Synthesis, Expression and Biological Activity of the Prohormone for Gastrin Releasing Peptide (ProGRP)

Abstract

Gastrin-releasing peptide (GRP) has a widespread distribution and multiple stimulating effects on endocrine and exocrine secretions and metabolism. The prohormone for GRP (ProGRP, 125 amino acids) is processed to the amidated, biologically active end products GRP(1-27) and GRP(18-27). Amidated forms of GRP are putative autocrine or paracrine growth factors in a number of cancers including colorectal cancer. However, the potential role and biological activity of proGRP has not been investigated. Using a newly developed antisera directed to the N terminus of human proGRP, proGRP immunoreactivity was detected in all of the endometrial, prostate, and colon cancer cell lines tested and in nine of 10 resected colorectal carcinomas. However, no amidated forms were detected, suggesting an attenuation of processing in tumors. Recombinant proGRP was expressed as a His-tag fusion protein and purified by metal affinity chromatography and HPLC. ProGRP stimulated proliferation of a colon cancer cell line and activated MAPK, but unlike GRP(18-27)amide had no effect on inositol phosphate production. ProGRP did not compete with iodinated bombesin in binding assays on Balb-3T3 cells transfected with the known GRP receptors, GRP-R or BRS-3. We conclude that proGRP is present in a number of cancer cell lines and in resected colorectal tumors and is biologically active. Our results suggest that antagonists to GRP precursors rather than the amidated end products should be developed as a treatment for colorectal and other cancers that express proGRP-derived peptides.

International Union of Pharmacology. LXVIII. Mammalian Bombesin Receptors: Nomenclature, Distribution, Pharmacology, Signaling, and Functions in Normal and Disease States

“…Furthermore, pro-GRP immunoreactivity is reported in 90% of resected colorectal carcinomas and all endometrial, prostate, and colon cancer cell lines tested, without any amidated forms being present (Dumesny et al, 2006). Recombinant pro-GRP stimulated proliferation of the colon cancer cell line DLD-1, activating MAP kinase, but did not stimulate phospholipase C activity nor bind to known bombesin receptors, suggesting it was stimulating the tumor growth through a novel receptor (Dumesny et al, 2006). At present no receptor has been isolated that mediates these actions, but they are not inhibited by BB 2 receptor antagonists, raising the possibility they could be mediated by a novel receptor.…”

“…COOH-terminal precursor forms of GRP are reported to stimulate the proliferation and migration of the human colorectal carcinoma cell line DLD-1 (Patel et al, 2007a,b) through a BB 2 receptor-independent mechanism and the growth of the prostate cancer cell line DU145 (Patel et al, 2007b). Furthermore, pro-GRP immunoreactivity is reported in 90% of resected colorectal carcinomas and all endometrial, prostate, and colon cancer cell lines tested, without any amidated forms being present (Dumesny et al, 2006). Recombinant pro-GRP stimulated proliferation of the colon cancer cell line DLD-1, activating MAP kinase, but did not stimulate phospholipase C activity nor bind to known bombesin receptors, suggesting it was stimulating the tumor growth through a novel receptor (Dumesny et al, 2006).…”

Bombesin-related peptides and their receptors: recent advances in their role in physiology and disease states

“…However, recently a number of studies have provided evidence that nonamidated precursor forms of GRP stimulate proliferation and migration of human colorectal cancer cells [14][15][16] through a novel receptor, which is at present uncategorized.…”

Automated Time-Resolved Immunofluorometric Assay for Progastrin-Releasing Peptide

“…Protease processing of the peptide results in several products: a signal peptide (residues Ϫ23-1), the 2 mature GRP forms (residues 1-27 and 18 -27), and the proGRP moiety (residues 31-125) (4 ). Several cleaved products of the proGRP peptide have also been detected.…”