Pan-cancer Analysis Of FYN And Low FYN Expression Inhibited Proliferation Of Daudi Cells By Akt/mTOR

[Objective] 1.Pan-cancer analysis was carried to reveal the position of FYN as an important immunomodulatory factor in tumors,which provides a new target for tumor prognosis prediction and immunotherapy.2.Daudi cell lines with stable and low expression of FYN were constructed to observe the effects of interferencing with the expression of FYN on the proliferation of Daudi cells.The study preliminarily explored the regulatory relationship between FYN and Akt/mTOR network regulation,provided experimental basis for the study on the molecular mechanism of Epstein-Barr virus(EBV)-related tumors.[Methods] 1.The relevant information of 33 common tumors was obtained from GEO database,the pan-cancer analysis were performed using Sanger Box online analysis tool on the relation of the expression of FYN in tumors and prognosis and immunity of tumors.2.Bioinformatics methods were used to analyze the correlation between the expression of FYN and clinicopathological data from GSE31312 data sets in diffuse large B lymphoma(DLBCL)patients.3.The expression of FYN was detected by RT-q PCR and Western blot in Burkitt lymphoma cell lines such as Raji cells,Daudi cells and Ramos cells.4.Through "enzyme digestion-ligation-transformation-screening of positive clones",the interference vector with p LKO-FYN-sh RNA was successfully constructed by Tet-p LKO-puro vector and FYN sh RNA interference sequence.The p LKO-FYN-sh RNA vector was packaged and transferred into 293 T cells to obtain the virus supernatant,which was transfected into Daudi cells to establish Daudi cell lines with stable and low expression of FYN.5.The interference effect of FYN was verified by RT-q PCR and Western Blot.6.CCK-8 assay and soft AGAR clone formation assay were used to observe the effect of low expression of FYN on the proliferation of EBV positive lymphoma Daudi cell line.7.Western Blot was used to detect the expression of Proliferating Cell Nuclear Antigen(PCNA)and the protein regulated by Akt/mTOR network.[Results]: 1.FYN showed low expression in lots of tumors.FYN was associated with survival periods in only a few tumor types.The expression of FYN was associated with tumor mutation load and microsatellite instability in a variety of tumors.The expression of FYN is significantly associated with multiple immunoregulatory factors in a variety of tumors.2.GEO data analysis showed that in patients with DLBCL lymphoma(from GSE31312),the Overall survival time(OS)and Progression-free survival time(PFS)were longer in patients with high expression of FYN.The expression of FYN was correlated with age,IPI score,and presence of extraordinal invasion(P=0.0096,P=0.0129,P=0.0115,respectively),but not with gender,Ann Arbor stage,LDH,and gene expression profile(P=0.9696,P=0.5814,P=0.1100,P=0.1974,respectively).3.The expression level of FYN was lower in Raji and Daudi cells with EBV positive lymphoma than Ramos cells with EBV negative lymphoma.4.Interfering with the expression of FYN can inhibit the proliferation of Daudi cells.5.The expression of PCNA was down-regulated in Daudi cells with low expression of FYN.6.The low expression of FYN suppressed the regulation of Akt/mTOR network in Daudi cells.[conclusion] 1.Interfering with FYN expression can inhibit the proliferation of Daudi cells with EBV positive lymphoma.2.The low expression of FYN may regulate and inhibit the proliferation of Daudi cells by inhibiting the Akt/mTOR network.