Screening and identification of a novel target specific for hepatoma cell line HepG2 from the FliTrx bacterial peptide library
Wenhan Li
Department of Immunology, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, China
Institute of Immunology, University of Heidelberg, 69120 Heidelberg, Germany
These authors contributed equally to this work
Search for more papers by this authorPing Lei
Department of Immunology, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, China
These authors contributed equally to this work
Search for more papers by this authorBing Yu
Department of Immunology, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, China
Search for more papers by this authorSha Wu
Department of Immunology, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, China
Search for more papers by this authorJilin Peng
Department of Immunology, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, China
Search for more papers by this authorXiaoping Zhao
Department of Immunology, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, China
Search for more papers by this authorHuifen Zhu
Department of Immunology, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, China
Search for more papers by this authorMichael Kirschfink
Institute of Immunology, University of Heidelberg, 69120 Heidelberg, Germany
Search for more papers by this authorCorresponding Author
Guanxin Shen
Department of Immunology, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, China
*Corresponding author: Tel, 86-27-83692611; E-mail, [email protected]Search for more papers by this authorWenhan Li
Department of Immunology, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, China
Institute of Immunology, University of Heidelberg, 69120 Heidelberg, Germany
These authors contributed equally to this work
Search for more papers by this authorPing Lei
Department of Immunology, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, China
These authors contributed equally to this work
Search for more papers by this authorBing Yu
Department of Immunology, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, China
Search for more papers by this authorSha Wu
Department of Immunology, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, China
Search for more papers by this authorJilin Peng
Department of Immunology, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, China
Search for more papers by this authorXiaoping Zhao
Department of Immunology, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, China
Search for more papers by this authorHuifen Zhu
Department of Immunology, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, China
Search for more papers by this authorMichael Kirschfink
Institute of Immunology, University of Heidelberg, 69120 Heidelberg, Germany
Search for more papers by this authorCorresponding Author
Guanxin Shen
Department of Immunology, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, China
*Corresponding author: Tel, 86-27-83692611; E-mail, [email protected]Search for more papers by this authorThis work was supported by the grants from the Hi-tech Research and Development Program of China (No. 2006AA02Z158) and the Science Foundation of the Ministry of Education of China (No. 20060487024)
Abstract
To explore new targets for hepatoma research, we used a surface display library to screen novel tumor cell-specific peptides. The bacterial FliTrx system was screened with living normal liver cell line L02 and hepatoma cell line HepG2 successively to search for hepatoma-specific peptides. Three clones (Hep1, Hep2, and Hep3) were identified to be specific to HepG2 compared with L02 and other cancer cell lines. Three-dimensional structural prediction proved that peptides inserted into the active site of Escherichia coli thioredoxin (TrxA) formed certain loop structures protruding out of the surface. Western blot analysis showed that FliC/TrxA-peptide fusion proteins could be directly used to detect HepG2 cells. Three different FliC/TrxA-peptide fusion proteins targeted the same molecule, at approximately 140 kDa, on HepG2 cells. This work presented for the first time the application of the FliTrx library in screening living cells. Three peptides were obtained that could be potential candidates for targeted liver cancer therapy.
References
- 1 Bosch FX, Ribes J, Díaz M, Cléries R. Primary liver cancer: worldwide incidence and trends. Gastroenterology 2004, 127: S5–S16.
- 2 Ferlay J, Bray F, Pisani P, Parkin DM. GLOBOCAN 2000: Cancer Incidence, Mortality and Prevalence Worldwide. Version 1.0. Lyon : IARC Press 2001.
- 3 El-Houseini ME, Mohammed SM, Wael ME, Tarek DH, Omar SD, Anwar AE. Enhanced detection of hepato-cellular carcinoma. Cancer Control 2005, 12: 248–253.
- 4 Zhang B, Zhang Y, Wang J, Zhang Y, Chen J, Pan Y, Ren L et al. Screening and identification of a targeting peptide to hepatocarcinoma from a phage display peptide library. Mol Med 2007, 13: 246–254.
- 5 Pan B, Li B, Russell SJ, Tom JY, Cochran AG, Fairbrother WJ. Solution structure of a phage-derived peptide antagonist in complex with vascular endothelial growth factor. J Mol Biol 2002, 316: 769–787.
- 6 Sun LY, Chu TW, Wang Y, Wang XY. Radiolabeling and biodistribution of a nasopharyngeal carcinoma-targeting peptide identified by in vivo phage display. Acta Biochim Biophys Sin 2007, 39: 624–632.
- 7 Lu Z, Murray KS, Van Cleave V, LaVallie ER, Stahl ML, McCoy JM. Expression of thioredoxin random peptide libraries on the Escherichia coli cell surface as functional fusions to flagellin: a system designed for exploring protein-protein interactions. Biotechnology 1995, 13: 366–372.
- 8 Lu Z, LaVallie ER, McCoy JM. Using bio-panning of FLITRX peptide libraries displayed on E. coli cell surface to study protein-protein interactions. Methods Mol Biol 2003, 205: 267–280.
- 9 Tripp BC, Lu Z, Bourque K, Sookdeo H, McCoy JM. Investigation of the “switch-epitope” concept with random peptide libraries displayed as thioredoxin loop fusions. Protein Eng 2001, 14: 367–377.
