Interaction of an alpha-melanocyte-stimulating hormone-diphtheria toxin fusion protein with melanotropin receptors in human melanoma metastases.
A hybrid toxin targeted to melanotropin receptors and selectively cytotoxic to melanoma cell lines in vitro has recently been developed. The toxin, a recombinant fusion protein (designated DAB389-MSH), contains the peptide sequences of alpha-melanocyte-stimulating hormone (alpha-MSH) and the catalytic (cytotoxic; Fragment A) and lipophilic (part of Fragment B) domains of diphtheria toxin. In the present study, binding of DAB389-MSH to melanotropin receptors in biopsy specimens of human and mouse melanoma metastases was assessed by measuring its ability to inhibit binding of a radiolabeled, superpotent analogue of alpha-MSH (125I-[Nle4,D-Phe7]-alpha-MSH; 125I-NDP-MSH) and comparing its potency in this system with those of the established ligands NDP-MSH and alpha-MSH. Radioligand binding to tissue sections in vitro was localized and quantified by autoradiography and image analysis. DAB389-MSH inhibited binding of 125I-NDP-MSH to experimental murine B16-F1C23 melanoma metastasis tissue and to melanoma metastases of three patients. In both mouse and human melanoma tissues, concentration-response relationships for DAB389-MSH-mediated inhibition of 125I-NDP-MSH binding were parallel, and its maximal effects were comparable in magnitude, to those of NDP-MSH and alpha-MSH. Half-maximal peptide concentrations for inhibition of 125I-NDP-MSH binding to mouse melanoma tissue sections were: NDP-MSH, 0.63 nM; alpha-MSH, 3.14 nM; and DAB389-MSH, 10.1 nM. In human melanoma tissues, the respective half-maximal peptide concentrations for inhibition of 125I-NDP-MSH binding to mouse melanoma tissue sections were: NDP-MSH, 1.80 nM; alpha-MSH, 2.43 nM; and DAB389-MSH, 11.9 nM. Taken together, these results suggest that NDP-MSH, alpha-MSH, and DAB389-MSH bind to a common melanotropin receptor in human metastatic melanoma cells. Since previous work has shown that melanotropin receptors are detectable in melanoma metastases of about 80% of human patients, malignant melanoma cells of many patients may be susceptible to killing by the melanotropin receptor-targeted cytotoxin DAB389-MSH.
Tatro
J B
JB
Division of Endocrinology, Diabetes, Metabolism and Molecular Medicine, Tufts University School of Medicine, Boston, Massachusetts.
Wen
Z
Z
Entwistle
M L
ML
Atkins
M B
MB
Smith
T J
TJ
Reichlin
S
S
Murphy
J R
JR
D000818:Animals; D004167:Diphtheria Toxin / Q000378:metabolism*; D004354:Drug Screening Assays, Antitumor; D006801:Humans; D008297:Male; D008545:Melanoma / Q000378:metabolism* / Q000556:secondary*; D008546:Melanoma, Experimental / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D011979:Receptors, Pituitary Hormone / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000378:metabolism*; D000521:alpha-MSH / Q000378:metabolism*
D004167:Diphtheria Toxin; D011979:Receptors, Pituitary Hormone; D011993:Recombinant Fusion Proteins; C069655:diphtheria toxin-related-alpha-melanocyte-stimulating hormone fusion toxin; D000521:alpha-MSH; C029446:MSH receptor