Abstract
Galectin-3 (Gal-3) is a β-galactosidase binding protein that modulates various cellular processes including cell adhesion, and metastasis. We evaluated the tumor targeting and imaging properties of a galectin-3 binding peptide originally selected from bacteriophage display, in a mouse model of human breast carcinoma expressing galectin-3. A galectin-3 binding peptide, ANTPCGPYTHDCPVKR, was synthesized with a Gly-Ser-Gly (GSG) spacer and 1,4,7,10, tetraazacyclododecane-N,N’,N’’,N’’’-tetracetic acid (DOTA) or 4,11-bis(carboxymethyl)-1,4,8,11 tetrazabicyclo[6.6.2]hexadecane 4,11-diacetic acid (CB-TE2A), and radiolabeled with 64Cu. The synthesized peptides 64Cu-DO3A-(GSG)-ANTPCGPYTHDCPVKR (64Cu-DO3A- pep) and 64Cu-CB-TE2A-(GSG)-ANTPCGPYTHDCPVKR(64Cu-CB-TE2A-pep) demonstrated an IC50 value of 97 ± 6.7 nM and 130 ± 10.2 nM, respectively, to cultured MDA-MB-435 breast carcinoma cells in vitro in a competitive displacement binding study. The tumor tissue uptake in SCID mice bearing MDA-MB-435 tumors was 1.2 ± 0.18 %ID/g (64Cu-DO3A-pep) and 0.85 ± 0.0.9 %ID/g (64Cu-CB-TE2A-pep) at 30 min, respectively. While liver retention was moderate with both radiolabeled peptides the kidney retention was observed to be high. Radiation dose delivered to the tumor was estimated to be 42 mGy/mCi and 129 mGy/ mCi with CB-TE2A and DO3A peptides, respectively. Imaging studies demonstrated tumor uptake with both 64Cu-DO3A- and 64Cu-CB-TE2A-(GSG)-ANTPCGPYTHDCPVKR after 2 h post injection. These studies suggest that gal-3 binding peptide could be developed into a PET imaging agent for galectin-3-expressing breast tumors.
Highlights
Galectin-3 (Gal-3) is a 31 kDa protein, which has high affinity for β-galactosides, and possesses a highly conserved carbohydrate recognition domain (CRD) [1]
The peptides were labeled with 64Cu in ammonium acetate buffer at 85 ̊C for 1 h, and further purified by Reverse phase high pressure liquid chromatography (RP-HPLC) in order to separate the radiolabeled peptides from their non-radiolabeled counterparts. 64Cu-DO3A- and 64Cu-CB-TE2A-peptide eluted with a retention time of 13.2 min and 13.8 min, respectively, a minute after the elution of their respective non-radiolabeled counterparts
MDA-MB-435 human breast carcinoma cells were originally derived from a pleural effusion of a woman with metastatic ductal breast carcinoma [27]
Summary
Galectin-3 (Gal-3) is a 31 kDa protein, which has high affinity for β-galactosides, and possesses a highly conserved carbohydrate recognition domain (CRD) [1]. Metastatic breast carcinoma cells express higher levels of gal-3 and have significantly increased adhesion to monolayers of endothelial cells compared with their non-metastatic counterparts [4]. Previous studies from our laboratory demonstrated that gal-3 is involved in heterotypic (carcinoma-endothelial cells) and homotypic (carcinoma-carcinoma) cellular adhesion via interactions with the tumor-specific Thomsen-Friedenreich (TF) mucin-type disaccharide, Gal 1-3GalNAc, expressed on most human adenocarcinomas [11]. Carbohydrate-based inhibitors that block gal-3-TF interactions [13,14], via binding to the CRD of gal-3 have previously been reported. Such inhibitors bound other galectin family members due to their conserved carbohydrate binding residues which
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