trypsinogen and Neoplasm-Metastasis

Serine protease 2 (PRSS2) is upregulated in gastric cancer tissues, correlates with poor prognosis and promotes migration and invasion of gastric cancer cells. However, the exact mechanism by which PRSS2 promotes metastasis in gastric cancer is unclear. We examined serum PRSS2 levels in healthy controls and gastric cancer patients by enzyme linked immunosorbent assay (ELISA) and analyzed the correlation between PRSS2 serum level with the clinicopathological characteristics of gastric cancer patients and matrix metalloproteinase-9 (MMP-9) expression. A lentiviral MMP-9 overexpression vector was constructed and used to transfect gastric cancer cells with stable silencing of PRSS2, and migration, invasion and epithelial-mesenchymal transition (EMT) of gastric cancer cells were examined. High serum PRSS2 levels were detected in gastric cancer patients and associated with lymphatic metastasis and TNM stage. Serum PRSS2 was positively correlated with serum MMP-9 level. PRSS2 silencing inhibited EMT, and knock-down of PRSS2 partially abrogated cell metastasis and EMT caused by overexpression of MMP-9. These results suggest that PRSS2 promotes the migration and invasion of gastric cancer cells through EMT induction by MMP-9. Our findings suggest that PRSS2 may be a potential early diagnostic marker and therapeutic target of gastric cancer.

Topics: Cell Line, Tumor; Cell Movement; Epithelial-Mesenchymal Transition; Gene Expression Regulation, Neoplastic; Humans; Lymphatic Metastasis; Matrix Metalloproteinase 9; Neoplasm Invasiveness; Neoplasm Metastasis; Peptide Hydrolases; Stomach Neoplasms; Trypsin; Trypsinogen

Proteolytic enzymes have shown efficacy in cancer therapy. We present a combination of the two pro-enzymes Trypsinogen and Chymotrypsinogen A with potent in vitro and in vivo anti-tumour efficacy. A synergetic anti-tumour effect for Trypsinogen and Chymotrypsinogen A was determined at a ratio 1:6 (named PRP) using 24 human cancer cell lines. The antiangiogenic effect of PRP was analysed by matrigel-based tube formation and by fibrous capsule formation assays. Furthermore, cell invasion and wound healing assays together with qRT-PCR determination of epithelial-to-mesenchymal transition (EMT) markers were performed on human cancer cells treated with PRP. Additionally, in vivo pharmacokinetic studies were implemented and the PRP's anti-tumour efficacy was explored against orthotopic pancreatic and ovarian cancer tumours. PRP formulation was proven to inhibit in vitro angiogenesis, tumour growth, cancer cell migration and invasiveness; and to be an effective and well tolerated in vivo anti-tumour treatment. Finally, the clinical efficacy of a suppository formulation containing both pancreatic pro-enzymes in the context of a UK Pharmaceuticals Special Scheme was evaluated in advanced cancer patients. Consequently, PRP could have relevant oncological clinical applications for the treatment of advanced or metastatic pancreatic adenocarcinoma and advanced epithelial ovarian cancer.

Topics: Animals; Apoptosis; Cell Proliferation; Chymotrypsinogen; Enzyme Precursors; Female; Humans; Mice; Mice, Nude; Neoplasm Metastasis; Ovarian Neoplasms; Pancreas; Pancreatic Neoplasms; Pilot Projects; Trypsinogen; Tumor Cells, Cultured; Xenograft Model Antitumor Assays

Activity-based protein profiling has emerged as a valuable technology for labeling, enriching, and assessing protein activities from complex mixtures. This is primarily accomplished via a two-step identification and quantification process. Here we show a highly quantitative and streamlined method, termed catch-and-release activity profiling of enzymes (CAPE), which reduces this procedure to a single step. Furthermore the CAPE approach has the ability to detect small quantitative changes that may have been missed by alternative mass spectrometry-based techniques.

