以聚乙烯亞胺修飾聚醣體作為非病毒型基因載體之評估

Polyethylene imine modified polysaccharide as a non-viral gene carrier

彭郁翔(Yu-Shiang Peng)

指導教授 ： 朱一民

國立清華大學/工學院/化學工程學系/博士(2014年)

摘要

基因治療在現今許多遺傳疾病及癌症的治療上展現了新的契機，因此基因載體在基因治療中扮演不可或缺的角色，而理想的基因載體須具備低細胞毒性並且維持高效率的轉染效能。近年來，以陽離子高分子作為基因載體逐漸受到重視，但此系統在高轉染效能的同時，也伴隨著高細胞毒性的缺點。因此本研究利用陽離子高分子聚乙烯亞胺(polyethylene imine, PEI)分別修飾聚醣類高分子如幾丁聚醣(chitosan, CHI)和硫酸化軟骨素(chondroitin sulfate, CS)，而設計出兩種共聚物，並進行基因轉染測試的探討。
在PEI 修飾CHI的研究中，我們分別以分子量600 和1800 g/mol 的PEI 以ethylene glycol diglycidyl ether (EX-810)進行開環反應接枝到CHI，在溶解度測試中，PEI顯著提升了CHI的水溶性，而體外測試(in vitro)中，我們發現經由PEI (Mw=600 g/mol) 修飾後的CHI明顯提升了基因轉染的效能。
在第二階段的研究，由於PEI造成細胞毒性的主要原因是具有大量正電荷，因此，我們將PEI接枝到帶有陰離子結構的CS，PEI 接枝CS (PEI-g-CS)同樣具有良好的DNA 結合能力，在體外測試中，也大幅改善細胞毒性的缺失，並維持其優異的基因表現。

關鍵字

幾丁聚醣 ； 硫酸化軟骨素 ； 基因載體 ； 聚乙烯亞胺

並列摘要

The purpose of this study was to design a gene vector for gene therapy. Chitosan (CHI) is a naturally derived material that has been used for gene transfer. However, its low water solubility often leads to decreased transfection efficiency. Grafting of highly water-soluble polyethylene imine (PEI) and polyethylene glycol (PEG) onto polymers can increase their solubility. This study reports a non-viral gene carrier with improved water solubility as well as enhanced transfection efficiency. Two molecular weights (Mw= 600 and 1,800 g/mol) of PEI were grafted onto CHI (PEI600-g-CHI and PEI1.8K-g-CHI, respectively) by opening the epoxide ring of ethylene glycol diglycidyl ether (EX-810). The transfection efficiency of PEI600-g-CHI/deoxyribonucleic acid (DNA) polyplexes was significantly higher than either PEI1.8K-g-CHI/DNA or CHI/DNA polyplexes.
Polyethylene imine (PEI) is commonly used for gene transfer. However, the high cytotoxicity often leads to decreased transfection efficiency. Solutions to this problem have been widely discussed, one of which being the grafting of highly biocompatible polymer such as chondroitin sulfate (CS). In this study, different molecular weight (1.8K and 25K Da) of PEI was grafted onto CS (PEI1.8K-g-CS and PEI25K-g-CS) through ethyl(dimethylaminopropyl) carbodiimide (EDC)/N-Hydroxysuccinimide (NHS) reaction. This modification resulted in a non-viral gene carrier with improved biocompatibility. The transfection efficiency of BMSCs using PEI25K-g-CS/DNA polyplexes was significantly higher than PEI25K/DNA polyplexes.

並列關鍵字

chitosan ； chondroitin sulfate ； gene carrier ； polyethylene imine

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