高通量筛选mTORC2复合体特异性小分子抑制剂
|
摘要
目的：建立针对mTORC2特异性的下游靶点p-AKT (Ser473)的细胞分子模型,与华北制药新药研究开发公司合作,从微生物菌种代谢文库中筛选出mTORC2特异性的天然小分子抑制剂；
方法：以mTORC2特异性的下游靶点p-AKT (Ser473)为第一轮的筛选靶点,分别以DMSO、Rapamycin、PI-103、HRP作为阴性对照、激活性阳性对照、抑制性阳性对照和底物空白对照,在正常的支气管上皮细胞系BEAS-2B中进行Cytoblot筛选。每种样品设计8种浓度梯度,各梯度之间呈3倍稀释。第一批检测样品为华北制药新药筛选中心拥有的1693种单体化合物；
结果：将信号值低于PI-103孔或相当,重复性和浓度依赖性好的样品,进一步进行Western验证,获得3种对p-AKT (Ser473)有抑制活性的小分子抑制剂；将信号值高于Rapamycin或相当,重复性和浓度依赖性好的样品,同样进行Western验证,获得1种对p-AKT (Ser473)有激活作用的小分子激动剂；
结论：针对p-AKT (Ser473)磷酸化水平的Cytoblot筛选方法,是一种基于活细胞整体,简单、快捷、灵敏度和特异性都比较高的方法,可以作为一种筛选mT0RC2特异性抑制剂的初筛模型；
目的：初步探讨小分子化合物HA选择性诱导Rictor蛋白发生降解的分子机制，为筛选mTORC2特异性抑制剂提供一种新的思路；
方法：Western进一步检测HA对多种肿瘤细胞中Rictor蛋白的影响；在HA诱导的Rictor蛋白降解的同时，加入蛋白酶体抑制剂MG-132，抑制蛋白酶体降解途径，发现Rictor蛋白的降解可以被逆转；将Rictor分成5段，分别构建GST融合蛋白，建立体外泛素模型，进一步确证Rictor蛋白可以被泛素化修饰；
结果：HA可以选择性的诱导MM细胞中Rictor蛋白发生泛素．蛋白酶体途径降解；同时不会诱导mTOR蛋白和mTORC1复合体中的Raptor蛋白发生降解；在HA诱导的Rictor蛋白降解模型中，可以看到：Rictor蛋白被降解，mTORC2功能受抑，其特异性的底物p-AKT(Ser473)位点磷酸化水平降低；加用蛋白酶体抑制剂时，逆转Rictor降解，p-AKT(Ser473)位点磷酸化水平得以恢复；体外泛素模型中，发现Rictor蛋白可以被泛素化修饰；
结论：基于HA诱导Rictor蛋白的降解现象，深入研究选择性降解Rictor蛋白的分子机制，将为筛选mTORC2特异性抑制剂提供一种全新的策略。
Objective:To establish the cellular and molecular models for phosphorylation-AKT (Ser473) which is the specific downstream target of mTORC2, cooperating with North China Pharmaceutical Group (NCPC) New Drug Research and Development Co.Ltd, and screening specific small molecule inhibitor of mTORC2 from the microbial strain metabolism library;
Methods:We used phosphorylation-AKT (Ser473) which was the specific downstream target of mTORC2 as the screening targets of first round. Using DMSO, Rapamycin, PI-103, HRP as negative control, activation positive control, suppression positive control and substrate blank control respectively, we conducted Cytoblot screening in normal bronchial epithelial cell line BEAS-2B. Eight concentration gradients were designed in each sample,3-fold dilution between two adjacent gradients. The first test samples were 1693 kinds of monomeric compounds owned by North China Pharmaceutical Group (NCPC) Center for New Drug Screening;
Results:When the signal value of a compound was lower than the value of PI-103 or equivalent, and presenting preferable reproducibility and concentration-dependent manner, we took further Western identification, and finally obtained three kinds of small molecule inhibitors which had suppressive activity of phosphorylation-AKT (Ser473); When the signal value of a compound was higher than the value Rapamycin or equivalent, and also presenting preferable reproducibility and concentration-dependent manner, we took further Western identification, and finally obtained one kind of small molecule agonist which had activation on phosphorylation-AKT (Ser473).
Conclusions:Phosphorylation-AKT (Ser473) Cytoblot screening method, is based on the whole living cells, which is simple and fast, having high sensitivity and specificity. It's could be used as a preliminary screening model to screen specific inhibitors of mTORC2;
Objective: To explore molecular mechanism of the phenomenon that small moleculecompounds HA induced Rictor protein degradation selectivly, and providing a novel threadfor screening specific inhibitors of mTORC2 complex;
Methods: We further tested the effects of HA induced Rictor protein degradation in avariety of tumor cell lines by western blot; in HA-induced Rictor protein degradation model,when we added the proteasome inhibitor MG-132, which could inhibit the proteasomedegradation pathway, Rictor protein degradation could be reversed; and then we dividedRictor into five clips, for the Rictor protein is too large, to reconstruct GST fusion proteinsrespectively, established In vitro Ubiquitination model, further confirmed Rictor proteincould be ubiquitylated;
Results: HA could selectively induce Rictor protein degradation occurring on MM cellline in ubiquitin-proteasome pathway; at the same time, HA could not induce mTORprotein and Raptor protein degradation, both of which are the important constituents of thecomplex of mTORC1; in HA-induced Rictor protein degradation model, we could see thatRictor protein was degraded, mTORC2 function was inhibited, the phosphorylation level ofits specific substrate p-AKT (Ser473) sites reduced; when combining with proteaseinhibitor, the Rictor protein degradation was reversed, the phosphorylation level of p-AKT(Ser473) sites was recovered;
Conclusion: Based on the induced Rictor degradation model by the compound of HA, bythorough research the molecular mechanism of this selective degradation, we will get apromising strategy to screen specific inhibitors of mTORC2.
方法：以mTORC2特异性的下游靶点p-AKT (Ser473)为第一轮的筛选靶点,分别以DMSO、Rapamycin、PI-103、HRP作为阴性对照、激活性阳性对照、抑制性阳性对照和底物空白对照,在正常的支气管上皮细胞系BEAS-2B中进行Cytoblot筛选。每种样品设计8种浓度梯度,各梯度之间呈3倍稀释。第一批检测样品为华北制药新药筛选中心拥有的1693种单体化合物；
结果：将信号值低于PI-103孔或相当,重复性和浓度依赖性好的样品,进一步进行Western验证,获得3种对p-AKT (Ser473)有抑制活性的小分子抑制剂；将信号值高于Rapamycin或相当,重复性和浓度依赖性好的样品,同样进行Western验证,获得1种对p-AKT (Ser473)有激活作用的小分子激动剂；
结论：针对p-AKT (Ser473)磷酸化水平的Cytoblot筛选方法,是一种基于活细胞整体,简单、快捷、灵敏度和特异性都比较高的方法,可以作为一种筛选mT0RC2特异性抑制剂的初筛模型；
目的：初步探讨小分子化合物HA选择性诱导Rictor蛋白发生降解的分子机制，为筛选mTORC2特异性抑制剂提供一种新的思路；
方法：Western进一步检测HA对多种肿瘤细胞中Rictor蛋白的影响；在HA诱导的Rictor蛋白降解的同时，加入蛋白酶体抑制剂MG-132，抑制蛋白酶体降解途径，发现Rictor蛋白的降解可以被逆转；将Rictor分成5段，分别构建GST融合蛋白，建立体外泛素模型，进一步确证Rictor蛋白可以被泛素化修饰；
结果：HA可以选择性的诱导MM细胞中Rictor蛋白发生泛素．蛋白酶体途径降解；同时不会诱导mTOR蛋白和mTORC1复合体中的Raptor蛋白发生降解；在HA诱导的Rictor蛋白降解模型中，可以看到：Rictor蛋白被降解，mTORC2功能受抑，其特异性的底物p-AKT(Ser473)位点磷酸化水平降低；加用蛋白酶体抑制剂时，逆转Rictor降解，p-AKT(Ser473)位点磷酸化水平得以恢复；体外泛素模型中，发现Rictor蛋白可以被泛素化修饰；
结论：基于HA诱导Rictor蛋白的降解现象，深入研究选择性降解Rictor蛋白的分子机制，将为筛选mTORC2特异性抑制剂提供一种全新的策略。
Objective:To establish the cellular and molecular models for phosphorylation-AKT (Ser473) which is the specific downstream target of mTORC2, cooperating with North China Pharmaceutical Group (NCPC) New Drug Research and Development Co.Ltd, and screening specific small molecule inhibitor of mTORC2 from the microbial strain metabolism library;
Methods:We used phosphorylation-AKT (Ser473) which was the specific downstream target of mTORC2 as the screening targets of first round. Using DMSO, Rapamycin, PI-103, HRP as negative control, activation positive control, suppression positive control and substrate blank control respectively, we conducted Cytoblot screening in normal bronchial epithelial cell line BEAS-2B. Eight concentration gradients were designed in each sample,3-fold dilution between two adjacent gradients. The first test samples were 1693 kinds of monomeric compounds owned by North China Pharmaceutical Group (NCPC) Center for New Drug Screening;
Results:When the signal value of a compound was lower than the value of PI-103 or equivalent, and presenting preferable reproducibility and concentration-dependent manner, we took further Western identification, and finally obtained three kinds of small molecule inhibitors which had suppressive activity of phosphorylation-AKT (Ser473); When the signal value of a compound was higher than the value Rapamycin or equivalent, and also presenting preferable reproducibility and concentration-dependent manner, we took further Western identification, and finally obtained one kind of small molecule agonist which had activation on phosphorylation-AKT (Ser473).
