In Vitro Study Of Aspirin Combined With Osimertinib In Inhibiting Of Non-small Cell Lung Cancer

ObjectiveIn this study,non-small cell lung cancer(NSCLC)H1975 and A549 cell lines were studied.By comparing the effects of aspirin and Osimertinib alone and the combination of the two drugs on the proliferation,apoptosis and protein expression levels of H1975 and A549 cell lines.To verify that the combination drug is more effective than the drug alone to enhance the anti-tumor effect of the drug.Whether the two drugs have a synergistic effect on tumor proliferation inhibition.And preliminary discussion of its related mechanisms.This will provide a theoretical basis for the treatment and prevention of non-small cell lung cancer.MethodsMTT colorimetric assay was used to detect non-small cell lung cancer H1975 cell lines with different concentrations of aspirin(1,2.5,5,7.5,10,12.5,15 mmol/L)and osimertinib(0.05,0.1,0.5,1,5,10,50 ?mol/L)and non-small cell lung cancer A549 cell lines were treated with different concentrations of aspirin(1,2.5,5,10,20 mmol/L)and osimertinib(0.5,1,5,10,50 ?mol/L)absorbance after 24 h,48 h,and 72 h,respectively.Calculated of proliferation inhibition rate of H1975 and A549 cells by aspirin and osimertinib.The combination of 5 ?mol/L osimertinib and(2.5,5.0,and 7.5 mmol/L)aspirin with H1975 cells for 48 h.5 ?mol/L osimertinib was combined with(2.5,5.0,10.0 mmol/L)aspirin for 48 h with A549 cells.Measure absorbance and calculate the proliferation inhibition rate of H1975 and A549 cells by aspirin and osimertinib.The Kim's formula was used to judge whether the combination of the two drugs had a synergistic effect on cell proliferation inhibition.Hoechst33342 staining was used to observe the morphological changes of aspirin and osimertinib alone and in combination with H1975 nuclei.Annexin V-FITC/PI staining assay for the effect of aspirin and osimertinib alone and in combination on apoptosis of H1975 cells.Calculated the rate of apoptosis in H1975 cells.Western blot analysis of the effects of aspirin and osimertinib alone and in combination on the expression of Bcl-2,Bax,Cox-2 and NF-?B proteins in H1975 cells.To investigate the mechanism of apoptosis induced by aspirin combined with osimertinib in H1975 cells.Preparation of compound aspirin and osimertinib tablets.Screening prescriptions,preparation of compound tablets according to the best production conditions.And in accordance with the Chinese Pharmacopoeia tablet inspections under the relevant provisions of the inspection.ResultsMTT results showed that aspirin and osimertinib alone had significant inhibitory effect on proliferation of H1975 cells in a concentration-dependent manner(P<0.05).The inhibitory effect was more obvious with time.Different concentrations of aspirin alone significantly inhibited the proliferation of A549 cells in a concentration-dependent manner(P<0.05).The inhibitory effect was more pronounced with time.Different concentrations of osimertinib alone had no significant inhibitory effect on the proliferation of A549 cells(P?0.05).The combination of aspirin and osimertinib has a synergistic effect on the inhibition of proliferation of H1975 cells compared to the two drugs alone(Q>1.15).Aspirin combined with osimertinib alone had no synergistic effect on proliferation inhibition of A549 cells comparing with aspirin alone(Q<1.15)Observed by Hoechst33342 staining fluorescence inverted microscope H1975 cells found aspirin and osimertinib alone and in combination with a large number of apoptotic bodies visible,dense staining of nuclei,the effect of combination use of more significant.Annexin V-FITC/PI staining showed that aspirin and osimertinib alone and in combination could induce apoptosis in H1975 cells.The rate of apoptosis was significantly higher in the drug-associated cells than in the drug alone,and the combination drug had a synergistic effect(Q>1.15).Western blot results showed that aspirin and osimertinib treated H1975 cells alone or in combination,and the expression levels of Bcl-2,Cox-2,and NF-?B protein gradually decreased.The effect of aspirin combined with osimertinib was significantly lower than that of the two drugs alone(P<0.05).Bax protein expression gradually increased.The effect of aspirin combined with osimertinib was significantly higher than that of the two drugs alone(P<0.05).The preparation of the compound aspirin and osimertinib tablets verified the feasibility of the prescription process,and the quality inspection met the standards and achieved the established requirements.ConclusionsAspirin and osimertinib alone and in combination can inhibit the proliferation of non-small cell lung cancer H1975,combined with significant potentiation and synergistic inhibition.Aspirin combined with osimertinib did not significantly enhance A549 cell proliferation inhibition compared to aspirin alone.Aspirin combined with osimertinib has no synergistic effect on A549 cell proliferation inhibition.The combination of aspirin and osimertinib has a synergistic effect on the apoptosis of non-small cell lung cancer H1975 cells,which may be related to the coordinated regulation of the expression of Bax? Bcl-2,Cox-2 and NF-?B.Aspirin combined with osimertinib provides a theoretical basis for clinical combination therapy and effective strategies for the prevention and treatment of non-small cell lung cancer.