gastrin-releasing-peptide and Carcinoma--Squamous-Cell

Testing for circulating biomarkers in lung cancer is hampered by the insufficient specificity. We aimed to assess the relative diagnostic accuracy of pro-gastrin-releasing peptide (ProGRP) for the differential diagnosis of small cell lung cancer and compare it with more conventional biomarkers.. We enrolled a cohort of 390 patients with a clinical suspicion of lung cancer and for whom a histologic assessment was available. Serum or plasma samples were assessed for ProGRP, carcinoembryonic antigen, CYFRA 21-2, and neuron-specific enolase. The performance of each biomarker in discriminating the small cell lung cancer and squamous cell carcinoma/adenocarcinoma from non-malignant lung disease, and small cell lung cancer from squamous cell carcinoma/adenocarcinoma, was assayed by receiver operating characteristic curve analysis.. At the cut-off levels suggested by the manufacturers, ProGRP and neuron-specific enolase showed an almost identical sensitivity of 55.2% and 55.6%, respectively, in discriminating small cell lung cancer with respect to non-malignant lung disease. In order to quantify the added value of ProGRP to other conventional markers, we ran a multivariable logistic regression analysis, but the results showed that no markers improve the performance of ProGRP.. ProGRP and neuron-specific enolase individually appear more accurate than other conventional biomarkers for small cell lung cancer, but the union of two markers does not increase the accuracy. The very small target group of patients with small cell lung cancer is a limitation of this study, which can explain why ProGRP alone does not show a sensitivity higher than neuron-specific enolase, as reported by other authors.

Topics: Adenocarcinoma; Aged; Antigens, Neoplasm; Biomarkers, Tumor; Carcinoembryonic Antigen; Carcinoma, Non-Small-Cell Lung; Carcinoma, Squamous Cell; Diagnosis, Differential; Female; Follow-Up Studies; Gastrin-Releasing Peptide; Humans; Keratin-19; Lung Neoplasms; Male; Peptide Fragments; Prognosis; Recombinant Proteins; ROC Curve; Small Cell Lung Carcinoma

To evaluate five serum tumor markers used alone or in combination for the diagnosis of lung cancer.. The level of five serum tumor markers: NSE, pro-GRP, CYFRA21-1, p53 antibody and CEA was detected by ELISA in 50 healthy adults, 170 lung cancer patients and 60 patients with respiratory infection.. The level of the five serum tumor markers in lung cancer patients was significantly higher than that of healthy adults and patients with respiratory infection (P < 0.01). The level of NSE and pro-GRP in patients with small-cell lung cancer was significantly higher than those of the other subtypes of lung cancer (P < 0.01); The level of CYFRA21-1 in patients with squamous-cell carcinoma was significantly higher than that of other subtypes (P < 0.01). The specificity of p53 antibody was 100% in diagnosing lung cancer and the sensitivity of NSE, pro-GRP was much higher for small-cell lung cancer than for other subtypes (P < 0.01); The same was observed in CYFRA21-1 for the diagnosis of squamous-cell carcinoma (P < 0.01). The sensitivity of the tumor markers in diagnosing lung cancer was significantly enhanced if used in combination (P < 0.01).. These five tumor markers are valuable auxiliary parameters in diagnosing lung cancer. The combination of NSE and pro-GRP is more appropriate than other combinations in diagnosing small-cell lung cancer; the combination of CYFRA21-1, CEA and p53 antibody is the most valuable combination for diagnosing non-small-cell lung cancer. p53 antibody has the highest specificity for diagnosing lung cancer; CYFRA21-1 is the most valuable parameter for diagnosing squamous carcinoma.

