Melanoma-targeting Properties of Technetium-labeled Cyclic a-Melanocyte- stimulating Hormone Peptide Analogues

Preliminary reports have demonstrated that technetium (Tc)-labeled cyclic [Cys, D-Phe]a-MSH3–13 (CCMSH) exhibits high tumor uptake and retention values in a murine melanoma mouse model. In this report, the tumor targeting mechanism of Tc-CCMSH was studied and compared with four other radiolabeled a-melanocyte stimulating hormone (a-MSH) peptide analogues: I-(Tyr)-[Nle, D-Phe]a-MSH [I(Tyr)-NDP]; Tc-CGCG-NDP; Tc-Gly-CCMSH; and TcNle-CCMSH. In vitro receptor binding, internalization, and cellular retention of radiolabeled a-MSH analogues in B16/F1 murine cell line demonstrated that >70% of the receptor-bound radiolabeled analogues were internalized together with the receptor. Ninety % of the internalized I-(Tyr)-NDP, whereas only 36% of internalized Tc-CCMSH, was released from the cells into the medium during a 4-h incubation at 37°C. Two mouse models, C57 mice and severe combined immunodeficient (Scid) mice, inoculated s.c. with B16/F1 murine and TXM-13 human melanoma cells were used for the in vivo studies. Tumor uptake values of 11.32 and 2.39 [% injected dose (ID)/g] for Tc-CCMSH at 4 h after injection, resulted in an uptake ratio of tumor:blood of 39.0 and 11.5 in murine melanoma-C57 and human melanoma-Scid mouse models, respectively. Two strategies for decreasing the nonspecific kidney uptake of Tc-CCMSH, substitution of Lys in CCMSH with Gly or Nle, and lysine coinjection, were evaluated. The biodistribution data for the modified peptides showed that Lys replacement dramatically decreased the kidney uptake, whereas the tumor uptakes of Tc-Nleand TcGly-CCMSH were significantly lower than that of Tc-CCMSH. Lysine coinjection significantly decreased the kidney uptake (e.g., from 14.6% ID/g to 4.5% ID/g at 4 h after injection in murine melanoma-C57 mice) without significantly changing the value of tumor uptake of TcCCMSH. In conclusion, the compact cyclic structure of Tc-CCMSH, its resistance to degradation, and its enhanced intracellular retention are the major contributing factors to the superior in vivo tumor targeting properties of Tc-CCMSH. Lys residue in Tc-CCMSH is critical to the tumor targeting in vivo, and lysine coinjection rather than lysine replacement can significantly decrease the nonspecific renal radioactivity accumulation without impeding the high melanoma-targeting properties of Tc-CCMSH. The metal-cyclized CCMSH molecule displays excellent potential for the development of melanoma-specific diagnostic and therapeutic agents.