RT Journal Article SR Electronic T1 Pharmacology and Intracellular Signaling Mechanisms of the Native Human Orphan Receptor BRS-3 in Lung Cancer Cells JF Journal of Pharmacology and Experimental Therapeutics JO J Pharmacol Exp Ther FD American Society for Pharmacology and Experimental Therapeutics SP 366 OP 380 VO 287 IS 1 A1 Richard R. Ryan A1 H. Christian Weber A1 Samuel A. Mantey A1 Wei Hou A1 Mary E. Hilburger A1 Tapas K. Pradhan A1 David H. Coy A1 Robert T. Jensen YR 1998 UL http://jpet.aspetjournals.org/content/287/1/366.abstract AB Neither the native ligand nor the cell biology of the bombesin (Bn)-related orphan receptor subtype 3 (BRS-3) is known. In this study, we used RT-PCR to identify two human lung cancer lines that contain sufficient numbers of native hBRS-3 to allow study: NCI-N417 and NCI-H720. In both cell lines, [dPhe6,βAla11,Phe13,Nle14]Bn(6-14) stimulates [3H]inositol phosphate. In NCI-N417 cells, binding of125I-[dTyr6,βAla11,Phe13,Nle14]Bn(6-14) was saturable and high-affinity. [dPhe6,βAla11,Phe13,Nle14]Bn(6-14) stimulated phospholipase D activity and a concentration-dependent release of [3H]inositol phosphate (EC50 = 25 nM) and intracellular calcium (EC50 = 14 nM); the increases in intracellular calcium were primarily from intracellular stores. hBRS-3 activation was not coupled to changes in adenylate cyclase activity, [3H]-thymidine incorporation or cell proliferation. No naturally occurring Bn-related peptides bound or activated the hBRS-3 with high affinity. Four different bombesin receptor antagonists inhibited increases in [3H]inositol phosphate. Using cytosensor microphysiometry, we found that [dPhe6,βAla11,Phe13, Nle14]Bn(6-14) caused concentration-dependent acidification. The results show that native hBRS-3 receptors couple to phospholipases C and D but not to adenylate cyclase and that they stimulate mobilization of intracellular calcium and increase metabolism but not growth. The discovery of human cell lines with native, functional BRS-3 receptors, of new leads for a more hBRS-3-specific antagonist and of the validity of microphysiometry as an assay has yielded important tools that can be used for the identification of a native ligand for hBRS-3 and for the characterization of BRS-3-mediated biological responses. The American Society for Pharmacology and Experimental Therapeutics