設計並合成具標靶傳送與增進癌細胞內累積量之長效型艾黴素製劑

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姓名

盧泉潓(Chuan-Hui Lu) 查詢紙本館藏

畢業系所

化學工程與材料工程學系

論文名稱

設計並合成具標靶傳送與增進癌細胞內累積量之長效型艾黴素製劑
(Design and Synthesis of Long-Acting Doxorubicin Derivatives Aiming Targeted Delivery and Enhanced Drug Translocation)

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摘要(中)

在此項研究中，我們設計一個具有專一性傳遞與增加藥物在細胞內累積量的長效型艾黴素(DOX)載體。從先前的研究中發現，IL類似物可攜帶小分子螢光物質FITC進到細胞裡; 根據這樣的發現，我們進一步想利用此條胜肽與小分子藥物-艾黴素進行共價接合增加攜帶藥物進入癌細胞中的累積量。更進一步，我們將一具有生物相容性的高分子聚乙二醇(polyethylene glycol, PEG) 藉以保護胜肽，避免酵素將胜肽切割進而達到延長體內循環的時間。此外我們也設計一段蛋白酶分解片段，使載體能在特定細胞周圍釋放。研究結果中發現，在聚乙二醇保護下，切割一定量的胜肽從十分鐘延長至四小時，可證明聚乙二醇可有效的保護胜肽。同時，在蛋白酶切割後，我們觀察到藥物在細胞內累積量上升。因此，我們認為此一藥物載體設計可應用於實際藥物傳遞，並且能增進藥物的累積量。

摘要(英)

In this study, we designed and synthesized the novel doxorubicin (DOX) formulation for enhanced drug delivery and cancer cell targeting. An Indolicidin analogue was covalently conjugated to DOX and acted as a transmembrane carrier for enhancing drug translocation into HepG2 cells. The bioinert polymer, polyethylene glycol (PEG), was conjugated with peptide for prolonging in vivo drug circulation. The protease cleavable sequences were designed for cancer cell recogition. The PEG segment could be detached from DOX-peptide in the presence of protease. It was found that the half-life in trypsin after PEG protection could be increased from 10 minutes to 4 hours. And the enhanced penetration could be observed after protease cleavage. Thus, this DOX formulation is able to be a potential to apply in pharmaceutical use.

關鍵字(中)

★ 抗生胜肽
★ 藥物載體

關鍵字(英)