Conclusions:Phosphorylation-AKT (Ser473) Cytoblot screening method, is based on the whole living cells, which is simple and fast, having high sensitivity and specificity. It's could be used as a preliminary screening model to screen specific inhibitors of mTORC2;
Objective: To explore molecular mechanism of the phenomenon that small moleculecompounds HA induced Rictor protein degradation selectivly, and providing a novel threadfor screening specific inhibitors of mTORC2 complex;
Methods: We further tested the effects of HA induced Rictor protein degradation in avariety of tumor cell lines by western blot; in HA-induced Rictor protein degradation model,when we added the proteasome inhibitor MG-132, which could inhibit the proteasomedegradation pathway, Rictor protein degradation could be reversed; and then we dividedRictor into five clips, for the Rictor protein is too large, to reconstruct GST fusion proteinsrespectively, established In vitro Ubiquitination model, further confirmed Rictor proteincould be ubiquitylated;
Results: HA could selectively induce Rictor protein degradation occurring on MM cellline in ubiquitin-proteasome pathway; at the same time, HA could not induce mTORprotein and Raptor protein degradation, both of which are the important constituents of thecomplex of mTORC1; in HA-induced Rictor protein degradation model, we could see thatRictor protein was degraded, mTORC2 function was inhibited, the phosphorylation level ofits specific substrate p-AKT (Ser473) sites reduced; when combining with proteaseinhibitor, the Rictor protein degradation was reversed, the phosphorylation level of p-AKT(Ser473) sites was recovered;
Conclusion: Based on the induced Rictor degradation model by the compound of HA, bythorough research the molecular mechanism of this selective degradation, we will get apromising strategy to screen specific inhibitors of mTORC2.
引文
1. Manning BD, Cantley LC. AKT/PKB Signaling:Navigating Downstream. Cell.2007, 129(7):1261-74
2. Sabatini DM. mTOR and cancer:insights into a complex relationship. Nat Rev Cancer. 2006,6(9):729-34
3. Bhaskar PT, Hay N. The Two TORCs and Akt. Dev Cell.2007,12(4):487-502
4. Guertin DA, Sabatini DM. Defining the role of mTOR in cancer. Cancer Cell.2007, 12(1):9-22
5. Laplante M, Sabatini DM. mTOR signaling at a glance. J Cell Sci.2009 Oct 15;122(Pt 20):3589-94
6. Faivre S, Kroemer Q Raymond E. Current development of mTOR inhibitors as anticancer agents. Nat Rev Drug Discov.2006,5(8):671-88.
7. Park S, Chapuis N, Bardet V, et al. PI-103, a dual inhibitor of Class IA phosphatidylinositide 3-kinase and mTOR, has antileukemic activity in AML. Leukemia.2008,22(9):1698-706
8. David A. Guertin and David M. Sabatini. The Pharmacology of mTOR Inhibition. Sci Signal.2009 Apr 21;2(67)
9. Shor B, Gibbons JJ, Abraham RT, et al. Targeting mTOR globally in cancer:thinking beyond rapamycin. Cell Cycle.2009 Dec;8(23):3831-7
10. Alessi DR, Pearce LR, Garcia-Martinez JM. New Insights into mTOR Signaling: mTORC2 and Beyond. Sci Signal.2009 Apr 21;2(67)
11. Bentzinger CF, Romanimo K, Cloetta D, et al. Skeletal muscle-specific ablation of raptor, but not of rictor, causes metabolic changes and results in muscle dystrophy. Cell Metab.2008 Nov;8(5):411-24
12. Feldman ME, Apsel B, Uotila A, et al. Active-site inhibitors of mTOR target rapamycin-resistant outputs of mTORC1 and mTORC2. PLOS Biol.2009 Feb 10; 7(2):e38
13. Sarbassov DD, Guertin DA, Ali SM, et al. Phosphorylation and regulation of Akt/PKB by the rictor-mTOR complex. Science.2005,307(5712):1098-101
14. Ikenoue T, Inoki K, Yang Q, et al. Essential function of TORC2 in PKC and Akt turn motif phosphorylation, maturation and signalling. EMBO J.2008,27(14):1919-31
15. Guertin DA, Stevens DM, Saitoh M, et al. mTOR complex 2 is required for the development of prostate cancer induced by Pten loss in mice. Cancer Cell.2009, 15(2):148-59
16. Yang J, Shamji A, Matchacheep S, et al. Identification of a small-molecule inhibitor of class Ⅰa PI3Ks with cell-based screening. Chem Biol.2007 Apr; 14(4):371-7.
17. Koike-Kumagai M, Yasunaga K, Morikawa R, et al. The target of rapamycin complex 2 controls dendritic tiling of Drosophila sensory neurons through the tricornered kinase signalling pathway. EMBO J.2009 Dec 16; 28(24):3879-92.
18. Risson V, Mazelin L, Roceri M, et al. Muscle inactivation of mTOR causes metabolic and dystrophin defects leading to severe myopathy. J Cell Biol.2009 Dec 14; 187(6):859-74.
1、 Bhaskar PT, Hay N. The Two TORCs and Akt. Dev Cell.2007,12(4):487-502
2、 Guertin DA, Sabatini DM. Defining the role of mTOR in cancer. Cancer Cell.2007, 12(1):9-22
3、 Laplante M, Sabatini DM. mTOR signaling at a glance. J Cell Sci.2009 Oct 15;122(Pt 20):3589-94
4、 Yilmaz OH, Valdez R, Theisen BK, et al. Pten dependence distinguishes haematopoietic stem cells from leukemia-initiating cells. Nature.2006, 441(7092):475-82
5、 Zhang J, Grindley JC, Yin T, et al. PTEN maintains haematopoietic stem cells and acts in lineage choice and leukaemia prevention. Nature.2006 May 25; 441 (7092):518-22
6、 David A. Guertin and David M. Sabatini. The Pharmacology of mTOR Inhibition. Sci Signal.2009 Apr 21;2(67)
7、 Risson V, Mazelin L, Roceri M, et al. Muscle inactivation of mTOR causes metabolic and dystrophin defects leading to severe myopathy. J Cell Biol.2009 Dec 14; 187(6):859-74.