Topics: Adenocarcinoma; Antibodies, Neoplasm; Antigens, Neoplasm; Biomarkers, Tumor; Carcinoembryonic Antigen; Carcinoma, Squamous Cell; Female; Gastrin-Releasing Peptide; Humans; Keratin-19; Keratins; Lung Neoplasms; Male; Middle Aged; Phosphopyruvate Hydratase; Tumor Suppressor Protein p53

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

Head and neck squamous cell carcinomas (HNSCCs) are characterized by up-regulation of the epidermal growth factor receptor (EGFR). We previously reported that a gastrin-releasing peptide/gastrin-releasing peptide receptor (GRP/GRPR) autocrine growth pathway is activated early in HNSCC carcinogenesis. GRP can induce rapid phosphorylation of EGFR and p42/44 mitogen-activated protein kinase (MAPK) activation in part via extracellular release of transforming growth factor alpha (TGF-alpha) by matrix metalloproteinases (MMPs). It has been reported that Src family kinases are activated by G-protein-coupled receptors (GPCRs), followed by downstream EGFR and MAPK activation. To further elucidate the mechanism of activation of EGFR by GRP in HNSCC, we investigated the role of Src family kinases. Blockade of Src family kinases using an Src-specific tyrosine kinase inhibitor A-419259 decreased GRP-induced EGFR phosphorylation and MAPK activation. GRP also failed to induce MAPK activation in dominant-negative c-Src-transfected HNSCC cells. Invasion and growth assays showed that c-Src was required for GRP-induced proliferation or invasion of HNSCC cells. In addition to TGF-alpha release, GRP induced amphiregulin, but not EGF, secretion into HNSCC cell culture medium, an effect that was blocked by the MMP inhibitor marimastat. TGF-alpha and amphiregulin secretion by GRP stimulation also was inhibited by blockade of Src family kinases. These results suggest that Src family kinases contribute to GRP-mediated EGFR growth and invasion pathways by facilitating cleavage and release of TGF-alpha and amphiregulin in HNSCC.

Topics: Amphiregulin; Carcinoma, Squamous Cell; Cell Division; Cell Line, Tumor; EGF Family of Proteins; Enzyme Activation; Epidermal Growth Factor; ErbB Receptors; Gastrin-Releasing Peptide; Glycoproteins; Head and Neck Neoplasms; Humans; Intercellular Signaling Peptides and Proteins; Mitogen-Activated Protein Kinases; Neoplasm Invasiveness; Phosphorylation; Receptors, Bombesin; src-Family Kinases; Transforming Growth Factor alpha

Head and neck squamous cell carcinomas (HNSCC) are characterized by upregulation of the epidermal growth factor receptor (EGFR), where EGFR serves as a potential therapeutic target. We previously reported that a gastrin-releasing peptide/gastrin-releasing peptide receptor (GRP/GRPR) autocrine growth pathway is activated early in HNSCC carcinogenesis. In the present study, we examined the mechanism of EGFR activation by GRP/GRPR in HNSCC proliferation. In HNSCC cells that express elevated levels of both GRPR and EGFR, we found that GRP induced rapid phosphorylation of EGFR as well as p44/42-MAPK activation. Using several EGFR-specific tyrosine kinase inhibitors and cells derived from EGFR knockout mice, we demonstrated that GRP-induced p44/42-MAPK activation was dependent upon EGFR activation. Further investigation demonstrated that cleavage of transforming growth factor-alpha (TGF-alpha) by matrix metalloproteinases mediated GRP-induced MAPK activation. In addition, HNSCC proliferation stimulated by GRP was eliminated upon specific inhibition of EGFR or MEK, and GRP failed to stimulate proliferation in EGFR-deficient cells. These results imply that the mitogenic effects of GRP in HNSCC are mediated by extracellular release of TGF-alpha and require the activation of an EGFR-dependent MEK/MAPK-dependent pathway.

