Autocrine Extra-Pancreatic Trypsin 3 Secretion Promotes Cell Proliferation and Survival in Esophageal Adenocarcinoma

Abstract

Trypsin or Tumor associated trypsin (TAT) activation of Protease-activated receptor 2 (PAR-2) promotes tumor cell proliferation in gastrointestinal cancers. The role of the trypsin/PAR-2 network in esophageal adenocarcinoma (EA) development has not yet been investigated. The aim of this study is to investigate the role of trypsin/PAR-2 activation in EA tumorogenesis and therapy. We found that esophageal adenocarcinoma cells (EACs) and Barrett’s Metaplasia (BART) expressed high levels of type 3 extra-pancreatic trypsinogen (PRSS3), a novel type of TAT. Activity of secreted trypsin was detected in cultured media from EA OE19 and OE33 cultures but not from BART culture. Surface PAR-2 expression in BART and EACs was confirmed by both flow cytometry and immunofluorescence. Trypsin induced cell proliferation (∼ 2 fold; P<0.01) in all tested cell lines at a concentration of 10 nM. Inhibition of PAR-2 activity in EACs via the PAR-2 antagonist ENMD (500 µM), anti-PAR2 antibody SAM-11 (2 µg/ml), or siRNA PAR-2 knockdown, reduced cell proliferation and increased apoptosis by up to 4 fold (P<0.01). Trypsin stimulation led to phosphorylation of ERK1/2, suggesting involvement of MAPK pathway in PAR-2 signal transduction. Inhibition of PAR-2 activation or siRNA PAR-2 knockdown in EACs prior to treatment with 5 FU reduced cell viability of EACs by an additional 30% (P<0.01) compared to chemotherapy alone. Our data suggest that extra-pancreatic trypsinogen 3 is produced by EACs and activates PAR-2 in an autocrine manner. PAR-2 activation increases cancer cell proliferation, and promotes cancer cell survival. Targeting the trypsin activated PAR-2 pathway in conjunction with current chemotherapeutic agents may be a viable therapeutic strategy in EA.

Inhibition of autophagosome-lysosome fusion by ginsenoside Ro via the ESR2-NCF1-ROS pathway sensitizes esophageal cancer cells to 5-fluorouracil-induced cell death via the CHEK1-mediated DNA damage checkpoint

“…43 Consistently, we found that ECA-109 and TE-1 cells are more resistant to 5-Fu than other chemosensitive esophageal cancer cells such as OE19 and OE33 cells (Fig. S3A) [44][45][46] and that 5-Fu induces prosurvival autophagy, and caspase-independent and autophagy-associated cell death (Fig. 5D and 6C).…”

“…S3A), which was consistent with previous reports. 44,45 Unexpectedly, these cells were more 5-Fu-resistant than other chemosensitive cells such as OE19 (IC 50 = 3.03 μg/ml) and OE33 (IC 50 = 1.24 μg/ml), 46 implying that ECA-109 and TE-1 cells were 5-Fu-resistant. A recent study showed that 5-Fu-induced cell death in chemoresistant esophageal cancer cells is apoptosis-independent, and that autophagy exerts a protective effect.…”

Re: Shi et al. Protease‐activated receptor 2 suppresses lymphangiogenesis and subsequent lymph node metastasis in a murine pancreatic cancer model. J Pathol 2014;234: 398‐409

“…In addition to the well-known role of PAR-2 in the primary proliferation of various cancer cells [2,3], including pancreatic cancer [4], the results presented highlighted convincingly the importance of stromal host receptors relative to local progression in the orthotopic pancreatic cancer model. The authors recorded an increase in the average volume of primary tumours in wild-type animals compared with PAR-2 −/− knockouts.…”

PRSS3 expression is associated with tumor progression and poor prognosis in epithelial ovarian cancer

“…One study has proposed that PRSS3 serves as a robust growth factor for HT29 colon cancer cells via the activation of PAR2 and the downstream phosphorylation of ERK, which results in cell proliferation [26]. Another significant study suggested that secreted PRSS3 acts on the PAR2 receptor as an autocrine survival signal; targeting this axis increased sensitivity to chemotherapy in esophageal adenocarcinoma cell lines [27]. Similarly, PRSS3 regulation of cellular adhesion and proliferation mediated by PAR2/G-protein signaling has been reported in other malignancies, such as breast cancer and gastric cancer [28,29].…”