8、 Shor B, Gibbons JJ, Abraham RT, et al. Targeting mTOR globally in cancer:thinking beyond rapamycin. Cell Cycle.2009 Dec;8(23):3831-7
10、 Alessi DR, Pearce LR, Garcia-Martinez JM. New Insights into mTOR Signaling: mTORC2 and Beyond. Sci Signal.2009 Apr 21; 2(67)
11、 Guertin DA, Stevens DM, Saitoh M, et al. mTOR complex 2 is required for the development of prostate cancer induced by Pten loss in mice. Cancer Cell.2009, 15(2):148-59
12、 Masri J, Bernath A, Martin J, et al. mTORC2 activity is elevated in gliomas and promotes growth and cell motility via overexpression of rictor. Cancer Res.2007 Dec 15; 67 (24):11712-20
13、 Sabatini DM. mTOR and cancer:insights into a complex relationship. Nat Rev Cancer. 2006,6(9):729-34
14、 Peterson TR, Laplante M, Thoreen CC, et al. DEPTOR is an mTOR inhibitor frequently overexpressed in multiple myeloma cells and required for their survival. Cell.2009 May 29;137(5):873-86
15、 Pearce LR, Huang X, Boudeau J, et al. Identification of Protor as a novel Rictor-binding component of mTOR complex-2. Biochem J.2007 Aug 1;405(3):513-22.
16、 Sarbassov DD, Ali SM, Kim DH, et al. Rictor, a Novel Binding Partner of mTOR, defines a Rapamycin-Insensitive and Raptor-Independent Pathway that Regulates the Cytoskeleton. Curr Biol.2004 Jul 27;14(14):1296-302.
17、 Feldman ME, Apsel B, Uotila A, et al. Active-site inhibitors of mTOR target rapamycin-resistant outputs of mTORC1 and mTORC2. PLOS Biol.2009 Feb 10; 7(2):e38
18、 Thoreen CC, Kang SA, Chang JW, et al. An ATP-competitive mammalian target of rapamycin inhibitor reveals rapamycin-resistant functions of mTORC1.J Biol Chem. 2009 Mar 20; 284 (12):8023-32.
19、 Zask A, Verheijen JC, Curran K, et al. ATP-Competitive Inhibitors of the Mammalian Target of Rapamycin:Design and Synthesis of Highly Potent and Selective Pyrazolopyrimidines. J Med Chem.2009 Jul 31.
20、 Maira SM, Stauffer F, Brueggen J, et al. Identification and characterization of NVP-BEZ235, a new orally available dual phosphatidylinositol 3-kinase/mammalian target of rapamycin inhibitor with potent in vivo antitumor activity. Mol Cancer Ther. 2008 Jul; 7(7):1851-63.
21、 Faivre S, Kroemer G, Raymond E. Current development of mTOR inhibitors as anticancer agents. Nat Rev Drug Discov.2006,5(8):671-88.
22、 Tamburini J, Chapuis N, Bardet V,et al. Mammalian target of rapamycin (mTOR) inhibition activates phosphatidylinositol 3-kinase/Akt by up-regulating insulin-like growth factor-1 receptor signaling in acute myeloid leukemia:rationale for therapeutic inhibition of both pathways. Blood.2008 Jan 1; 111(1):379-82.
23、 Gupta M, Ansell SM, Novak AJ, et al. Inhibition of histone deacetylase overcomes rapamycin-mediated resistance in diffuse large B-cell lymphoma by inhibiting Akt signaling through mTORC2. Blood.2009 Oct 1;114(14):2926-35
24、 Koike-Kumagai M, Yasunaga K, Morikawa R, et al. The target of rapamycin complex 2 controls dendritic tiling of Drosophila sensory neurons through the Tricornered kinase signalling pathway. EMBO J.2009 Dec 16; 28(24):3879-92.
1、 Guertin DA, Sabatini DM. Defining the role of mTOR in cancer. Cancer Cell.2007, 12(1):9-22
2、 Laplante M, Sabatini DM. mTOR signaling at a glance. J Cell Sci.2009 Oct 15;122(Pt 20):3589-94
3、 Yilmaz OH, Valdez R, Theisen BK, et al. Pten dependence distinguishes haematopoietic stem cells from leukemia-initiating cells. Nature.2006,441 (7092):475-82
4、 Zhang J, Grindley JC, Yin T, et al. PTEN maintains haematopoietic stem cells and acts in lineage choice and leukaemia prevention. Nature.2006 May 25; 441(7092):518-22
5、 Guertin DA, Stevens DM, Saitoh M, et al. mTOR complex 2 is required for the development of prostate cancer induced by Pten loss in mice. Cancer Cell.2009, 15(2):148-59
6、 Sabatini DM. mTOR and cancer:insights into a complex relationship. Nat Rev Cancer. 2006,6(9):729-34
7、 Bhaskar PT, Hay N. The Two TORCs and Akt. Dev Cell.2007,12(4):487-502
8、 Jacinto E, Facchinetti V, Liu D, et al. SIN1/MIP1 maintains rictor-mTOR complex integrity and regulates Akt phosphorylation and substrate specificity. Cell.2006 Oct 6; 127(1):125-37.
9、 Richter JD, Sonenberg N. Regulation of cap-dependent translation by eIF4E inhibitory proteins. Nature.2005 Feb 3; 433(7025):477-80.
10、 Ma XM, Blenis J. Molecular mechanisms of mTOR-mediated translational control.Nat Rev Mol Cell Biol.2009 May; 10(5):307-18.
11、 Ganley IG, Lam du H, Wang J, et al. ULK1.ATG13.FIP200 complex mediates mTOR signaling and is essential for autophagy. J Biol Chem.2009 May 1; 284(18):12297-305.
12、 Hosokawa N, Hara T, Kaizuka T, et al. Nutrient-dependent mTORCl association with the ULK1-Atg13-FIP200 complex required for autophagy. Mol Biol Cell.2009 Apr; 20(7):1981-91.
13、 Jung CH, Jun CB, Ro SH, et al. ULK-Atg13-FIP200 complexes mediate mTOR signaling to the autophagy machinery. Mol Biol Cell.2009 Apr; 20(7):1992-2003.
14、 Thoreen CC, Kang SA, Chang JW, et al. An ATP-competitive mammalian target of rapamycin inhibitor reveals rapamycin-resistant functions of mTORC1. J Biol Chem. 2009 Mar 20; 284(12):8023-32.
15、 Porstmann T, Santos CR, Griffiths B, et al. SREBP activity is regulated by mTORC1 and contributes to Akt-dependent cell growth. Cell Metab.2008 Sep; 8(3):224-36.
16、 Bentzinger CF, Romanimo K, Cloetta D, et al. Skeletal muscle-specific ablation of raptor, but not of rictor, causes metabolic changes and results in muscle dystrophy. Cell Metab.2008 Nov; 8(5):411-24
17、 Cunningham JT, Rodgers JT, Arlow DH, et al. mTOR controls mitochondrial oxidative function through a YY1-PGC-1 alpha transcriptional complex. Nature.2007 Nov 29; 450(7170):736-40.
18、 Sancak Y, Thoreen CC, Peterson TR, et al. PRAS40 is an insulin-regulated inhibitor of the mTORC1 protein kinase. Mol Cell.2007 Mar 23; 25(6):903-15.
19、 Konstantinopoulos PA, Papavassiliou AG The tuberous sclerosis complex. N Engl J Med.2007 Jan 4; 356(1):92-3.
20、 Vander Haar E, Lee SI, Bandhakavi S, et al. Insulin signalling to mTOR mediated by the Akt/PKB substrate PRAS40. Nat Cell Biol.2007 Mar; 9(3):316-23.
