Knockdown of EPCR inhibits the proliferation and migration of human gastric cancer cells via the ERK1/2 pathway in a PAR-1-dependent manner

Abstract

Endothelial protein C receptor (EPCR) has been implicated in the carcinogenesis of diverse tumor types. This tumor-promoting effect of EPCR is associated with the upregulation of activated protein C and the activation of protease-activated receptor 1 (PAR-1). However, the exact role of EPCR in gastric cancer (GC) and the mechanisms underlying the regulation of EPCR remain elusive. In the present study, we investigated the effects of EPCR on human GC cells, as well as the underlying mechanisms. An siRNA inference system was used to knock down the expression of EPCR in GC cells, and CCK-8, colony formation and Transwell assays were performed to determine the effects of EPCR knockdown on the proliferation and migration of the tumor cells. Additionally, cell cycle distribution and apoptosis were assessed by flow cytometry, and activated PAR-1 levels were determined by cell ELISA. The results indicated that the proliferation, clonogenicity and migration were significantly reduced and that the cell cycle was arrested in the Gap 1 phase by EPCR knockdown in SGC7901 and AGS cells. Meanwhile, apoptosis was promoted by EPCR knockdown in the two cell lines. The activation of PAR-1 on the cell surface of SGC7901 and AGS cells was significantly reduced after the knockdown of EPCR. By contrast, blockade of PAR-1 reduced the proliferation and migration of gastric cells in vitro. Additionally, after the knockdown of EPCR or treatment with PAR-1 antibody, the expression of pERK1/2 was significantly downregulated in the SGC7901 and AGS cells, while the expression levels of p-AKT (S473) and p-AKT (T308) were unchanged. The findings of the present study demonstrated that EPCR exerts pro-carcinogenic effects in GC cells in a PAR-1-dependent manner via the ERK1/2-MAPK pathway. Thus, EPCR may be a potential molecular diagnostic or therapeutic target for GC.

A recombinant signalling‐selective activated protein C that lacks anticoagulant activity is efficacious and safe in cutaneous wound preclinical models

“…Furthermore, CRISPR knockout of EPCR reduces human trophoblast proliferation and invasion, 46 and siRNA knockdown of EPCR in human gastric cancer cells inhibits proliferation and migration. 47 3K3A-APC also increased activation of MMP-2 in mouse fibroblasts, as previously demonstrated by APC In human fibroblasts. 12 That it did so in both phenotypes may be explained by our previous finding that APC can directly activate MMP-2 even in the absence of cells, 32 suggesting 3K3A-APC similarly activates MMP-2 suspended in media without the need for surface membrane proteins such as EPCR.…”

Ankyrin‐repeat and SOCS box‐containing protein 9 (ASB9) regulates ovarian granulosa cells function and MAPK signaling

“…The localization of prothrombin and PAR1 in GC suggests that these factors may be important mediators of cellular function in the ovarian follicle (Roach et al, 2002). Similarly, it has been reported that in human gastric cancer cells, in vitro knockdown of EPCR inhibits cell proliferation and migration by inhibiting the activation of PAR1 and MAPK3/1 (Q. Wang et al, 2018). Also, EPCR knockdown or treatment with PAR1 antibody significantly decreased MAPK3/1 phosphorylation (Q. Wang et al, 2018).…”

“…Similarly, it has been reported that in human gastric cancer cells, in vitro knockdown of EPCR inhibits cell proliferation and migration by inhibiting the activation of PAR1 and MAPK3/1 (Q. Wang et al, 2018). Also, EPCR knockdown or treatment with PAR1 antibody significantly decreased MAPK3/1 phosphorylation (Q. Wang et al, 2018). In ovalbumin‐allergic rats, thrombin promotes airway remodeling via PAR1, while MAPK3/1 signaling pathway plays an important role in this process since pMAPK3/1 inhibitors effectively inhibit the airway remodeling in these rats (Bi et al, 2015).…”

“…Cleavage of the N‐terminus unmasks a tethered ligand that binds to the receptor and triggers intracellular signaling (Heuberger & Schuepbach, 2019; Vu et al, 1991). Activation of PAR1 by thrombin has been shown to lead to the activation of MAPK3/1 signaling pathway resulting in increased proliferation (H. Wang et al, 2002), while inhibition of the activation of PAR1 and MAPK3/1 inhibits cell proliferation and migration (Q. Wang et al, 2018). Although the molecular mechanisms of the interaction between ASB9 and PAR1 need to be further investigated and detailed, we could nonetheless hypothesize that ASB9 could negatively affect MAPK3/1 phosphorylation through binding to PAR1 followed by PAR1 degradation via the ubiquitination/proteasome pathway, which would lead to proliferation blockade and initiation of GC differentiation (Figure 7).…”

Endothelial Protein C Receptor (EPCR), Protease Activated Receptor-1 (PAR-1) and Their Interplay in Cancer Growth and Metastatic Dissemination

“…There are contradictory findings that are of paramount importance with respect to the role of the APC/EPCR/PAR-1 axis in cancer progression [42,75,76]. Wang et al [42] demonstrated that gastric cancer tissue expresses elevated levels of EPCR and that EPCR-mediated APC activation induces proliferation and migration of the MGC803 gastric cancer cells.…”