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 [Leu(15)]G17 and [Leu(15)]G17-Gly to DLD-1 cell membranes in competition with [(3)H]G17-Gly was examined. Binding of [(3)H]cholecystokinin-8 (CCK8) to DLD-1 cell membranes was also assessed. Whole cell binding experiments were carried out using [(125)I-Tyr(12),Leu(15)]G17-Gly. In addition, the ability of [Leu(15)]G17 and [Leu(15)]G17-Gly to stimulate cell growth, as determined by cell counting, was tested. [Leu(15)]G17 and [Leu(15)]G17-Gly competed with [(3)H]G17-Gly at both a high and a low affinity site on DLD-1 membranes. The IC(50) values for [Leu(15)]G17 were 6.0 x 10(-8) M and 6.9 x 10(-6) M while those for [Leu(15)]G17-Gly were 3.2 x 10(-9) M and 4.9 x 10(-6) M. [(3)H]CCK8 did not bind to either site. [Leu(15)]G17-Gly also competed with [(125)I-Tyr(12),Leu(15)]G17-Gly at both a high and a low affinity site on DLD-1 cells with similar affinities as observed with membranes. [Leu(15)]G17 and [Leu(15)]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 CCK(1) or CCK(2) receptors.
Ahmed
Shawn
S
Creighton University School of Medicine, Department of Biomedical Sciences, 2500 California Plaza, Omaha, NE 68137, USA.
Budai
Barna
B
Herédi-Szabó
Krisztina
K
Farkas
Judit
J
Tóth
Géza
G
Murphy
Richard F
RF
Lovas
Sándor
S
D000818:Animals; D016466:CHO Cells; D045744:Cell Line, Tumor; D002462:Cell Membrane / Q000378:metabolism; D015179:Colorectal Neoplasms / Q000378:metabolism* / Q000473:pathology; D006224:Cricetinae; D004305:Dose-Response Relationship, Drug; D005755:Gastrins / Q000737:chemistry / Q000502:physiology*; D006801:Humans; D008934:Mitogens / Q000737:chemistry / Q000502:physiology*; D011869:Radioligand Assay; D043887:Receptor, Cholecystokinin B / Q000378:metabolism*; D011949:Receptors, Cholecystokinin / Q000378:metabolism
D005755:Gastrins; D008934:Mitogens; D043887:Receptor, Cholecystokinin B; D011949:Receptors, Cholecystokinin