21、 Wang L, Harris TE, Roth RA, et al. PRAS40 regulates mTORC1 kinase activity by functioning as a direct inhibitor of substrate binding. J Biol Chem.2007 Jul 6; 282(27):20036-44.
22、 Hong-Brown LQ, Brown CR, Kazi AA, et al. Alcohol and PRAS40 knockdown decrease mTOR activity and protein synthesis via AMPK signaling and changes in mTORC1 interaction. J Cell Biochem.2010 Apr 15; 109(6):1172-84.
23、 Harrington LS, Findlay GM, Lamb RF. Restraining PI3K:mTOR signalling goes back to the membrane. Trends Biochem Sci.2005 Jan; 30(1):35-42.
24、Zhang H, Bajraszewski N, Wu E, et al. PDGFRs are critical for PI3K/Akt activation and negatively regulated by mTOR. J Clin Invest.2007 Mar; 117(3):730-8.
25、Hardie DG AMP-activated/SNF1 protein kinases:conserved guardians of cellular energy.Nat Rev Mol Cell Biol.2007 Oct; 8(10):774-85.
26、 Gwinn DM, Shackelford DB, Egan DF, et al. AMPK phosphorylation of raptor mediates a metabolic checkpoint. Mol Cell.2008 Apr 25; 30(2):214-26.
27、 Wouters BG, Koritzinsky M. Hypoxia signalling through mTOR and the unfolded protein response in cancer. Nat Rev Cancer.2008 Nov; 8(11):851-64.
28、 DeYoung MP, Horak P, Sofer A, et al. Hypoxia regulates TSC1/2-mTOR signaling and tumor suppression through REDD1-mediated 14-3-3 shuttling. Genes Dev.2008 Jan 15;22(2):239-51.
29、 Li Y, Wang Y, Kim E, et al. Bnip3 mediates the hypoxia-induced inhibition on mammalian target of rapamycin by interacting with Rheb. J Biol Chem.2007 Dec 7; 282(49):35803-13.
30、 Nicklin P, Bergman P, Zhang B, et al. Bidirectional transport of amino acids regulates mTOR and autophagy. Cell.2009 Feb 6; 136(3):521-34.
31、 Nobukuni T, Joaquin M, Roccio M, et al. Amino acids mediate mTOR/raptor signaling through activation of class 3 phosphatidylinositol 3OH-kinase. Proc Natl Acad Sci U S A.2005 Oct 4; 102(40):14238-43.
32、 Juhasz G, Hill JH, Yan Y, et al. The class Ⅲ PI(3)K Vps34 promotes autophagy and endocytosis but not TOR signaling in Drosophila. J Cell Biol.2008 May 19; 181(4):655-66.
33、 Kim DH, Sarbassov DD, Ali SM, et al. mTOR interacts with raptor to form a nutrient-sensitive complex that signals to the cell growth machinery. Cell.2002 Jul 26; 110(2):163-75.
34、 Kim E, Goraksha-Hicks P, Li L, et al. Regulation of TORC1 by Rag GTPases in nutrient response. Nat Cell Biol.2008 Aug; 10(8):935-45.
35、 Sancak Y, Peterson TR, Shaul YD, et al. The Rag GTPases bind raptor and mediate amino acid signaling to mTORC1. Science.2008 Jun 13; 320(5882):1496-501
36、 Feng Z, Zhang H, Levine AJ, et al. The coordinate regulation of the p53 and mTOR pathways in cells. Proc Natl Acad Sci U S A.2005 Jun 7; 102(23):8204-9.
37、 Lee DF, Kuo HP, Chen CT, et al. IKK beta suppression of TSC1 links inflammation and tumor angiogenesis via the mTOR pathway. Cell.2007 Aug 10; 130(3):440-55.
38、 Inoki K, Li Y, Zhu T, et al. TSC2 is phosphorylated and inhibited by Akt and suppresses mTOR signalling. Nat Cell Biol.2002 Sep; 4(9):648-57.
39、 Toschi A, Lee E, Xu L, et al. Regulation of mTORC1 and mTORC2 complex assembly by phosphatidic acid:competition with rapamycin. Mol Cell Biol.2009 Mar; 29(6):1411-20.
40、 Sarbassov DD, Guertin DA, Ali SM, et al. Phosphorylation and regulation of Akt/PKB by the rictor-mTOR complex. Science.2005,307(5712):1098-101
41、 Calnan DR, Brunet A. The FoxO code. Oncogene.2008 Apr 7; 27(16):2276-88.
42、 Peterson TR, Laplante M, Thoreen CC, et al. DEPTOR is an mTOR inhibitor frequently overexpressed in multiple myeloma cells and required for their survival. Cell.2009 May 29;137(5):873-86
43、 Jacinto E, Loewith R, Schmidt A, et al. Mammalian TOR complex 2 controls the actin cytoskeleton and is rapamycin insensitive. Nat Cell Biol.2004 Nov; 6(11):1122-8.
44、 Guertin DA, Stevens DM, Saitoh M, et al. mTOR complex 2 is required for the development of prostate cancer induced by Pten loss in mice. Cancer Cell.2009, 15(2):148-59
45、 Masri J, Bernath A, Martin J, et al. mTORC2 activity is elevated in gliomas and promotes growth and cell motility via overexpression of rictor. Cancer Res.2007 Dec 15； 67 (24):11712-20
46、 Guertin DA, Sabatini DM. Defining the role of mTOR in cancer. Cancer Cell.2007, 12(1):9-22
47、 Q. Yang, K. L. Guan. Expanding mTOR signaling. Cell Res.2007 Aug; 17(8):666-81.
48、 Peterson TR, Laplante M, Thoreen CC, et al. DEPTOR is an mTOR inhibitor frequently overexpressed in multiple myeloma cells and required for their survival. Cell.2009 May 29;137(5):873-86
49、 J. Huang, B. D. Manning. A complex interplay between Akt, TSC2 and the two mTOR complexes. Biochem Soc Trans.2009 Feb; 37(Pt 1):217-22.
50、 Smith EM, Finn SG, Tee AR, et al. The tuberous sclerosis protein TSC2 is not required for the regulation of the mammalian target of rapamycin by amino acids and certain cellular stresses. J Biol Chem.2005 May 13; 280(19):18717-27.
51、 Sancak Y, Peterson TR, Shaul YD, et al. The Rag GTPases bind raptor and mediate amino acid signaling to mTORC1. Science.2008 Jun 13; 320(5882):1496-501
52、 Kim E, Goraksha-Hicks P, Li L, et al. Regulation of TORC1 by Rag GTPases in nutrient response. Nat Cell Biol.2008 Aug; 10(8):935-45.
53、 Yuan TL, Cantley LC. PI3K pathway alter-ations in cancer:Variations on a theme. Oncogene.2008 Sep 18; 27(41):5497-510.
54、 Krymskaya VP, Goncharova EA. PI3K/mTORCl activation in hamartoma syn-dromes: Therapeutic prospects. Cell Cycle.2009 Feb 1; 8(3):403-13.
55、 Lane HA, Breuleux M. Optimal targeting of the mTORCl kinase in human cancer. Curr Opin Cell Biol.2009 Apr; 21(2):219-29.
56、 Hudes G, Carducci M, Tomczak P, et al. Temsirolimus, interferon a, or both for advanced renal-cell carcinoma. N Engl J Med.2007 May 31; 356(22):2271-81.
57、 Choo AY, Yoon SO, Kim SG, et al. Rapamycin differentially inhibits S6Ks and 4E-BP1 to mediate cell-type-specific repression of mRNA translation. Proc Natl Acad Sci U S A.2008 Nov 11; 105(45):17414-9.
