gastrin-releasing-peptide and Cell-Transformation--Neoplastic

Gastrin is a pro-proliferative, anti-apoptotic hormone with a central role in acid secretion in the gastric mucosa and a long-standing association with malignant progression in transgenic mouse models. However, its exact role in human gastric malignancy requires further validation. Gastrin expression is tightly regulated by two closely associated hormones, somatostatin and gastrin-releasing peptide, and aspects of their interaction may be deregulated during progression to gastric adenocarcinoma. Furthermore, agonists and antagonists of the receptors for all three hormones have shown modest clinical efficacy against gastric adenocarcinoma, which might provide useful information on the future combined use of these agents.

Topics: Animals; Antineoplastic Agents, Hormonal; Apoptosis; Cancer Vaccines; Cell Differentiation; Cell Movement; Cell Transformation, Neoplastic; Gastrin-Releasing Peptide; Gastrins; Gene Expression Regulation, Neoplastic; Helicobacter Infections; Humans; Mice; Neoplasm Invasiveness; Neoplasms, Experimental; Neovascularization, Pathologic; Precancerous Conditions; Risk Factors; Somatostatin; Stomach Neoplasms

Small-cell lung cancer (SCLC) is a particularly aggressive cancer, which metastasises early. Despite initial sensitivity to radio- and chemo-therapy, it invariably relapses, so that the 2-year survival remains less than 5%. Neuropeptides particularly arginine vasopressin (AVP) and gastrin-releasing peptide (GRP) act as autocrine and paracrine growth factors and the expression of these and their receptors are a hallmark of the disease. Substance-P analogues including [D-Arg1,D-Phe5,D-Trp7,9,Leu11]-substance-P (SP-D) and [Arg6,D-Trp7,9,NmePhe8]-substance-P (6-11) (SP-G) inhibit the growth of SCLC cells by modulating neuropeptide signalling. We show that GRP and V1A receptors expression leads to the development of a transformed phenotype. Addition of neuropeptide provides some protection from etoposide-induced cytotoxicity. Receptor expression also leads to an increased sensitivity to substance-P analogue-induced growth inhibition. We show that SP-D and SP-G act as biased agonists at GRP and V1A receptors causing blockade of Gq-mediated Ca2+ release while directing signalling to activate ERK via a pertussis toxin-sensitive pathway. This is the first description of biased agonism at V1A receptors. This unique pharmacology governs the antiproliferative properties of these agents and highlights their potential therapeutic potential for the treatment of SCLC and particularly in tumours, which have developed resistance to chemotherapy.

Topics: Animals; Arginine Vasopressin; Cell Division; Cell Transformation, Neoplastic; Cricetinae; Cricetulus; Etoposide; Gastrin-Releasing Peptide; Humans; Peptide Fragments; Receptors, Neuropeptide; Substance P; Transfection; Tumor Cells, Cultured

The mammalian gastrin-releasing peptide receptor (GRP-R) belongs to the superfamily of G protein-coupled receptors and mediates actions of the regulatory GRP and bombesin, the amphibian homolog of GRP. Owing to its frequent ectopic expression in some epithelial human malignancies, such as cancers of the colon, lung, and prostate, ligand-specific receptor activation may initiate intracellular signals of cell proliferation, differentiation and migration in this context. Because the underlying molecular mechanisms of aberrant human GRP-R (hGRP-R) expression in tumorigenesis remain unknown, we examined in this study the transcriptional activation of hGRP-R in gastrointestinal and prostate cancer cells, which natively express functional hGRP-R. Using various hGRP-R promoter mutants cloned into a luciferase reporter plasmid, transient transfection studies demonstrated robust transcriptional activation in gastrointestinal and prostate cancer cells. Although our study revealed distinct patterns of transcriptional hGRP-R activation in gastrointestinal and prostate cancer cells, genomic sequences between 97 and 247 bp upstream of the major RNA initiation site appear to be of particular significance for basal transcriptional hGRP-R activation. Based on this study, future examination of transcription factor interaction with the hGRP-R promoter will be important to identify molecular mechanisms of hGRP-R regulation relevant in human cancers that express functional receptor sites

Topics: Carcinoma; Cell Line, Tumor; Cell Transformation, Neoplastic; Epithelial Cells; Gastrin-Releasing Peptide; Gastrointestinal Neoplasms; Gene Expression Regulation, Neoplastic; Humans; Luciferases; Male; Mutation; Promoter Regions, Genetic; Prostatic Neoplasms; Receptors, Bombesin; RNA, Messenger; Transcriptional Activation

Gastrin-releasing peptide (GRP) is known as an autocrine growth factor for a number of gastrointestinal cancers. There is, however, little information on the expression of GRP in the squamous epithelia and squamous cell carcinoma, particularly in the esophagus. With a differential display approach, up-regulated GRP was observed in human esophageal squamous cell carcinoma (ESCC) samples obtained from a high-risk area for esophageal cancer, Linzhou in northern China. Up-regulation of phosphoglycerate mutase and P311 HUM (3.1) and down-regulation of keratin 13, cystatin B, endoglin and annexin I were observed. Using a reverse transcription-polymerase chain reaction (RT-PCR) method, significant over-expression of GRP was observed in 10 out of 12 ESCC samples (83.3%) and all four ESCC cell lines. With in situ hybridization, GRP mRNA expression was detected in nine out of 21 (42.8%) samples with basal cell hyperplasia (BCH), five out of seven (71.4%) samples with dysplasia (DYS) and 17 out of 24 (70.9%) ESCC samples. In contrast, GRP was expressed only in three out of 16 (18.7%) normal epithelium. Digital image analysis revealed that the mean value of GRP expression index, determined by intensity and area ratio of staining, was 0.19 in normal epithelium, 1.23 in BCH, 2.94 in DYS and 2.38 in ESCC, showing a progressive increase. Studies on ESCC cell lines showed GRP increased cell growth in a dose-dependent pattern in GRP receptor-positive ESCC cells, but not in GRP receptor-negative ESCC cells. GRP (1 mM) also increased cyclooxygenase-2 protein expression by 3.4-fold. This is the first demonstration that GRP is over-expressed in ESCC, and its over-expression may play a role in ESCC development and growth.

