以穀胱甘肽敏感之阿黴素/糖基化牛血清白蛋白偶合奈米粒子作為乳腺癌標靶藥物遞送載體之研究

Glutathione-Sensitive Doxorubicin-Coupled Glycated Bovine Serum Albumin Nanoparticles as Targeted Drug Delivery Carriers for Breast Cancer

徐暐翔(Wei-Hsiang Hsu)

指導教授 ： 王勝仕

國立臺灣大學/工學院/化學工程學研究所/碩士(2021年)

https://doi.org/10.6342/NTU202101672

摘要

牛血清白蛋白(BSA)具有生物相容性、生物可降解性、無毒性、低成本和良好的水溶性等特性，再加上其具有多種官能基，如羥基、羧基和硫醇基等等，可用於交聯和藥物的偶聯，因此BSA被認為是一種有潛力的藥物載體。而在乳腺癌細胞系中，能發現一種過度表達的果糖轉運蛋白－GLUT5，此受體被證明能夠應用於乳腺癌的標靶藥物治療上。透過D-glucose的還原胺化反應，能對BSA進行修飾，在其表面上形成果糖胺的結構，藉此使BSA能被GLUT5識別。
阿黴素(doxorubicin, DOXO)是治療乳腺癌常見的化療藥物。在本研究中，首先製備了具有果糖胺結構的糖基化牛血清白蛋白(F-BSA)，然後使用6-maleimido-hexanoic acid將阿黴素改質，並透過maleimide-thiol reaction使阿黴素與F-BSA的硫醇基結合，形成穀胱甘肽敏感的maleimide-thiol group。然而，有證據顯示BSA分子太小而無法受惠於腫瘤組織的enhanced permeability and retention (EPR) effect，因此我們透過desolvation method將阿黴素/糖基化牛血清白蛋白偶合物(F-BSA-DOXO)進一步製成粒徑較大的奈米粒子(NPs)。
本研究所製備出的奈米粒子為球狀，平均粒徑為60.74 ± 18.38 nm，ζ-potential為-42.20 ± 5.75 mV，而loading capacity (LC%)則為3.57%。製備出的F-BSA-DOXO NPs是以傅立葉轉換紅外線光譜(FTIR)來分析化學結構；接著使用體外藥物釋放來研究F-BSA-DOXO NPs的藥物釋放行為；最後對MCF-7以及CHO-K1細胞進行MTT試驗以評估F-BSA-DOXO NPs的抗癌活性以及選擇性，並透過共軛焦雷射掃描顯微鏡來觀察細胞對藥物的攝取。本研究的結果表明，F-BSA-DOXO NPs為乳腺癌標靶藥物治療的潛力候選藥物。

關鍵字

牛血清白蛋白 ； 阿黴素 ； 乳腺癌 ； 奈米粒子 ； 生物材料

並列摘要

Bovine serum albumin (BSA) is considered a potential drug carrier due to its nontoxicity, biocompatibility, biodegradability in vivo, low cost, and good solubility in water. Also, BSA has various functional groups, such as hydroxyl groups, carboxylic acid groups, and thiol/sulfhydryl groups, which can be applied to cross-linkage and drug coupling. A type of fructose transporter, GLUT5, is overexpressed in breast cancer lines and has been shown to be a promising receptor for targeted drug delivery in breast cancer therapy. Upon glycation by the reductive amination of D-glucose, the fructosamine structure was formed on the glycated serum albumin (e.g., bovine serum albumin; BSA) which can be recognized by GLUT5.
Doxorubicin (DOXO) is known as a commonly used chemotherapy drug to treat breast cancer. In this study, the fructosamine structure-carrying glycated bovine serum albumin (F-BSA) was first prepared, followed by coupling with DOXO using 6-maleimidohexanoic acid to form the glutathione-sensitive maleimide-thiol group. Evidence indicates that the drug-encapsulated glycated proteins are inclined to be rapidly removed from tumors; therefore, efforts have been devoted to synthesizing the drug-encapsulated glycated protein nanoparticles via desolvation method to augment the enhanced permeability and retention (EPR) effect.
The nanoparticles were spherical with a mean size of 60.74 ± 18.38 nm (mean ± SD) and a ζ-potential of -42.20 ± 5.75 mV. The loading capacity was found to be 3.57%. Fructosamine assay was applied to the determination of the amount of fructosamine structures. The formation of synthesized F-BSA-DOXO nanoparticles (NPs) conjugates were verified by Fourier-transform infrared spectroscopy (FTIR). Next, the drug release behavior of the synthesized F-BSA-DOXO conjugates was further evaluated using the in vitro dialysis release method. The anticancer activity and selectivity of F-BSA-DOXO NPs were evaluated by performing MTT assay on MCF-7 cell lines and CHO-K1 cell lines. Finally, the cellular uptake was measure by confocal laser scanning microscopy (CLSM). The outcome from this study suggests that F-BSA-DOXO NPs serve as potential candidates for targeted drug delivery carriers used in breast cancer therapy.

並列關鍵字

bovine serum albumin ； doxorubicin ； breast cancer ； nanoparticles ； biomaterials

參考文獻

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