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完整後設資料紀錄
DC 欄位 | 值 | 語言 |
---|---|---|
dc.contributor.advisor | 張以承 | |
dc.contributor.author | Meng-Lun Hsieh | en |
dc.contributor.author | 謝孟倫 | zh_TW |
dc.date.accessioned | 2021-06-16T09:15:56Z | - |
dc.date.available | 2022-08-28 | |
dc.date.copyright | 2017-08-28 | |
dc.date.issued | 2017 | |
dc.date.submitted | 2017-07-17 | |
dc.identifier.citation | [ 1 ] Global burden of diabetes. International Diabetes federation. Diabetic atlas 7thedition 2017, Brussels. Available at http://www.idf.org/diabetesatlas (Accessed 4th April 2017).
[ 2 ] Chen L, Magliano DJ, Zimmet PZ. The worldwide epidemiology of type 2 diabetes mellitus--present and future perspectives. Nat Rev Endocrinol. 2011 Nov 8;8(4):228-36. doi: 10.1038/nrendo.2011.183. [ 3 ] Genetic basis of type 1 and type 2 diabetes, obesity and their complications. Advances and emerging opportunities in diabetes research: a strategic planning report of the DMICC. 2011. www2.niddk.nih.gov/NR 2011. [ 4 ] Kahn CR. Banting Lecture. Insulin action, diabetogenes, and the cause of type II diabetes. Diabetes 1994. Aug;43(8):1066-1084 [ 5 ] Fujioka K. Pathophysiology of type 2 diabetes and the role of incretin hormones and beta-cell dysfunction. JAAPA. 2007 Dec;Suppl:3-8. [ 6 ] Ahmadian M, Suh JM, Hah N, Liddle C, Atkins AR, Downes M, Evans RM. PPARγ signaling and metabolism: the good, the bad and the future. Nat Med. 2013 May;19(5):557-66. [ 7 ] Alarcón de la Lastra C, Sánchez-Fidalgo S, Villegas I, Motilva V. New pharmacological perspectives and therapeutic potential of PPAR-gamma agonists. Curr Pharm Des. 2004;10(28):3505-24 [ 8 ] Barak Y, Nelson MC, Ong ES, Jones YZ, Ruiz-Lozano P, Chien KR, Koder A, Evans RM. PPAR gamma is required for placental, cardiac, and adipose tissue development. Mol Cell. 1999 Oct;4(4):585-95. [ 9 ] Imai T, Takakuwa R, Marchand S, Dentz E, Bornert JM, Messaddeq N, Wendling O, Mark M, Desvergne B, Wahli W, Chambon P, Metzger D. Peroxisome proliferator-activated receptor gamma is required in mature white and brown adipocytes for their survival in the mouse. Proc Natl Acad Sci U S A. 2004 Mar 30;101(13):4543-7. Epub 2004 Mar 16. [ 10 ] Hevener AL, He W, Barak Y, Le J, Bandyopadhyay G, Olson P, Wilkes J, Evans RM, OlefskyJ. Muscle-specific Pparg deletion causes insulin resistance. Nat Med. 2003 Dec;9(12):1491-7. [ 11 ] He W, Barak Y, Hevener A, Olson P, Liao D, Le J, Nelson M, Ong E, Olefsky JM, Evans RM. Adipose-specific peroxisome proliferator-activated receptor gamma knockout causes insulin resistance in fat and liver but not in muscle. ProcNatlAcadSci U S A. 2003 Dec 23;100(26):15712-7. [ 12 ] Gavrilova O, Haluzik M, Matsusue K, Cutson JJ, Johnson L, Dietz KR, Nicol CJ, Vinson C, Gonzalez FJ, Reitman ML. Liver peroxisome proliferator-activated receptor gamma contributes to hepatic steatosis, triglyceride clearance, and regulation of body fat mass. JBiol Chem. 2003 Sep 5;278(36):34268-76. [ 13 ] Odegaard J, Ricardo-Gonzalez RR, Goforth MH, Morel CR, Subramanian V, Mukundan L, Red Eagle A, Vats D, Brombacher F, Ferrante AW, Chawla A. Macrophage-specific PPARgamma controls alternative activation and improves insulin resistance. Nature. 