Radioiodination of Rhenium Cyclized α-Melanocyte-Stimulating Hormone Resulting in Enhanced Radioactivity Localization and Retention in Melanoma

Radiohalogenated α-melanocyte-stimulating hormone (α-MSH) analogs were proposed for melanoma imaging and potential radiotherapy because α-MSH receptors are overexpressed on both mouse and human melanoma cell lines. However, biodistribution studies in tumor-bearing mice with radiohalogenated α-MSH peptides showed very rapid tumor radioactivity wash out due to lysosomal degradation of the radiohalogenated complex after internalization, which decreased the therapeutic efficacy significantly (R. Stein et al., Cancer Res., 55: 3132–3139, 1995; P. K. Garg et al., Bioconjugate Chem., 6: 493–501, 1995.). The melanoma-targeting metallopeptide ReO[Cys3,4,10,d-Phe7]α-MSH3–13 (ReCCMSH) was shown to possess high tumor uptake and retention properties (J. Chen et al., Cancer Res., 60: 5649–5658, 2000). Therefore, three peptides, Ac-Lys-ReCCMSH(Arg11), Ac-d-Lys-ReCCMSH(Arg11), and [Nle4,d-Phe7]α-MSH (NDP) (for comparison), labeled with N-succinimidyl 4-[125I]iodobenzoate (125I-PIB), were prepared and evaluated in vitro and in vivo to develop radiohalogenated α-MSH peptide analogs with high tumor uptake, retention, and favorable biodistribution characteristics. In vitro cell binding and internalization data showed that approximately 90% of radioiodinated peptides were internalized at 2 h in cultured B16/F1 melanoma cells. Cellular retention studies showed that the receptor-bound radioiodinated linear α-MSH analog NDP was released from the cells into the medium very quickly, whereas significant amounts of cell-associated radioactivity remained in the cells for Ac-Lys(125I-3- or 4-iodobenzoate (IBA))-ReCCMSH(Arg11) and Ac-d-Lys(125I-IBA)-ReCCMSH(Arg11). The in vitro data clearly demonstrate that rhenium cyclization significantly enhanced peptide trapping in the cells, as did d-amino acid incorporation. The combination of these two effects resulted in a 2.9-fold increase in the retention of radioactivity for Ac-d-Lys(125I-IBA)-ReCCMSH(Arg11) relative to 125I-IBA-NDP at 4 h. In vivo studies also showed that Ac-d-Lys(125I-IBA)-ReCCMSH(Arg11) exhibited extremely high radioactivity accumulation and prolonged retention in the tumor. Ac-d-Lys(125I-IBA)-ReCCMSH(Arg11) and Ac-Lys(125I-IBA)-ReCCMSH(Arg11) exhibited much higher tumor uptake at 24 h after injection compared with 125I-IBA-NDP [7.18% injected dose/gram (ID/g), 4.92% ID/g, and 0.26% ID/g, respectively]. Ac-d-Lys(125I-IBA)-ReCCMSH(Arg11) also showed very fast whole body clearance and low nonspecific radioactivity accumulation in normal tissues compared with 125I-IBA-NDP and Ac-Lys(125I-IBA)-ReCCMSH(Arg11). A tumor:blood ratio of 34.3 was observed for Ac-d-Lys(125I-IBA)-ReCCMSH(Arg11) at 24 h postinjection, whereas values of 4.3 and 2.0 were observed for Ac-Lys(125I-IBA)-ReCCMSH(Arg11) and 125I-IBA-NDP, respectively. The biodistribution data clearly demonstrate that both rhenium cyclization and d-Lys incorporation enhanced the tumor localization and retention of the radiolabel. Therefore Ac-d-Lys-ReCCMSH(Arg11) is an excellent candidate for additional therapeutic studies.