Development of Human Hepatocellular Carcinoma Cell-Targeted Protein Cages
Development of Human Hepatocellular Carcinoma Cell-Targeted Protein Cages
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DOI：
10.1021/bc300015f
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发表时间：
2012-07-01
影响因子：
4.7
通讯作者：
Hashizume, Makoto
中科院分区：
文献类型：
--
作者：
Toita, Riki;Murata, Masaharu;Hashizume, Makoto
我们在本文中描述了一种人肝细胞癌（HCC）细胞靶向蛋白笼，其中称为SP 94的HCC结合肽在天然存在的热休克蛋白（Hsp）笼的表面进行了修饰。使用具有不同长度的两种类型的异双功能接头（SM（PEG）（n））和两种类型的SP 94化学合成六种类型的HCC靶向Hsp笼。肽，其在肽的N-或C-末端含有独特的Cys残基。使用基质辅助激光解吸/电离飞行时间质谱（MALDI-ToF MS）分析、十二烷基硫酸钠-聚丙烯酰胺凝胶电泳（SDS-PAGE）分析和动态光散射（DLS）测量来表征这些Hsp笼。荧光显微镜观察显示，所有工程化蛋白笼选择性地结合HCC细胞，但不结合测试的其它细胞系（包括正常肝细胞）。此外，Hsp笼上SP 94肽的数量、SP 94肽的缀合位点以及Hsp笼和SP 94肽之间的接头长度对工程化Hsp笼与HCC细胞的结合具有重要影响。通过较长的接头分子与SP 94肽的N-末端缀合并含有高SP 94肽水平的工程化Hsp笼显示出对HCC细胞的更大结合。令人惊讶的是，通过优化这三个因子，实现了对HCC细胞高达10倍的亲和力。这些结果不仅对于使用SP 94肽开发HCC细胞靶向装置至关重要，而且对于创建利用其他肽配体的其他细胞靶向材料也至关重要。
We described herein a human hepatocellular carcinoma (HCC) cell-targeted protein cage for which the HCC-binding peptide termed SP94 was modified at the surface of a naturally occurred heat shock protein (Hsp) cage. Six types of HCC-targeted Hsp cages were chemically synthesized using two types of heterobifunctional linker (SM(PEG)(n)) with different lengths and two types of SP94. peptide, which contained a unique Cys residue at the N- or C-terminus of the peptide. These Hsp cages were characterized using matrix assisted laser desorption/ionization-time-of-flight mass spectrometry (MALDI-ToF MS) analyses, sodium dodecylsulfate-polyacrylamide gel electrophoresis (SDS-PAGE) analyses, and dynamic light scattering (DLS) measurement Fluorescence microscopic observations revealed that all the engineered protein cages bind selectively to HCC cells but not to the other cell lines tested (including normal liver cell). Moreover, the number of SP94 peptides on Hsp cages, conjugation site of SP94 peptide, and linker length between a Hsp cage and a SP94 peptide had important effects upon the binding of engineered Hsp cages to HCC cells. An engineered Hsp cage conjugated to the N-terminus of SP94 peptide via a longer linker molecule and containing high SP94 peptide levels showed greater binding toward HCC cells. Surprisingly, through optimization of these three factors, up to 10 fold greater affinity toward HCC cells was achieved. These results are critically important not only for the development of HCC cell targeting devices using SP94 peptide, but also to create other cell-targeting materials that utilize other peptide ligands.