多柔比星负载 AS1411 适体对结直肠癌细胞的抗增殖作用。
Anti-Proliferative Effect of Doxorubicin-Loaded AS1411 Aptamer on Colorectal Cancer Cell.
机构信息
Division of Biochemistry, Department of Preclinical Science, Faculty of Medicine, Thammasat University, Pathumthani, 12120, Thailand.
Department of Chemistry, Faculty of Science and Technology, Thammasat University, Pathumthani, 12120, Thailand.
出版信息
Asian Pac J Cancer Prev. 2021 Jul 1;22(7):2209-219. doi: 10.31557/APJCP.2021.22.7.2209.
BACKGROUND
Doxorubicin (Dox) inhibits DNA replication and causes DNA damage resulting in cell death. It is a common drug for treatment of many cancers. Treatment efficacy and side effects of Dox are critical issues in using it because the drug lacks of specificity. The objective of this study was to improve the specificity of Dox by the incorporation of this drug with AS1411 aptamer (ASA).
METHODS
Dox was intercalated into the duplex sites of ASA, a recognition molecule for a number of cancer cells, and formed Dox-loaded ASA. The recognition ability proceeded through specific binding between the aptamer and nucleolin overexpressed in the cancer cells. The tested cells were human colorectal adenocarcinoma cell line (SW480) and human normal colon cell CCD841 CoN (CCD841). Binding of ASA to the cells was tested using flow cytometer and fluorescence microscope. Intercalation of Dox into DNA duplex was confirmed by fluorescence spectrometry. Effect of ASA, Dox, and Dox-loaded ASA on cell viability was examined by cell proliferation assay. Caspase-3 activation was analyzed by western blotting.
RESULTS
ASA bound specifically to SW480 cells via interaction between the aptamer and nucleolin because the nucleolin was highly expressed in SW480 cells. ASA decreased the viability of SW480 cells in a dose-dependent manner. Dox was more toxic than ASA. Fluorescence quenching revealed that Dox was able to intercalate in base pairing sites of the aptamer. Dox-loaded ASA inhibited the proliferation of SW480 cells, because the aptamer facilitated the Dox uptake into these cells which caused the cell apoptosis, indicated by the significant decrease in procaspase-3, apoptosis marker protein.
CONCLUSION
This study succeeded to prepare Dox-loaded ASA by intercalation of the drug that inherited the binding function from the aptamer and anti-cancer activity from Dox. Dox-loaded ASA showed promise for effective cancer treatment with lower level of side effects.
背景
多柔比星（多柔比星）抑制 DNA 复制并导致 DNA 损伤，从而导致细胞死亡。它是治疗许多癌症的常用药物。由于药物缺乏特异性，因此使用多柔比星时，其治疗效果和副作用是关键问题。本研究的目的是通过将该药物与 AS1411 适体（ASA）结合来提高多柔比星的特异性。
方法
将多柔比星插入 ASA 的双链体位点中，ASA 是许多癌细胞的识别分子，并形成负载多柔比星的 ASA。该测试细胞是人结肠直肠腺癌细胞系（SW480）和人正常结肠细胞 CCD841 CoN（CCD841）。通过流式细胞仪和荧光显微镜测试 ASA 与细胞的结合。通过荧光光谱法证实了多柔比星插入 DNA 双链体中。通过细胞增殖测定法检查 ASA、多柔比星和负载多柔比星的 ASA 对细胞活力的影响。通过蛋白质印迹法分析 caspase-3 激活。
结果
ASA 通过适体与核仁蛋白之间的相互作用特异性地与 SW480 细胞结合，因为核仁蛋白在 SW480 细胞中高度表达。ASA 以剂量依赖性方式降低 SW480 细胞的活力。多柔比星比 ASA 更有毒。荧光猝灭表明，多柔比星能够插入适体的碱基配对位点中。负载多柔比星的 ASA 抑制了 SW480 细胞的增殖，因为适体促进了多柔比星进入这些细胞，导致细胞凋亡，表明 procaspase-3，凋亡标志物蛋白显着减少。
结论
本研究通过插入药物成功制备了负载多柔比星的 ASA，该药物继承了适体的结合功能和多柔比星的抗癌活性。负载多柔比星的 ASA 有望具有较低副作用的有效癌症治疗。
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