CD148 Serves As S Prognostic Marker Of Gastric Cancer And Hinders Tumor Progression By Dephosphorylating EGFR

Objective:Gastric cancer(GC)is the fourth most common malignancies and the second-leading cause of cancer-related deaths worldwide.Difficulties for early detection,high rates of recurrence and/or distant metastasis and poorly responds to the traditional radiotherapy/chemotherapy lead to a poor overall prognosis of GC patients.The median overall survival of patients with distant metastasis is less than 12 months.At present,compared to the conventional therapy methods such as surgery,chemotherapy and radiotherapy,targeting therapy and immune therapy that have favorable effects in other malignancies have showed limited survival benefits for advanced GC patients.Therefore,understanding of the molecular mechanisms underlying the oncogenesis and progression is critical for identifying early detection markers and candidate intervention targets to improve the overall survival and life quality in patients with GC.By dephosphorylating target proteins,PTPRs(protein tyrosine phosphatase receptors)could potentially participate in reducing signaling transductions and influencing the malignant biological processes of tumors.CD148,also called DEP-1 or PTPRJ,is a member of receptor-type Protein Tyrosine Phosphatase family encoded by PTPRJ gene and has controversial impacts on signaling transductions in cancers via dephosphorylating proteins that play vital roles in the etiology of tumors,such as EGFR,ERK1/2,VEGFR,SRC and so on.Previous researches have shown controversial effects in specific tumors.On one hand,CD 148 inhibited the proliferation and metastasis of colon cancer cells,the growth of breast or thyroid cancer cells.On the other hand,CD148 boost breast cancer progression through promoting migration or invasion of cells.The role of CD 148 in gastric cancer has not been reported yet.Besides,the mechanism of aberration of CD148 in malignancies is still largely unclear.Although previous papers have reported that miRNA-328 and miRNA-155 negatively regulated the expression of CD148 and consequently influenced the malignant processes in cancers,the mechanism regulating the expression of CD 148 in GC is worthy of further investigation.Our research focused on the expression of CD148 in GC and its possible upstream mechanism,how CD148 regulated the malignant biological behaviors and its downstream mechanism,which may help to provide new strategies for early diagnosis and intervention.Methods:1.IHC staining was used to detect the expression of CD148 of the tissues from 109 GC patients and subsequently to analysis the correlation between the expression of CD 148 and features of clinicopathology,recurrence-free survival and overall survival;2.Construction of CD148 knockdown/knockout/overexpression in BGC,MKN45,SGC cell lines by transfected with siRNA/CRISPER/Cas9/overexpression plasmid and validated via western blotting;3.Investigation of the impacts of knockdown/overexpression of CD148 on proliferation,motility and invasiveness of GC cells in vitro;4.Investigation of the influences of knockout of CD148 on sizes and weights of the subcutaneous transplantation of tumors;5.Western blotting was used to validate whether CD148 regulate the phosphorylation of EGFR and downstream PI3K/AKT and MEK/ERK pathways;6 In silico analysis revealed the potential upstream mechanism resulting in the depression of CD 148 expression.Results:1.Reduced CD148 expression indicates adverse pathological features of GC,including tumor stages,lymph node or distant metastases,vascular invasions,and differentiations,as well as shorter survival;2.Overexpression of CD 148 impeded tumor proliferation,motility,and invasiveness,while CD148 knock-down promoted the ability of gastric cancer cells to grow and metastasize in vitro;3.Knockout of CD148 promoted the growth of subcutaneous transplantation of tumors in vivo;4.Overexpression of CD 148 negatively regulated EGFR phosphorylation of multiple tyrosine residues,including Y1173,Y1068,and Y1092,and downstream phosphorylation of PI3K,AKT,MEK1/2,ERK1/2,but did not change the total expression of EGFR;CD148 knocked-down in vitro or knocked-out in vivo upregulated EGFR phosphorylation of multiple tyrosine residues,including Y1173,Y1068,and Y1092,and downstream phosphorylation of PI3K,AKT,MEK1/2,ERK1/2,but did not change the total expression of EGFR;5;Bioinformatics analysis revealed a 3’UTR-specific methylation may regulate CD148 expression and the potential regulators are TET2 and TET3.Conclusion:Reduced expression of CD148 in GC correlated with poor RFS and OS.CD148 inhibited the activation of PI3K/AKT and MEK/ERK pathway through dephosphorylating EGFR and consequently suppressed the malignant processes such as proliferation,migration and invasion.CD 148 is a convincing prognostic marker as well as a potentially therapeutic target for gastric cancer.