2007 Jun 28;447(7148):1116-20. [ 14 ] Lu M, Sarruf DA, Talukdar S, Sharma S, Li P, Bandyopadhyay G, Nalbandian S, Fan W, Gayen JR, Mahata SK, Webster NJ, Schwartz MW, Olefsky JM. Brain PPAR-γ promotes obesity and is required for the insulin-sensitizing effect of thiazolidinediones. Nat Med. 2011 May;17(5):618-22. [ 15 ] Soccio RE, Chen ER, Lazar MA. Thiazolidinediones and the promise of insulin sensitization in type 2 diabetes. Cell Metab. 2014 Oct 7;20(4):573-91. [ 16 ] Funk CD. Prostaglandins and leukotrienes: advances in eicosanoid biology. Science. 2001 Nov 30;294(5548):1871-5. [ 17 ] Laurence Brunton, Bruce Chabner, Bjorn Knollman. Goodman and Gilman's The Pharmacological Basis of Therapeutics. 2011; 12th Revised edition chapter 33. [ 18 ] Shiraki T, Kamiya N, Shiki S, Kodama TS, Kakizuka A, Jingami H. Alpha,beta-unsaturated ketone is a core moiety of natural ligands for covalent binding to peroxisome proliferator-activated receptor gamma. J Biol Chem. 2005 Apr 8;280(14):14145-53. [ 19 ] Chou WL, Chuang LM, Chou CC, Wang AH, Lawson JA, FitzGerald GA, Chang ZF. Identification of a novel prostaglandin reductase reveals the involvement of prostaglandin E2 catabolism in regulation of peroxisome proliferator-activated receptor gamma activation. J Biol Chem. 2007 Jun 22;282(25):18162-72. [ 20 ] Harmon GS, Dumlao DS, Ng DT, Barrett KE, Dennis EA, Dong H, Glass CK.. Pharmacological correction of a defect in PPAR-gamma signaling ameliorates disease severity in Cftr-deficient mice. Nat Med. 2010 Mar;16(3):313-8 [ 21 ] Rizos CV, Elisaf MS, Mikhailidis DP, Liberopoulos EN. How safe is the use of thiazolidinediones in clinical practice? Expert Opin Drug Saf. 2009 Jan;8(1):15-32. doi: 10.1517/14740330802597821. [ 22 ] Kliewer SA, Lenhard JM, Willson TM, Patel I, Morris DC, Lehmann JM. A prostaglandin J2 metabolite binds peroxisome proliferator-activated receptor gamma and promotes adipocyte differentiation. Cell. 1995 Dec 1;83(5):813-9. [ 23 ] Forman BM, Tontonoz P, Chen J, Brun RP, Spiegelman BM, Evans RM. 15-Deoxy-Δ12, 14-prostaglandin J2 is a ligand for the adipocyte determination factor PPARγ. Cell. 1995 Dec 1;83(5):803-12 [ 24 ] Nagy L1, Tontonoz P, Alvarez JG, Chen H, Evans RM. Oxidized LDL regulates macrophage gene expression through ligand activation of PPARγ. Cell. 1998 Apr 17;93(2):229-40. [ 25 ] Huang JT, Welch JS, Ricote M, Binder CJ, Willson TM, Kelly C, Witztum JL, Funk CD, Conrad D, Glass CK. Interleukin-4-dependent production of PPAR-gamma ligands in macrophages by 12/15-lipoxygenase. Nature. 1999 Jul 22;400(6742):378-82. [ 26 ] McIntyre TM1, Pontsler AV, Silva AR, St Hilaire A, Xu Y, Hinshaw JC, Zimmerman GA, Hama K, Aoki J, Arai H, Prestwich GD. Identification of an intracellular receptor for lysophosphatidic acid (LPA): LPA is a transcellular PPARgamma agonist. Proc Natl Acad Sci U S A. 2003 Jan 7;100(1):131-6. [ 27 ] Schopfer FJ1, Lin Y, Baker PR, Cui T, Garcia-Barrio M, Zhang J, Chen K, Chen YE, Freeman BA. Nitrolinoleic acid: an endogenous peroxisome proliferator-activated receptor gamma ligand. Proc Natl Acad Sci U S A. 2005 Feb 15;102(7):2340-5. [ 28 ] Shibata T, Kondo M, Osawa T, Shibata N, Kobayashi M, Uchida K. 15-deoxy-delta 12,14-prostaglandin J2. A prostaglandin D2 metabolite generated during inflammatory