★ Doxorubicin
★ PEGylation
★ Cell-penetrating peptide

論文目次

Abstract (English) I

Abstract (Chinese) II

List of Figures V

List of Tables VII

Chapter 1 Introduction 1

1.1 Research motivation 1

1.2 Research purpose 3

Chapter 2 Literature survey 4

2.1 Importance of drugs or biologics delivery 4

2.1.1 Small molecule drugs 4

2.1.2 Peptide and protein drug 6

2.1.3 Genetic drugs 7

2.2 Carriers in drug delivery 7

2.2.1 Metal-based Nanoparticles 9

2.2.2 Polymer-based carriers 11

2.2.3 Liposomal-based carriers 12

2.2.3.1 Enhanced permeability and retention (EPR) effect 13

2.2.4 Peptide-based carriers 14

2.2.4.1 Antimicrobial peptides (AMPs) 14

2.2.4.2 Cell-penetrating peptides (CPPs) 15

2.3 DOX delivery 16

2.3.1 Structure of Doxil® 17

2.3.2 Improved PEGylated liposomal doxorubicin 18

2.4 Targeting design for drug delivery 20

2.4.1 Antibody 20

2.4.2 Matrix metalloproteinase-2/9 21

2.5 Biostability in drug delivery system 23

2.6 Molecular design for DOX delivery 24

Chapter 3 Materials and Methods 25

3.1 Chemical compounds and biomolecules 25

3.2 Experiment framework 26

3.2.1 PEGylation process 27

3.2.2 Protease digestion 28

3.2.3 Analysis of high-performance liquid chromatography 29

3.2.4 Peptide conjugated with DOX using sulfo-SMPB 29

3.2.5 Complete formulation synthesis process 29

3.2.6 Cytotoxicity assay 30

Chapter 4 Delivery System Design 31

4.1 Role of CPP for DOX intracellular translocation 31

4.1.1 The role of free CPP on DOX intracellular translocation 32

4.1.2 Synthesis of CPP and DOX conjugates 34

4.1.3 The role of conjugated CPP on DOX intracellular translocation 36

4.2 Shortest MMP-2 targeting sequence design 38

4.3 Enzyme digestion of peptides and its PEGylated form 42

4.3.1 Peptide PEGylation and purification 43

4.3.2 Effect of PEG length on the protease stability of CPP 46

4.3.3 MMP-2 digestion of PEGylated MMP-2 recognizable peptide 48

4.4 Novel formulation of PEG-GPLGI-P17-DOX conjugate 49

4.4.1 Synthesis of PEG-GPLGI-P17-DOX conjugate 49

4.4.2 Cell toxicity of PEG-GPLGI-P17-DOX conjugate 50

Chapter 5 Conclusions 52

Reference 53

參考文獻

1. Cortes-Funes, H.; Coronado, C., Role of anthracyclines in the era of targeted therapy. Cardiovascular toxicology 2007, 7 (2), 56-60.

2. Zhao, Q.; Han, B.; Wang, Z.; Gao, C.; Peng, C.; Shen, J., Hollow chitosan-alginate multilayer microcapsules as drug delivery vehicle: doxorubicin loading and in vitro and in vivo studies. Nanomedicine : nanotechnology, biology, and medicine 2007, 3 (1), 63-74.

3. Zhu, L.; Kate, P.; Torchilin, V. P., Matrix metalloprotease 2-responsive multifunctional liposomal nanocarrier for enhanced tumor targeting. ACS nano 2012, 6 (4), 3491-8.

4. Liang, J. F.; Yang, V. C., Synthesis of doxorubicin-peptide conjugate with multidrug resistant tumor cell killing activity. Bioorganic & medicinal chemistry letters 2005, 15 (22), 5071-5.

5. Frenkel, V.; Etherington, A.; Greene, M.; Quijano, J.; Xie, J.; Hunter, F.; Dromi, S.; Li, K. C., Delivery of liposomal doxorubicin (Doxil) in a breast cancer tumor model: investigation of potential enhancement by pulsed-high intensity focused ultrasound exposure. Academic radiology 2006, 13 (4), 469-479.

6. Sapra, P.; Allen, T. M., Internalizing antibodies are necessary for improved therapeutic efficacy of antibody-targeted liposomal drugs. Cancer research 2002, 62 (24), 7190-4.

7. Shi, N. Q.; Gao, W.; Xiang, B.; Qi, X. R., Enhancing cellular uptake of activable cell-penetrating peptide-doxorubicin conjugate by enzymatic cleavage. International journal of nanomedicine 2012, 7, 1613-21.

8. Langer, R., Drug delivery and targeting. Nature 1998, 392 (6679 Suppl), 5-10.

9. Sneader, W., The discovery of aspirin: a reappraisal. BMJ 2000, 321 (7276), 1591-4.

10. Frederick, C. A.; Williams, L. D.; Ughetto, G.; van der Marel, G. A.; van Boom, J. H.; Rich, A.; Wang, A. H., Structural comparison of anticancer drug-DNA complexes: adriamycin and daunomycin. Biochemistry 1990, 29 (10), 2538-49.

11. González, I. D.; Saez, R. S.; Rodilla, E. M.; Yges, E.; Toledano, F., Hypersensitivity reactions to chemotherapy drugs. Alergol Immunol Clin 2000, 15, 161-181.

12. Craik, D. J.; Fairlie, D. P.; Liras, S.; Price, D., The future of peptide-based drugs. Chemical biology & drug design 2013, 81 (1), 136-47.

13. Quattrocchi, E.; Kourlas, H., Teriparatide: a review. Clinical therapeutics 2004, 26 (6), 841-54.

14. Kjeldsen, T., Yeast secretory expression of insulin precursors. Applied microbiology and biotechnology 2000, 54 (3), 277-86.

