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Antiproliferative gastrin/cholecystokinin receptor antagonists target the 78-kDa gastrin-binding protein
1Ludwig Institute for Cancer Research, Melbourne Tumour Biology Branch P.O. Royal Melbourne Hospital, Victoria, Australia.
Abstract:
Inhibition of colon carcinoma cell growth by the nonselective gastrin/cholecystokinin (CCK) receptor antagonists proglumide and benzotript provided evidence that gastrin functions as an autocrine growth factor. However, the molecular properties of the receptor mediating the antagonist effects have not been identified. A 78-kDa gastrin-binding protein (GBP), the sequence of which is related to the family of enzymes possessing enoyl-CoA hydratase and 3-hydroxyacyl-CoA dehydrogenase activities, has been previously purified from porcine gastric mucosal membranes. I now report that covalent cross-linking of 125I-labeled [Nle15]gastrin2,17 to the 78-kDa GBP is inhibited by crotonyl-CoA and by acetoacetyl-CoA. Gastrin, CCK, and their analogues also inhibit cross-linking, and the spectrum of analogue affinities correlates better with the values previously reported for binding to the gastrin/CCK-C receptor than with the values reported for binding to either the CCK-A or the gastrin/CCK-B receptor. Cross-linking is also inhibited by proglumide and benzotript, but no inhibition is seen with either the CCK-A receptor-selective antagonist L364,718 or the gastrin/CCK-B receptor-selective antagonist L365,260. The affinities of antagonists for the GBP correlate well with their affinities for the gastrin/CCK-C receptor and with their potencies for inhibition of colon carcinoma cell growth. I conclude that the 78-kDa gastrin-binding protein is (i) a member of the hydratase/dehydrogenase family of fatty acid oxidation enzymes, (ii) the gastrin/CCK-C receptor, and (iii) the target for the antiproliferative action of two gastrin/CCK receptor antagonists.
Insights
Gastrin acts as a colon cancer growth factor. Researchers identified a 78-kDa gastrin-binding protein as the gastrin/cholecystokinin-C receptor, crucial for gastrin
Area of Science:
- Biochemistry
- Molecular Biology
- Oncology
Background:
- Gastrin and cholecystokinin (CCK) are implicated in colon carcinoma cell growth.
- The specific receptor mediating these effects, particularly the gastrin/CCK-C receptor, remained molecularly uncharacterized.
- Previous studies suggested gastrin functions as an autocrine growth factor in colon cancer.
Purpose of the Study:
- To identify the molecular identity of the gastrin/CCK-C receptor.
- To characterize the gastrin-binding protein (GBP) purified from porcine gastric mucosa.
- To determine if the GBP is the target for gastrin/CCK receptor antagonists in colon cancer.
Main Methods:
- Covalent cross-linking of radiolabeled gastrin analogue to the 78-kDa GBP.
- Assessing inhibition of cross-linking by various ligands, including gastrin analogues, fatty acid metabolites, and known receptor antagonists.
- Correlating binding affinities of antagonists to GBP with their known affinities for different CCK receptor subtypes and their antiproliferative potencies.
Main Results:
- Covalent cross-linking of gastrin to the 78-kDa GBP was inhibited by fatty acid oxidation intermediates (crotonyl-CoA, acetoacetyl-CoA).
- The binding affinities of gastrin analogues to the 78-kDa GBP closely matched those reported for the gastrin/CCK-C receptor.
- Selective CCK-A and CCK-B receptor antagonists did not inhibit gastrin cross-linking, while nonselective antagonists (proglumide, benzotript) did, correlating with their antiproliferative effects.
Conclusions:
- The 78-kDa gastrin-binding protein is identified as the gastrin/CCK-C receptor.
- This receptor belongs to the hydratase/dehydrogenase family involved in fatty acid oxidation.
- The 78-kDa gastrin-binding protein is the molecular target mediating the antiproliferative effects of gastrin/CCK receptor antagonists in colon carcinoma cells.