58、 Thoreen CC, Kang SA, Chang JW, et al. An ATP-competitive mTOR inhibitor reveals rapamycin-insensitive functions of mTORCl. J Biol Chem.2009 Mar 20; 284(12):8023-32.
59、 Feldman ME, Apsel B, Uotila A, et al. Active-site inhibitors of mTOR target rapamycin-resistant outputs of mTORC1 and mTORC2. PLoS Biol.2009 Feb 10; 7(2):e38.
60、 Dorrello NV, Peschiaroli A, Guardavaccaro D, et al. S6K1-and βTRCP-mediated degradation of PDCD4 promotes protein translation and cell growth. Science.2006 Oct 20;314(5798):467-71.
61、 Holz MK, Ballif BA, Gygi SP, et al. mTOR and S6K1 mediate assembly of the translation preinitiation complex through dynamic protein interchange and ordered phosphorylatio events. Cell.2005 Nov 18; 123(4):569-80.
62、 Jacinto E, Facchinetti V, Liu D, et al. SIN1/MIP1 maintains rictor-mTOR complex integrity and regulates Akt phosphorylation and substrate specificity. Cell.2006 Oct 6; 127(1):125-37.
63、Shiota C, Woo JT, Lindner J, et al. Multiallelic disruption of the rictor gene in mice reveals that mTOR complex 2 is essential for fetal growth and viability. Dev Cell.2006 Oct; 11(4):583-9.
64、 Guertin DA, Stevens DM, Thoreen CC, et al. Ablation in mice of the mTORC components raptor, rictor, or mLST8 reveals that mTORC2 is required for signaling to Akt-FOXO and PKCa, but not S6K1. Dev Cell.2006 Dec; 11(6):859-71.
65、 Garcia-Martinez JM, Alessi DR. mTOR complex-2 (mTORC2) controls hydrophobic motif phosphorylation and activation of serum-and glucocorticoid-induced protein kinase 1 (SGK1). Biochem J.2008 Dec 15; 416(3):375-85.
66、 Sarbassov DD, Guertin DA, Ali SM, et al. Phosphorylation and regulation of Akt/PKB by the rictor-mTOR complex. Science.2005 Feb 18; 307 (5712):1098-101.
67、 Yap TA, Garrett MD, Walton MI, et al. Targeting the PI3K-AKT-mTOR pathway: Progress, pitfalls, and promises. Curr Opin Pharmacol.2008 Aug; 8(4):393-412.
68、 Maira SM, Stauffer F, Brueggen J, et al. Identification and characterization of NVP-BEZ235, a new orally available dual phosphatidylinositol 3-kinase/mammalian target of rapamycin inhibitor with potent in vivo antitumor activity. Mol Cancer Ther. 2008 Jul;7(7):1851-63.
69、 Serra V, Markman B, Scaltriti M, et al. NVP-BEZ235, a dual PI3K/mTOR inhibitor, prevents PI3K signaling and inhibits the growth of cancer cells with activating PI3K mutations. Cancer Res.2008 Oct 1; 68(19):8022-30.
70、 Schnell CR, Stauffer F, Allegrini PR, et al. Effects of the dual phosphatidylinositol 3-kinase/mammalian target of rapamycin inhibitor NVP-BEZ235 on the tumor vasculature:Implications for clinical imaging. Cancer Res.2008 Aug 15; 68(16):6598-607.
71、 Engelman JA, Chen L, Tan X, et al. Effective use of PI3K and MEK inhibitors to treat mutant K-ras G12D and PIK3CAH1047R murine lung cancers. Nat Med.2008 Dec; 14(12):1351-6.
72、 Sarbassov DD, Ali SM, Sengupta S, et al. Prolonged rapamycin treatment inhibits mTORC2 assembly and Akt/PKB. Mol Cell.2006 Apr 21; 22(2):159-68.
73、 Copp J, Manning G, Hunter T. TORC-specific phosphorylation of mammalian target of rapamycin (mTOR):Phospho-Ser2481 is a marker for intact mTOR signaling complex 2. Cancer Res.2009 Mar 1; 69(5):1821-7.
74、 Shor B, Zhang WG, Toral-Barza L, et al. A new pharmacologic action of CCI-779 involves FKBP12-independent inhibition of mTOR kinase activity and profound repression of global protein synthesis. Cancer Res.2008 Apr 15; 68(8):2934-43.
75、 Barquilla A, Crespo JL, Navarro M. Rapamycin inhibits trypanosome cell growth by preventing TOR complex 2 formation. Proc Natl Acad Sci U S A.2008 Sep 23; 105(38):14579-84.
76、 Nardella C, Carracedo A, Alimonti A, et al. Differential requirement of mTOR in postmitotic tissues and tumorigenesis. Sci Signal.2009 Jan 27; 2(55):ra2.
77、 Guertin DA, Stevens DM, Saitoh M, et al. mTOR complex 2 is required for the development of prostate cancer induced by Pten loss in mice. Cancer Cell.2009 Feb 3; 15(2):148-59.
78、 Bentzinger CF, Romanino K, Cloetta D, et al. Skeletal muscle-specific ablation of raptor, but not of rictor, causes metabolic changes and results in muscle dystrophy. Cell Metab.2008 Nov; 8(5):411-24.
79、 Kumar A, Harris TE, Keller SR, et al. Muscle-specific deletion of rictor impairs insulin-stimulated glucose transport and enhances basal glycogen synthase activity. Mol Cell Biol.2008 Jan; 28(1):61-70.
80、 Beevers CS, Chen L, Liu L, et al. Curcumin disrupts the mammalian target of rapamycin-raptor complex. Cancer Res.2009 Feb 1; 69(3):1000-8.
81、 Knight ZA, Gonzalez B, Feldman ME, et al. A pharmacological map of the PI3-K family defines a role for p110α in insulin signaling. Cell.2006 May 19; 125(4):733-47.
2. Sabatini DM. mTOR and cancer:insights into a complex relationship. Nat Rev Cancer. 2006,6(9):729-34
3. Bhaskar PT, Hay N. The Two TORCs and Akt. Dev Cell.2007,12(4):487-502
4. Guertin DA, Sabatini DM. Defining the role of mTOR in cancer. Cancer Cell.2007, 12(1):9-22
5. Laplante M, Sabatini DM. mTOR signaling at a glance. J Cell Sci.2009 Oct 15;122(Pt 20):3589-94
6. Faivre S, Kroemer Q Raymond E. Current development of mTOR inhibitors as anticancer agents. Nat Rev Drug Discov.2006,5(8):671-88.
7. Park S, Chapuis N, Bardet V, et al. PI-103, a dual inhibitor of Class IA phosphatidylinositide 3-kinase and mTOR, has antileukemic activity in AML. Leukemia.2008,22(9):1698-706
8. David A. Guertin and David M. Sabatini. The Pharmacology of mTOR Inhibition. Sci Signal.2009 Apr 21;2(67)
9. Shor B, Gibbons JJ, Abraham RT, et al. Targeting mTOR globally in cancer:thinking beyond rapamycin. Cell Cycle.2009 Dec;8(23):3831-7
10. Alessi DR, Pearce LR, Garcia-Martinez JM. New Insights into mTOR Signaling: mTORC2 and Beyond. Sci Signal.2009 Apr 21;2(67)
11. Bentzinger CF, Romanimo K, Cloetta D, et al. Skeletal muscle-specific ablation of raptor, but not of rictor, causes metabolic changes and results in muscle dystrophy. Cell Metab.2008 Nov;8(5):411-24
12. Feldman ME, Apsel B, Uotila A, et al. Active-site inhibitors of mTOR target rapamycin-resistant outputs of mTORC1 and mTORC2. PLOS Biol.2009 Feb 10; 7(2):e38
13. Sarbassov DD, Guertin DA, Ali SM, et al. Phosphorylation and regulation of Akt/PKB by the rictor-mTOR complex. Science.2005,307(5712):1098-101
14. Ikenoue T, Inoki K, Yang Q, et al. Essential function of TORC2 in PKC and Akt turn motif phosphorylation, maturation and signalling. EMBO J.2008,27(14):1919-31
15. Guertin DA, Stevens DM, Saitoh M, et al. mTOR complex 2 is required for the development of prostate cancer induced by Pten loss in mice. Cancer Cell.2009, 15(2):148-59
16. Yang J, Shamji A, Matchacheep S, et al. Identification of a small-molecule inhibitor of class Ⅰa PI3Ks with cell-based screening. Chem Biol.2007 Apr; 14(4):371-7.
