PT - JOURNAL ARTICLE AU - Richard R. Ryan AU - H. Christian Weber AU - Samuel A. Mantey AU - Wei Hou AU - Mary E. Hilburger AU - Tapas K. Pradhan AU - David H. Coy AU - Robert T. Jensen TI - Pharmacology and Intracellular Signaling Mechanisms of the Native Human Orphan Receptor BRS-3 in Lung Cancer Cells DP - 1998 Oct 01 TA - Journal of Pharmacology and Experimental Therapeutics PG - 366--380 VI - 287 IP - 1 4099 - http://jpet.aspetjournals.org/content/287/1/366.short 4100 - http://jpet.aspetjournals.org/content/287/1/366.full SO - J Pharmacol Exp Ther1998 Oct 01; 287 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