processes. J Biol Chem. 2002 Mar 22;277(12):10459-66. [ 29 ] Tzameli I, Fang H, Ollero M, Shi H, Hamm JK, Kievit P, Hollenberg AN, Flier JS. Regulated production of a peroxisome proliferator-activated receptor-γ ligand during an early phase of adipocyte differentiation in 3T3-L1adipocytes. J Biol Chem. 2004 Aug 20;279(34):36093-102. [ 30 ] Hansen LH, Knudsen S, Sørensen SJ. The effect of the lacY gene on the induction of IPTG inducible promoters, studied in Escherichia coli and Pseudomonas fluorescens. Curr Microbiol. 1998 Jun;36(6):341-7. [ 31] Smith LM, Sanders JZ, Kaiser RJ, Hughes P, Dodd C, Connell CR, Heiner C, Kent SB, Hood LE. Fluorescence detection in automated DNA sequence analysis. Nature. 1986 Jun 12-18;321(6071):674-9. [ 32 ] Structural Genomics Consortium. et al. Protein production and purification. Nat Methods. 2008 Feb;5(2):135-46. doi: 10.1038/nmeth.f.202. [ 33 ] Bradford, M.M. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem. 1976 May 7;72:248-54. [ 34 ] Mahmood T, Yang PC. Western blot: technique, theory, and trouble shooting. N Am J Med Sci. 2012 Sep;4(9):429-34. doi: 10.4103/1947-2714.100998. [ 35 ] Michaelis L, Menten ML, Johnson KA, Goody RS. The original Michaelis constant: translation of the 1913 Michaelis-Menten paper. Biochemistry. 2011 Oct 4;50(39):8264-9. doi: 10.1021/bi201284u. [ 36 ] Angel TE, Aryal UK, Hengel SM, Baker ES, Kelly RT, Robinson EW, Smith RD Mass spectrometry-based proteomics: existing capabilities and future directions. Chem Soc Rev. 2012 May 21;41(10):3912-28. doi: 10.1039/c2cs15331a. [ 37 ] Cao E, et al. Effect of freezing and thawing rates on denaturation of proteins in aqueous solutions. Biotechnol Bioeng. 2003 Jun 20;82(6):684-90. [ 38 ] Sahdev S, Khattar SK, Saini KS (2008) Production of active eukaryotic proteins through bacterial expression systems: a review of the existing biotechnology strategies. Mol Cell Biochem. 2008 Jan;307(1-2):249-64. [ 39] Wu, Y.H., Ko, T.P., Guo, R.T., Hu, S.M., Chuang, L.M., Wang, A.H. Structural basis for catalytic and inhibitory mechanisms of human prostaglandin reductase PTGR2. Structure. 2008 Nov 12;16(11):1714-23. doi: 10.1016/j.str.2008.09.007. [ 40 ] Douglas KT. Mechanism of action of glutathione-dependent enzymes. Adv Enzymol Relat Areas Mol Biol. 1987;59:103-67. [ 41 ] Tommy Liljefors, Povl Krogsgaard-Larsen, Ulf Madsen. Textbook of Drug Design and Discovery, Third Edition. July 25, 2002 by CRC Press. [ 42 ] Reynolds CH, Merz KM, Ringe D, eds.. Drug Design: Structure- and Ligand-Based Approaches (1 ed.). 2010.Cambridge, UK: Cambridge University Press. [ 43 ] Caraus I, Alsuwailem AA, Nadon R, Makarenkov V. Detecting and overcoming systematic bias in high-throughput screening technologies: a comprehensive review of practical issues and methodological solutions. Brief Bioinform. 2015 Nov;16(6):974-86. doi: 10.1093/bib/bbv004. [ 44 ] Lengauer T, Rarey M. Computational methods for biomolecular docking.. Curr Opin Struct Biol. 1996 Jun;6(3):402-6. [ 45 ] Goldman BB1, Wipke WT. QSD quadratic shape descriptors. 2. Molecular docking using quadratic shape descriptors (QSDock). Proteins. 