17. Koike-Kumagai M, Yasunaga K, Morikawa R, et al. The target of rapamycin complex 2 controls dendritic tiling of Drosophila sensory neurons through the tricornered kinase signalling pathway. EMBO J.2009 Dec 16; 28(24):3879-92.
18. Risson V, Mazelin L, Roceri M, et al. Muscle inactivation of mTOR causes metabolic and dystrophin defects leading to severe myopathy. J Cell Biol.2009 Dec 14; 187(6):859-74.
1、 Bhaskar PT, Hay N. The Two TORCs and Akt. Dev Cell.2007,12(4):487-502
2、 Guertin DA, Sabatini DM. Defining the role of mTOR in cancer. Cancer Cell.2007, 12(1):9-22
3、 Laplante M, Sabatini DM. mTOR signaling at a glance. J Cell Sci.2009 Oct 15;122(Pt 20):3589-94
4、 Yilmaz OH, Valdez R, Theisen BK, et al. Pten dependence distinguishes haematopoietic stem cells from leukemia-initiating cells. Nature.2006, 441(7092):475-82
5、 Zhang J, Grindley JC, Yin T, et al. PTEN maintains haematopoietic stem cells and acts in lineage choice and leukaemia prevention. Nature.2006 May 25; 441 (7092):518-22
6、 David A. Guertin and David M. Sabatini. The Pharmacology of mTOR Inhibition. Sci Signal.2009 Apr 21;2(67)
7、 Risson V, Mazelin L, Roceri M, et al. Muscle inactivation of mTOR causes metabolic and dystrophin defects leading to severe myopathy. J Cell Biol.2009 Dec 14; 187(6):859-74.
8、 Shor B, Gibbons JJ, Abraham RT, et al. Targeting mTOR globally in cancer:thinking beyond rapamycin. Cell Cycle.2009 Dec;8(23):3831-7
10、 Alessi DR, Pearce LR, Garcia-Martinez JM. New Insights into mTOR Signaling: mTORC2 and Beyond. Sci Signal.2009 Apr 21; 2(67)
11、 Guertin DA, Stevens DM, Saitoh M, et al. mTOR complex 2 is required for the development of prostate cancer induced by Pten loss in mice. Cancer Cell.2009, 15(2):148-59
12、 Masri J, Bernath A, Martin J, et al. mTORC2 activity is elevated in gliomas and promotes growth and cell motility via overexpression of rictor. Cancer Res.2007 Dec 15; 67 (24):11712-20
13、 Sabatini DM. mTOR and cancer:insights into a complex relationship. Nat Rev Cancer. 2006,6(9):729-34
14、 Peterson TR, Laplante M, Thoreen CC, et al. DEPTOR is an mTOR inhibitor frequently overexpressed in multiple myeloma cells and required for their survival. Cell.2009 May 29;137(5):873-86
15、 Pearce LR, Huang X, Boudeau J, et al. Identification of Protor as a novel Rictor-binding component of mTOR complex-2. Biochem J.2007 Aug 1;405(3):513-22.
16、 Sarbassov DD, Ali SM, Kim DH, et al. Rictor, a Novel Binding Partner of mTOR, defines a Rapamycin-Insensitive and Raptor-Independent Pathway that Regulates the Cytoskeleton. Curr Biol.2004 Jul 27;14(14):1296-302.
17、 Feldman ME, Apsel B, Uotila A, et al. Active-site inhibitors of mTOR target rapamycin-resistant outputs of mTORC1 and mTORC2. PLOS Biol.2009 Feb 10; 7(2):e38
18、 Thoreen CC, Kang SA, Chang JW, et al. An ATP-competitive mammalian target of rapamycin inhibitor reveals rapamycin-resistant functions of mTORC1.J Biol Chem. 2009 Mar 20; 284 (12):8023-32.
19、 Zask A, Verheijen JC, Curran K, et al. ATP-Competitive Inhibitors of the Mammalian Target of Rapamycin:Design and Synthesis of Highly Potent and Selective Pyrazolopyrimidines. J Med Chem.2009 Jul 31.
20、 Maira SM, Stauffer F, Brueggen J, et al. Identification and characterization of NVP-BEZ235, a new orally available dual phosphatidylinositol 3-kinase/mammalian target of rapamycin inhibitor with potent in vivo antitumor activity. Mol Cancer Ther. 2008 Jul; 7(7):1851-63.
21、 Faivre S, Kroemer G, Raymond E. Current development of mTOR inhibitors as anticancer agents. Nat Rev Drug Discov.2006,5(8):671-88.
22、 Tamburini J, Chapuis N, Bardet V,et al. Mammalian target of rapamycin (mTOR) inhibition activates phosphatidylinositol 3-kinase/Akt by up-regulating insulin-like growth factor-1 receptor signaling in acute myeloid leukemia:rationale for therapeutic inhibition of both pathways. Blood.2008 Jan 1; 111(1):379-82.
23、 Gupta M, Ansell SM, Novak AJ, et al. Inhibition of histone deacetylase overcomes rapamycin-mediated resistance in diffuse large B-cell lymphoma by inhibiting Akt signaling through mTORC2. Blood.2009 Oct 1;114(14):2926-35
24、 Koike-Kumagai M, Yasunaga K, Morikawa R, et al. The target of rapamycin complex 2 controls dendritic tiling of Drosophila sensory neurons through the Tricornered kinase signalling pathway. EMBO J.2009 Dec 16; 28(24):3879-92.
1、 Guertin DA, Sabatini DM. Defining the role of mTOR in cancer. Cancer Cell.2007, 12(1):9-22
2、 Laplante M, Sabatini DM. mTOR signaling at a glance. J Cell Sci.2009 Oct 15;122(Pt 20):3589-94
3、 Yilmaz OH, Valdez R, Theisen BK, et al. Pten dependence distinguishes haematopoietic stem cells from leukemia-initiating cells. Nature.2006,441 (7092):475-82
4、 Zhang J, Grindley JC, Yin T, et al. PTEN maintains haematopoietic stem cells and acts in lineage choice and leukaemia prevention. Nature.2006 May 25; 441(7092):518-22
5、 Guertin DA, Stevens DM, Saitoh M, et al. mTOR complex 2 is required for the development of prostate cancer induced by Pten loss in mice. Cancer Cell.2009, 15(2):148-59
6、 Sabatini DM. mTOR and cancer:insights into a complex relationship. Nat Rev Cancer. 2006,6(9):729-34
7、 Bhaskar PT, Hay N. The Two TORCs and Akt. Dev Cell.2007,12(4):487-502
8、 Jacinto E, Facchinetti V, Liu D, et al. SIN1/MIP1 maintains rictor-mTOR complex integrity and regulates Akt phosphorylation and substrate specificity. Cell.2006 Oct 6; 127(1):125-37.
9、 Richter JD, Sonenberg N. Regulation of cap-dependent translation by eIF4E inhibitory proteins. Nature.2005 Feb 3; 433(7025):477-80.
10、 Ma XM, Blenis J. Molecular mechanisms of mTOR-mediated translational control.Nat Rev Mol Cell Biol.2009 May; 10(5):307-18.
