利用噬菌体肽库筛选高转移潜能人肝癌细胞结合肽

目的筛选高转移潜能人肝癌细胞特异性结合肽。方法以高转移潜能人肝癌细胞系HCCLM3作为靶细胞，低转移潜能人肝癌细胞系MHCC97L为吸附细胞对噬菌体随机7肽库进行差减筛选，采用噬菌体结合实验和抗噬菌体免疫细胞化学染色对阳性克隆的特异性进行鉴定。结果经3轮筛选，随机挑选48个噬菌体克隆进行DNA序列分析，展示AWYPLPP肽的噬菌体被高度富集77％（37／48）；噬菌体结合实验显示，与低转移潜能人肝癌细胞系MHCC97L、PLC／PRE／5和无转移潜能入肝癌细胞系Hep3B以及正常入肝细胞系CCLl3相比，AWYPLPP噬菌体特异性与高转移潜能人肝癌细胞系HCCLM3、HCCLM6、MHCC97H和MHCC97结合（P〈0．01）；抗噬菌体免疫细胞化学染色证实AWYPLPP噬菌体特异性靶向高转移潜能人肝癌细胞系HCCLM3、HCCLM6、MHCC97H和MHCC97。结论从噬菌体肽库中获得与高转移潜能人肝癌细胞特异性结合肽，可作为肝癌转移复发靶向治疗研究的载体。

Screening high metastatic potential human hepatocellular carcinoma cell-binding peptides from a phage-displayed peptide library

To screen peptides specifically binding to high metastatic potential human hepatocellular carcinoma (HCC) cells from a phage-displayed peptide library. Methods Two human HCC cell lines with high and low metastatic potentials, HCCLM3 and MHCC97L, were used as the target cells for subtractive biopanning from a 7-mer peptide phage-displayed library. The specificity of selected phage clone binding to high metastatic potential human HCC cells was identified by phage binding assay and immunocytochemistry studies. Results After 3 rounds of biopanning, DNA sequencing of 48 finally selected phage revealed that 77% of them encoded an identical peptide AWYPLPP (37/48). Phage binding assay showed that the AWYPLPP phage preferentially bound to several high metastatic potential HCC cell lines (HCCLM3, HCCLM6, MHCC97H, and MHCC97) as compared with low metastatic potential HCC cell lines (MHCC97L and PLC/PRE/5), no metastatic potential HCC cell line Hep3B, and normal human liver cell lineCCL13 (P< 0.01). Immunocytochemistry demonstrated that the AWYPLPP phage was able to specifically target high metastatic potential HCC cell lines (HCCLM3, HC-CLM6, MHCC97H, and MHCC97). Conclusion A novel peptide specifically binding to high metastatic potential human HCC cells was obtained successfully from a phage-displayed peptide library. It might help design new peptide-targeted therapy for metastatic recurrence of HCC.