Identification of a high affinity TAG-72 binding peptide by phage display selection
Authors
Xiao, NanCheng, Dengfeng
Wang, Yi
Chen, Ling
Liu, Xinrong
Dou, Shuping
Liu, Guozheng
Liang, Min Min
Hnatowich, Donald J.
Rusckowski, Mary
UMass Chan Affiliations
Department of Medicine, Divison of Hematology/OncologyDepartment of Radiology
Document Type
Journal ArticlePublication Date
2011-01-01Keywords
Amino Acid SequenceAnimals
Antigens, Neoplasm
Cell Line, Tumor
Colonic Neoplasms
Glycoproteins
HT29 Cells
Humans
Male
Mice
Mice, Nude
Molecular Sequence Data
Peptide Library
Peptides
Protein Binding
Technetium
Tissue Distribution
Tomography, Emission-Computed, Single-Photon
Xenograft Model Antitumor Assays
phage display
TAG-72 antigen
peptide
colon cancer tumor cell LS-174T
Cancer Biology
Neoplasms
Oncology
Radiology
Therapeutics
Metadata
Show full item recordAbstract
PURPOSE: Phage display was used to select novel peptides that specifically bind the TAG-72 antigen and with properties suitable for imaging TAG-72 positive cancers. RESULTS: After three rounds of selection against TAG-72 and using two different elution conditions including a long elution, the consensus sequences FRERCDKHPQKCTKFL and DPRHCQKRVLPCPAWL were expressed on phages G3-15 and T3-15 respectively. ELISA, fluorescence-activated cell sorting analysis and fluorescence microscopy provided evidence that both phages specifically bound TAG-72 in vitro. Both peptides are stable in 37oC serum. By a cell binding competition assay, the IC50 for T3-15 was measured as 0.29 nM and therefore 36-fold higher affinity than G3-15 at 10.32 nM. The biodistribution in mice carrying LS-174T tumors in one thigh were similar for both 99mTc-peptides at 30 min, but at 90 min the 99mTc-T3-15 peptide accumulated almost three times higher in the tumor. The SPECT/CT images were consistent with the biodistribution results. PROCEDURES: The f88-4/Cys6 phage library and two different elution conditions were used to identify two new higher affinity binding peptides for the TAG-72 antigen. One, was a single brief elution with pH 2.2 glycine buffer, and the second began with the glycine elution but was followed with a longer elution with Tris buffered saline (TBS) at pH 7.4. The phages that bound TAG-72 were evaluated by fluorescence-activated cell sorting analysis using TAG-72 positive LS-174T cells and confirmed by immunofluorescence imaging. The consensus peptides displayed on the selected phages were synthesized and conjugated with NHS-MAG3 for radiolabeling with 99mTc. The IC50 for TAG-72 binding was evaluated by cell binding competition in vitro while binding affinity was evaluated in vivo by necropsy and SPECT/CT imaging in a tumor mouse model. CONCLUSION: We have identified a peptide with a sub nanomolar inhibition constant for the TAG-72 antigen that may have application in cancer imaging.Source
Cancer Biol Ther. 2011 Jan 1;11(1):22-31. DOI: 10.4161/cbt.11.1.13797. Link to article on publisher's websiteDOI
10.4161/cbt.11.1.13797Permanent Link to this Item
http://hdl.handle.net/20.500.14038/48378PubMed ID
20980835Related Resources
Link to Article in PubMedae974a485f413a2113503eed53cd6c53
10.4161/cbt.11.1.13797