11、 Ganley IG, Lam du H, Wang J, et al. ULK1.ATG13.FIP200 complex mediates mTOR signaling and is essential for autophagy. J Biol Chem.2009 May 1; 284(18):12297-305.
12、 Hosokawa N, Hara T, Kaizuka T, et al. Nutrient-dependent mTORCl association with the ULK1-Atg13-FIP200 complex required for autophagy. Mol Biol Cell.2009 Apr; 20(7):1981-91.
13、 Jung CH, Jun CB, Ro SH, et al. ULK-Atg13-FIP200 complexes mediate mTOR signaling to the autophagy machinery. Mol Biol Cell.2009 Apr; 20(7):1992-2003.
14、 Thoreen CC, Kang SA, Chang JW, et al. An ATP-competitive mammalian target of rapamycin inhibitor reveals rapamycin-resistant functions of mTORC1. J Biol Chem. 2009 Mar 20; 284(12):8023-32.
15、 Porstmann T, Santos CR, Griffiths B, et al. SREBP activity is regulated by mTORC1 and contributes to Akt-dependent cell growth. Cell Metab.2008 Sep; 8(3):224-36.
16、 Bentzinger CF, Romanimo K, Cloetta D, et al. Skeletal muscle-specific ablation of raptor, but not of rictor, causes metabolic changes and results in muscle dystrophy. Cell Metab.2008 Nov; 8(5):411-24
17、 Cunningham JT, Rodgers JT, Arlow DH, et al. mTOR controls mitochondrial oxidative function through a YY1-PGC-1 alpha transcriptional complex. Nature.2007 Nov 29; 450(7170):736-40.
18、 Sancak Y, Thoreen CC, Peterson TR, et al. PRAS40 is an insulin-regulated inhibitor of the mTORC1 protein kinase. Mol Cell.2007 Mar 23; 25(6):903-15.
19、 Konstantinopoulos PA, Papavassiliou AG The tuberous sclerosis complex. N Engl J Med.2007 Jan 4; 356(1):92-3.
20、 Vander Haar E, Lee SI, Bandhakavi S, et al. Insulin signalling to mTOR mediated by the Akt/PKB substrate PRAS40. Nat Cell Biol.2007 Mar; 9(3):316-23.
21、 Wang L, Harris TE, Roth RA, et al. PRAS40 regulates mTORC1 kinase activity by functioning as a direct inhibitor of substrate binding. J Biol Chem.2007 Jul 6; 282(27):20036-44.
22、 Hong-Brown LQ, Brown CR, Kazi AA, et al. Alcohol and PRAS40 knockdown decrease mTOR activity and protein synthesis via AMPK signaling and changes in mTORC1 interaction. J Cell Biochem.2010 Apr 15; 109(6):1172-84.
23、 Harrington LS, Findlay GM, Lamb RF. Restraining PI3K:mTOR signalling goes back to the membrane. Trends Biochem Sci.2005 Jan; 30(1):35-42.
24、Zhang H, Bajraszewski N, Wu E, et al. PDGFRs are critical for PI3K/Akt activation and negatively regulated by mTOR. J Clin Invest.2007 Mar; 117(3):730-8.
25、Hardie DG AMP-activated/SNF1 protein kinases:conserved guardians of cellular energy.Nat Rev Mol Cell Biol.2007 Oct; 8(10):774-85.
26、 Gwinn DM, Shackelford DB, Egan DF, et al. AMPK phosphorylation of raptor mediates a metabolic checkpoint. Mol Cell.2008 Apr 25; 30(2):214-26.
27、 Wouters BG, Koritzinsky M. Hypoxia signalling through mTOR and the unfolded protein response in cancer. Nat Rev Cancer.2008 Nov; 8(11):851-64.
28、 DeYoung MP, Horak P, Sofer A, et al. Hypoxia regulates TSC1/2-mTOR signaling and tumor suppression through REDD1-mediated 14-3-3 shuttling. Genes Dev.2008 Jan 15;22(2):239-51.
29、 Li Y, Wang Y, Kim E, et al. Bnip3 mediates the hypoxia-induced inhibition on mammalian target of rapamycin by interacting with Rheb. J Biol Chem.2007 Dec 7; 282(49):35803-13.
30、 Nicklin P, Bergman P, Zhang B, et al. Bidirectional transport of amino acids regulates mTOR and autophagy. Cell.2009 Feb 6; 136(3):521-34.
31、 Nobukuni T, Joaquin M, Roccio M, et al. Amino acids mediate mTOR/raptor signaling through activation of class 3 phosphatidylinositol 3OH-kinase. Proc Natl Acad Sci U S A.2005 Oct 4; 102(40):14238-43.
32、 Juhasz G, Hill JH, Yan Y, et al. The class Ⅲ PI(3)K Vps34 promotes autophagy and endocytosis but not TOR signaling in Drosophila. J Cell Biol.2008 May 19; 181(4):655-66.
33、 Kim DH, Sarbassov DD, Ali SM, et al. mTOR interacts with raptor to form a nutrient-sensitive complex that signals to the cell growth machinery. Cell.2002 Jul 26; 110(2):163-75.
34、 Kim E, Goraksha-Hicks P, Li L, et al. Regulation of TORC1 by Rag GTPases in nutrient response. Nat Cell Biol.2008 Aug; 10(8):935-45.
35、 Sancak Y, Peterson TR, Shaul YD, et al. The Rag GTPases bind raptor and mediate amino acid signaling to mTORC1. Science.2008 Jun 13; 320(5882):1496-501
36、 Feng Z, Zhang H, Levine AJ, et al. The coordinate regulation of the p53 and mTOR pathways in cells. Proc Natl Acad Sci U S A.2005 Jun 7; 102(23):8204-9.
37、 Lee DF, Kuo HP, Chen CT, et al. IKK beta suppression of TSC1 links inflammation and tumor angiogenesis via the mTOR pathway. Cell.2007 Aug 10; 130(3):440-55.
38、 Inoki K, Li Y, Zhu T, et al. TSC2 is phosphorylated and inhibited by Akt and suppresses mTOR signalling. Nat Cell Biol.2002 Sep; 4(9):648-57.
39、 Toschi A, Lee E, Xu L, et al. Regulation of mTORC1 and mTORC2 complex assembly by phosphatidic acid:competition with rapamycin. Mol Cell Biol.2009 Mar; 29(6):1411-20.
40、 Sarbassov DD, Guertin DA, Ali SM, et al. Phosphorylation and regulation of Akt/PKB by the rictor-mTOR complex. Science.2005,307(5712):1098-101
41、 Calnan DR, Brunet A. The FoxO code. Oncogene.2008 Apr 7; 27(16):2276-88.
42、 Peterson TR, Laplante M, Thoreen CC, et al. DEPTOR is an mTOR inhibitor frequently overexpressed in multiple myeloma cells and required for their survival. Cell.2009 May 29;137(5):873-86
43、 Jacinto E, Loewith R, Schmidt A, et al. Mammalian TOR complex 2 controls the actin cytoskeleton and is rapamycin insensitive. Nat Cell Biol.2004 Nov; 6(11):1122-8.
44、 Guertin DA, Stevens DM, Saitoh M, et al. mTOR complex 2 is required for the development of prostate cancer induced by Pten loss in mice. Cancer Cell.2009, 15(2):148-59
45、 Masri J, Bernath A, Martin J, et al. mTORC2 activity is elevated in gliomas and promotes growth and cell motility via overexpression of rictor. Cancer Res.2007 Dec 15； 67 (24):11712-20
46、 Guertin DA, Sabatini DM. Defining the role of mTOR in cancer. Cancer Cell.2007, 12(1):9-22
47、 Q. Yang, K. L. Guan. Expanding mTOR signaling. Cell Res.2007 Aug; 17(8):666-81.
