聚脯胺酸三螺旋環肽骨架的合成及其於G蛋白耦聯受體之應用

簡易檢索 / 詳目顯示

回結果列表

研究生：	柯建瑜 Ko, Chien-Yu
論文名稱：	聚脯胺酸三螺旋環肽骨架的合成及其於G蛋白耦聯受體之應用 Synthesis of Polyproline tri-Helix Macrocycle Scaffold and Their Applications for G Protein-Coupled Receptors
指導教授：	王聖凱 Wang, Sheng-Kai
口試委員:	洪嘉呈 Horng, Jia-Cherng 劉雅雯 Liu, Ya-Wen
學位類別：	碩士 Master
系所名稱：	理學院 - 化學系 Department of Chemistry
論文出版年：	2022
畢業學年度：	110
語文別：	中文
論文頁數：	135
中文關鍵詞：	G蛋白耦聯受體、聚脯胺酸螺旋、聚脯胺酸三螺旋環肽、多巴胺D2受體、多價交互作用、正交保護
外文關鍵詞：	G protein-coupled receptor, polyproline helix, polyproline tri-helix macrocycle, dopamine D2 receptors, multivalent interaction, orthogonal protection
相關次數：	點閱：1 下載：0
分享至:	分享至facebook 分享至twitter

查詢本校圖書館目錄查詢臺灣博碩士論文知識加值系統勘誤回報

摘要
帕金森氏症是一種影響神經中樞的慢性退化疾病，目前其成因還不清楚。近年來的研究發現，G蛋白偶聯受體中的腺苷酸A2A受體、多巴胺D2受體和其之間的交互作用可緩和神經傳遞，進而於藥理學上用於治療帕金森氏症。若能使藥物和此數個受體更好的結合，將可提升其治療效果。
對於具多價性的藥物而言，若剛性骨架上配基的距離與受體之間的距離匹配，其則親和性和選擇性將優於非剛性骨架。因此本研究將開發把一至多個腺苷酸A2A受體之拮抗劑與多巴胺D2受體之促效劑，接上非天然聚脯胺酸單螺旋多肽的方法，藉此達到兩藥物的距離控制。接著利用在固相上的模組化組裝反應合成保護基排列方式不同的聚脯胺酸三螺旋環肽，並將藥物接上後，研究其之間於G蛋白偶聯受體親合度的差異。未來再透過此分子骨架正交保護的技術，將可以研發出具有強結合能力與高選擇性的複合藥物。

Abstract
Parkinson's disease is a chronic degenerative disease that affects the center nerve system from unclear cause. Recent studies have found that the interaction between adenosine A2A receptors and dopamine D2 receptors, which belong to G protein-coupled receptors, can mediate neurotransmission and can further to be used as the improved treatment of Parkinson's disease. Therefore, if the ligands can simultaneously act on several receptors, the therapeutic effect can be enhanced.
For multivalent ligands design, if the distance between receptors is matched by the ligands on a rigid scaffold, the avidity and selectivity will be better than flexible scaffolds. Therefore, in this thesis, we developed the strategy to control the distance between adenosine A2A receptor antagonists and dopamine D2 receptor agonists by connecting them onto the rigid polyproline scaffolds separately. Furthermore, we synthesized polyproline tri-helix macrocycles with different arrangement of protecting groups by the modular assembly of peptide on solid support, study the avidity to G protein-coupled receptors after conjugated them with ligands. In the future, through the technique of orthogonal protection and ligand conjugation on polyproline scaffolds, the drug with strong avidity and high selectivity can be developed.