Topics: Base Sequence; Carcinoma, Squamous Cell; Cell Transformation, Neoplastic; DNA Primers; Esophageal Neoplasms; Gastrin-Releasing Peptide; Humans; In Situ Hybridization; Reverse Transcriptase Polymerase Chain Reaction

The neuropeptide bombesin stimulates tumour cell proliferation in vitro. Through pharmacological testing, 20-40% of human colorectal tumours have been shown to be equipped with bombesin/gastrin releasing peptide receptor (GRP-R). The aim of the present study was to test whether GRP-R expression is correlated with tumour characteristics and usual prognostic factors in colorectal adenocarcinomas. A sensitive reverse transcription (RT)-competitive polymerase chain reaction (PCR) method was validated by studying GRP-R mRNA in separated layers of normal colonic wall, and GRP-R mRNA levels (in parallel with binding studies) in colon cancer cell lines LoVo and Caco-2. GRP-R mRNA levels were then determined in 29 surgical tumour specimens and the results compared with tumour histology and, using histochemistry, with the accumulation of p53 protein and a Ki-67 cell proliferation index. The mRNA was not detected in normal colonic epithelium, whereas a distinct signal was observed after amplification in 27/29 (93%) tumour specimens. Estimates of mRNA levels in the 27 positive tumours ranged from 52 to 8000 amol/0.25 microgram total RNA, and were significantly higher in poorly/moderately differentiated tumours (P < 0.05) and in tumours with lymphatic vessel invasion (P < 0.01). There was no relationship with p53 accumulation or to the proliferation index. Our results show that GRP-R mRNA can be detected in most colorectal tumour specimens, and suggest a link between high mRNA levels and both tumour dedifferentiation and lymph vessel invasion, but not proliferation.

Topics: Aged; Cell Transformation, Neoplastic; Colonic Neoplasms; Female; Gastrin-Releasing Peptide; Humans; Immunohistochemistry; Lymphatic Diseases; Neoplasm Invasiveness; Neoplasm Proteins; RNA, Messenger

Gastrin-releasing peptide (GRP) is a neuropeptide with growth factor activity in vitro for a variety of tumors including neuroblastoma. If GRP is secreted by neuroblastomas, its detection in serum might be an excellent way to both diagnose and monitor this tumor in patients.. Small portions of resected tumor specimens were maintained in tissue culture as tumor explants for 24 hours. The tumors included: 3 ganglioneuromas, 1 neuroblastoma, 1 primitive neuroectodermal tumor, 1 Wilms' tumor, 1 rhabdoid tumor, and 1 benign brachial plexus tumor. Control flasks were maintained simultaneously under identical conditions. After 24 hours of incubation, the tumor-conditioned media and the control media were assayed in duplicate for [GRP] using a radioimmunoassay.. All the conditioned media from the benign tumors contained < 25 pg/mL net GRP, whereas all the malignant tumor-conditioned media contained > or = 45 pg/mL (P = 0.003).. These data suggest that GRP is secreted by pediatric retroperitoneal tumors and that the amount secreted varies directly with the degree of malignancy of the tumor. This study suggests that GRP may be a candidate tumor marker for pediatric retroperitoneal tumors.

Topics: Biomarkers, Tumor; Cell Transformation, Neoplastic; Cells, Cultured; Child; Culture Media, Conditioned; Gastrin-Releasing Peptide; Gastrointestinal Hormones; Humans; Peptides; Retroperitoneal Neoplasms

1. The ability of bombesin or platelet-derived growth factor (PDGF) to stimulate Ca2+ inflow (assessed by measuring changes in the intracellular free Ca2+ concentration in cells loaded with fura-2) in NIH-3T3 cells transformed with the EJ/T24-Ha-ras-1 oncogene is inhibited when compared with the action of the agonists on wild-type cells. 2. The effects of transformation with the ras oncogene are associated with complete inhibition of the ability of bombesin to release Ca2+ from intracellular stores, a substantial decrease in the number of bombesin receptors, no change in the ability of foetal calf serum or ionomycin to release Ca2+ from intracellular stores and the activation of protein kinase C. 3. The effects of transformation with the H-ras oncogene on the ability of bombesin or PDGF to stimulate Ca2+ inflow were mimicked by a 30 min exposure of wild-type cells to phorbol dibutyrate. This action of phorbol dibutyrate was completely blocked by prior treatment of wild-type cells for 24 h with the phorbol ester. 4. It is concluded that one of the actions of the H-ras oncogene in fibroblasts is to inhibit agonist-stimulated Ca2+ inflow by a mechanism which involves the activation of protein kinase C.

Topics: Animals; Bombesin; Calcium; Cell Line, Transformed; Cell Transformation, Neoplastic; Egtazic Acid; Enzyme Activation; Fibroblasts; Gastrin-Releasing Peptide; Genes, ras; Mice; Peptides; Phorbol 12,13-Dibutyrate; Platelet-Derived Growth Factor; Protein Kinase C; Receptors, Bombesin; Receptors, Neurotransmitter