EVIDENCE FOR AUTOCRINE GROWTH-STIMULATION BY A GASTRIN CCK-LIKE PEPTIDE OF THE GASTRIC-CANCER HGT-1 CELL-LINE

Gastrin has been shown to promote the growth of some colonic tumor cell lines. To evaluate the involvement of this hormone in the proliferation of gastric tumors, we studied the effects of gastrin/CCK-receptor antagonists (L365,260 and L364,718), proglumide and C terminal-specific gastrin antibodies on the human gastric adenocarcinoma cell line HGT-1. L365,260, but not L364,718, dose-dependently inhibited cell proliferation. (72% after 4 days at 10 nM) and [H-3]thymidine incorporation (68% after 2 days at 10 nM) in serum-free medium. No cytotoxic effects of proglumide or L365,260 on this cell line were detected. Proglumide inhibited cell proliferation in serum-free medium (40% and 66.5% after 2 and 4 days of treatment; IC50 = 1.4 mM) and in 5% fetal calf serum (FCS)-supplemented medium (30% and 22% after 2 and 4 days of treatment; IC50 = 3.25 mM). [H-3]Thymidine incorporation was also inhibited by proglumide in serum-free medium (IC50 = 2.3 mM) and 5% FCS-supplemented medium (IC50 = 3.35 mM). Gastrin did not induce cell proliferation or increase [H-3]thymidine incorporation and no high-affinity gastrin binding sites were observed. However, C terminal-specific gastrin antibodies, even at low concentration, caused a dramatic decrease in both cell number (IC50 = 1:4000 antiserum dilution) and [H-3]thymidine incorporation (IC50 = 1:400 antiserum dilution) in the HGT-1 cell line. In addition, immunofluorescence analysis revealed that these antibodies specifically bind HGT-1 cells and radioimmunoassay analysis confirms the presence of gastrin/CCK-like peptide in cell extracts. Gastrin mRNA was analyzed by Northern blotting with a oligonucleotide probe corresponding to the natural sequence of the human hormone. We found the presence of three main transcripts (1.4, 3.3 and 3.7 kb), two of which were also found in extracts from the antral mucosa. These data strongly support the hypothesis of the presence in the cell of a gastrin/CCK-like peptide which is involved in autocrine or intracrine growth control of this cell line.