代謝組學在腫瘤研究中應用

1、2012-3-13 1-1 代謝組學的定義代謝組學的定義 代謝組學代謝組學 (metabonomics): 某一生物或細胞所有低分子質量代謝產(chǎn)物進行定性和定量檢測某一生物或細胞所有低分子質量代謝產(chǎn)物進行定性和定量檢測, 分析活細胞中代謝物譜變化的研究領域分析活細胞中代謝物譜變化的研究領域 生物標本生物標本: 血液、尿液等體液血液、尿液等體液, 組織、細胞提取物、細胞培養(yǎng)液組織、細胞提取物、細胞培養(yǎng)液 1-2 代謝組學分析技術代謝組學分析技術-NMR-NMR NMR光譜光譜: NMR技術是最早被用于代謝組學研究的技術之一其利用原子核技術是最早被用于代謝組學研究的技術之一其利用原子核 在磁場中的能

2、量變化來獲得相關信息。目前常用的有在磁場中的能量變化來獲得相關信息。目前常用的有 1H-NMR、 13C-NMR和 和31P-NMR 1-2 代謝組學分析技術代謝組學分析技術-NMR-NMR 1-2 代謝組學分析技術代謝組學分析技術-NMR-NMR 1-2 代謝組學分析技術代謝組學分析技術-NMR-NMR Warburg, O. On the origin of cancer cells. Science 123, 309314 (1956) The Warburg Effect 1-2 代謝組學分析技術代謝組學分析技術-MS-MS MS: Mass Spectrometry, 質譜質譜 基本

3、原理是基本原理是: 將樣品中各組分電離成離子束將樣品中各組分電離成離子束, 進入質量分析器聚集進入質量分析器聚集 而得到而得到MS圖譜，以確定其質量。圖譜，以確定其質量。 1-2 代謝組學分析技術代謝組學分析技術-MS-MS 1-2 代謝組學分析技術代謝組學分析技術-MS-MS MS: Mass Spectrometry, 質譜質譜 需聯(lián)合色譜技術對樣品進行前期分離需聯(lián)合色譜技術對樣品進行前期分離 ( 1 ) GC MS聯(lián)用聯(lián)用 ： GC技術是以氣體作為流動相的色技術是以氣體作為流動相的色 譜法常用于分離揮發(fā)性化合物。譜法常用于分離揮發(fā)性化合物。 ( 2 ) LC MS聯(lián)用聯(lián)用 ： LC技技

4、術是術是 以液以液 體作體作 為流為流 動相動相 的的 色色 譜譜 法適用于分離低法適用于分離低 揮發(fā)性或非揮發(fā)性、熱穩(wěn)定性揮發(fā)性或非揮發(fā)性、熱穩(wěn)定性 差差 的物質。的物質。 2-1 Tumor glucose metabolic phenotypes: Glucose glycolysis and oxidative phosphorylation Warburg, O. On the origin of cancer cells. Science 123, 309314 (1956) The Warburg Effect 2-2 Tumor glucose metabolic phenot

5、ypes: The pentose phosphate pathway 2-3 Tumor lipid metabolic phenotypes: Simplified overview of tumor lipid metabolism Journal of Nuclear Medicine 2008, Vol. 49 No. Suppl_2 43S-63S 2-4 Tumor amino acid metabolic phenotypes: Regulation of glycolysis, glutaminolysis and de novo nucleotide biosynthesi

6、s in tumor cells. Current Opinion in Genetics 20(1):51-6. 2-2 HIF-1 in tumor cell metabolism HIF-1: activating transcription of genes encoding glucose transporters and glycolytic enzyme Expression of genes encoding glucose transporters and glycolytic enzymes. The glycolytic pathway is shown at left.