Topics: Amino Acid Sequence; Animals; Cattle; Cell Line, Tumor; Humans; Isotope Labeling; Mass Spectrometry; Molecular Sequence Data; Neoplasm Metastasis; Neoplasm Proteins; Organophosphonates; Peptides; Trypsin; Trypsinogen

Although it is accepted that regulatory T cells (T regs) contribute to cancer progression, most studies in the field consider nonantigen-specific suppression. Here, we show the presence of tumor antigen-specific CD4(+) T regs in the blood of patients with metastatic melanoma. These CD4(+) T regs recognize a broad range of tumor antigens, including gp100 and TRP1 (melanoma tissue differentiation antigens), NY-ESO-1 (cancer/testis antigen) and survivin (inhibitor of apoptosis protein (IAP) family antigen). These tumor antigen-specific T regs proliferate in peripheral blood mononuclear cells (PBMC) cultures in response to specific 15-mer peptides, produce preferentially IL-10 and express high levels of FoxP3. They suppress autologous CD4(+)CD25(-) T cell responses in a cell contact-dependent manner and thus share properties of both naturally occurring regulatory T cells and type 1 regulatory T cells. Such tumor antigen-specific T regs were not detected in healthy individuals. These tumor antigen-specific T regs might thus represent another target for immunotherapy of metastatic melanoma.

Topics: Adult; Aged; Antigens, Neoplasm; CD4-Positive T-Lymphocytes; Female; Forkhead Transcription Factors; gp100 Melanoma Antigen; Humans; Inhibitor of Apoptosis Proteins; Interleukin-10; Interleukin-2 Receptor alpha Subunit; Leukocytes, Mononuclear; Male; Melanoma; Membrane Glycoproteins; Membrane Proteins; Microtubule-Associated Proteins; Middle Aged; Neoplasm Metastasis; Neoplasm Proteins; Survivin; T-Lymphocytes, Regulatory; Trypsin; Trypsinogen

Distant metastasis is the predominant cause of death in early-stage non-small cell lung cancer (NSCLC). Currently, it is impossible to predict the occurrence of metastasis at early stages and thereby separate patients who could be cured by surgical resection alone from patients who would benefit from additional chemotherapy. In this study, we applied a comparative microarray approach to identify gene expression differences between early-stage NSCLC patients whose cancer ultimately did or did not metastasize during the course of their disease. Transcriptional profiling of 82 microarrays from two patient groups revealed differential expression of several gene families including known predictors of metastasis (e.g., matrix metalloproteinases). In addition, we found S100P, S100A2, trypsinogen C (TRY6), and trypsinogen IVb (PRSS3) to be overexpressed in tumors that metastasized during the course of the disease. In a third group of 42 patients, we confirmed the induction of S100 proteins and trypsinogens in metastasizing tumors and its significant correlation with survival by real-time quantitative reverse transcription-PCR. Overexpression of S100A2, S100P, or PRSS3 in NSCLC cell cultures led to increased transendothelial migration, corroborating the role of S100A2, S100P, and PRSS3 in the metastatic process. Taken together, we provide evidence that expression of S100 proteins and trypsinogens is associated with metastasis and predicts survival in early stages of NSCLC. For the first time, this implicates a role of S100 proteins and trypsinogens in the metastatic process of early-stage NSCLC.

Topics: Carcinoma, Non-Small-Cell Lung; Cell Movement; Gene Expression Regulation, Neoplastic; Humans; Lung Neoplasms; Neoplasm Metastasis; Oligonucleotide Array Sequence Analysis; Predictive Value of Tests; S100 Proteins; Trypsinogen

Trypsinogen/trypsin is one of the major serine proteases and is produced by pancreatic acinar cells. Tumor-associated trypsinogen (TAT) has been reported to be produced by several cancer cell lines. The biological roles and activation mechanisms of both TAT and pancreatic acinar trypsinogen (PAT) have not been elucidated in the context of cancer extension, in particular at the stage of invasion and metastasis. In this study, we investigate the roles played by PAT and TAT in pancreatic cancer invasion. In addition, we determined their mechanisms of activation and identified a trypsinogen activity-stimulating factor (TASF) produced by pancreatic cancer cells. TAT expression and high TAT activity were associated with high invasive and liver metastatic potential in SW1990 and CAPAN-2 cells. Moreover, a trypsinogen activating effect and activity prolonging effect was observed in a mixture of these supernatants with trypsinogen. These cells revealed significantly enhanced invasiveness upon invasion assay and in the presence of PAT. TAT and PAT were activated by TASF, active u-PA, produced by pancreatic cancer cells. Activated TAT and PAT can degrade not only ECM proteins but they can also activate other latent proteases. This ECM-protease-network may form a vicious cycle, thereby promoting tumor cell invasion.

