In vitro panning of a targeting peptide to hepatocarcinoma from a phage display peptide library
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 panning 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.
Available from: PubMed Central
- "Triplicate determinations were done at each data point. Selectivity was determined using a formula as follows : Selectivity = ODM13 - ODC1/ODS2 - ODC2. Here, ODM13 and OD C1 represent the OD values from the selected phages and control phages binding to A498 cells, respectively. "
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ABSTRACT: Specific peptide ligands to cell surface receptors have been extensively used in tumor research and clinical applications. Phage display technology is a powerful tool for the isolation of cell-specific peptide ligands. To screen and identify novel markers for renal cell carcinoma, we evaluated a peptide that had been identified by phage display technology.
A renal carcinoma cell line A498 and a normal renal cell line HK-2 were used to carry out subtractive screening in vitro with a phage display peptide library. After three rounds of panning, there was an obvious enrichment for the phages specifically binding to the A498 cells, and the output/input ratio of phages increased about 100 fold. A group of peptides capable of binding specifically to the renal carcinoma cells were obtained, and the affinity of these peptides to the targeting cells and tissues was studied.
Through a cell-based ELISA, immunocytochemical staining, immunohistochemical staining, and immunofluorescence, the Phage ZT-2 and synthetic peptide ZT-2 were shown to specifically bind to the tumor cell surfaces of A498 and incision specimens, but not to normal renal tissue samples.
A peptide ZT-2, which binds specifically to the renal carcinoma cell line A498 was selected from phage display peptide libraries. Therefore, it provides a potential tool for early diagnosis of renal carcinoma or targeted drug delivery in chemotherapy.
Journal of Experimental & Clinical Cancer Research 11/2011; 30(1):105. DOI:10.1186/1756-9966-30-105 · 4.43 Impact Factor
Available from: ibp.ac.cn
- "Protein display technologies such as phage, bacteria, and yeast display have been developed for isolating antibodies from large libraries of recombinant antibodies and have become major tools for generating monoclonal antibodies for research and the clinic (Hoogenboom 2005). Phage display antibody libraries can be readily screened for binding to antigens presented on whole cells by " panning " (Hoogenboom et al. 1991; Du et al. 2006). On the other hand, yeast and bacterial surface displays have been mainly employed to screen antibodies for soluble antigens (Boder et al. 2000; Daugherty et al. 1999; Boder and Wittrup 1997; Georgiou et al. 1997). "
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ABSTRACT: Anchored periplasmic expression (APEx) is a method for isolating high affinity ligand-binding proteins from large combinatorial libraries, and antibodies highly specific for soluble antigens were successfully isolated from APEx antibody libraries in combination with flow cytometric sorting (Harvey et al., Proc Natl Acad Sci USA 101(25):9193-9198, 2004). However, many disease markers and drug targets are localized on the cell surface, and often, unique posttranslational modifications and/or properly folded epitopes are lost when they were expressed and isolated in soluble form. In this study, we demonstrate that Escherichia coli spheroplasts, displaying antibodies and screened by a combination of plate-panning and flow cytometric sorting, can be used for isolating antibodies specific for antigens on the human cell surface. Two rounds of plate-panning followed by one round of flow cytometric sorting resulted in 7,200-fold enrichment of antibodies specific for the protective antigen of Bacillus anthracis from a large excess of spheroplasts expressing a scFv antibody to digoxin (a negative control). There is the potential to use this technique for library screening to find novel antibodies against disease cell surface antigens.
Applied Microbiology and Biotechnology 09/2010; 88(6):1385-91. DOI:10.1007/s00253-010-2861-3 · 3.34 Impact Factor
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ABSTRACT: ObjectiveTo identify and localize the synthesized targeting peptide A54 to liver cancer cell line BEL-7502 in vivo and in vitro for confirming the potential clinical application of peptide A54 in hepatocarcinoma targeting therapy.
MethodsPhage A54 was confirmed by ELISA. Biotin and FAM labeled A54 peptides were identified and localized by means of immunohistochemistry
ResultsA54 peptide could target the liver-tumor tissue in vivo and adhere to several liver-tumor cells in vitro. FAM-labeled A54 peptides were localized on the membrane surface of liver-tumor cells.
ConclusionSynthesized A54 peptide obtained from in vivo phage display technology still kept special ability to adhere liver-tumor cell in vivo and in vitro. The A54 peptide could be a candidate carrier for hepatocarcinoma targeting therapy.
Chinese Journal of Cancer Research 11/2006; 18(4):241-245. DOI:10.1007/s11670-006-0241-4 · 1.94 Impact Factor
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