- 10 Craik DJ, Simonsen S, Daly NL The cyclotides: novel macrocyclic peptides as scaffolds in drug design. Curr Opin Drug Discov Devel 2002, 251–260.
- 11 Li P, Roller PP. Cyclization strategies in peptide derived drug design. Curr Top Med Chem 2002, 2: 325–341.
- 12 Ibrahim GF, Fleet GH, Lyons MJ, Walker RA. Method for the isolation of highly purified Salmonella flagellins. J Clin Microbiol 1985, 22: 1040–1044.
- 13 Nilsson F, Tarli L, Viti F, Neri D. The use of phage display for the development of tumor targeting agents. Adv Drug Deliv Rev 2000, 43: 165–196.
- 14 Bussolati B, Grange C, Tei L, Deregibus MC, Ercolani M, Aime S, Camussi G. Targeting of human renal tumor-derived endothelial cells with peptides obtained by phage display. J Mol Med 2007, 85: 897–906.
- 15 Javitt NB. HepG2 cells as a resource for metabolic studies: lipoprotein, cholesterol, and bile acids. FASEB J 1990, 4: 161–168.
- 16 She YM, Narindrasorasak S, Yang S, Spitale N, Roberts EA, Sarkar B. Identification of metal-binding proteins in human hepatoma lines by immobilized metal affinity chromatography and mass spectrometry. Mol Cell Proteomics 2003, 2: 1306–1318.
- 17 Pasqualini R, Ruoslahti E. Organ targeting in vivo using phage display peptide libraries. Nature 1996, 380: 364–366.
- 18 Zurita AJ, Troncoso P, Cardó-Vila M, Logothetis CJ, Pasqualini R, Arap W. Combinatorial screenings in patients: the interleukin-11 receptor α as a candidate target in the progression of human prostate cancer. Cancer Res 2004, 64: 435–439.
- 19 Rasmussen UB, Schreiber V, Schultz H, Mischler F, Schughart K. Tumor cell-targeting by phage-displayed peptides. Cancer Gene Ther 2002, 9: 606–612.
- 20 Dane KY, Chan LA, Rice JJ, Daugherty PS. Isolation of cell specific peptide ligands using fluorescent bacterial display libraries. J Immunol Methods 2006, 309: 120–129.
- 21 Ladner RC. Polypeptides from phage display. A superior source of in vivo imaging agents. Q J Nucl Med 1999, 43: 119–124.
- 22 Park JW, Hong K, Kirpotin DB, Colbern G, Shalaby R, Bseiga J, Shao Y et al. Anti-Her2 immunoliposomes: enhanced efficacy attributable to targeted delivery. Clin Cancer Res 2002, 8: 1172–1181.
- 23 Zitzmann S, Krämer S, Mier W, Mahmut M, Fleig J, Altmann A, Eisenhut M et al. Identification of a new prostate-specific cyclic peptide with the bacterial FliTrx system. J Nucl Med 2005, 46: 782–785.
- 24 Barry MA, Dower WJ, Johnston SA. Toward cell-targeting gene therapy vectors: selection of cell-binding peptides from random peptide-presenting phage libraries. Nat Med 1996, 2: 299–305.
- 25 Zhang J, Spring H, Schwab M. Neuroblastoma tumor cell-binding peptides identified through random peptide phage display. Cancer Lett 2001, 171: 153–164.
- 26 Giordano RJ, Cardo-Vila M, Lahdenranta J, Pasqualini R, Arap W. Biopanning and rapid analysis of selective interactive ligands. Nat Med 2001, 7: 1249–1253.
- 27 Romanov VI, Durand DB, Petrenko VA. Phage display selection of peptides that affect prostate carcinoma cells attachment and invasion. Prostate 2001, 47: 239–251.
- 28 Binetruy-Tournaire R, Demangel C, Malavaud B, Vassy R, Rouyre S, Kraemer M. Identification of a peptide blocking vascular endothelial growth factor (VEGF)-mediated angiogenesis. EMBO J 2000, 19: 1525–1533.
- 29 Folgori A, Tafi R, Meola A, Felici F, Galfré G, Cortese R, Monaci P et al. A general strategy to identify mimotopes of pathological antigens using only random peptide libraries and human sera. EMBO J 1994, 13: 2236–2243.
- 30 Daugherty PS, Olsen MJ, Iverson BL, Georgiou G. Development of an optimized expression system for the screening of antibody libraries displayed on the Escherichia coli surface. Protein Eng 1999, 12: 613–621.
- 31 Van Regenmortel MH. Antigenicity and immunogenicity of synthetic peptides. Biologicals 2001, 29: 209–213.
- 32 Geysen HM, Mason TJ, Rodda SJ. Cognitive features of continuous antigenic determinants. J Mol Recognit 1988, 1: 32–41.
- 33 Deshayes K, Schaffer ML, Skelton NJ, Nakamura GR, Kadkhodayan S, Sidhu SS. Rapid identification of small binding motifs with high-throughput phage display: discovery of peptidic antagonists of IGF-1 function. Chem Biol 2002, 9: 495–505.
