CHARACTERIZATION OF A BOMBESIN GASTRIN-RELEASING PEPTIDE RECEPTOR ON A HUMAN GASTRIC-CANCER CELL-LINE/

This study examined the expression of receptors of the bombesin (BBS) family in human gastric-cancer cell lines. Of 5 cell lines screened on ly one, St42, demonstrated specific binding sites for I-125-Tyr(4)-BBS , which have been further characterized. This binding was saturable, a nd temperature- and time-dependent. Scatchard analysis of displacement data performed at 37 degrees C revealed 2 binding sites: a high-affin ity, low-capacity site (K-D = 0.13 nM, B-max = 1500 sites/cell) and a lower-affinity, higher-capacity site (K-D = 11 nM, B-max = 35,000 site s/cell); the latter was lost when internalization of peptide was preve nted, suggesting that it may be an artefact. Displacement assays with gastrin-releasing peptide (GRP) and neuromedin B (NMB) revealed that t he receptro was of the GRP-preferring sub-type (GRP IC50 = 0.35 nM; NM B IC50 = 112 nM). Co-valent cross-linking of I-125-Tyr(4)-BBS to the r eceptor demonstrated the presence of a single band corresponding to a molecular weight of 37 to 44 kDa on SDS-PAGE, similar to that of the c loned GRP receptor protein core. G-protein linkage of this receptor wa s demonstrated by selective inhibition of I-125-Tyr(4)-BBS binding by guanosine nucleotides. The binding of BBS to the receptor resulted in a rise in intracellular calcium. Three of four structurally distinct B BS antagonists bound to the receptor with high affinity, but [DPhe(12) , Leu(14)]-bombesin did not cause any displacement of I-125-Tyr(4)-BBS even at 10 mM. The functional significance of GRP receptors on human gastric-cancer cells is as yet unknown, but further studies may determ ine whether such receptors have importance in the therapy of gastric c ancer. (C) 1994 Wiley-Liss, Inc.