摘要    i
目錄    iii
圖目錄    v
表目錄    vi
流程目錄    vii
縮寫對照表    viii
第一章、緒論    1
1. 受體    1
2. G蛋白偶聯受體    2
2.1. G蛋白偶聯受體簡介    2
2.2. G蛋白偶聯受體之生物測試與研究發展    4
3. 多價交互作用    7
4. 聚脯胺酸多肽    11
5. 正交保護基團    13
6. 研究動機    16
7. 實驗設計    16
第二章、結果與討論    19
1. 脯胺酸構築單元之合成與開發    19
1.1. Alloc構築單元之合成    19
1.2. Cbz構築單元之開發與合成    20
2. 多巴胺D2受體促效劑之合成以及其官能基的修飾    22
2.1. (±)-PPHT-NH2之合成    22
2.2. (±)-PPHT-NH2官能基之轉換    23
3. 具異價配基的聚脯胺酸多肽之合成    24
3.1. 聚脯胺酸單螺旋多肽與聚脯胺酸三螺旋環肽之合成策略    24
3.2. 聚脯胺酸單螺旋多肽的合成與修飾    25
3.3. 聚脯胺酸單螺旋多肽與配基的耦合    26
3.4. 聚脯胺酸三螺旋環肽的合成以及與配基的耦合    28
4. 具多巴胺D2受體促效劑的聚脯胺酸多肽合成及生物測試    34
4.1. 具多價促效劑的聚脯胺酸單螺旋多肽之合成與生物測試    34
4.2. 具多價促效劑的聚脯胺酸三螺旋環肽之合成策略與生物測試    37
5. 結論    42
第三章、實驗方法與材料    44
1. General Methods for Synthesis and Characterization    44
2. Synthesis of Peptide Building Blocks    45
3. Synthesis of modified-(±)-PPHT    55
4. Synthesis and Analytical data of Peptide Scaffolds    65
4.1. General methods for peptide synthesis and analysis    65
4.1.1. Materials and peptide analysis    65
4.1.2. Peptide acid synthesis    65
4.1.3. Peptide amide synthesis    67
4.1.4. Polyproline N-terminus azido modification    68
4.1.5. Polyproline C-terminus alkynyl modification    68
4.1.6. Peptide assembly by CuAAC reaction on resins    69
4.1.7. Peptide cyclization    69
4.1.8. N-Alloc deprotection    69
4.1.9. N-Cbz deprotection    70
4.1.10. GPCRs ligands coupling on scaffold    70
4.2. Synthesis of peptide monomers: peptide amides    71
4.3. Synthesis of peptide monomers: peptide acids    73
4.4. Synthesis of peptide monomers: peptide alkynes    77
4.5. Synthesis of linear peptide trimers: peptide acids    81
4.6. Synthesis of linear peptide trimers: peptide alkynes    84
4.7. Synthesis of cyclic peptide trimers    87
4.8. Synthesis of scaffold 26, 37, 42, 43 and 72–75    92
4.8.1. N-Alloc deprotection    92
4.8.2. Adenosine A2AR antagonist 22 installation     94
4.8.3. N-Cbz deprotection    96
4.8.4. Dopamine D2R agonist 20 installation     103
第四章、參考資料    111
附錄    115

                                

