The Anti-proliferative Effect And Mechanism Of Ibr-7,an Ibrutinib Derivative,on Non-small Cell Lung Cancer

Objective The aim of our study is to explore the role of Ibr-7 alone and in combination with ABT-199 against non-small cell lung cancer,and provide a reliable basis and development direction for the screening of candidate drug for non-small cell lung cancer.Methods CCK-8 was used to detect the effects of Ibr and Ibr-7,Ibr-7 and ABT-199 alone or in combination on proliferation inhibition of non-small cell lung cancer cells.DAPI staining and Annexin-FITC/PI staining were used to detect apoptosis.Western Blotting was used to detect the expression of apoptosis-related protein and phosphorylated protein.Wound Healing Assay was used to detect the effects of Ibr-7 on migration.Proteomic Quantification of protein phosphorylation after the treatment of Ibr-7.We used Immunofluorescence to detect the co-localization of p-s6（ser235,236）and LARP1 after treatment with Ibr-7.Furthermore,Co-Immunoprecipitaiton（Co-IP）were used to determine the effect of Ibr-7 treatment on the interaction of p-s6 with LARP1.Results Ibr-7 inhibited the proliferation of non-small cell lung cancer cells.The IC₅₀of Ibr-7 against EGFR wild-type lung cancer cells A549 and H460 was 4.04μM and 1.93μM.The IC₅₀of Ibr-7 against EGFR mutant-type lung cancer cells H1975 and PC9 was 1.33μM and 1.11μM.The IC₅₀of Ibr against EGFR wild-type lung cancer cells A549 and H460 was28.99μM and 17.25μM.The IC₅₀of Ibr against EGFR mutant-type lung cancer cells was6.12μM and 0.06μM.DAPI staining,Annexin-FITC/PI staining and western blotting showed Ibr-7 induce apoptosis through caspase cascade.Ibr-7 inhibited the migration of A549 and H1975 cells.We also found that Ibr-7 significantly inhibited the phosphorylation of AKT-m TOR-S6 and may disrupt the binding of p-s6 to LARP1 in A549 cells.The IC50of ABT-199 at 48h on non-small cell lung cancer A549 and H1975 was 23.08μM and 10.23μM,respectively.When Ibr-7 is at a concentration of 2μM,it has a strong synergistic effect with 20μM ABT-199,the combination index can reach 0.25,suggesting a strong synergistic effect in A549 cells.In combination,Ibr-7 can enhance the apoptotic effect of ABT-199 and reduce the high expression level of Mcl-1,thereby Ibr-7 enhancing the anti-lung cancer effect of ABT-199.Conclusion Ibr-7 can significantly inhibit the proliferation of EGFR wild-type and mutant-type lung cancer cells.Ibr-7 induces apoptosis by inhibiting the phosphorylation of AKT-m TOR-S6 pathway;Ibr-7 may regulate the level of Mcl-1 to sensitize the antitumor effect of ABT-199.