Metformin Suppresses Self-Renewal Ability and Tumorigenicity of Osteosarcoma Stem Cells via Reactive Oxygen Species-Mediated Apoptosis and Autophagy

Abstract

Osteosarcoma is the most frequently diagnosed primary malignant bone sarcoma in children and adolescents. Recent studies have shown that cancer stem cells (CSCs), a cluster of tumor cells with the ability to self-renew, play an essential role in tumor recurrence and metastasis. Thus, it is necessary to develop therapeutic strategies specifically targeting CSCs. Metformin, the first-line drug for type 2 diabetes, exhibits antineoplastic activities in various kinds of tumors. New evidence has suggested that metformin may target CSCs and prevent their recurrence. However, the underlying specific mechanisms remain unclear. In this study, we found that metformin significantly suppressed the self-renewal ability of osteosarcoma stem cells (OSCs) and induced G0/G1 phase arrest by blocking the activity of cyclin-dependent kinases. Furthermore, metformin induced apoptosis through a mitochondria-dependent pathway, leading to the collapse of the mitochondrial transmembrane potential and the production of reactive oxygen species (ROS). Importantly, metformin acted directly on the mitochondria, which resulted in decreased ATP synthesis. This change allowed access to the downstream AMPK kinase, and the activation of AMPK led to the reversal of the mTOR pathway, triggering autophagy. Particularly, metformin-mediated autophagy disturbed the homeostasis of stemness and pluripotency in the OSCs. Additionally, our mouse xenograft model confirmed the potential therapeutic use of metformin in targeting OSCs. In conclusion, our findings suggest that metformin suppresses the self-renewal ability and tumorigenicity of OSCs via ROS-mediated apoptosis and autophagy.

Metformin induces apoptosis and inhibits migration by activating the AMPK/p53 axis and suppressing PI3K/AKT signaling in human cervical cancer cells

“…Moreover, the results suggested that metformin induced cytotoxicity by promoting cell cycle arrest in the G 2 /M phase and increasing apoptosis in CaSki and HeLa cells, and promoting G 0 /G 1 phase cell cycle arrest in c33a and Hela cells. Previous studies have illustrated the effects of metformin on the cell cycle in human osteosarcoma, demonstrating increased K7M2 and MG63 cell numbers in the G 0 /G 1 phase, and increased u20S and 143B cell numbers in the G 2 /M phase (49,56). Additionally, metformin has been reported to enhance anticancer effects by arresting human colon carcinoma cells in the G 0 /G 1 phase (SW480 cells) or the G 2 /M phase (HCT116 p53 −/− cells) (57,58).…”

Effect of metformin on stem cells: Molecular mechanism and clinical prospect

“…Courtois et al [ 76 ] confirmed the anti-proliferative effect of metformin on gastric carcinoma cell lines by regulating the expression of genes implicated in cell-cycle regulation ( GADD45, p21, E2F1, and PCNA )[ 76 ]. Zhao et al [ 77 ] demonstrated that metformin inhibits the proliferation of osteosarcoma stem cells (OSCs) by inducing G0/G1 phase cell cycle arrest[ 77 ]. In a recent study, Barbieri et al [ 78 ] stated that metformin and other biguanides exert anti-proliferative effects in glioblastoma CSCs by interfering with the activity of the extracellular portion of the active transmembrane chloride ion channel.…”

“…A recent study highlighted the strong dependence on energy-producing pathways of colorectal cancer CSCs, suggesting that modulation of AMPK activity is an effective therapeutic approach[ 81 ]. Zhao et al [ 77 ] showed that metformin induced caspase-mediated apoptosis in OSCs by inducing mitochondrial dysfunction. In addition, metformin influenced the capacity of OSCs to self-renew via enhanced autophagy, which was suppressed by 3-methyladenine, an inhibitor of autophagy[ 77 ].…”

“…Zhao et al [ 77 ] showed that metformin induced caspase-mediated apoptosis in OSCs by inducing mitochondrial dysfunction. In addition, metformin influenced the capacity of OSCs to self-renew via enhanced autophagy, which was suppressed by 3-methyladenine, an inhibitor of autophagy[ 77 ].…”

Apelin Rejuvenates Aged Human Mesenchymal Stem Cells by Regulating Autophagy and Improves Cardiac Protection After Infarction

“…These results further confirmed that Apelin overexpression rejuvenates AMSCs by activating autophagy. Accumulating evidence indicates that AMPK signaling plays a critical role in regulating autophagy ( Zhao et al, 2019 ; Khorraminejad-Shirazi et al, 2020 ). Next, we investigated whether Apelin mediates autophagy levels in AMSCs via this pathway.…”