High and low affinity receptors mediate growth effects of gastrin and gastrin-Gly on DLD-1 human colonic carcinoma cells.

Gastrin (G17) and N-carboxymethylgastrin (G17-Gly) have been shown to stimulate the growth of colon cancer cells both in vivo and in vitro. The identity of the receptor mediating these effects is controversial. A recent study demonstrated the presence of a low affinity binding site for G17 and G17-Gly on the DLD-1 human colon cancer cell line. The goal of the current study was to further investigate the role of this receptor in mediating the growth-promoting effects of gastrin peptides. Binding of [Leu15]G17 and [Leu15]G17-Gly to DLD-1 cell membranes in competition with [3H]G17-Gly was examined. Binding of [3H]cholecystokinin-8 (CCK8) to DLD-1 cell membranes was also assessed. Whole cell binding experiments were carried out using [125I-Tyr12,Leu15]G17-Gly. In addition, the ability of [Leu15]G17 and [Leu15]G17-Gly to stimulate cell growth, as determined by cell counting, was tested. [Leu15]G17 and [Leu15]G17-Gly competed with [3H]G17-Gly at both a high and a low affinity site on DLD-1 membranes. The IC50 values for [Leu15]G17 were 6.0×10−8 M and 6.9×10−6 M while those for [Leu15]G17-Gly were 3.2×10−9 M and 4.9×10−6 M. [3H]CCK8 did not bind to either site. [Leu15]G17-Gly also competed with [125I-Tyr12,Leu15]G17-Gly at both a high and a low affinity site on DLD-1 cells with similar affinities as observed with membranes. [Leu15]G17 and [Leu15]G17-Gly significantly stimulated the growth of DLD-1 cells in a dose-dependent and biphasic manner. The binding profiles of the peptides tested suggest that these sites are different from previously identified wild-type and mutant CCK1 or CCK2 receptors.