開發奈米載體應用於化學治療合併基因治療對抗藥性細胞之評估

Abstract

為了實現合併基因治療與化學治療在對付多重抗藥性癌細胞上，我們開發可同時輸送基因與化療藥物的奈米載體。藉由將mPEG-PCL共聚高分子接枝在PEI高分子上，合成出mPEG-PCL-PEI三團聯高分子。我們使用核磁共振儀與凝膠滲透層析儀分析所合成高分子聚合物，並使用pyrene螢光物質來研究合成的高分子聚合物形成奈米粒子其臨界微胞濃度(CMC)。mPEG-PCL-PEI三團聯高分子形成微胞後使用動態雷射光散射儀(DLS)與穿透式電子顯微鏡(TEM) 來觀察奈米粒子大小、表面電位、形態，發現包覆太平洋紫杉醇的大小為226 nm，與DNA混合後的奈米粒子為238 nm。我們進一步利用流式細胞儀分析mPEG-PCL-PEI三團聯高分子微胞的基因轉殖效率,結果發現在轉殖綠色螢光蛋白基因方面，在抗藥性細胞MCF-7/ADR細胞株轉殖效果會比單獨PEI 25K好。在實驗中，我們轉殖 MDR-1 小干擾RNA 能夠抑制 50% P-glycoprotein 表現。我們並使用MCF-7wt 與 MCF-7/ADR 細胞株進行雙功能微胞毒殺效果測試。由這些結果顯示出所合成mPEG-PCL-PEI三團聯高分子載體同時具有基因治療與化學治療的潛力。

To realize gene therapy and chemotherapy in multi-drug-resistant cancer cell, we develop carriers which can co-delivery gene and chemotherapy drug. The mPEG-PCL-PEI (M510i) tri-block polymers were synthesized by use mMPEG-PCL copolymer modified to PEI. The characteristic of these tri-block polymers were evaluated by 1H nuclear magnetic resonance and gel permeation chromatography. The critical micelle concentration (CMC) of micelle was evaluated by using pyrene as fluorescence probe. The particle size, zeta potential, and morphology of micelle was studied by dynamic light scattering and transmission electron microscopy. The results indicate that the paclitaxel loaded micelles and DNA complexes with micelles were 226 nm and 238 nm. The gene transfection efficiency was evaluated by used flow cytometry to evaluate green fluorescence protein (GFP) expression. The gene transfection efficiency performed better than PEI 25K in MCF-7 ADR cell. In siRNA experiments, we can transfect MDR-1 siRNA to silence P-glycoprotein expression 50%. In viro cytotoxicity, dual agent micelle of were tested of MCF-7 wt and MCF-7 ADR by MTT assy. These results suggested the PEG-PCL-PEI tri-block polymer as potential carriers for gene therapy and chemotherapy.