Topics: Enzyme Activators; Humans; Immunoblotting; Liver Neoplasms; Neoplasm Invasiveness; Neoplasm Metastasis; Pancreatic Neoplasms; Reverse Transcriptase Polymerase Chain Reaction; Trypsinogen; Urokinase-Type Plasminogen Activator

Trypsinogen (TRY), the precursor to the serine protease trypsin, is found in the pancreas and mediates digestive proteolysis in the small intestine. Differential display of cDNAs expressed by human colorectal tumor tissues compared with adjacent normal colonic mucosa identified an isoform of TRY (TRY2) up-regulated in colorectal cancers. Northern blot analysis of RNA isolated from a series of 28 malignant colon tumors and corresponding normal mucosa showed that TRY transcripts were up-regulated 2- to 33-fold in 29% of tumors. Further, TRY mRNA was expressed in 6 colorectal cancer cell lines, with highest levels detected in the metastatic tumor lines SW620 and HT29. Immunostaining for TRY protein expression showed intense immunoreactivity in the supranuclear cytoplasm of colon tumors in 16% of tissue specimens. To evaluate the relative contributions of 2 isoforms of TRY, TRY1 and TRY2, to total TRY mRNA expression, a semi-quantitative multiplex RT-PCR assay was developed. TRY2 mRNA was detected in all 6 colorectal tumor cell lines, whereas TRY1 mRNA was expressed only in the metastatic tumor lines, showing that the high levels of TRY expression in the metastatic tumor lines are likely due to up-regulation of TRY1. Evaluation of TRY1 and TRY2 mRNA expression by multiplex RT-PCR in a series of 20 colon tumor tissues representative of the range of tumor progression showed that TRY2 mRNA was expressed much more commonly than TRY1 mRNA in normal mucosa (26% vs. 6%) as well as in primary tumor tissues (65% vs. 15%). These data demonstrate that TRY2 is the dominant TRY in colon tissue and suggest that up-regulation of TRY1 expression in colon tumors may be associated with a metastatic phenotype.

Topics: Colorectal Neoplasms; Gene Expression Profiling; Gene Expression Regulation, Enzymologic; Gene Expression Regulation, Neoplastic; Humans; Neoplasm Metastasis; Neoplasm Staging; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Transcription, Genetic; Trypsin; Trypsinogen; Tumor Cells, Cultured

The purpose of this study was to evaluate the trypsinogen expression in human colorectal cancers. We observed the trypsinogen-1 mRNA expression in five of ten human colorectal cancer cell lines by reverse transcriptase-polymerase chain reaction (RT-PCR), and gelatinolytic activity in conditioned medium of cancer cells at acidic pH by gelatin zymography. Furthermore, trypsinogen protein expression was observed in 68 out of 154 (44.2%) surgical specimens of colorectal cancer immunohistochemically. However, there was no statistically significant relationship between the trypsinogen expression and the clinicopathological findings. These findings suggest that trypsinogen might be involved in cancer invasion and metastasis throughout the cancer progression.

Topics: Colorectal Neoplasms; Culture Media, Conditioned; Gelatin; HT29 Cells; Humans; Hydrogen-Ion Concentration; Immunohistochemistry; Neoplasm Metastasis; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Trypsinogen; Tumor Cells, Cultured

Pancreatic carcinomas electively induced by immunological mechanism [(1), (2)] keep their exocrine secretory specificity along the various stages of their evolution: (a) during the transformation phase from adenoma to carcinoma; (b) in the evolved carcinoma; (c) in its metastasis; (d) in the ascitic carcinomatous cells formed. They are called: immuno-inducted carcinoma. The carcinomatous cells of the constantly deadly ascites cease their production of secretion granules after passages by intraperitoneal graft; but this secretion reappears in the solid carcinomas they induce by subcutaneous graft and contains trypsinogen and chymotrypsinogen, even after the 92nd passage, at the 757 day. Besides, the antisera (1) enhance the growth and the affinity for pancreas and adipose tissue of the carcinomatous ascitic strains they induced. They, sometimes, produce nodular hepatic carcinomas.

Topics: Antibodies; Ascites; Carcinoma; Chymotrypsinogen; Lipoprotein Lipase; Lymph Nodes; Lymphatic Metastasis; Neoplasm Metastasis; Neoplasm Transplantation; Neoplasms, Experimental; Pancreatic Neoplasms; Trypsinogen