Key functions of mTOR signal pathway

 Rapamycin (also known as sirolimus), a macrolide compound first discovered in the 1960s in soil samples from Easter Island, was found to have anti-tumor and immunosuppressive properties. However, it wasn’t until 1994 that target protein mTOR (mammalian target of rapamycin) was identified (Brown et al. 1994; Sabatini et al. 1994). The mTOR protein is a serine/threonine protein kinase in the PI3K-related kinase family. It is evolutionarily deeply conserved and integrates many intracellular and extracellular signals. The mTOR pathway is regulated by growth factors, hormones, glucose, DNA damage, and hypoxia.  The mTOR pathway integrates these varied signals to regulate cell growth, proliferation, and survival as well as cytoskeletal organization. The protein mTOR is the catalytic component of mTOR complex 1 (mTORC1) and mTOR complex 2 (mTORC2). mTORC1 is involved in regulation of cell growth by increasing protein synthesis, lipid and nucleotide synthesis, and reduced autophagy. mTORC2 is involved in cytoskeletal remodeling, cell proliferation and cell survival. With such critical roles in fundamental cellular processes it is important that we continue to better elucidate the mechanisms of mTOR signaling and identify potential targets for therapies of diseases related to these functions.

 Key molecules involved in mTOR signal pathway

 The two known mTOR complexes (mTORC1 and mTORC2) have distinct, but related, functions and subunit assemblies. mTORC1 is generally better understood than TORC2. This is due, in part, to the relative insensitivity of mTORC2 to rapamycin and lethality caused by deletion of mTORC2 components complicating the job of identifying mTORC2 functions. In spite of this, research continues to provide insights into the activators and effectors of both mTOR complexes.

 mTORC1 is composed of five proteins: mTOR, PRAS40, DEPTOR, Raptor, and mLST8 (also called GβL),. PRAS40 and DEPTOR are both negative regulators of mTORC1 (Peterson et al. 2009; Sancak et al. 2007; Wang et al. 2007).  Raptor recruits substrates to mTORC1 (Nojima et al. 2003) and mLST8 binds and stabilizes the catalytic kinase domain of mTOR (Yang et al. 2013). 

 mTORC1 has several downstream targets (reviewed by: Peterson et al. 2009; Saxton and Sabatini 2017). Rheb is an activator of mTORC1 which is inhibited by TSC2. AMPK and p53, which are activated by DNA damage and low cellular energy, both inhibit mTORC1 by activating TSC2. AMPK also inhibits mTORC1 directly by phosphorylating Raptor. The PI3K/AKT pathway and the Ras/Raf/Mek/Erk pathway, each activated by growth factors and insulin or EGFR, activate mTORC1 by inhibiting TSC2. Further, leucine and arginine, acting through the Rag - GATOR1 - GATOR2 pathway, can activate mTORC1. Thus factors like DNA damage and low energy will reduce cell growth while growth factor and free nucleotides (arginine and leucine) increase cell growth. When activated, mTORC1 increases protein synthesis through activation of CAD, eEF2 eIF4B and eIF4E. Autophagy is inhibited by inhibiting ULK1 and transcription factor EB.

 mTORC2 is composed of six proteins: DEPTOR, Rictor, mLST8, mSin1, and Protor1. As in mTORC1, DEPTOR is inhibitory and mLST8 stabilizes the kinase loop of mTOR. Rictor may serve a substrate binding function, similar to Raptor. Protor1 (Pearce et al. 2007; Woo et al. 2007) and mSin1 (Jacinto et al. 2006; Yang et al. 2006) serve regulatory functions. mTORC2, activated by growth factors like insulin, phosphorylates AKT, SGK, and PKC to increase metabolism and cell survival. Phosphorylated AKT increases metabolism and decreases apoptosis by several means including by inhibiting TSC2 to activate, among other things, mTORC1. mTORC2 also regulates cytoskeletal reorganization and cell migration via PKC phosphorylation.

 mTOR signal pathway in human cancer

 The mTOR pathway has been implicated in the potential etiology and treatment of several human diseases, including cancers (reviewed by: Saxton and Sabatini 2017; Watanabe, Wei, and Huang 2011). The PI3K/AKT and Ras pathways are mutated in many human cancers, leading to increased mTORC1 activity in the tumor cells. Before the mTORC1 was identified as the target of rapamycin it was known that this compound had tumor suppressive properties. Additionally, LKB1 and p53, upstream inhibitors of mTORC1 are tumor suppressor genes. The capacity of mTORC2 to phosphorylate AKT, which decreases apoptosis and increases cell proliferation in a number of ways, including activation of mTORC1, implicates this complex in cancer development as well. Functional mTORC2 is required for tumorigenisis in some PTEN deficient models of prostate cancer (Guertin et al. 2009). 

