两种叶酸偶联物的合成及体外靶向性研究

张文典; 崔杰; 夏一帆; 张欣; 段少峰

doi:10.11665/j.issn.1000-5048.20210407

两种叶酸偶联物的合成及体外靶向性研究

1.
河南大学第一附属医院，开封 457001
2.
河南大学药学院，开封 457001

基金项目: 河南省医学科技攻关计划联合共建项目资助(No.2018020306)；开封市科技发展计划项目资助(No.1903024)

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出版历程
- 收稿日期: 2021-01-13
- 修回日期: 2021-05-10
- 刊出日期: 2021-08-24

Synthesis of two folate conjugates and their targeting effect in vitro

1.
The First Affiliated Hospital of Henan University, Kaifeng 475001
2.
School of Pharmacy, Henan University, Kaifeng 475001, China

Funds: This study was supported by the Joint Construction Project of Henan Medical Science and Technology Research Project (No.2018020306); and the Program of Kaifeng Science and Technology Development Project (No.1903024)

摘要

摘要: 合成叶酸偶联物并将其应用于叶酸靶向脂质体制备，考察其在体外肝癌HepG2细胞中的靶向性。通过酰胺反应将叶酸、胆固醇琥珀酸单酯与两种不同相对分子质量的聚氧乙烯二胺材料连接，合成两种两亲性的叶酸-聚乙二醇-胆固醇琥珀酸单酯（Folate-PEG₂₀₀₀-CHEMS和Folate-PEG₄₀₀₀-CHEMS），并利用核磁共振氢谱（¹H NMR）和超高分辨率复杂体分离鉴定质谱系统对其进行了结构表征。选取钙黄绿素为模型药物，采用薄膜分散法分别利用Folate-PEG₂₀₀₀-CHEMS和Folate-PEG₄₀₀₀-CHEMS制备了钙黄绿素脂质体FA-PEG₂₀₀₀-L与FA-PEG₄₀₀₀-L。利用激光粒度仪检测FA-PEG₂₀₀₀-L与FA-PEG₄₀₀₀-L的粒径、Zeta电位；利用流式细胞仪和激光共聚焦扫描显微镜，考察了脂质体FA-PEG₂₀₀₀-L与FA-PEG₄₀₀₀-L在HepG2细胞体外摄取实验中的药物递送效果。结果显示，钙黄绿素脂质体的平均粒径为（205.8 ± 10.2） nm，经电位测试脂质体的Zeta电位为-（1.19 ± 0.31） mV。FA-PEG₄₀₀₀-L靶向脂质体的荧光强度分别约为普通脂质体、FA-PEG₂₀₀₀-L的3.6和3.1倍（P < 0.01），FA-PEG₄₀₀₀-L在HepG2细胞中的递送效率明显高于FA-PEG₂₀₀₀-L与非靶向组，说明Folate-PEG₄₀₀₀-CHEMS可应用于脂质体制备，并可促进体外HepG2细胞对脂质体的摄取。
- 脂质体 /
- 叶酸 /
- 聚乙二醇 /
- 肝癌 /
- 细胞摄取 /
- 偶联物
Abstract: The aim of this study was to study the synthesis of two folate conjugates and their application in the preparation of folate targeted liposome, and to investigate their targeting effect in hepatocellular carcinoma HepG2 cell line in vitro. In this study, Folate-PEG-Cholesteryl hemisuccinate(Folate-PEG₂₀₀₀-CHEMS and Folate-PEG₄₀₀₀-CHEMS)were synthesized by linking folate and cholesterol succinate with two kinds of PEG materials. Structures of Folate-PEG₂₀₀₀-CHEMS and Folate-PEG₄₀₀₀-CHEMS were characterized by ¹H NMR and ultra-high resolution mass spectrometry. Calcein was selected as the model drug, and calcein liposomes FA-PEG₂₀₀₀-L and FA-PEG₄₀₀₀-L were prepared by film dispersion method using Folate-PEG₂₀₀₀-CHEMS and Folate-PEG₄₀₀₀-CHEMS, respectively. The particle size and Zeta potential of FA-PEG₂₀₀₀-L and FA-PEG₄₀₀₀-L were measured by laser particle size analyzer. The drug delivery effect of FA-PEG₂₀₀₀-L and FA-PEG₄₀₀₀-L was evaluated by cellular uptake experiment in HepG2 cell line in vitro. Flow cytometry and laser confocal scanning microscope were used to determine fluorescence in HepG2 cells in vitro. The results showed that the average particle size of calcein liposome was (205.8 ± 10.2) nm, and the Zeta potential of calcein liposome was -(1.19 ± 0.31) mV.There was no significant difference in particle size and Zeta potential between FA-PEG₂₀₀₀-L and FA-PEG₄₀₀₀-L. The fluorescence intensity of FA-PEG₄₀₀₀-L liposome group was about 3.6 and 3.1 times higher than that of non-targeted liposome group and FA-PEG₂₀₀₀-L liposome group, with statistically significant difference (P < 0.01). The drug delivery efficiency of FA-PEG₄₀₀₀-L group in HepG2 cells was higher than that in FA-PEG₂₀₀₀-L and non-targeted groups, and the results indicated that Folate-PEG₄₀₀₀-CHEMS can promote the uptake of liposome by HepG2 cells in vitro. All in all, Folate-PEG₄₀₀₀-CHEMS could be applied in the preparation of folate targeted liposome, which could promote the uptake of liposome by HepG2 cells.
- liposome /
- folate /
- PEG /
- hepatocellular carcinoma /
- cellular uptake experiment /
- conjugate

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