The Function And Mechanism Of FAM135B In The Development Of Esophageal Squamous Cell Cancer

Esophageal cancer is the eighth most common cancer in the world with more than 450,000 new cases every year.According to pathological characteristics,esophageal cancer could be divided into two main types,esophageal squamous cell carcinoma(ESCC)and esophageal adenocarcinoma(EAC).In China,most of esophageal cancers are ESCCs.Both morbidity and cancer related death rate of ESCC ranks fourth among all cancers.Recently,taking advantage of the whole genome sequencing(WGS),whole-exome sequencing(WES)and comparative genomic hybridization(CGH)conducted in our previous work,we presented a landscape of genomic alterations in ESCC,and found that FAM135B was a new cancer related gene for the first time.This work is based on our previous achievement,and focused on the new gene FAM135B.We researched on the role and functional mechanism of FAM135B in the development and progress of ESCC,as well as its clinical significance which might provide theoretical basis for further research on clinical applications.Firstly,we compared levels of protein FAM135B in human ESCC and normal adjacent tissues by immunohistochemical analysis.The result showed that cytoplasmic level of FAM135B was significantly higher in ESCC tissues than in normal tissues,and that the level of FAM135B in ESCC tissues was correlated with patients' prognosis.Next,we constructed stable FAM135B overexpression or knockdown cell lines and verified that FAM135B promoted ESCC cells malignant phenotype,including proliferation,clone formation,migration and invasion,and resistance to cisplatin.In vivo experiment also proved the oncogenic role of FAM135B in nude mice.Next,we step forward to investigate the functional mechanism of FAM135B.We applied a 2D-LC-MS/MS based approach and identified GRN as the interacting protein of FAM135B.Signaling pathway analysis showed that overexpression of FAM135B promoted the secretion of GRN which then activated the PI3K/AKT/mTOR pathway.Eventually,FAM135B,GRN and the critical pathway formed a loop to promote the oncogenic role of each other.As a growth factor,GRN was rarely researched in ESCC.Here,we proved that GRN could also promote ESCC cells proliferation and resistance to cisplatin.More importantly,we analyzed the expression of GRN in ESCC tissues and found that GRN was significantly highly expressed in ESCC than normal tissues.What's more,patients with both FAM135B and GRN overexpression survived tremendously shorter after treatment on ESCC.Cox regression analysis indicated that overexpression of both FAM135B and GRN could be an independent prognosis factor for ESCC.Furthermore,FAM13 5B transgenic mice were generated and treated with 4-NQO to induce esophageal squamous cell carcinoma.The result showed that FAM13 5B transgenic mice bore heavier tumor burden than wild-type and survived a relatively shorter lifespan after 4-NQO treatment.FAM135B transgenic mice also showed more spontaneous tumors than wild type mice.Besides,levels of GRN in both serum of transgenic mice and medium of FAM135B overexpression cells were significantly higher than control groups,which suggested that serum GRN level may provide diagnostic or recurrent discriminations for esophageal squamous cell carcinoma patients.Taken together,these results suggested that FAM135B and GRN play critical roles in the regulation of esophageal squamous cell carcinoma and could be potential therapeutic targets and prognostic factors in ESCC.