臺灣博碩士論文加值系統

近年來生物性高分子被廣泛的使用在藥物傳導的領域上，因為它們具有好的生物相容性以及能夠減少藥物副作用，並且增加藥物的傳導效率，像是soluble polymer conjugates、micelles以及polymer-protein等都是，而其中polymer-drug conjugate也是被廣為討論的一種方式，因為polymer-drug conjugate必須藉由內吞作用(endocytosis)被輸送到細胞內，也因此控制了藥物在人體內的分布。Polymer-drug conjugate主要包含了一段用來作為主幹的水溶性高分子、一段生物可分解的支鏈以用來連接藥物並促進藥物在腫瘤組織中釋放，以及一段生物不可分解的支鏈以用來增加藥物專一性。而本篇研究的目的著重在methacryloxy succinimide(MAOS)的合成並且期望它能被應用在化學治療的領域上。我們經由核磁共振(NMR)以及紅外線光譜儀(IR)來分析MAOS單體的化學結構，以及藉由凝膠滲透層析(gel permeation chromatography)來分析聚合物的分子量以及分子量分布(Polydispersity)。首先，我們使用dicyclohexylcarbodiimide(DCC)作為MAOS單體合成的試劑，其產率可達到65%；因為聚合物的分子量對於它在人體內傳遞的過程有顯著的影響，因此我們選用了原子轉移自由基聚合法(Atom Trasfer Radical Polymerization)來聚合MAOS單體，以期望得到低分子量分布的聚合物，並且嘗試使用不同的觸媒來探討它對聚合的影響；此外，我們還使用了自由基聚合法(Free Radical Polymerization)來聚合MAOS單體，並且與原子轉移自由基聚合法作為對照。而MAOS高分子與五倍當量的1-amino-2-propanol反應可以得到一類稱為N-(2-hydroxypropyl)methacrylamide(HPMA)的水溶性高分子，而這一類高分子目前被廣泛的使用在藥物傳導上，並且正在進行臨床實驗第二期的測試，藉由這種方式，我們期望得到具有低分子量分布的HPMA高分子。在我們的研究當中，我們還成功的合成出了一個新型不可被生物分解的支鏈，稱為N-(4-aminobutyl)maleimide以用來增進polymer-drug conjugate對腫瘤細胞的專一性；我們並選用了thiophenol來作為支鏈對抗體鍵結能力的測試，並且我們成功的將新型支鏈鍵結到高分子上。最後，在我們的研究當中，我們成功的合成了MAOS單體，並利用原子轉移自由基聚合法聚合MAOS單體，以得到低分子量分布的MAOS高分子；此外，我們還成功合成出用來與抗體作鍵結的新型支鏈，並且成功的將支鏈接上了MAOS高分子。在未來，我們期望MAOS高分子能夠成為一個多功能性的高分子，並在化學治療上扮演一個重要的角色。

Polymers with intrinsic antitumour properties, soluble polymer conjugates, and micelles and polymer-protein adducts have been known for many years. These compounds are considered as new chemical entities. Conjugation of drugs to these polymeric carriers can restrict cellular uptake of the drug to the endocytic route and significantly change the biodistribution of conjugated drug. The polymer drug conjugate consists of a water soluble polymer as carrier, biodegradable linkers to facilitate controlled release of drug at the tumor site and non-biodegradable linkers to facilitate receptor mediated targeting. In this study, we attempt to address the formation of methacryloxy succinimide (MAOS) for chemotherapy. The chemical structure of MAOS monomer and polymers were identified by NMR and IR and the molecular weight of MAOS polymer was analyzed by gel permeation chromatography (GPC). The MAOS monomer was synthesized by dicyclohexylcarbodiimide (DCC) approach and the yield was to 65%. The MAOS polymerization was processed by Atom Transfer Radical Polymerization (ATRP) and the influence of different reaction condition was studied. The free radical polymerization approach was used to examine the molecular weight distribution of poly(MAOS) compared to ATRP. The N-(2-hydroxypropyl)methacrylamide (HPMA), a water-soluble polymer used in polymeric therapy, was converted from poly(MAOS) by reacting with 5 equiv of 1-amino-2-propanol. By this method, we expected to obtain a low polydispersity index (PDI) poly(HPMA). A novel non-biodegradable linker, N-(4-aminobutyl) maleimide, for receptor mediate targeting was synthesized in our study. The binding activity of N-(4-aminobutyl) maleimide to antibodies was tested by reacting with thiophenol. Finally, N-(4-aminobutyl) maleimide was attached to poly(MAOS) with the ratio approximately 5%. In conclusion, MAOS synthesis, MAOS polymerization, analysis of MAOS polymer, synthesis of novel non-biodegradable linker, and attachment of linker to MAOS polymer were successfully studied. In the future, we expect poly(MAOS) could be a key pre-polymer used in chemotherapy.

