Discussions On Low Expression Of PBRM1 Enhances The Sensitivity Of Immunotherapy In Colorectal Cancer Cells

[Objective]To explore the role of PBRM1(Polybromo-1)gene in immunotherapy of human colorectal cancer,and to provide a new potential biomarker for evaluating the efficacy of immunotherapy for colorectal cancer[Methods]1.The expression of PBRM1 mRNA was detected by RT-PCR in 94 human colorectal cancer tissues,paired adjacent tissues and normal tissues,and the correlation between PBRM1 mRNA expression and baseline data of colorectal cancer patients was analyzed;2.The expression of PBRM1 mRNA in human normal pulmonary epithelial cell Beas-2b and colorectal cancer cells RKO,SW620,SW480 and HCT116 were detected by RT-PCR;3.Human colorectal cancer cell line SW620 with low expression of PBRM1 was selected to construct stable cell line with PBRM1 interfering silence/overexpression and negative control cell line by lentivirus packaging technology.The establishment of stable cell line was verified by RT-PCR and Western Blotting;4.After CTL was successfully prepared,the drug components were divided into CTL group and CTL combined with PD-L1 monoclonal antibody group.The CTL killing activity of different cell lines was detected by MTS reagent after 72 hours of treatment with the above four stable cell lines respectively;5.Tunel kit was used to detect the apoptosis of different target cells 72 hours after drug treatment;6.The expression levels of IFN-?,IL-12 and IL-10 in cell culture supernatant were detected by ELISA kit.[Result]1.Clinical sample detection:PBRM1 expression in 94 cases of human colorectal cancer tissues,adjacent tissues and normal tissues increased gradually,and there were significant differences compared with normal tissues(F=46.727,P<0.001).The expression of PBRM1 was positively correlated with the depth of infiltration(Kendall's tau-b=0,223,P=0.009);2.Cell lines detection:The expression of PBRM1 in RKO,SW620,SW480 and HCT116 colorectal cancer cells was lower than that in human normal lung epithelial cells Beas-2b,and the difference was statistically significant(P<0.01);3.RT-PCR and Western Blotting validated the successful construction of PBRM1 interference silencing/over-expression SW620 and corresponding negative control stable cell lines;4.The killing effect of CTL in PBRM1 interference silencing SW620 was significantly higher than that in negative control group.The killing effect of CTL in PBRM1 overexpression SW620 was significantly lower than that in negative control group(P<0.001);5.There was no significant difference in apoptotic effect of CTL or CTL combined with PD-L1 monoclonal antibody on PBRM1 silencing/overexpression SW620 compared with the control group(P>0.05);6.Compared with the negative control group,the IFN-? secreted by PBRM1 silencing SW620 combined with CTL or CTL and PD-L1 monoclonal antibody increased significantly,while the IL-12 and IL-10 levels decreased;The levels of IFN-gamma secreted by overexpressed colorectal cancer cells were significantly lower than those of negative control group,and the levels of IL-12 and IL-10 were significantly higher,with statistical significance(P<0.05).[Conclusions]1.The expression of PBRM1 mRNA was low in colorectal cancer tissues and cells,and was positively correlated with colorectal cancer infiltration depth.2.SW620 with low expression of PBRMI enhances the sensitivity of CTL or CTL combined with PD-L1 monoclonal antibody to immunotherapy.