Topics: Animals; Carcinoma, Squamous Cell; Cell Division; Enzyme Activation; Enzyme Inhibitors; ErbB Receptors; Fibroblasts; Gastrin-Releasing Peptide; Head and Neck Neoplasms; Humans; MAP Kinase Kinase Kinase 1; Mice; Mice, Knockout; Mitogen-Activated Protein Kinase 3; Mitogen-Activated Protein Kinases; Mitogens; Phosphorylation; Protein Serine-Threonine Kinases; Receptors, Bombesin; Signal Transduction; Transcriptional Activation; Tumor Cells, Cultured

Gastrin-releasing peptide receptor (GRPR)-mediated autocrine growth appears to be an early marker of susceptibility to tobacco-related lung cancers. Because expression of GRPR, however, has not been reported in squamous cell carcinoma of the head and neck (SCCHN), we investigated its expression and that of its ligand GRP in normal mucosa and SCCHN tissues and the involvement of these proteins in the proliferation of SCCHN cells.. We assessed GRPR messenger RNA (mRNA) expression in specimens from 25 patients with SCCHN, six control noncancer patients, and 14 SCCHN cell lines by use of quantitative reverse transcriptase-polymerase chain reaction. We used neutralizing GRP monoclonal antibody 2A11 to block the GRP-GRPR interaction in SCCHN cell lines and xenografts and assessed the antibody's effect on proliferation by counting cultured cells or measuring xenograft tumor volume in vivo. All statistical tests were two-sided.. Tumor and mucosa tissues, respectively, from SCCHN patients expressed sixfold and fourfold higher levels of GRPR mRNA than normal mucosa tissue from noncancer patients (P<.001). The levels of GRPR expression in the tumor and adjacent normal epithelium of individual patients with SCCHN were correlated (r =.652; P =.001), suggesting that increased GRPR expression is an early event in SCCHN formation. SCCHN cells expressed fivefold higher levels of GRPR mRNA than did cultured normal mucosal epithelial cells (P =.005). GRP stimulated proliferation of SCCHN cells in a dose-dependent fashion (P =.006). Neutralizing GRP monoclonal antibody 2A11 inhibited SCCHN cell proliferation in vitro and in vivo. Median survival was 54 months in patients with higher levels of GRPR mRNA and was not reached in those with lower levels.. GRP and GRPR appear to participate in an autocrine regulatory pathway in SCCHN. Thus, strategies that specifically target GRP and/or GRPR may be effective therapeutic approaches for this disease.

Topics: Adult; Aged; Aged, 80 and over; Animals; Antibodies, Monoclonal; Autocrine Communication; Blotting, Western; Carcinoma, Squamous Cell; Female; Gastrin-Releasing Peptide; Gene Expression Regulation, Neoplastic; Head and Neck Neoplasms; Humans; Male; Mice; Mice, Nude; Middle Aged; Receptors, Bombesin; Respiratory Mucosa; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; RNA, Neoplasm; Survival Analysis; Time Factors

Neutral endopeptidase (NEP; CALLA, CD10, EC 3.4.24.11) is a cell surface endopeptidase that hydrolyses bioactive peptides, including the bombesin-like peptides, as well as other neuropeptides. Bombesin-like peptides and other neuropeptides are autocrine growth factors for both small cell lung cancer (SCLC) and non-small cell lung cancer (NSCLC). Low expression of NEP has been reported in SCLC and NSCLC cell lines. NEP inhibition has been shown to increase proliferation in one cell line. To date, NEP expression has not been quantitatively evaluated in normal adult lung, SCLC or NSCLC tumors, paired uninvolved lung from the same patient, or in other pulmonary neoplasms such as mesotheliomas and carcinoids. We examined the expression of NEP in these tissues and human cell lines using immunohistochemistry, flow cytometry, enzyme activity, ELISA, Western blot, and reverse transcription (RT)-PCR. Uninvolved lung tissue from different individuals displayed considerable variation in NEP activity and protein. By immunohistochemistry, NEP expression was detectable in alveolar and airway epithelium, fibroblasts of normal lung, and in mesotheliomas, whereas it was undetectable in most SCLC, adenocarcinoma, squamous cell carcinoma, and carcinoid tumors of the lung. NEP activity and protein levels were lower in all SCLC and adenocarcinoma tumors when compared to adjacent uninvolved lung, often at levels consistent with expression derived from contaminating stroma. NEP expression and activity were reduced or undetectable in most SCLC and lung adenocarcinoma cell lines. NEP mRNA by RT-PCR was not expressed or was in low abundance in the majority of lung cancer cell lines. The majority of lung tumors did not express NEP by RT-PCR as compared with normal adjacent lung. In addition, recombinant NEP abolished, whereas an NEP inhibitor potentiated, the calcium flux generated by neuropeptides in some lung cancer cell lines, demonstrating potential physiological significance for low NEP expression. NEP, therefore, is a signal transduction and possibly a growth modulator for both SCLC and NSCLC, emphasizing the role of neuropeptides in the pathogenesis of the major histological forms of lung cancer.

