In vitro panning of a targeting peptide to hepatocarcinoma from a phage display peptide library q Bing Du a , Min Qian a,b, * , Zhongliang Zhou a , Peng Wang a , Lei Wang a , Xiaoping Zhang a , Miao Wu a , Ping Zhang a , Bing Mei a a School of Life Sciences, East China Normal University, Shanghai, PR China b Immunology Division, E-institutes of Shanghai University (EISU), China Received 12 January 2006 Available online 20 February 2006 Abstract Phage display technology has been used as a powerful tool in the discovery of ligands specific to receptor(s) on the surface of a cancer cell and could also impact clinical issues including functional diagnosis and cell-specific drug delivery. After three rounds of in vitro pan- ning and two rounds of reverse absorption, a group of phages capable of addressing BEL-7402 enormously were obtained for further analysis. Through a cell-based ELISA, immunofluorescence, FACS, and in vivo binding study, WP05 (sequence TACHQHVRMVRP) was demonstrated to be the most effective peptide in targeting four kinds of liver cancer cell lines (BEL-7402, BEL-7404, SMMC-7721, and HepG2), but not the normal liver cell line HL-7702. In conclusion, the peptide WP05 which was screened by in vitro phage display technology was proved to be a targeting peptide to several common hepatocellular carcinoma cell lines. Ó 2006 Elsevier Inc. All rights reserved. Keywords: Phage display library; In vitro; Hepatocellular carcinoma; Targeting peptides Hepatocarcinoma is one of the most challenging malig- nancies with a very high mortality [12]. To date, no preop- erative treatment has been proven useful [6]. The major drawback of current cytotoxics and gene therapy vectors is their lack of selectivity [14,16]. Thus, the targeting ther- apy has become a potential effective choice in the curing of hepatocarcinoma. However, a major complication of this approach is delivering sufficient amount of these ther- apeutics to the tumor tissues. Traditional targeting mole- cules mostly made up of monoclonal antibody or other ligands, so the specifically binding ability has become noth- ing the matter. But the immunogenicity to human body and the penetration to tumor tissues still cannot be treated ideally. In principle, the small size of peptides makes them attractive for tumor-targeting applications [11]. So, more and more scientists began to give their attention to low molecular weight protein and peptides [5]. Phage display was originally described in 1985 by Smith [15] who presented the use of the non-lytic filamentous bac- teriophage fd for the display of specific binding peptides on the phage coat. Phage display is a molecular diversity tech- nology that allows the presentation of a large number of peptides or proteins on the surface of filamentous phage for various applications. These libraries permit the selec- tion of peptides or proteins with high affinity and specificity for almost any target. The most important feature of this technology is the direct link between the experimental phe- notype and its encapsulated genotype [3]. Since 1985 [15], phage display has been an important tool for both basic 0006-291X/$ - see front matter Ó 2006 Elsevier Inc. All rights reserved. doi:10.1016/j.bbrc.2006.02.050 q We thank Wei Shi for expert assistance with FACS analyses, Quanzhi Lu, for assistance and access to fluorescence microscopy and Microplate Reader, and Yunbo Gao, Hongyan Tan, and Lisha Kuang for zealous help. This work was supported by Shanghai Scientific Development Foundation (04JC14033), National Natural Science Foundation (30440039), the ‘‘Shu Guang’’ project of Shanghai Municipal Education Commission and Shanghai Education Development Foundation (02SG21), and the Special Fund of Nanotechnology of Shanghai Scientific Development Foundation (0352nm113). * Corresponding author. Fax: +86 021 62233754. E-mail address: mqian@bio.ecnu.edu.cn (M. Qian). www.elsevier.com/locate/ybbrc Biochemical and Biophysical Research Communications 342 (2006) 956–962 BBRC