Enhanced invasiveness of pancreatic adenocarcinoma cells stably transfected with cationic trypsinogen cDNA

Abstract

Various studies have described increased expression of cationic trypsinogen in malignant tumor cells. To explore the role of secreted cationic trypsinogen in invasion by cancer cells, we introduced cationic trypsinogen cDNA into Panc-1, a pancreatic adenocarcinoma-derived cell line that lacks expression of endogeneous trypsinogen. Four independent clones (designated Panc-1-Try-7, -9, -11 and -24) stably expressing cationic trypsinogen mRNA were isolated and processed for further study. In a zymographic analysis, gelatinolytic activity for cationic trypsinogen was detectable in serum-free conditioned media obtained from all 4 transfectants but not in media from mock-transfected or parental Proteolytic degradation of extracellular matrix components is a process essential for tumor invasion and metastasis. 1-4 A variety of proteases produced by tumor cells are involved in matrix degradation. 1-4 These enzymes include 2 major categories: matrix metalloproteinases (MMPs) and serine proteases. Although the role of MMPs in tumor malignancy has been studied extensively, 5-9 much less is known about serine proteases, except for urokinase-type plasminogen activator (u-PA). 10 -13 Although trypsin, another member of the serine protease family, also has been implicated in the spread of malignant tumor cells, the underlying molecular mechanism has not been fully elucidated. Several previous reports have described increased amounts of trypsinogen-like enzymes in malignant tumor cells. 14 -21 Koshikawa et al. 22 identified 1-and 2-chain forms of cationic trypsinogen in a gastric carcinoma cell line. Under the acidic condition that prevails in the extracellular space between malignant tumor cells, cationic trypsinogen is spontaneously converted to trypsin, an active form. 23 Trypsin not only degrades extracellular matrix glycoproteins 22 but also activates proenzymes of various MMP and serine proteases. 19 We previously reported that under acidic conditions cationic trypsinogen produced by human pancreatic ductal cancer spontaneously becomes activated and acquires gelatinolytic activity. 24 Furthermore, we found that serine protease inhibitors had a suppressive effect on invasion and metastasis by human pancreatic cancer cell lines. 25 However, the importance of trypsin in initiating invasion of tumor cells is still unclear; 1 major reason is that no available serine protease inhibitor can produce sufficiently selective inhibition of trypsin.To obtain more direct evidence that trypsin potentiates tumor invasion, we transfected cationic trypsinogen cDNA into Panc-1, a pancreatic adenocarcinoma-derived cell line that normally is deficient in expression of trypsinogen. The resulting transfectants acquired enhanced invasiveness. MATERIAL AND METHODS Cell cultureHuman pancreatic adenocarcinoma cell lines, (Capan-1, BxPC-3, AsPC-1, Panc-1 and MIAPaCa-2), were obtained from the American Type Culture Collection (Rockville, MD). These cell lines were maintained at 37°C in a 5% CO 2 incubator and grown in RPMI-1640 medium supplemented w...

Tumor-derived trypsin enhances proliferation of intrahepatic cholangiocarcinoma cells by activating protease-activated receptor-2

“…5). This band was consistent with that reported for the activated form of trypsinogen-1 (14,16). In addition, the supernatant exhibited gelatinolytic activity with bands at ~72 and 92 kDa.…”

“…Active trypsin, produced and secreted by cancer cells, plays an essential role in facilitating invasion and metastasis by digesting components of the extracellular matrix (8)(9)(10)(11)(12)(13)(14)16,17). At the same time, the contribution of PAR-2 and the effect of active trypsin on signaling pathways influencing malignant tumors are still being studied (22)(23)(24)(25)(26)(27).…”

“…Under acidic conditions that prevail in the extracelluar space between malignant tumor cells, trypsinogen-1 is spontaneously converted to trypsin (15). We previously reported that when a pancreatic cancer cell line that lacked expression of trypsinogen mRNA was transfected with trypsinogen-1 cDNA, the cells secreted trypsinogen and acquired significantly greater invasive ability (16). In addition, we demonstrated in pancreatic cancer cell lines that hepatic metastasis and invasive ability were correlated with the amount of trypsinogen-1 and were inhibited by a serine protease inhibitor (17 plays an essential role in facilitating the invasion and metastasis of the cancer cells.…”

Angiotensin II induces tumor progression and fibrosis in intrahepatic cholangiocarcinoma through an interaction with hepatic stellate cells

“…Human pancreatic cancer cells expressed and secreted pancreatic cationic-type trypsinogen in vitro, which is spontaneously converted into active trypsin at acidic pH (pH 4.5-5.5), in contrast to anionic-type trypsinogen, which is not. Cationic-type trypsinogen could increase the invasive ability and cell proliferation of pancreatic cancer cells (3). In previous reports, it was described that the high expression of a specific G-protein-coupled receptor, protease-activated receptor-2 (PAR-2), was observed in pancreatic cancer cells and fibroblasts around tumor tissue (4,5).…”

Bryostatin I inhibits growth and proliferation of pancreatic cancer cells via suppression of NF-κB activation

“…It is known that activation of NF-κB is inhibited during the process of inflammation [20,21]. The chemotherapeutic inhibition of NF-κB expression prevents proliferation, invasion and metastasis of cells in various types of cancers [22][23][24]. Although, a number of agents have been identified which play an important role in the inhibition of NF-κB expression.…”