7、 Symbols for genes encoding the respective enzymes are coded by font according to the mRNA expression pattern (normalized to 18S rRNA) in ES cells cultured under nonhypoxic (N) or hypoxic (H) conditions for 16 hr (lanes 16 at right) as follows: (1) (bold) increased expression in hypoxic Hif1a+/+ cel

8、ls, loss of induction in Hif1a+/ cells, and loss of basal and induced expression in Hif1a/ cells; (2) (bold and italicized) no effect of hypoxia on expression in Hif1a+/+ cells but decreased expression in hypoxic Hif1a+/ and Hif1a/ cells; (3) (italicized) no effect of hypoxia on expression in Hif1a+

9、/+ cells but decreased expression in hypoxic and nonhypoxic Hif1a/ cells; (4) (plain) no effect of hypoxia or HIF-1 deficiency on expression. mRNA expression in Hep3B cells was also assayed (lanes 7,8). The indicated genes encode the following proteins: (GLUT1 and GLUT3) glucose transporter 1 and 3;

10、 (HK1 and HK2) hexokinase 1 and 2; (GPI) glucosephosphate isomerase; (PFKL) phosphofructokinase L; (ALDA and ALDC) aldolase A and C; (TPI) triosephosphate isomerase; (GAPDH) glyceraldehyde-3-phosphate dehydrogenase; (PGK1) phosphoglycerate kinase 1; (PGM) phosphoglucomutase; (ENO1) enolase 1; (PKM)

11、pyruvate kinase M; (LDHA) lactate dehydrogenase A. GLUCOSE (EXT) and GLUCOSE (INT) refer to extracellular and intracellular glucose, respectively, Genes Dev. 1998 Jan 15;12(2):149-62. 2-2 HIF-1 in tumor cell metabolism HIF-1: up-regulated the plasma membrane lactate transporter MCT4 The plasma membr

12、ane lactate transporter MCT4, but not MCT1, is up-regulated by hypoxia through a HIF-1a-dependent mechanism. J Biol Chem 2006, 281:9030-9037. 2-2 HIF-1 in tumor cell metabolism PDK PDH HIF-1-mediated expression of pyruvate dehydrogenase kinase: A metabolic switch required for cellular adaptation to

13、hypoxia CELL METABOLISM 3, 177185, MARCH 2006 HIF-1mediates adaptation to hypoxia by actively downregulating mitochondrial oxygen consumption CELL METABOLISM 3, 187197, MARCH 2006 2-2 HIF-1 in tumor cell metabolism Figure 1. HIF-1-dependent induction of PDK1 expression in hypoxic cells Figure 6. Coo

14、rdinate regulation of hypoxia- induced metabolic switches by HIF-1 2-2 HIF-1 in tumor cell metabolism 2-3 mTOR in tumor cell metabolism Mammalian target of rapamycin up-regulation of pyruvate kinase isoenzyme type M2 is critical for aerobic glycolysis and tumor growth 2-3 mTOR in tumor cell metaboli

15、sm: PPP/lipid synthesis Molecular Cell, Volume 39, Issue 2, 171-183, 30 July 2010 Highlights: 1. mTORC1 stimulates glucose uptake and glycolysis through HIF1 2. mTORC1 stimulates the pentose phosphate pathway and lipid biosynthesis through SREBP1 SREBP: sterol regulatory element-binding protein, SRE

16、BPs belong to the basic helix- loop-helixleucine zipper (bHLH-Zip) family of transcription factors 2-3 mTOR in tumor cell metabolism: PPP/lipid synthesis J Clin Invest. 2002;109(9):11251131 2-3 mTOR in tumor cell metabolism: lipid synthesis mTORC1 signaling will contribute to tumor development and p

17、rogression, the stimulation of ribosome biogenesis leading to an overall increase in protein synthesis is likely to be a major mechanism by which mTORC1 promotes anabolic cell growth and proliferation in tumor cells. 2-3 mTOR in tumor cell metabolism J Mol Med (2011) 89:221228 2-4 Summary ? LDHA MCT

18、4 PDH Glutaminase FAS SREBP1/2 二氯乙酸DCA 丙酮酸脫氫酶 臨床實驗 3-1 代謝組學與腫瘤診斷代謝組學與腫瘤診斷 3-1 代謝組學與腫瘤診斷代謝組學與腫瘤診斷 3-1 代謝組學與腫瘤診斷代謝組學與腫瘤診斷 3-1 代謝組學與腫瘤診斷代謝組學與腫瘤診斷 Fig. 6 Lactate-glucose molar ratio for each patient sample. a normal tissue (NT); b cirrhotic tissue (CIR); c hepatocellular carcinoma (HCC); d liver metastasis from colorectal carcinoma (ME