Radiolabeling approaches for cholecystokinin B receptor imaging

Regulatory peptides and their analogs are being extensively investigated as radiopharmaceuticals for cancer imaging. In particular, cholecystokinin (CCK) receptors of the, subtype B (CCK-BR) have been shown to be overexpressed in certain neuroendocrine tumors including medullary thyroid cancer. Our recent work, has focused on new methods to radiolabel the CCK8 peptide with In-111 or Tc-99m for the purpose of developing for in vivo CCK-B receptor imaging, Labeling of CCK8 with In-111 was achieved at the N-terminus of the peptide by adding, in solid phase, a glutamate coupled diethylenetriaminepentaacetic acid (DTPA) moiety through a glycine linker, yielding DTPA-Glu-G-CCK8. For labeling with Tc-99m, the, CCK8 peptide was modified at its N-terminus by introducing, in the following order-cysteine, glycine, and a diphenylphosphinopropionyl moiety-giving a 10-residue peptide derivative, Phos-GC-CCK8. A cell culture model was developed for the purpose of evaluating the binding properties of these two ligands. The human epidermoid carcinoma cell line, A431, was transfected with a plasmid containing the full coding sequence of the human CCK-BR under a strong viral promoter, obtaining a number of receptors in the range of 2-5 x 10(6) per cell. Control cells were transfected with vector alone. An animal tumor model utilizing these two cell lines was developed to evaluate the specificity of interaction with the CCK-BR and biodistribution properties of the compounds, CCK-BR positive and control cells were subcutaneously injected in opposite flanks of CD1 female nude mice in order to obtain xenografts differing only in their ability to express CCK-B receptors. High performance liquid chromatography (HPLC) and other chromatographic methods were utilized to assess stability of the radiolabeled compounds after injection. Both In-111-DTPA-Glu-G-CCK8 and Tc-99m-Phos-GC-CCK8 showed similar binding affinities for cultured CCK-BR expressing cells, with dissociation constants in the range of 20-40 nM. With the two xenograft approach. we were able to demonstrate specific interaction with the receptor of both CCK analogs in our animal model. The deta obtained shows rapid specific, localization of both compounds on the CCK-BR overexpressing xenografts. Both tracers show rapid plasma clearance of unbound peptide. Clearance of In-111-DTPA-Glu-G-CCK8 appears to be preferentially through the kidneys, whereas Tc-99m-Phos-GC-CCK8 clearance occurs both through kidneys and the hepatobiliary system. Both our labeling approaches appear adequate for clinical use of peptide based radiopharmaceuticals, although Tc-99m-Phos-GC-CCK8 shows elevated accumulation in the gastrointestinal tract, which causes high background activity. (C) 2003 Wiley Periodicals, Inc.