Topics: Adenocarcinoma; Adult; Base Sequence; Blotting, Western; Bradykinin; Calcium; Carcinoma, Non-Small-Cell Lung; Carcinoma, Small Cell; Carcinoma, Squamous Cell; Cell Line; DNA Primers; Enzyme Inhibitors; Gastrin-Releasing Peptide; Gene Expression; Glycopeptides; Humans; Immunohistochemistry; Lung; Lung Neoplasms; Mesothelioma; Molecular Sequence Data; Neoplasm Metastasis; Neprilysin; Peptides; Polymerase Chain Reaction; Pulmonary Alveoli; Recombinant Proteins; RNA, Messenger; Signal Transduction; Tumor Cells, Cultured

Gastrin-releasing peptide is an important growth-modulating factor in developing lung epithelium. It is known to be produced by small cell carcinomas of the lung, and an autocrine loop involving gastrin-releasing peptide and its receptor has been demonstrated in many small cell lung tumors. We investigated whether such an autocrine loop could also be demonstrated in non-small cell lung carcinoma, since gastrin-releasing peptide is known to stimulate human bronchial epithelial cells, from which non-small cell tumors should emerge. We report here that gastrin-releasing peptide is produced by a bronchiolo-alveolar carcinoma cell line (A549) adapted to serum-free and growth factor-free conditions. A549 cells adapted to these conditions, termed A549-R0 cells, display extensive membrane interdigitations, Golgi apparatus, and secretory-like granules, and grow as a mixture of attached colonies and floating cells. Gastrin-releasing peptide is present in the conditioned medium produced by A549-R0 cells. Colony formation of cells derived from a squamous cell carcinoma of the lung, 239T, was stimulated 9-fold by A549-R0 conditioned medium or by authentic gastrin-releasing peptide, measured in serum-free conditions. The growth stimulatory activity was inhibited by a monoclonal antibody to gastrin-releasing peptide. Transcripts for receptors for the bombesin family of peptides were also demonstrated in A549-R0 cells and 239T cells. These results demonstrate that non-small cell lung carcinomas can secrete gastrin-releasing peptide and can also respond to the peptide.

Topics: Base Sequence; Bombesin; Carcinoma, Non-Small-Cell Lung; Carcinoma, Squamous Cell; Cell Line; Culture Media, Conditioned; Culture Media, Serum-Free; Culture Techniques; DNA Primers; Gastrin-Releasing Peptide; Gastrointestinal Hormones; Gene Expression; Growth Substances; Humans; Immunohistochemistry; Lung Neoplasms; Microscopy, Electron; Molecular Sequence Data; Neuropeptides; Peptide Biosynthesis; Peptides; Protein Precursors; Radioimmunoassay; Receptors, Bombesin; Tumor Cells, Cultured; Tumor Stem Cell Assay

Serial changes in serum gastrin level were detected by radioimmunoassay in 58 lung cancer patients before and after operation. In comparing these tests with those of 40 cases of noncancerous thoracic lesions and 151 normal adults, the serum gastrin from lung cancer patients is significantly higher than that of noncancerous thoracic lesions and normal individuals (P less than 0.01). The gastrin level is closely related to stage of cancer, size of primary tumor, presence of lymph node metastasis, and type of histological classification. The serum gastrin was found to decrease gradually after the removal of the tumor and to return to normal on the 14th postoperative day. Those patients whose serum gastrin level can return to normal on the 14th postoperative day will have a good prognosis; if not, their prognosis will be very poor. These results suggest that serum from patients with lung cancer contains a high concentration of gastrin that can help differentiate benign from malignant thoracic lesions and evaluate prognosis of patients with lung cancer. Therefore, the cause of high serum gastrin in patients with lung cancer is likely due to the gastrin-producing property of the lung cancer cells.

