title: Coexistence of novel amylin-binding sites with calcitonin receptors in human breast carcinoma MCF-7 cells.
abstract: Amylin, calcitonin (CT) and calcitonin gene-related peptide (CGRP) share limited structural homology including amino-terminal ring structures linked by a disulfide bridge and amidated carboxy-termini. Here, we have compared [125I]Bolton-Hunter-[Lys1] rat amylin ([125I]amylin) binding and the stimulation of cyclic AMP accumulation by human (h) amylin, hCT and hCGRP-I in the human breast carcinoma cell lines MCF-7 and T47D, which predominantly express hCT1a and hCT1b receptor isoforms (hCTR1a, hCTR1b) at a similar total number of hCT-binding sites. In MCF-7 cells, half-maximal inhibition (IC50) of [125I]amylin binding by human amylin was observed at 3.6 +/- 0.8 nM (n = 6). hCT and hCGRP-I displaced [125I]amylin binding with 22 and 66 times higher IC50. [125I]hCT binding was inhibited by hCT with an IC50 of 8.1 +/- 1.9 nM (n = 5), and human amylin and hCGRP-I were over 100 times less potent. In T47D cells, on the other hand, specific binding of [125I]amylin was not observed, but hCT inhibited [125I]hCT binding with an IC50 of 3.2 +/- 0.4 nM (n = 3), and human amylin and hCGRP-I had over 200 times higher IC50. In MCF-7 cells, half-maximal stimulation (EC50) of cyclic AMP accumulation by human amylin, hCT and hCGRP-I occurred at 1.4 +/- 0.2, 1.7 +/- 0.4 and 6.3 +/- 1.3 nM respectively. In T47D cells, the EC50 of hCT was 0.32 +/- 0.02 nM (n = 3), and 30- and 1900-fold higher with human amylin and hCGRP-I. In conclusion, the expression of hCTR1a and hCTR1b and [125I]hCT binding were indistinguishable in MCF-7 and T47D cells. Yet, [125I]amylin binding was only recognized in MCF-7 cells, consistent with a distinct amylin receptor.
authors.affiliation: Research Laboratory for Calcium Metabolism, Department of Orthopaedic Surgery, Zurich, Switzerland.
authors.forename: U
authors.initials: U
authors.lastname: Zimmermann
authors.forename: B
authors.initials: B
authors.lastname: Fluehmann
authors.forename: W
authors.initials: W
authors.lastname: Born
authors.forename: J A
authors.initials: JA
authors.lastname: Fischer
authors.forename: R
authors.initials: R
authors.lastname: Muff
mesh_terms: D000682:Amyloid / Q000378:metabolism / Q000494:pharmacology*; D001665:Binding Sites; D001667:Binding, Competitive; D001943:Breast Neoplasms / Q000378:metabolism*; D015740:Calcitonin Gene-Related Peptide / Q000494:pharmacology; D000242:Cyclic AMP / Q000096:biosynthesis; D005260:Female; D006801:Humans; D007457:Iodine Radioisotopes; D058228:Islet Amyloid Polypeptide; D016133:Polymerase Chain Reaction; D011869:Radioligand Assay; D018003:Receptors, Calcitonin / Q000235:genetics / Q000378:metabolism*; D058287:Receptors, Islet Amyloid Polypeptide; D018000:Receptors, Peptide / Q000378:metabolism*; D013268:Stimulation, Chemical; D014407:Tumor Cells, Cultured
chemical_list: D000682:Amyloid; D007457:Iodine Radioisotopes; D058228:Islet Amyloid Polypeptide; D018003:Receptors, Calcitonin; D058287:Receptors, Islet Amyloid Polypeptide; D018000:Receptors, Peptide; D000242:Cyclic AMP; D015740:Calcitonin Gene-Related Peptide