2000 Jan 1;38(1):79-94. [ 46 ] Feig M, Onufriev A, Lee MS, Im W, Case DA, Brooks CL 3rd. Performance comparison of generalized born and Poisson methods in the calculation of electrostatic solvation energies for protein structures. J Comput Chem. 2004 Jan 30;25(2):265-84. [ 47] Sies H. Oxidative stress: Oxidants and antioxidants. Exp Physiol. 1997 Mar;82(2):291-5. [ 48 ] Smale ST. Luciferase assay. Cold Spring Harb Protoc. 2010 May;2010(5):pdb.prot5421. doi: 10.1101/pdb.prot5421. [ 49 ] Macielag MJ. Chemical properties of antibacterials and their uniqueness. In Dougherty TJ, Pucci MJ. Antibiotic Discovery and Development. 2012. pp. 801–2. ISBN 978-1-4614-1400-1 [ 50 ] Veber DF, Johnson SR, Cheng HY, Smith BR, Ward KW, Kopple KD. Molecular properties that influence the oral bioavailability of drug candidates. J Med Chem. 2002 Jun 6;45(12):2615-23. [ 51 ] Xie W1, Wood AR, Lyssenko V, Weedon MN, Knowles JW, Alkayyali S, Assimes TL, Quertermous T, Abbasi F, Paananen J, Häring H, Hansen T, Pedersen O, Smith U, Laakso M; MAGIC Investigators; DIAGRAM Consortium; GENESIS Consortium; RISC Consortium, Dekker JM, Nolan JJ, Groop L, Ferrannini E, Adam KP, Gall WE, Frayling TM, Walker M. Genetic variants associated with glycine metabolism and their role in insulin sensitivity and type 2 diabetes. Diabetes. 2013 Jun;62(6):2141-50. doi: 10.2337/db12-0876. Epub 2013 Feb 1. | |
dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/59092 | - |
dc.description.abstract | 過氧化物酶體增殖物活化受體γ(PPARγ)主要表現在脂肪組織中,透過與配體結合,在全身能量代謝、葡萄糖恆定以及胰島素阻抗性中扮演重要的調節功能。由於它在脂肪及葡萄糖中重要的代謝功能,所以PPARγ被視為治療疾病的重要標的,例如:第二型糖尿病、脂肪肝等疾病。目前廣泛使用的抗糖尿病藥物,是一種稱為thiazolidinedione(TZD)藥物,此藥物是人工合成之PPARγ強力配體。然而,目前人工合成的PPARγ配體,會有水分滯留、體重增加、及骨質疏鬆的副作用。因此需要開發更有效能的藥物去調控PPARγ活化以調控脂肪及葡萄糖之代謝。現今,PPARγ天然的內生性配體仍不確定。我們團隊過去研究發現15-keto-PGE2為PPARγ內生性的配體, 吾人更盡一步發現前列腺素還原酶-2(PTGR2)是一種可將15-keto-PGE2代謝成為無活性13,14-dihydro-15- keto-PGE2的酵素。Ptgr2基因剔除鼠餵食在高脂高糖飲食之下,相較於對照組,變得更瘦、胰島素敏感性增加、葡萄糖耐受性增加,但沒有水分滯留及骨質疏鬆的副作用,證實抑制PTGR2為治療糖尿病與肥胖的重要新途徑。本研究,目的為開發PTGR2的小分子抑制劑,吾人利用理性藥物設計及高通量化合物篩選方式以篩選出PTGR2小分子抑制物,並進一步在細胞株透過驗證其抑制PTGR2酵素活性、活化PPARγ的能力、以及其細胞毒性。綜合以上分析,找尋最適合的抑制物以進行日後藥物最佳化與活體實驗。 | zh_TW |
dc.description.abstract | Peroxisome proliferator-activated receptor γ (PPARγ) is a master regulator of whole-body energy metabolism, glucose homeostasis, and insulin resistance mainly expressed in adipose tissue. PPARγ acts through transcriptional regulation of genes involved in glucose and energy homeostasis upon ligand binding. The widely used anti-diabetic agent thiazolidinediones (TZD) are potent synthetic PPARγ agonist. However, most PPARγ agonists are associated with significant side effects, such as water retention, increased adiposity, and osteoporosis. There is an urgent need to develop effective way to modulate PPARγ activity without unwanted side effects. Nowadays, the natural ligands for PPARγ are still not certain (mostly oxidized fatty acids). Our team previously found prostaglandin reductase-2 (PTGR2) can catalyze15-keto-PGE2 to 13,14-dihydro-15-keto-PGE2, and provided further evidence that15-keto-PGE2 is an endogenous PPARγ ligand. Our team further found that Ptgr2 knockout mice are leaner, more insulin sensitive, and more glucose tolerant than control in high-fat high-sucrose (HFHS) diet without fluid retention and osteoporosis, indicating PTGR2 inhibition is a novel therapeutic approach for treating type 2 diabetes and obesity.
