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Adiponectin receptor agonist AdipoRon induces apoptotic cell death and suppresses proliferation in human ovarian cancer cells

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Abstract

We tested the hypothesis that stimulation of adiponectin receptors with the synthetic agonist AdipoRon suppresses proliferation and induces apoptotic death in human high grade serous ovarian tumor cell lines and in ex vivo primary tumors, mediated by activation of 5′ AMP-activated protein kinase (AMPK) and inhibition of mechanistic target of rapamycin (mTOR). We determined the effect of AdipoRon on high grade serous ovarian tumor cells lines (OVCAR3, OVCAR4, A2780) and ex vivo primary tumor tissue. Western blotting analysis was performed to examine changes in activation of AMPK and mTOR signaling and flow cytometry was utilized to examine changes in cell cycle progression. Immunofluorescence of cleaved caspase-3 positive cells and flow cytometry of annexin V positive cells were used to determine changes in apoptotic response. The CyQUANT proliferation assay was used to assess cell proliferation. AdipoRon treatment increased AMPK phosphorylation (OVCAR3 P = 0.01; A2780 P = 0.02) but did not significantly alter mTOR activity. AdipoRon induced G1 cell cycle arrest in OVCAR3 (+ 12.1%, P = 0.03) and A2780 (+ 12.0%, P = 0.002) cells. OVCAR3 and OVCAR4 cells treated with AdipoRon underwent apoptosis based on cleaved caspase-3 and annexin V staining. AdipoRon treatment resulted in a dose dependent decrease in cell number versus vehicle treatment in OVCAR3 (−61.2%, P < 0.001), OVCAR4 (−79%, P < 0.001), and A2780 (−56.9%, P < 0.001). Ex vivo culture of primary tumors treated with AdipoRon resulted in an increase in apoptosis measured with cleaved caspase-3 immunohistochemistry. AdipoRon induces activation of AMPK and exhibits an anti-tumor effect in ovarian cancer cell lines and primary tumor via a mTOR-independent pathway.

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Acknowledgements

This research was supported in part by University of Colorado Department of Obstetrics and Gynecology Academic Enrichment Fund grant. The authors appreciate the contribution to this research made by E. Erin Smith, HTL(ASCP)CM QIHC, Allison Quador, HTL(ASCP)CM, and Jessica Arnold, HTL(ASCP)CM of the University of Colorado Denver Tissue Biobanking and Histology Shared Resource. This resource is supported in part by the Cancer Center Support Grant (P30CA046934). Flow cytometry was performed by the University of Colorado Cancer Center Flow Cytometry Shared Resource (supported by NCI Cancer Center Support Grant P30CA046934). We would also like to acknowledge that imaging was made possible by the University of Colorado Advanced Light Microscopy Core, supported in part by NIH/NCRR CCTSI grant UL1 RR025780. Contents are the authors’ sole responsibility.

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  1. Division of Gynecologic Oncology, Department of Obstetrics and Gynecology, University of Colorado School of Medicine, 12631 East 17th Avenue B198-6, Aurora, CO, 80045, USA

    Amin A. Ramzan & Kian Behbakht

  2. Division of Reproductive Sciences, Department of Obstetrics and Gynecology, University of Colorado School of Medicine, Aurora, CO, USA

    Benjamin G. Bitler, Douglas Hicks, Kelsey Barner, Lubna Qamar, Theresa Powell, Thomas Jansson & Heidi Wilson

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Ramzan, A.A., Bitler, B.G., Hicks, D. et al. Adiponectin receptor agonist AdipoRon induces apoptotic cell death and suppresses proliferation in human ovarian cancer cells. Mol Cell Biochem 461, 37–46 (2019). https://doi.org/10.1007/s11010-019-03586-9

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