Abstract
Rapamycin and its analogues inhibit mTOR, which leads to decreased protein synthesis and decreased cancer cell proliferation in many experimental systems. Adenosine 5′- monophosphate-activated protein kinase (AMPK) activators such as metformin have similar actions, in keeping with the TSC2/1 pathway linking activation of AMPK to inhibition of mTOR. As mTOR inhibition by rapamycin is associated with attenuation of negative feedback to IRS-1, rapamycin is known to increase activation of AKT, which may reduce its anti-neoplastic activity. We observed that metformin exposure decreases AKT activation, an action opposite to that of rapamycin. We show that metformin (but not rapamycin) exposure leads to increased phosphorylation of IRS-1 at Ser789, a site previously reported to inhibit downstream signaling and to be an AMPK substrate phosphorylated under conditions of cellular energy depletion. siRNA methods confirmed that reduction of AMPK levels attenuates both the IRS-1 Ser789 phosphorylation and the inhibition of AKT activation associated with metformin exposure. Although both rapamycin and metformin inhibit mTOR (the former directly and the latter through AMPK signaling), our results demonstrate previously unrecognized differences between these agents. The data are consistent with the observation that maximal induction of apoptosis and inhibition of proliferation are greater for metformin than rapamycin.
Similar content being viewed by others
References
Rini B, Kar S, Kirkpatrick P (2007) Temsirolimus. Nat Rev Drug Discov 6:599–600
Sun SY, Rosenberg LM, Wang X, Zhou Z, Yue P, Fu H, Khuri FR (2005) Activation of Akt and eIF4E survival pathways by rapamycin-mediated mammalian target of rapamycin inhibition. Cancer Res 65:7052–7058
O’Reilly KE, Rojo F, She QB, Solit D, Mills GB, Smith D, Lane H, Hofmann F, Hicklin DJ, Ludwig DL, Baselga J, Rosen N (2006) mTOR inhibition induces upstream receptor tyrosine kinase signaling and activates Akt. Cancer Res 66:1500–1508
Bhaskar PT, Hay N (2007) The two TORCs and Akt. Dev Cell 12:487–502
Guertin DA, Sabatini DM (2007) Defining the role of mTOR in cancer. Cancer Cell 12:9–22
Wang X, Yue P, Kim YA, Fu H, Khuri FR, Sun SY (2008) Enhancing mammalian target of rapamycin (mTOR)-targeted cancer therapy by preventing mTOR/raptor inhibition-initiated, mTOR/rictor-independent Akt activation. Cancer Res 68:7409–7418
Easton JB, Kurmasheva RT, Houghton PJ (2006) IRS-1: auditing the effectiveness of mTOR inhibitors. Cancer Cell 9:153–155
Ozes ON, Akca H, Mayo LD, Gustin JA, Maehama T, Dixon JE, Donner DB (2001) A phosphatidylinositol 3-kinase/Akt/mTOR pathway mediates and PTEN antagonizes tumor necrosis factor inhibition of insulin signaling through insulin receptor substrate-1. Proc Natl Acad Sci USA 98:4640–4645
Tzatsos A, Kandror KV (2006) Nutrients suppress phosphatidylinositol 3-kinase/Akt signaling via raptor-dependent mTOR-mediated insulin receptor substrate 1 phosphorylation. Mol Cell Biol 26:63–76
Harrington LS, Findlay GM, Gray A, Tolkacheva T, Wigfield S, Rebholz H, Barnett J, Leslie NR, Cheng S, Shepherd PR, Gout I, Downes CP, Lamb RF (2004) The TSC1-2 tumor suppressor controls insulin-PI3K signaling via regulation of IRS proteins. J Cell Biol 166:213–223
Guertin DA, Sabatini DM (2009) The pharmacology of mTOR inhibition. Sci Signal 2:e24
Hardie DG (2008) AMPK and Raptor: matching cell growth to energy supply. Mol Cell 30:263–265
Gwinn DM, Shackelford DB, Egan DF, Mihaylova MM, Mery A, Vasquez DS, Turk BE, Shaw RJ (2008) AMPK phosphorylation of raptor mediates a metabolic checkpoint. Mol Cell 30:214–226
Shaw RJ (2009) LKB1 and AMP-activated protein kinase control of mTOR signalling and growth. Acta Physiol (Oxf) 196:65–80
Zakikhani M, Dowling R, Fantus IG, Sonenberg N, Pollak M (2006) Metformin is an AMP kinase-dependent growth inhibitor for breast cancer cells. Cancer Res 66:10269–10273