1. Alberts, B.; Bray, D.; Hopkin, K.; Johnson, A.; Lewis, J.; Raff, M.; Roberts, K.; Walter, P. Essential Cell Biology (Fourth ed.). 2014. New York, NY, USA: Garland Science. p. 534.
2. Agnati, L. F.; Tarakanov, A.O.; Ferre, S.; Fuxe, K.; Guidolin, D. Receptor-receptor interactions, receptor mosaics, and basic principles of molecular network organization. J. Mol. Neurosci. 2005, 26, 193–208.
3. Guidolin, D.; Marcoli, M.; Tortorella, C.; Maura, G.; Agnati, L. F. Receptor-Receptor Interactions as a Widespread Phenomenon: Novel Targets for Drug Development? Front. Endocrinol. 2019, 10, 53.
4. Escuela, D. O. B.; Romero-Fernandez, W.; Rivera, A.; Fuxe, K. On the G-Protein-Coupled Receptor Heteromers and Their Allosteric Receptor-Receptor Interactions in the Central Nervous System: Focus on Their Role in Pain Modulation. Evidence-Based Complementary Altern. Med. 2013, 9, 1–17.
5. Hanlon, C. D.; Andrew, D. J. Outside-in signaling--a brief review of GPCR signaling with a focus on the Drosophila GPCR family. J. Cell Sci. 2015, 128, 3533–3542.
6. King, N.; Hittinger, C. T.; Carroll, S. B. Evolution of key cell signaling and adhesion protein families predates animal origins. Science. 2003, 301, 361–363.
7. Filmore, D. It's a GPCR World. Mod. Drug Discovery. 2004, 7, 24–28.
8. Hauser, A. S.; Attwood, M. M.; Anderson, M. R.; Schiöth, H. B.; Gloriam, D. E. Trends in GPCR drug discovery: new agents, targets and indications. Nat. Rev. Drug Discovery. 2017, 16, 829–842.
9. Hurowitz, E. H.; Melnyk, J. M.; Chen, Y.-J.; Kouros-Mehr, H.; Simon, M. I.; Shizyua, H. Genomic Characterization of the Human Heterotrimeric G Protein , , and  Subunit Genes. DNA Res. 2000, 7, 111–120.
10. Jeremic, D.; Sanchez-Rodriguez, I.; Jimenez-Diaz, L.; Navarro-Lopez, J. D. Therapeutic potential of targeting G protein-gated inwardly rectifyingpotassium (GIRK) channels in the central nervous system. Pharmacol. Ther. 2021, 223, 107808.
11. Liu, K.; Noel, S.; Titus, S. A.; Inglese, J.; Eskay, R. L.; Shinn, P.; Austin, C. P.; Heilig, M. A.; Zheng, W. A Multiplex Calcium Assay for Identification of GPCR Agonists and Antagonists. Assay Drug Dev. Technol. 2010, 8, 367–379.
12. Kamai, M.; Jockers, R. Biological significance of GPCR heteromerization in the
neuro‑endocrine system. Front. Endocrinol. 2011, 2, 1–14.
13. Fuxe, K.; Marcellino, D.; Genedani, S.; Agnati, L. Adenosine A2A Receptors, Dopamine D2 Receptors and Their Interactions in Parkinson’s Disease Mov. Disord. 2007, 22, 1990–2017.
14. Fernandes, S. M.; Lee, Y. S.; Gillies, R. J.; Hruby, V. J. Synthesis and evaluation of bivalent ligands for binding to the human melanocortin-4 receptor. Bioorg. Med. Chem. 2014, 22, 6360–6365.
15. Mammen, M.; Choi, S. K.; Whitesides, G. M. Polyvalent Interactions in Biological Systems: Implications for Design and Use of Multivalent Ligands and Inhibitors. Angew. Chem. Int. Ed. 1998, 37, 2754–2794.
16. Dam, T. K.; Gerken, T. A.; Brewer, C. F. Thermodynamics of Multivalent Carbohydrate-Lectin Cross-linking Interactions: Importance of Entropy in the Bind and Jump Mechanism. Biochemistry 2009, 48, 3822–3827.
17. Cecioni, S.; Imberty, A.; Vidal, S. Glycomimetics versus multivalent glycoconjugates for the design of high affinity lectin ligands. Chem. Rev. 2015, 115 , 1, 525–561.
18. 溫興荃。聚脯胺酸環肽的合成及其在DC-SIGN和Langerin之應用。碩士論文，國立清華大學化學系，新竹市，2018。
19. Ruggireo, M. T.; Sibik, J.; Orlando, R.; Zeitler, J. A.; Korter, T. M. Measuring the Elasticity of Poly-L-Proline Helices with Terahertz Spectroscopy. Angew. Chem. Int. Ed. 2016, 55, 6877–6881.
20. Kumin, M.; Sonntag, L.-S.; Wennemers, H.; J. Am. Soc. Chem. Azidoproline Containing Helices: Stabilization of the Polyproline II Structure by a Functionalizable Group. 2007, 129, 466–467.
21. Wilhelm, P.; Lewandowski, B.; TrappN.; Wennemers, H. A Crystal Structure of an Oligoproline PPII-Helix, at Last. J. Am. Chem. Soc. 2014, 136, 15829–15832.