Topics: Adenocarcinoma; Adult; Aged; Carcinoma, Small Cell; Carcinoma, Squamous Cell; Female; Gastrin-Releasing Peptide; Gastrins; Humans; Lung Neoplasms; Lymphatic Metastasis; Male; Middle Aged; Neoplasm Staging; Peptides; Postoperative Period; Prognosis; Thoracic Diseases

Levels of gastrin-releasing peptide (GRP) were determined by radioimmunoassay in human normal main and lobar bronchus and parenchymal lung tissue extracts. It was found that the level of GRP differed significantly between all 3 areas. The concentration of GRP was statistically higher in main bronchus (median 6.74 ng/g) compared to both lobar bronchus (median 4.79 ng/g) and parenchymal lung (median 1.73 ng/g), and also statistically higher in lobar bronchus compared to parenchymal lung. Chromatographically, GRP-immunoreactivity in both main and lobar bronchial extracts corresponded to GRP1-27 and GRP18-27, while in lung tissue only one major species was identified which corresponded in retention time to GRP18-27. No significant difference was detected when the levels of GRP in normal lobar bronchus and normal lung tissue were compared to the levels in lobar bronchus and lung taken from patients with lung carcinoma, at a site adjacent to the carcinoma. However, a significant difference was observed between the GRP content of normal main bronchus compared to main bronchus from patients with carcinoma. GRP was measured in 26/56 lung carcinomas examined. The levels ranged from 42,000 ng/g in a carcinoid tumour to 0.18 ng/g in a squamous-cell carcinoma, though only in 6 tumours were the levels outside the range determined for normal pulmonary tissue. Chromatography of selected tumour extracts of different histopathologies showed that there were differences in the GRP products present.

Topics: Adenocarcinoma; Adult; Aged; Carcinoid Tumor; Carcinoma; Carcinoma, Small Cell; Carcinoma, Squamous Cell; Chromatography, High Pressure Liquid; Female; Gastrin-Releasing Peptide; Gastrointestinal Hormones; Humans; Lung; Lung Neoplasms; Male; Middle Aged; Peptides; Radioimmunoassay

Twenty-seven cases of surgically resected large cell carcinoma of the lung including nine cases of giant cell carcinoma were examined ultrastructurally and immunohistochemically. Ultrastructurally, of 18 large cell carcinomas other than giant cell carcinoma eight showed characteristic differentiation toward adenocarcinoma, four toward adenosquamous carcinoma, and one each toward squamous cell carcinoma and neuroendocrine cell carcinoma, but the remaining four were undifferentiated. Six of the nine giant cell carcinomas also showed features of adenocarcinoma, two showed features of squamous cell carcinoma, and one was undifferentiated carcinoma. Immunohistochemically, secretory component (SC) was observed in seven of 14 cases with features of adenocarcinoma and two of four cases with features of adenosquamous carcinoma. Carcinomas with only squamous cell differentiation did not stain for SC. Keratin staining was positive in five of the 14 with features of adenocarcinoma, three of the four cases with features of adenosquamous carcinoma and two of the three cases with features of squamous cell carcinoma. The numbers of tumor cells positive for keratin and/or SC were small. One carcinoma with neurosecretory type granules was stained positively for calcitonin. These findings indicate that many large cell carcinomas showed differentiation toward glandular cells and/or squamous cells, and some did not show any differentiation ultrastructurally or immunohistochemically, indicating that the majority of large cell carcinomas are poorly differentiated form of either adenocarcinomas or squamous cell carcinomas.

Topics: Adenocarcinoma; Calcitonin; Carcinoma, Small Cell; Carcinoma, Squamous Cell; Cytoplasmic Granules; Gastrin-Releasing Peptide; Histocytochemistry; Humans; Immunoenzyme Techniques; Keratins; Lung Neoplasms; Peptides; Secretory Component; Staining and Labeling