15. Couch, R. B., Seasonal inactivated influenza virus vaccines. Vaccine 2008, 26 Suppl 4, D5-9.

16. Neumann, G.; Fujii, K.; Kino, Y.; Kawaoka, Y., An improved reverse genetics system for influenza A virus generation and its implications for vaccine production. Proceedings of the National Academy of Sciences of the United States of America 2005, 102 (46), 16825-16829.

17. In Results of G004, a phase IIb actively controlled clinical trial with the TGF-b2 targeted compound AP 12009 for recurrent anaplastic astrocytoma, J Clin Oncol (Meeting Abstracts), 2006; p 1566.

18. Hughes, G. A., Nanostructure-mediated drug delivery. Disease-a-month : DM 2005, 51 (6), 342-61.

19. Liong, M.; Lu, J.; Kovochich, M.; Xia, T.; Ruehm, S. G.; Nel, A. E.; Tamanoi, F.; Zink, J. I., Multifunctional inorganic nanoparticles for imaging, targeting, and drug delivery. ACS nano 2008, 2 (5), 889-96.

20. Bagalkot, V.; Zhang, L.; Levy-Nissenbaum, E.; Jon, S.; Kantoff, P. W.; Langer, R.; Farokhzad, O. C., Quantum dot-aptamer conjugates for synchronous cancer imaging, therapy, and sensing of drug delivery based on bi-fluorescence resonance energy transfer. Nano letters 2007, 7 (10), 3065-70.

21. Cheng, Y.; A, C. S.; Meyers, J. D.; Panagopoulos, I.; Fei, B.; Burda, C., Highly efficient drug delivery with gold nanoparticle vectors for in vivo photodynamic therapy of cancer. Journal of the American Chemical Society 2008, 130 (32), 10643-7.

22. Namazi, H.; Adeli, M., Dendrimers of citric acid and poly (ethylene glycol) as the new drug-delivery agents. Biomaterials 2005, 26 (10), 1175-83.

23. Pasut, G.; Veronese, F., Polymer–drug conjugation, recent achievements and general strategies. Progress in Polymer Science 2007, 32 (8), 933-961.

24. Khandare, J.; Minko, T., Polymer–drug conjugates: progress in polymeric prodrugs. Progress in Polymer Science 2006, 31 (4), 359-397.

25. Bangham, A. D.; Horne, R. W., Negative Staining of Phospholipids and Their Structural Modification by Surface-Active Agents as Observed in the Electron Microscope. Journal of molecular biology 1964, 8, 660-8.

26. Torchilin, V. P., Recent advances with liposomes as pharmaceutical carriers. Nature Reviews Drug Discovery 2005, 4 (2), 145-160.

27. Greish, K., Enhanced permeability and retention (EPR) effect for anticancer nanomedicine drug targeting. Methods Mol Biol 2010, 624, 25-37.

28. Maruyama, K., Intracellular targeting delivery of liposomal drugs to solid tumors based on EPR effects. Advanced drug delivery reviews 2011, 63 (3), 161-9.

29. Juliano, R., Liposomes and the reticuloendothelial system: interactions of liposomes with macrophages and behavior of liposomes in vivo. In Targeting of drugs, Springer: 1982; pp 285-300.

30. Patankar, N.; Waterhouse, D., Nano-particulate Drug Delivery Systems for Camptothecins. Cancer Therapy 2012, 8, 90-104.

31. Hancock, R. E., Peptide antibiotics. Lancet 1997, 349 (9049), 418-22.

32. Zasloff, M., Magainins, a class of antimicrobial peptides from Xenopus skin: isolation, characterization of two active forms, and partial cDNA sequence of a precursor. Proceedings of the National Academy of Sciences of the United States of America 1987, 84 (15), 5449-53.

33. Frankel, A. D.; Pabo, C. O., Cellular uptake of the tat protein from human immunodeficiency virus. Cell 1988, 55 (6), 1189-93.

34. Derossi, D.; Joliot, A. H.; Chassaing, G.; Prochiantz, A., The third helix of the Antennapedia homeodomain translocates through biological membranes. The Journal of biological chemistry 1994, 269 (14), 10444-50.