22. Krimm, S.; Mark, J. E.; Tiffany, M. L. Influence of counterions on the helical conformations of charged polypeptide chains. Biopolymers 1969, 8, 695–697.
23. Adzhubei, A. A.; Sternberg, M. J. E.; Makarov, A. A. Polyproline-II Helix in Proteins: Structure and Function. J. Mol. Biol. 2013, 425, 2100–2132.
24. Kroll, C.; Mansi, R; Braun, F,; Dobitz, S.; Maecke, H.R.; Wennemers, H. Hybrid Bombesin Analogues: Combining an Agonist and an Antagonist in Defined Distances for Optimized Tumor Targeting. J. Am. Chem. Soc. 2013, 135, 16793–16796.
25. Lin, T.-H.; Lin, C.-H.; Liu, Y.-J.; Huang, C.-Y.; Lin, Y.-C.; Wang, S.-K. Controlling Ligand Spacing on Surface: Polyproline-Based Fluorous Microarray as a Tool in Spatial Specificity Analysis and Inhibitor Development for Carbohydrate–Protein Interactions. ACS Appl. Mater. Interfaces 2017, 9, 41691–41699.
26. Senoo, A.; Yamada, Y.; Ojima, K.; Doura, T.; Hamachi, I.; Kiyonaka, S. Orthogonal activation of metabotropic glutamate receptor using coordination chemogenetics. Front. Chem. 2022, 9, 1–10.
27. Iha, R. K.; Wooley, K. L.; Nyström, A. M.; Burke, D. J.; Kade, M. J.; Hawker, C. J. Applications of Orthogonal “Click” Chemistries in the Synthesis of Functional Soft Materials. Chem. Rev. 2009, 109, 5620–5686.
28. Iha, R. K.; Wooley, K. L.; Nyström, A. M.; Burke, D. J.; Kade, M. J.; Hawker, C. J. Applications of Orthogonal “Click” Chemistries in the Synthesis of Functional Soft Materials. Chem. Rev. 2009, 109, 5620–5686.
29. Wong, C. H.; Zimmerman, S. C. Orthogonality in organic, polymer, and supramolecular chemistry: from Merrifield to click chemistry. Chem. Commun. 2013, 49, 1679.
30. Isidro-Llobet, A.; Álvarez, M.; Albericio, F. Amino Acid-Protecting Groups Chem. Rev. 2009, 109, 2455–2504.
31. James. “Protecting Groups for Amines – Carbamates” Master Organic Chemistry. 7 Jun. 2018.
32. Mammen, M.; Choi, S. K.; Whitesides, G. M. Polyvalent Interactions in Biological Systems: Implications for Design and Use of Multivalent Ligands and Inhibitors. Angew. Chem. Int. Ed. 1998, 37, 2754–2794.
33. Romantini, N.; Alam, S.; Dobitz, S.; Spillmann, M.; Foresta, M. D.; Schibli, R.; Schertler, G. F. X.; Wennemers, H.; Deupi, X.; Behe, M.; Berger, P. Exploring the signaling space of a GPCR using bivalent ligands with a rigid oligoproline backbone. Proc. Natl. Acad. Sci. U.S.A. 2021, 118, 1–8.
34. Tsai, C.-L.; Wu, S.-Y.; Hsu, H.-K.; Huang, S.-B.; Lin, C.-H.; Chan, Y.-T.; Wang, S.-K. Preparation and conformational analysis of polyproline tri-helix macrocycle nanoscaffolds of varied sizes. Nanoscale 2021, 13, 4592–4601.
35. Soriano, A.; Ventura, R.; Molero, A.; Hoen, R.; Casado, V.; Cortes, A.; Fanelli, F.; Albericio, F.; Lluis, C.; Franco, R.; Royo, M. Adenosine A2A Receptor-Antagonist/Dopamine D2 Receptor-Agonist Bivalent Ligands as Pharmacological Tools to Detect A2A-D2 Receptor Heteromers. J. Med. Chem. 2009, 52, 5590–5602.
36. Wang, Y.; Yang, X.-F.; Zhong, Y.; Gong, X.; Li, Z.; Li, H. Development of a red fluorescent light-up probe for highly selective and sensitive detection of vicinal dithiol-containing proteins in living cells. Chem. Sci. 2016, 7, 518–524.
37. Lin, C. H.; Wen, H. C.; Chiang, C.C.; Huang, J.S.; Chen, Y.; Wang, S. K. Polyproline Tri‐Helix Macrocycles as Nanosized Scaffolds to Control Ligand Patterns for Selective Protein Oligomer Interactions. Small 2019, 1900561.
38. Ferré,S.; Casadó, V.; Devi, L.A.; Filizola, M.; Jockers, R.; Lohse, M.J.; Milligan, G.; Pin, J.P.; Guitart, X. G protein-coupled receptor oligomerization revisited: functional and pharmacological perspectives. Pharmacol Rev. 2014, 66, 413–434.
39. Roe, M. W.; Lemasters, J. J.; Herman, B. Assessment of Fura-2 for measurements of cytosolic free calcium. Cell Calcium 1990, 11, 63–73.
40. 林勤皓。聚脯胺酸環肽骨架的開發與其在選擇性多價蛋白質交互作用之應用。博士論文，國立清華大學化學系，新竹市，2019。

簡易檢索 / 詳目顯示

相關論文