 Unfortunately,
inhibition has not yet been as effective in cancer treatment as initially hoped. Analogs of rapamycin, called rapalogs (i.e., temsirolimus, everolimus, and ridaforolimus), have been developed, but clinically performed more poorly than expected based on pre-clinical research. This is likely due to the fact that these rapalogs block some, but not all, of mTORC1’s substrates, the inhibition of mTORC1 may remove negative feedback on the PI3K/AKT pathway, and long-term rapalog exposure provides selection pressure for proliferation of cells with mutations of the rapamycin binding domain (FRB domain). Furthermore, because mTORC1 prevents autophagy, inhibiting mTORC1 may increase autophagy. The role of autophagy in cancer is unclear, having been reported to as both pro-tumorigenic and anti-tumorigenic (White and DiPaola 2009). Somewhat paradoxically mTORC1 inhibition may actually support tumor cell survival and proliferation by providing nutrients through autophagy. 

 Latest Progress in the mTOR signal pathway

 Work continues to try to improve our understanding of the mTOR pathway and its potential treatment of health conditions like cancer. So called second-generation mTOR kinase inhibitors (e.g., AZD8055) have been developed and are being tested (Chresta et al. 2010). These second-generation inhibitors directly block mTOR catalytic activity in both complexes (Rodrik-Outmezguine et al. 2011) but these still seem run into a problem problems of developing resistance over long-term exposure. Some of this resistance seems to be due to selection for MTOR mutations that increase activity of the kinase domain. To overcome mutations of either the kinase domain of MTOR or the FRB domain researchers have developed a bivalent third-generation mTOR inhibitor called RapaLink that links the FRB binding domain to the kinase inhibitor of the second-generation inhibitors. This third-generation compound blocks mTORC1 and mTORC2 in cells resistant to either the first- or second-generation mTOR inhibitors (Rodrik-Outmezguine et al. 2016). RapaLink has shown tumor reduction in a glioblastoma model, but was followed by tumor regrowth (Fan et al. 2017). Work continues into efficacy of these inhibitors alone or in combination with other compounds, such and inhibitors of PI3K/AKT or autophagy.

 Additional exciting recent work has linked mTOR to regulation of longevity. Reduced expression of mTOR homologs has increase lifespan in yeast (Kaeberlein et al. 2005), C. elegans (Vellai et al. 2003), Drosophila (Kapahi et al. 2004), and mice (Wu et al. 2013).  While the mechanism isn’t clear, the propect of using this information to increase human longevity is intriguing. One difficulty is that prolonged inhibition of mTOR leads to immunosuppression and insulin resistance. However some research does support the efficacy of using mTOR inhibition to increase human lifespan without substantial side effects (Arriola Apelo et al. 2016; Mannick et al. 2014).

 We have learned a great deal about the mTOR pathway and there is much more to discover. The mTOR pathway integrates many intracellular and extracellular signals to regulate various aspects of cell growth, proliferation, survival, and motility. With a major role in such fundamental functions it is important that we continue trying to more fully elucidate this pathway. The prospect of discovering more about the etiology and potential treatments of cancers and other human diseases further increases the importance of studying the mTOR pathway. 

 Reference papers (add links)

 Arriola Apelo, Sebastian I. et al. 2016. “Alternative Rapamycin Treatment Regimens Mitigate the Impact of Rapamycin on Glucose Homeostasis and the Immune System.” Aging Cell 15(1):28–38.

Brown, Eric J. et al. 1994. “A Mammalian Protein Targeted by G1-Arresting Rapamycin–receptor Complex.” Nature 369:756.

Chresta, Christine M. et al. 2010. “AZD8055 Is a Potent, Selective, and Orally Bioavailable ATP-Competitive Mammalian Target of Rapamycin Kinase Inhibitor with in Vitro and in Vivo Antitumor Activity.” Cancer Research 70(1):288–98.

Fan, Qi Wen et al. 2017. “A Kinase Inhibitor Targeted to MTORC1 Drives Regression in Glioblastoma.” Cancer Cell 31(3):424–35.