In this study, we sought to identify PTGR2 small-molecule inhibitor through rational drug design and high-throughput compound screening. We further validated the enzymatic inhibitory activities, the PPARγ transactivating activities, and the cytotoxicity of these hits. Based on this study, we identified potential PTGR2 inhibitors for further optimization and animal experiment. | en |
dc.description.provenance | Made available in DSpace on 2021-06-16T09:15:56Z (GMT). No. of bitstreams: 1 ntu-106-R04455002-1.pdf: 2163769 bytes, checksum: 33ec9d285b3c0b320cf9f77dd18678e6 (MD5) Previous issue date: 2017 | en |
dc.description.tableofcontents | 誌謝 i
中文摘要 ii 英文摘要 iii Chapter I – Backgrounds and Rationale. ……………………………………..….…..1 1. Type 2 diabetes mellitus…………………………………………………......….….1 2. PPARγ is a mater regulator of energy metabolism…………………………….…..1 3. Synthetic thiazolidinedione have serious side effects………….…………….….…3 4. The search for endogenous PPARγ ligands…………………………………......…4 5. Metabolic phenotype of Ptgr2 knockout mice and experimental rationale….........7 6. Specific aim of this study………………………………………………….….…..10 Chapter II – Materials and Methods………………………………………..............11 1. cDNA cloning, expression, purification, and functional characterization of human prostaglandin reductase 2......................................................................................11 2. Enzymatic assay for human prostaglandin reductase 2..........................................13 3. Prostaglandin analysis...........................................................................................14 4. High-throughput compound screening..................................................................15 5. Validation of hit by measurement of the half maximal inhibitory concentration (IC50)....................................................................................................................17 6. Validation of hits by analyzing PPARγ transactivating activities in cultured cells.......................................................................................................................18 7. Evaluation of cytotoxicity using MTS assay........................................................19 Chapter III – Results……………………………………………….....……………...20 1. cDNA cloning, expression, purification, and functional characterization of human prostaglandin reductase 2.......................................................................................20 2. Identifying human prostaglandin reductase 2 inhibitors through rational drug design and high-throughput compound screening, and evaluating their toxicity...22 3. Analyzed PPARγ transactivaing activities in cell-base level ................................24 Chapter VI – Discussion……….……………………….……..…....…………...........25 Figures ……………………………………………….………..……………….......….30 Tables…………………………………………………………….…..…………......….52 References …………………………………………..……………..……….….......….57 Appendix……...............................................................................................................66 | |
dc.language.iso | en | |
dc.title | 透過高通量化合物篩選人類前列腺素還原酶-2小分子抑制劑 | zh_TW |
dc.title | Identification of small-molecule human PTGR-2 inhibitor through high-throughput compound screening | en |
dc.type | Thesis | |
dc.date.schoolyear | 105-2 | |
dc.description.degree | 碩士 | |
dc.contributor.oralexamcommittee | 莊立民,潘思樺 | |
dc.subject.keyword | 前列腺素還原?2,過氧化小體增生活化受體γ,第2型糖尿病,降血糖藥物,小分子抑制劑, | zh_TW |
dc.subject.keyword | PTGR-2,PPARγ,type 2 diabetes,oral anti-diabetic agents,enzyme inhibitor, | en |
dc.relation.page | 66 | |
dc.identifier.doi | 10.6342/NTU201701633 | |
dc.rights.note | 有償授權 | |
dc.date.accepted | 2017-07-18 | |
dc.contributor.author-college | 醫學院 | zh_TW |
dc.contributor.author-dept | 基因體暨蛋白體醫學研究所 | zh_TW |
顯示於系所單位: | 基因體暨蛋白體醫學研究所 |
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