Contents

1. Introduction…………………………………………………………1
1-1 Cancer…………………………………………………………….…1
1-2 Treatment of cancer……………………………………........1
1-3 Drug delivery systems…………………………………………..3
1-3.1 Delivery of free drugs…………………………………...…3
1-3.2 Macromolecules as drug carriers………………………..…4
1-4 EPR effect…………………………………………………….……4
1-5 Polymer-drug conjugate………………………………………...5
1-5.1 Introduction of polymer-drug conjugate……….…….….5
1-5.2 Structure of polymer-drug conjugate……………….…...7
1-5.2.1 Polymer backbone..............................…...8
1-5.2.2 Solublizing residue…………………..…………….….…9
1-5.2.3 Targeting moiety.............. ..............…...10
1-5.2.4 Spacer………………………………………..............11
1-5.2.5 Drug………………………………………........…….….14
1-6 Project motive and aim……………………………...……….15

2. Mechanism…………………....………………...……………….18
2-1 Methacryloxy succinimide (MAOS)……………………...…..18
2-2 Poly (N-methacryloxysuccinimide) (PMAOS)………………..20
2-3 N-(2-hydroxypropyl)methacrylamide (HPMA)………...…….24
2-3.1 Design of HPMA copolymers……………………….…………25
2-3.2 Biocompatibility and immunocompatibility of HPMA…..29
2-4 Atom transfer radical polymerization (ATRP)…………...31

3. Experiment and Instrument……………….…….……………..35
3-1 Materials………………………………...………………………35
3-1.1 Experiment reagent……………………………………………35
3-1.2 Experiment solvent……………………………………………36
3-2 Equipment………………………………………………….……..37
3-2.1 Nuclear Magnetic Resonance, NMR…………………..…….37
3-2.2 Fourier Transform Infrared Spectrometer, FT-IR……..37
3-2.3 Gel Permeation Chromatography, GPC………...………….37
3-3 Experiment procedure…………………………..………………38
3-3.1 Synthesis of MAOS monomer…………………………..…….38
3-3.2 Preparation of poly(MAOS) …………….……….…………39
3-3.3 Synthesis of (4-aminobutyl)carbamic acid tert-
butylester………………..............................40
3-3.4 Synthesis of N-(4-tert-butoxycarbonyl-aminobutyl)
maleamic acid……...................................41
3-3.5 Synthesis of N-(4-tert-butoxycarbonyl-aminobutyl)
maleimide……...................................……41
3-3.6 The cleavage of N-(4-tert-butoxycarbonyl-aminobutyl)
maleimide…….......................................43
3-3.7 Preparation of poly(HPMA)…………………...…………..44
3-3.8 Reaction of N-(4-tert-butoxycarbonyl-aminobutyl)
maleimide with thiophenol………………………………...44
3-3.9 Preparation of MAOS copolymer with 5%
aminomaleimide.…...................................45
3-3.10 Reaction of MAOS copolymer with 5% aminomaleimide
with thiophenol………………………………………………46

4. Results and discussion………………………………..……...47
4-1 Synthesis and characterization of MAOS monomer………..47
4-1.1 Synthesis of MAOS monomer…………………..…………...47
4-1.2 Structure identification by NMR……………….…………47
4-1.3 Structure identification by FT-IR…………..………...48
4-2 Preparation and characterization of poly (MAOS)……….49
4-2.1 Preparation of poly (MAOS)…………………………………49
4-2.2 Structure identification by NMR………………………….50
4-2.3 Structure identification by FT-IR……………………….52
4-2.4 Molecular weight measurement by GPC…………………….53
4-3 Preparation and characterization of poly (HPMA)……….53
4-3.1 Preparation of poly (HPMA)…………………………………53
4-3.2 Structure identification by NMR………………………….56
4-3.3 Structure identification by FT-IR……………………….57
4-3.4 Molecular weight measurement by GPC…………………….58
4-4 Synthesis and characterization of N-(4-aminobutyl)
maleimide………….....................................59
4-4.1 Synthesis of (4-aminobutyl)carbamic acid tert-butyl
ester………………...................................59
4-4.2 Structure identification by NMR………………………...59
4-4.3 Synthesis of N-(4-tert-butoxycarbonyl-aminobutyl)
maleamic acid……...................................61
4-4.4 Structure identification by NMR………………………….61
4-4.5 Synthesis of N-(4-tert-butoxycarbonyl-aminobutyl)
maleimide……...…..................................63
4-4.6 Structure identification by NMR………………………….63
4-4.7 Preparation of N-(4-aminobutyl) maleimide…………….64
4-4.8 Structure identification by NMR………………………….65
4-4.9 Reaction of N-(4-tert-butoxycarbonyl-aminobutyl)
maleimide with thiophenol………………………………….66
4-4.10 Structure identification by NMR…………………………67
4-5 Preparation and characterization of 5% aminomaleimide and MAOS copolymer……………………………………………………68
4-5.1 Preparation of MAOS copolymer with 5%
aminomaleimide……………............................68
4-5.2 Structure identification by NMR………………………….68
4-5.3 Reaction of MAOS copolymer with 5% aminomaleimide with thiophenol……………………………………………………….70
4-5.4 Structure identification by NMR………………………….70

5. Conclusion and future work………………………………….…72

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