α-MSH Receptor Autoradiography on Mouse and Human Melanoma Tissue Sections and Biopsies

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Abstract
MSH receptors and their binding characteristics of [125I]-labelled derivatives of alpha-MSH have been studied extensively on various mouse and human melanoma cell lines in culture. The aim of this study was to determine the binding characteristics of alpha-MSH radioligands to MSH receptors occurring in experimental mouse and human melanoma tumours as well as in human melanoma biopsies. For this reason, solid tumours were grown on experimental animals by inoculation of murine B16-F1 and human D10 and HBL melanoma cells. After excision and cryosectioning of the tumours, frozen tissue sections were incubated with [(125I)Tyr2]-alpha-MSH or [(125I)Tyr2,Nle4,D-Phe7]-alpha-MSH and specific alpha-MSH binding sites were visualized by subsequent autoradiography. The presence of increasing concentrations of unlabelled alpha-MSH during incubation with tracer led to a dose-dependent displacement of the radioligand. Quantitative analysis of the autoradiograms produced dissociation constants which were comparable with those obtained with cell binding assays: KD = 1.87 and 1.31 nmol/l for B16 tumours and cells, respectively; 0.32 and 0.33 nmol/l for D10, and 2.24 and 1.36 nmol/l for HBL tumours and cells, respectively. This indicates similar binding properties of alpha-MSH radioligands to both cultured melanoma cells and tissue sections of melanoma tumours from experimental animals. Similar binding characteristics were also observed with human melanoma tissue sections originating from biopsies of melanoma patients.