitory forms of PLN leads to decreases in the affinity of SERCA2a for Ca 2+ (2). These observations underscore the primary role of PLN as a regulator of SERCA2a activity and, therefore, as a crucial regulator of cardiac contractility. PLN inhibition of SERCA2a can be reversed by either external (i.e., activation of β-adrenergic receptors) or internal (i.e., increased...

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Section: Discussionmentioning

confidence: 87%

“…Metformin accelerates cardiac autophagy by promoting AMPK-mediated ULK1 phosphorylation (23) and also promotes p62-mediated selective autophagic flux in a dosage-dependent manner (24). Accordingly, we asked whether PLN degradation would be affected by metformin in CMNCs.…”

Section: Resultsmentioning

confidence: 99%

Metformin increases degradation of phospholamban via autophagy in cardiomyocytes

Teng

Miyake

Yokoe

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

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“…Cantrell et al showed for the first time that metformin potently inhibits growth in two EC cell lines [half maximal inhibitory concentration (IC 50 ) of 1 mM metformin] after 72 h of treatment in a normal glucose environment (17). Takahashi et al demonstrated that after 48 h of treatment with increasing concentrations of metformin in normal glucose conditions, ≥5 mM metformin significantly reduce the number of viable cells (IC 50 of 6.78 mM metformin) (21). Furthermore, the metformin inhibitory proliferation effect has been reported for certain other types of cancer.…”

Section: Discussionmentioning

confidence: 99%

“…However, the majority of these previous studies possessed methodological weaknesses and/or insufficient data. Additionally, experimental studies revealed that metformin is an effective antiestrogenic agent, inhibiting cell proliferation and leading to growth arrest, as well as inducing apoptosis in EC (17,(20)(21)(22). Furthermore, recent findings have suggested that metformin is important in suppressing the migration and invasion of cancer cells, which could prevent metastasis (23)(24)(25).…”

Section: Introductionmentioning

confidence: 99%

Proliferation and metastatic potential of endometrial cancer cells in response to metformin treatment in a high versus normal glucose environment

et al. 2016

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Abstract. In order to improve our understanding of the potential preventive and therapeutic role of metformin, the present study aimed to investigate the capability of low-dose metformin in the efficient inhibition of cancer development and the reduction of the metastasis of endometrial adenocarcinoma type I and primary endometrial epithelial cells (eEPs), with the drug acting as a treatment in a hyperinsulinemic environment exposed to high and normal glucose conditions. The Ishikawa endometrial adenocarcinoma cell line and primary eEPs were exposed to an environment with high (17 mM) or normal glucose (5 mM) and treated with insulin, low-dose metformin (0.1 mM) or a combined treatment. Metastatic potential was assessed by migration and invasion assays, and relative cell proliferation was determined. Metformin at a low dose potently inhibited the insulin action, decreasing the ability of the endometrial cancer (EC) cell line to migrate and invade in a high and normal glucose environment, and decreasing the migration ability of the primary eEPs. In the EC cell line, the insulin treatment increased the proliferation, without any subsequent reduction of proliferation by the addition of 0.1 mM metformin; however, relative cell proliferation sensitivity to metformin was observed in the range between 1 and 5 mM regardless of the glucose concentration present. Overall, metformin at 0.1 mM is not efficient enough to decrease the proliferation in an EC cell line. However, at this concentration, metformin can inhibit the insulin action in endometrial epithelial cancer cells, demonstrating an anti-metastatic effect in high and normal glucose environments.

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“…Treatment with metformin attenuates the estrogen-dependent proliferative expression of c-myc and c-fos in the obese rat endometrium, an effect which was accompanied by inhibition of the phosphorylation of insulin and IGF1 receptors, as well as Erk 1/2 (17,55). In vitro studies have indicated that metformin inhibits rat endometrial cell line proliferation and suppresses endometrial cancer cell growth via cell cycle arrest and concomitant autophagy and apoptosis (55,56).…”

Section: Metformin and Endometrial Cancermentioning

confidence: 99%

Clinical benefits of metformin in gynecologic oncology

2015

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Metformin impairs growth of endometrial cancer cells via cell cycle arrest and concomitant autophagy and apoptosis

Cited by 115 publications

References 37 publications

Metformin increases degradation of phospholamban via autophagy in cardiomyocytes

Metformin increases degradation of phospholamban via autophagy in cardiomyocytes

Proliferation and metastatic potential of endometrial cancer cells in response to metformin treatment in a high versus normal glucose environment

Clinical benefits of metformin in gynecologic oncology

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