1. Pharmacodynamics And Mechanism Of Selective S1P1 Agonist SYL927 In The Treatment Of Idiopathic Pulmonary Fibrosis 2. Pharmacodynamics And Mechanism Of Pan-PI3K Inhibitor In The Treatment Of Gastric Cancer

Idiopathic pulmonary fibrosis(IPF)is a chronic,progressive interstitial lung disease with short median survival time and less particularly effective medicines.At present,chronic inflammation and destruction of alveolar capillary endothelial barrier are closely related to the occurrence and development of IPF.Sphingosine-1-phosphate receptor 1(S1P1)is a kind of G protein coupled receptor,which plays an important role in regulating lymphocyte trafficking and vascular barrier protection.SYL927 is a novel selective S1P1 agonist synthesized by the Institute of Meterial Medica,Chinese Academy of Medical Sciences.Based on the important role of S1P1 in regulation of inflammation and vascular endothelial barrier,we conducted pharmacodynamic study and preliminary mechanism study on SYL927 in the treatment of IPF.We first established bleomycin-induced pulmonary fibrosis model in mice to investigate the therapeutic effect of SYL927 on IPF.The data showed that SYL927 could significantly improve the survival rate of mice with pulmonary fibrosis,improve kinds of lung functions,and remarkably alleviate the infiltration of inflammatory cells and collagen deposition in lung tissue.Western blot assay showed that SYL927 could significantly upregulate the expression of ZO-1 and E-cadherin,downregulate the expression of Snail and Fibronectin.Therefore,SYL927 exhibited a significant therapeutic effect on IPF,wihich may be related to its effects on anti-inflammation and alveolar capillary endothelial barrier protection.The alveolar capillary endothelial barrier consists of capillary endothelial cells,alveolar epithelial cells and their respective basement membrane.Therefore,we then investigated the effects of SYL927 on capillary endothelial cells and alveolar epithelium,respectively.The data of laser confocal microscopy in vitro showed that SYL927 could promote the formation of actin ring by inducing F-actin remodeling and could regulate the location of tight junction related protein ZO-1 on the inner side of cell membrane,which significantly promoted the tight junction of vascular endothelial cells.The study of vascular endothelial permeability through quantitative detection by FITC-BSA further showed that SYL927 could significantly inhibit the permeability of vascular endothelial cell barrier in vitro.In addition,the results of celiac capillary leakage of mice in vivo indicated that SYL927 could remarkably reduce the capillary leakage induced by acetic acid,and its efficacy was similar to that of Dexamethasone.It is suggested that SYL927 had a significant protective effect on vascular endothelial barrier.Alveolar epithelial mesenchymal transition(EMT)and vascular endothelial mesenchymal transition(EndoMT)are important sources of myofibroblasts,and are also important key factors of barrier damage.Western blot study showed that S YL927 could reverse the EMT/EndoMT induced by TGF-?1,which could upregulate the expression of of E-cadherin and downregulate the expression of Snail in A549 cell line,and significantly downregulate the expression of Fibronectin,Snail and Slug in vascular endothelial cell EA.hy926 cell line.In conclusion,this study indicated that SYL927 had a significant effect on bleomycin induced pulmonary fibrosis in mice,and its mechanism might be related to its immunosuppression effects and protection of alveolar capillary endothelial barrier,suggesting that targeting S1P1 may be a new strategy in the treatment of IPF.Gastric cancer is a kind of malignant tumor with high morbidity,especially in China,with poor prognosis and limited therapeutic drugs.PI3K/AKT signaling pathway is reported important for cell growth,proliferation,metabolism and apoptosis regulation,etc.At present,studies showed that this pathway is closely related to the occurrence and development of gastric cancer because of the overactivation,such as the PIK3CA and PTEN genes mutation as well.Hence,PI3K/AKT pathway inhibitors may be potential targets in the treatment of gastric cancer.Compounds 6a and 7a are novel Pan-PI3K inhibitors synthesized by the Institute of Meterial Medica,Chinese Academy of medical sciences.Therefore,we hope to investigate the pharmacodynamics and mechanism of 6a and 7a of gastric cancer cell HGC-27 with PIK3CA and PTEN mutation.We first investigated the inhibitory activity of 6a and 7a on HGC-27 by MTT viability assay.The data showed that compounds 6a and 7a could significantly inhibit the proliferation of HGC-27 cells in vitro.Then,the effects of 6a and 7a on cell cycle and apoptosis were detected by flow cytometry.The results showed that 6a and 7a could arrest HGC-27 cell cycle in GO/G1 phase and promote apoptosis.Western blot analysis showed that 6a and 7a could effectively inhibit the expression of P-AKT(S473),P-S6RP and P-4E-BP so to effectively inhibit the activation of PI3K/AKT pathway.Based on the above results,the xenograft model of HGC-27 cell line was established to further explore the inhibitory activity of compounds 6a and 7a on gastric cancer in vivo.The results showed that compounds 6a(40 mg/kg)and 7a(20/40 mg/kg)could significantly inhibit tumor growth,and the tumor inhibition rate of 7a(40 mg/kg)was up to 88.67%.Immunohistochemistry of tumor tissue showed that compounds 6a(40 mg/kg)and 7a(20/40 mg/kg)could remarkably inhibit the expression of tumor proliferation marker Ki67 and P-AKT(S473)in tumor tissues.In conclusion,this study showed that Pan-PI3K inhibitors 6a and 7a could effectively inhibit the proliferation of gastric cancer cell HGC-27 in vivo and vitro by inhibiting PI3K/Akt/mTOR signaling pathway,and induced HGC-27 cells G0/G1 phase arrest and apoptosis.