48、 Peterson TR, Laplante M, Thoreen CC, et al. DEPTOR is an mTOR inhibitor frequently overexpressed in multiple myeloma cells and required for their survival. Cell.2009 May 29;137(5):873-86
49、 J. Huang, B. D. Manning. A complex interplay between Akt, TSC2 and the two mTOR complexes. Biochem Soc Trans.2009 Feb; 37(Pt 1):217-22.
50、 Smith EM, Finn SG, Tee AR, et al. The tuberous sclerosis protein TSC2 is not required for the regulation of the mammalian target of rapamycin by amino acids and certain cellular stresses. J Biol Chem.2005 May 13; 280(19):18717-27.
51、 Sancak Y, Peterson TR, Shaul YD, et al. The Rag GTPases bind raptor and mediate amino acid signaling to mTORC1. Science.2008 Jun 13; 320(5882):1496-501
52、 Kim E, Goraksha-Hicks P, Li L, et al. Regulation of TORC1 by Rag GTPases in nutrient response. Nat Cell Biol.2008 Aug; 10(8):935-45.
53、 Yuan TL, Cantley LC. PI3K pathway alter-ations in cancer:Variations on a theme. Oncogene.2008 Sep 18; 27(41):5497-510.
54、 Krymskaya VP, Goncharova EA. PI3K/mTORCl activation in hamartoma syn-dromes: Therapeutic prospects. Cell Cycle.2009 Feb 1; 8(3):403-13.
55、 Lane HA, Breuleux M. Optimal targeting of the mTORCl kinase in human cancer. Curr Opin Cell Biol.2009 Apr; 21(2):219-29.
56、 Hudes G, Carducci M, Tomczak P, et al. Temsirolimus, interferon a, or both for advanced renal-cell carcinoma. N Engl J Med.2007 May 31; 356(22):2271-81.
57、 Choo AY, Yoon SO, Kim SG, et al. Rapamycin differentially inhibits S6Ks and 4E-BP1 to mediate cell-type-specific repression of mRNA translation. Proc Natl Acad Sci U S A.2008 Nov 11; 105(45):17414-9.
58、 Thoreen CC, Kang SA, Chang JW, et al. An ATP-competitive mTOR inhibitor reveals rapamycin-insensitive functions of mTORCl. J Biol Chem.2009 Mar 20; 284(12):8023-32.
59、 Feldman ME, Apsel B, Uotila A, et al. Active-site inhibitors of mTOR target rapamycin-resistant outputs of mTORC1 and mTORC2. PLoS Biol.2009 Feb 10; 7(2):e38.
60、 Dorrello NV, Peschiaroli A, Guardavaccaro D, et al. S6K1-and βTRCP-mediated degradation of PDCD4 promotes protein translation and cell growth. Science.2006 Oct 20;314(5798):467-71.
61、 Holz MK, Ballif BA, Gygi SP, et al. mTOR and S6K1 mediate assembly of the translation preinitiation complex through dynamic protein interchange and ordered phosphorylatio events. Cell.2005 Nov 18; 123(4):569-80.
62、 Jacinto E, Facchinetti V, Liu D, et al. SIN1/MIP1 maintains rictor-mTOR complex integrity and regulates Akt phosphorylation and substrate specificity. Cell.2006 Oct 6; 127(1):125-37.
63、Shiota C, Woo JT, Lindner J, et al. Multiallelic disruption of the rictor gene in mice reveals that mTOR complex 2 is essential for fetal growth and viability. Dev Cell.2006 Oct; 11(4):583-9.
64、 Guertin DA, Stevens DM, Thoreen CC, et al. Ablation in mice of the mTORC components raptor, rictor, or mLST8 reveals that mTORC2 is required for signaling to Akt-FOXO and PKCa, but not S6K1. Dev Cell.2006 Dec; 11(6):859-71.
65、 Garcia-Martinez JM, Alessi DR. mTOR complex-2 (mTORC2) controls hydrophobic motif phosphorylation and activation of serum-and glucocorticoid-induced protein kinase 1 (SGK1). Biochem J.2008 Dec 15; 416(3):375-85.
66、 Sarbassov DD, Guertin DA, Ali SM, et al. Phosphorylation and regulation of Akt/PKB by the rictor-mTOR complex. Science.2005 Feb 18; 307 (5712):1098-101.
67、 Yap TA, Garrett MD, Walton MI, et al. Targeting the PI3K-AKT-mTOR pathway: Progress, pitfalls, and promises. Curr Opin Pharmacol.2008 Aug; 8(4):393-412.
68、 Maira SM, Stauffer F, Brueggen J, et al. Identification and characterization of NVP-BEZ235, a new orally available dual phosphatidylinositol 3-kinase/mammalian target of rapamycin inhibitor with potent in vivo antitumor activity. Mol Cancer Ther. 2008 Jul;7(7):1851-63.
69、 Serra V, Markman B, Scaltriti M, et al. NVP-BEZ235, a dual PI3K/mTOR inhibitor, prevents PI3K signaling and inhibits the growth of cancer cells with activating PI3K mutations. Cancer Res.2008 Oct 1; 68(19):8022-30.
70、 Schnell CR, Stauffer F, Allegrini PR, et al. Effects of the dual phosphatidylinositol 3-kinase/mammalian target of rapamycin inhibitor NVP-BEZ235 on the tumor vasculature:Implications for clinical imaging. Cancer Res.2008 Aug 15; 68(16):6598-607.
71、 Engelman JA, Chen L, Tan X, et al. Effective use of PI3K and MEK inhibitors to treat mutant K-ras G12D and PIK3CAH1047R murine lung cancers. Nat Med.2008 Dec; 14(12):1351-6.
72、 Sarbassov DD, Ali SM, Sengupta S, et al. Prolonged rapamycin treatment inhibits mTORC2 assembly and Akt/PKB. Mol Cell.2006 Apr 21; 22(2):159-68.
73、 Copp J, Manning G, Hunter T. TORC-specific phosphorylation of mammalian target of rapamycin (mTOR):Phospho-Ser2481 is a marker for intact mTOR signaling complex 2. Cancer Res.2009 Mar 1; 69(5):1821-7.
74、 Shor B, Zhang WG, Toral-Barza L, et al. A new pharmacologic action of CCI-779 involves FKBP12-independent inhibition of mTOR kinase activity and profound repression of global protein synthesis. Cancer Res.2008 Apr 15; 68(8):2934-43.
75、 Barquilla A, Crespo JL, Navarro M. Rapamycin inhibits trypanosome cell growth by preventing TOR complex 2 formation. Proc Natl Acad Sci U S A.2008 Sep 23; 105(38):14579-84.
76、 Nardella C, Carracedo A, Alimonti A, et al. Differential requirement of mTOR in postmitotic tissues and tumorigenesis. Sci Signal.2009 Jan 27; 2(55):ra2.
77、 Guertin DA, Stevens DM, Saitoh M, et al. mTOR complex 2 is required for the development of prostate cancer induced by Pten loss in mice. Cancer Cell.2009 Feb 3; 15(2):148-59.
78、 Bentzinger CF, Romanino K, Cloetta D, et al. Skeletal muscle-specific ablation of raptor, but not of rictor, causes metabolic changes and results in muscle dystrophy. Cell Metab.2008 Nov; 8(5):411-24.
79、 Kumar A, Harris TE, Keller SR, et al. Muscle-specific deletion of rictor impairs insulin-stimulated glucose transport and enhances basal glycogen synthase activity. Mol Cell Biol.2008 Jan; 28(1):61-70.
80、 Beevers CS, Chen L, Liu L, et al. Curcumin disrupts the mammalian target of rapamycin-raptor complex. Cancer Res.2009 Feb 1; 69(3):1000-8.
81、 Knight ZA, Gonzalez B, Feldman ME, et al. A pharmacological map of the PI3-K family defines a role for p110α in insulin signaling. Cell.2006 May 19; 125(4):733-47.