35. Powers, J.-P. S.; Hancock, R. E., The relationship between peptide structure and antibacterial activity. Peptides 2003, 24 (11), 1681-1691.

36. Rozek, A.; Friedrich, C. L.; Hancock, R. E., Structure of the bovine antimicrobial peptide indolicidin bound to dodecylphosphocholine and sodium dodecyl sulfate micelles. Biochemistry 2000, 39 (51), 15765-74.

37. Tsai, C. W.; Hsu, N. Y.; Wang, C. H.; Lu, C. Y.; Chang, Y.; Tsai, H. H.; Ruaan, R. C., Coupling molecular dynamics simulations with experiments for the rational design of indolicidin-analogous antimicrobial peptides. Journal of molecular biology 2009, 392 (3), 837-54.

38. Storm, G., Liposomes as delivery system for doxorubicin in cancer chemotherapy. Pharmaceutisch weekblad. Scientific edition 1988, 10 (6), 288-90.

39. Janes, K. A.; Fresneau, M. P.; Marazuela, A.; Fabra, A.; Alonso, M. J., Chitosan nanoparticles as delivery systems for doxorubicin. Journal of controlled release : official journal of the Controlled Release Society 2001, 73 (2-3), 255-67.

40. Kovar, M.; Strohalm, J.; Etrych, T.; Ulbrich, K.; Rihova, B., Star structure of antibody-targeted HPMA copolymer-bound doxorubicin: a novel type of polymeric conjugate for targeted drug delivery with potent antitumor effect. Bioconjugate chemistry 2002, 13 (2), 206-15.

41. Ma, Y.; Manolache, S.; Denes, F. S.; Thamm, D. H.; Kurzman, I. D.; Vail, D. M., Plasma synthesis of carbon magnetic nanoparticles and immobilization of doxorubicin for targeted drug delivery. Journal of biomaterials science. Polymer edition 2004, 15 (8), 1033-49.

42. Bae, M.; Cho, S.; Song, J.; Lee, G. Y.; Kim, K.; Yang, J.; Cho, K.; Kim, S. Y.; Byun, Y., Metalloprotease-specific poly(ethylene glycol) methyl ether-peptide-doxorubicin conjugate for targeting anticancer drug delivery based on angiogenesis. Drugs under experimental and clinical research 2003, 29 (1), 15-23.

43. Working, P. K.; Dayan, A. D., Pharmacological-toxicological expert report. CAELYX. (Stealth liposomal doxorubicin HCl). Human & experimental toxicology 1996, 15 (9), 751-85.

44. Gabizon, A. A., Pegylated liposomal doxorubicin: metamorphosis of an old drug into a new form of chemotherapy. Cancer investigation 2001, 19 (4), 424-36.

45. Terada, T.; Iwai, M.; Kawakami, S.; Yamashita, F.; Hashida, M., Novel PEG-matrix metalloproteinase-2 cleavable peptide-lipid containing galactosylated liposomes for hepatocellular carcinoma-selective targeting. Journal of controlled release : official journal of the Controlled Release Society 2006, 111 (3), 333-42.

46. Chung, D. E.; Kratz, F., Development of a novel albumin-binding prodrug that is cleaved by urokinase-type-plasminogen activator (uPA). Bioorganic & medicinal chemistry letters 2006, 16 (19), 5157-63.

47. Veronese, F. M.; Pasut, G., PEGylation, successful approach to drug delivery. Drug discovery today 2005, 10 (21), 1451-8.

48. Katayama, S.; Hirose, H.; Takayama, K.; Nakase, I.; Futaki, S., Acylation of octaarginine: Implication to the use of intracellular delivery vectors. Journal of controlled release : official journal of the Controlled Release Society 2011, 149 (1), 29-35.

49. Turk, B. E.; Huang, L. L.; Piro, E. T.; Cantley, L. C., Determination of protease cleavage site motifs using mixture-based oriented peptide libraries. Nature biotechnology 2001, 19 (7), 661-7.

指導教授

阮若屈(Ruoh-Chyu Ruaan)

審核日期

2013-8-28

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