64Cu-Labeled Peptide for PET of Breast Carcinomas Expressing the Thomsen-Friedenreich Carbohydrate Antigen

UNLABELLED: Thomsen-Friedenreich (TF) antigen is a disaccharide, galactose β1-3 N-acetylgalactosamine (Galβ1-3GalNAc), expressed on the cell surfaces of most human carcinomas including breast. In this study, we synthesized and evaluated the in vitro and in vivo properties of a (64)Cu-radiolabeled TF antigen-specific peptide derived from bacteriophage display for the purpose of breast tumor targeting and PET of human breast tumors in xenografted mice. METHODS: The TF antigen-specific peptide IVWHRWYAWSPASRI was synthesized with the chelator 1,4,7-triazacyclononane-1,4,7-triacetic acid (NOTA) at the amino terminus, followed by a Gly-Ser-Gly (GSG) spacer. Amino acids Asp and Arg were introduced at both ends to enhance its solubility. Purified NO2A-GSG-DRD-IVWHRWYAWSPASRI-DRD (NO2A-TFpep) was radiolabeled with (64)Cu and evaluated for binding to human MDA-MB-435 breast cancer cells, 50% inhibitory concentration (IC(50)), and serum stability. In vivo pharmacokinetic and small-animal PET studies were performed using SCID mice bearing MDA-MB-435 tumor xenografts. RESULTS: (64)Cu-NO2A-TFpep bound to human MDA-MB-435 breast carcinoma cells, whereas almost no binding was observed to normal human breast 184A1 cells. The peptide exhibited an apparent IC(50) value of 70 ± 8.0 nM. In vivo biodistribution studies indicated radiolabeled peptide accumulation in tumors of MDA-MB-435 xenografted SCID mice of approximately 1.10 ± 0.20 percentage injected dose per gram (%ID/g) and 0.90 ± 0.12 %ID/g, at 0.5 and 1 h, respectively. Accumulation of radioactivity was low in other organs, with the exception of liver (1.52 ± 0.12 %ID/g) and kidneys (15.4 ± 1.73 %ID/g) at 1 h. Live imaging studies with (64)Cu-NO2A-TFpep (15 MBq) demonstrated good tumor uptake at 1 h after injection, whereas no tumor uptake was observed with a scrambled radiolabeled peptide (64)Cu-NO2A-GSG-DRD-RWSWWAVHRIPYSAI-DRD. CONCLUSION: (64)Cu-NO2A-TFpep may function as a noninvasive in vivo tumor imaging agent of human breast and other carcinomas expressing the TF carbohydrate antigen. This is the first such TF antigen-targeting peptide used in tumor imaging.