Dissociation characteristics of endothelin receptor agonists and antagonists in cloned human type-B endothelin receptor.
The human type-B endothelin receptor (h-ETB) was cloned from human lung poly A+RNA and stably expressed in CHO cells. Endothelin (ET) receptor binding and stimulation of PI hydrolysis demonstrated that the cloned h-ETB receptor is functional and linked to intracellular signal transduction pathways in CHO cells. The molecular mass of the h-ETB receptor was determined to be 65 KDa, and Bmax and Kd were 0.36 pmol/mg and 80 pM, respectively. Competition studies employing receptor ligands revealed that the potencies of the test ligands (IRL1620, PD142893, and Ro46-2005) were dependent on the length of the incubation time, whereas the natural agonists (ET-1 and ET-3) were not. When competing with ET-1 in the h-ETB receptor binding, the IC50 increased from 1.2 nM to 8.2 nM for IRL1620, 0.068 microM to 1.9 microM for PD142893, and 0.76 microM to 12.7 microM for Ro46-2005, as the incubation time increased from 1 hr to 24 hr. These time-induced changes are likely due to differences in the dissociation characteristics between the artificial ligands and the natural ligands.
Chiou
W J
WJ
Pharmaceutical Products Division, Abbott Laboratories, Abbott Park, IL 60064, USA.
Magnuson
S R
SR
Dixon
D
D
Sundy
S
S
Opgenorth
T J
TJ
Wu-Wong
J R
JR
D000818:Animals; D016466:CHO Cells; D006224:Cricetinae; D065128:Endothelin Receptor Antagonists; D016232:Endothelins / Q000378:metabolism*; D006801:Humans; D007700:Kinetics; D008970:Molecular Weight; D009842:Oligopeptides / Q000378:metabolism*; D010446:Peptide Fragments / Q000378:metabolism*; D011485:Protein Binding; D011743:Pyrimidines / Q000378:metabolism; D044023:Receptor, Endothelin B; D017466:Receptors, Endothelin / Q000819:agonists / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D013449:Sulfonamides / Q000378:metabolism; D000096926:Benzenesulfonamides
D065128:Endothelin Receptor Antagonists; D016232:Endothelins; D009842:Oligopeptides; D010446:Peptide Fragments; D011743:Pyrimidines; D044023:Receptor, Endothelin B; D017466:Receptors, Endothelin; D011993:Recombinant Fusion Proteins; D013449:Sulfonamides; C074308:sovateltide; C076213:PD 142893; C083751:Ro 46-2005; D000096926:Benzenesulfonamides