A Novel Small Molecule YJ-4-135 Inhibits Pancreatic Cancer Metastasis By Targeting EP4 Receptor

Pancreatic cancer is an extremely malignant digestive tract tumor,with a 5-year survival rate of approximately 9%.Most pancreatic cancer patients are diagnosed with distant metastasis in the advanced stage due to an inability to detect the disease early.Few shows a significantly response to surgery and radiation or chemotherapy therapy which ultimately leads to poor prognosis and death.Previous studies have shown that prostaglandin E₂（PGE₂）produced by COX2 promotes the migration of pancreatic cancer cells in vitro.The EP4receptor,a main prostaglandin receptor,overexpresses in pancreatic cancer and contributes to many kind of tumor metastasis.However,the role and mechanism of EP4 receptor in pancreatic cancer metastasis remains unknown.In this study we found that interfereing with EP4 receptor in pancreatic cancer inhibited PGE₂-induced cell migration.Therefore,we proposed the idea of developing small molecule EP4 receptor antagonists targeting pancreatic cancer metastasis.A novel small molecule compound YJ-4-135 could specifically inhibit human EP4 receptor with better activity(IC₅₀=0.98 n M)than E7046(IC₅₀=10.64 n M).Meanwhile it could inhibit monkey,mouse and rat EP4 receptors.Therefore,YJ-4-135 is a highly selective and potent EP4 receptor antagonist.Further,subsequent series of evaluation systems,including glosensor,CRE reporter gene,β-Arrestin assay and phosphorylation of ERK1/2 verified that YJ-4-135 could inhibit the accumulation of c AMP,EP4/Arrestin interaction and phosphorylation of ERK1/2.Then we found that YJ-4-135 inhibited the migration of pancreatic cancer cells in dose-dependent manner as determined by wound healing and Transwell in vitro.We further built mouse liver metastasis model of pancreatic cancer to evaluate efficacy of YJ-4-135 in vivo.Our results showed YJ-4-135（75 mg/kg）inhibited liver metastasis of pancreatic cancer as well as slowed down the loss of mice weight.Additionally,the drug pairs of YJ-4-135 and gemcitabine showed a strong synergy and significantly improve the survival rate.To explore the underlying mechanism of YJ-4-135 in inhibition of pancreatic cancer metastasis,we discovered YJ-4-135 could diminish the expression of YAP and the transcriptional activity of TEADs.We further found YJ-4-135 inhibited the Hippo pathway by decreasing the activity of PKA and phosphorylation of ERK.In addition,YJ-4-135 could restore the transcription of metastasis related gene（Vimentin,Snai1 and E-cadherin）of YAP downstream induced by PGE₂.Together,these results revealed a mechanism for YJ-4-135 in the regulation of pancreatic cancer metastasis through blocking EP4/Hippo pathway.In conclusion,we found a novel high-selective and potent EP4 receptor antagonist YJ-4-135,which can inhibit pancreatic cancer metastasis in vitro and vivo.We further found that YJ-4-135 inhibit EP4-mediated Hippo signaling pathway activation.In summary,we have identified that EP4 receptor is an important regulator for pancreatic cancer metastasis,providing a new target for pancreatic cancer treatment.