The bombesin-like peptides are a large family of peptides originally characterized in frog skin, but later found to have wide distribution and potent physiologic effect in mammals. One mammalian homolog of bombesin is gastrin-releasing peptide (GRP). GRP is widely distributed in the GI tract and CNS; but of greatest clinical importance, high levels of GRP are expressed by most small cell lung carcinomas (SCLC). Neuromedin B (NMB), the other known mammalian bombesin-like peptide, is similarly expressed in SCLC. In that many SCLC express receptors for bombesin-like peptides, and GRP and NMB stimulate the growth of SCLC, bombesin-like peptides are autocrine growth factors for lung cancer. Bombesin-like peptide also stimulate pancreatic hyperplasia and the growth of colon, breast and prostatic carcinoma cell lines. During the prior grant period, we characterized the genes for the known bombesin-like peptides and began to characterize their tissue specific expression. However, the factors that regulate GRP expression in neoplastic cell lines remains poorly understood. From these studies it has become apparent first, that bombesin-like peptides and their receptors are more widely expressed in human neoplasia than originally realized, and second, that the family of bombesin-like peptides is likely more diverse than originally realized. It is our basic hypothesis that these as yet uncharacterized mammalian bombesin-like peptides will also play a role in human neoplasia. Thus the broad objectives of this proposal are first, to characterized new bombesin-like peptides and second, to continue to define what regulates bombesin-lie peptide gene expression in neoplastic cell lines. We propose the following specific aims: (1) To characterize the structure, function and distribution of a potential new bombesin-like peptide that we have discovered (and named ARP) which arises by use of an alternate transcriptional start site within the GRP gene. Preliminary studies suggest RNAs encoding ARP are present in SCLC cell lines, GI tract, normal prostate and prostate carcinoma cell lines. (2) To characterize new bombesin-like peptides, specifically t identify true mammalian bombesin, distinct from mammalian GRP; to identify mammalian phyllolitorin; and to identify the ligand for a newly discovered bombesin-like peptide receptor. (3) To characterize the elements in the GRP gene promoter that including conventional protein chemistry, mass- spectrometry, low-stringency hybridization and polymerase chain reaction. The distribution of new peptides will be determined by immunohistochemistry and in situ hybridization, and pharmacologic specificity will be tested against the known bombesin-like peptide receptors expressed in fibroblast cell lines and Xenopus oocytes.
Specific Aim 3 will be accomplished by transfection of luminescent reporter gene constructs, gel shift analysis and DNase footprinting. The discovery of new bombesin-like peptides in human neoplasia will potentially lead to new diagnostic and therapeutic approaches.