Guertin, David A. et al. 2009. “MTOR Complex 2 Is Required for the Development of Prostate Cancer Induced by Pten Loss in Mice.” Cancer Cell 15(2):148–59.

Jacinto, Estela et al. 2006. “SIN1/MIP1 Maintains Rictor-MTOR Complex Integrity and Regulates Akt Phosphorylation and Substrate Specificity.” Cell 127(1):125–37.

Kaeberlein, Matt et al. 2005. “Regulation of Yeast Replicative Life Span by TOR and Sch9 in Response to Nutrients.” Science 310(5751):1193 LP-1196.

Kapahi, Pankaj et al. 2004. “Regulation of Lifespan in Drosophila by Modulation of Genes in the TOR Signaling Pathway.” Current Biology 14(10):885–90.

Mannick, Joan B. et al. 2014. “MTOR Inhibition Improves Immune Function in the Elderly.” Science Translational Medicine 6(268):268ra179.

Nojima, Hiroki et al. 2003. “The Mammalian Target of Rapamycin (MTOR) Partner, Raptor, Binds the MTOR Substrates P70 S6 Kinase and 4E-BP1 through Their TOR Signaling (TOS) Motif.” Journal of Biological Chemistry  278(18):15461–64.

Pearce, Laura R. et al. 2007. “Identification of Protor as a Novel Rictor-Binding Component of MTOR Complex-2.” The Biochemical Journal 405(3):513–22.

Peterson, Timothy R. et al. 2009. “DEPTOR Is an MTOR Inhibitor Frequently Overexpressed in Multiple Myeloma Cells and Required for Their Survival.” Cell 137(5):873–86.

Rodrik-Outmezguine, Vanessa S. et al. 2011. “MTOR Kinase Inhibition Causes Feedback-Dependent Biphasic Regulation of AKT Signaling.” Cancer Discovery 1(3):248 LP-259.

Rodrik-Outmezguine, Vanessa S. et al. 2016. “Overcoming MTOR Resistance Mutations with a New-Generation MTOR Inhibitor.” Nature 534:272.

Sabatini, David M., Hediye Erdjument-Bromage, Mary Lui, Paul Tempst, and Solomon H. Snyder. 1994. “RAFT1: A Mammalian Protein That Binds to FKBP12 in a Rapamycin-Dependent Fashion and Is Homologous to Yeast TORs.” Cell 78(1):35–43.

Sancak, Yasemin et al. 2007. “PRAS40 Is an Insulin-Regulated Inhibitor of the MTORC1 Protein Kinase.” Molecular Cell 25(6):903–15.

Saxton, Robert A. and David M. Sabatini. 2017. “MTOR Signaling in Growth, Metabolism, and Disease.” Cell 168(6):960–76.

Vellai, Tibor et al. 2003. “Influence of TOR Kinase on Lifespan in C. Elegans.” Nature 426:620.

Wang, Lifu, Thurl E. Harris, Richard A. Roth, and John C. Lawrence. 2007. “PRAS40 Regulates MTORC1 Kinase Activity by Functioning as a Direct Inhibitor of Substrate Binding.” Journal of Biological Chemistry  282(27):20036–44.

Watanabe, R., L. Wei, and J. Huang. 2011. “MTOR Signaling, Function, Novel Inhibitors, and Therapeutic Targets.” Journal of Nuclear Medicine 1:497–501.

White, Eileen and Robert S. DiPaola. 2009. “The Double-Edged Sword of Autophagy Modulation in Cancer.” Clinical Cancer Research 15(17):5308–16.

Woo, So-Yon et al. 2007. “PRR5, a Novel Component of MTOR Complex 2, Regulates Platelet-Derived Growth Factor Receptor Beta Expression and Signaling.” The Journal of Biological Chemistry 282(35):25604–12.

Wu, J. Julie et al. 2013. “Increased Mammalian Lifespan and a Segmental and Tissue-Specific Slowing of Aging after Genetic Reduction of MTOR Expression.” Cell Reports4(5):913–20.

Yang, Haijuan et al. 2013. “MTOR Kinase Structure, Mechanism and Regulation.” Nature 497:217.

Yang, Qian, Ken Inoki, Tsuneo Ikenoue, and Kun-Liang Guan. 2006. “Identification of Sin1 as an Essential TORC2 Component Required for Complex Formation and Kinase Activity.” Genes & Development 20(20):2820–32.