Zakikhani M, Dowling RJ, Sonenberg N, Pollak MN (2008) The effects of adiponectin and metformin on prostate and colon neoplasia involve activation of AMP-activated protein kinase. Cancer Prev Res (Phila Pa) 1:369–375
Dowling RJ, Zakikhani M, Fantus IG, Pollak M, Sonenberg N (2007) Metformin inhibits mammalian target of rapamycin-dependent translation initiation in breast cancer cells. Cancer Res 67:10804–10812
Algire C, Zakikhani M, Blouin M-J, Shuai JH, Pollak M (2008) Metformin attenuates the stimulatory effect of a high energy diet on in vivo H59 carcinoma growth. Endocr Relat Cancer 15:833–839
Pollak M (2008) Insulin and insulin-like growth factor signalling in neoplasia. Nat Rev Cancer 8:915–928
Tzatsos A, Tsichlis PN (2007) Energy depletion inhibits phosphatidylinositol 3-kinase/Akt signaling and induces apoptosis via AMP-activated protein kinase-dependent phosphorylation of IRS-1 at Ser-794. J Biol Chem 282:18069–18082
Jakobsen SN, Hardie DG, Morrice N, Tornqvist HE (2001) 5′-AMP-activated protein kinase phosphorylates IRS-1 on Ser-789 in mouse C2C12 myotubes in response to 5-aminoimidazole-4-carboxamide riboside. J Biol Chem 276:46912–46916
Choo AY, Yoon SO, Kim SG, Roux PP, Blenis J (2008) Rapamycin differentially inhibits S6Ks and 4E-BP1 to mediate cell-type-specific repression of mRNA translation. Proc Natl Acad Sci USA 105:17414–17419
Boura-Halfon S, Zick Y (2009) Phosphorylation of IRS proteins, insulin action, and insulin resistance. Am J Physiol Endocrinol Metab 296:E581–E591
Gual P, Marchand-Brustel Y, Tanti JF (2005) Positive and negative regulation of insulin signaling through IRS-1 phosphorylation. Biochimie 87:99–109
Manning BD, Cantley LC (2007) AKT/PKB signaling: navigating downstream. Cell 129:1261–1274
Vivanco I, Sawyers CL (2002) The phosphatidylinositol 3-Kinase AKT pathway in human cancer. Nat Rev Cancer 2:489–501
Hartley D, Cooper GM (2002) Role of mTOR in the degradation of IRS-1: regulation of PP2A activity. J Cell Biochem 85:304–314
Virshup DM, Shenolikar S (2009) From promiscuity to precision: protein phosphatases get a makeover. Mol Cell 33:537–545
Padmanabhan S, Mukhopadhyay A, Narasimhan SD, Tesz G, Czech MP, Tissenbaum HA (2009) A PP2A regulatory subunit regulates C. elegans insulin/IGF-1 signaling by modulating AKT-1 phosphorylation. Cell 136:939–951
Shu Y, Sheardown SA, Brown C, Owen RP, Zhang S, Castro RA, Ianculescu AG, Yue L, Lo JC, Burchard EG, Brett CM, Giacomini KM (2007) Effect of genetic variation in the organic cation transporter 1 (OCT1) on metformin action. J Clin Invest 117:1422–1431
Zhuang Y, Miskimins WK (2008) Cell cycle arrest in Metformin treated breast cancer cells involves activation of AMPK, downregulation of cyclin D1, and requires p27Kip1 or p21Cip1. J Mol Signal 3:18
Liu B, Fan Z, Edgerton SM, Deng XS, Alimova IN, Lind SE, Thor AD (2009) Metformin induces unique biological and molecular responses in triple negative breast cancer cells. Cell Cycle 8:2031–2040
Huang J, Wu S, Wu CL, Manning BD (2009) Signaling events downstream of mammalian target of rapamycin complex 2 are attenuated in cells and tumors deficient for the tuberous sclerosis complex tumor suppressors. Cancer Res 69:6107–6114
Acknowledgments
We thank Ms. L. Lui for valuable assistance in manuscript preparation. This study was supported by the Translational Acceleration grant #16512 from the Canadian Breast Cancer Research Alliance and the Translational Cancer Research Award ref: 2008-08 from the Terry Fox Research Institute.
Author information
Authors and Affiliations
Corresponding author
Rights and permissions
About this article
Cite this article
Zakikhani, M., Blouin, MJ., Piura, E. et al. Metformin and rapamycin have distinct effects on the AKT pathway and proliferation in breast cancer cells. Breast Cancer Res Treat 123, 271–279 (2010). https://doi.org/10.1007/s10549-010-0763-9
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s10549-010-0763-9