Anti-Breast Cancer Target Combination Mining And Synergistic Drug Combination Optimization Based On Artificial Intelligence

As a highly heterogeneous malignant tumor,the pathogenesis of breast cancer is complex and diverse,involving multiple gene mutations and dysregulation of multiple signaling pathways.Monotherapy has some problems,such as limited efficacy,large poisonous side effect,and easy to develop drug resistance.Therefore,it is an important strategy to systemically identify key targets with synergistic effect on breast cancer,so as to develop specific targeted therapeutic drugs and carry out combination therapy.However,the traditional wet-lab experimental method remains time-and cost-consuming to screen synergistic target combinations from a large number of target combinations.How to identify new superior target combinations is still a major challenge.In this study,deep learning algorithm will be combined with large-scale biomedical data to systematically,efficiently and accurately excavate superior anti-breast cancer target combination from a large number of target combinations,and the synergistic anti-tumor effect of the target combination will be experimentally verified.Furthermore,the specific targeted drugs for the superior target combination will be used for combination therapy,and the synergistic anti-tumor effect and safety of the drug combination will be evaluated,which is expected to provide reference for the development and treatment of synergistic anti-tumor targets and drug combination.Methods:Firstly,the targets of compounds with anti-tumor effects were collected and randomly combined to generate target combination prediction samples.The anti-tumor synergism prediction model based on deep learning algorithm and large-scale biomedical data was used to predict the synergistic anti-tumor effects of the target combinations on MCF7 and MDA-MB-231 cells,and to excavate the superior anti-breast cancer target combination.Subsequently,MCF7 and MDA-MB-231 cells were transfected with the specific siRNA of the superior target combination.The effective silencing of target genes was detected using q RT-PCR,and the effect of co-silencing of the superior target combination on the proliferative activity of MCF7 and MDA-MB-231 cells was investigated using CCK-8.Then,the specific small molecule inhibitors were used as the therapeutic drug combination of the superior target combination,and the proliferative inhibitory activity and synergistic anti-tumor effect of this drug combination on MCF7 and MDA-MB-231 cells under different combination ratios were investigated using CCK-8.Finally,a breast cancer bearing mice model was established to investigate the effects of specific targeted drugs and their combinations on tumor growth of mice,and to evaluate the anti-tumor activity of the drug combination in vivo.Furthermore,the effects of specific targeted drugs and their combinations on body weight,liver and kidney function indexes and blood routine indexes of mice were investigated to evaluate the safety of the drug combination.Results:The optimal anti-tumor synergism prediction model had good prediction performance,which got the R²for the test set was 0.74,the MSE was 2.00,the RMSE was1.41 and the MAE was 1.06.Based on the prediction model,the combination of BIRC5,NAMPT and TOP1 targets ranked first in the prediction results of MCF7 and MDA-MB-231cells and the combination of HSP72,NAMPT and m TOR targets ranked 850 and 37 in the prediction results of MCF7 and MDA-MB-231 cells were obtained.Subsequently,MCF7 and MDA-MB-231 cells were transfected with the specific siRNA of the superior target combinations.The results of q RT-PCR showed that single transfection of siRNA targeting each gene and co-transfection of siRNA targeting three genes could effectively reduce the mRNA expression of corresponding target genes in MCF7 and MDA-MB-231 cells.The results of CCK-8 assay showed that co-silencing of BIRC5,NAMPT and TOP1 had significant inhibitory effects on the proliferation of MCF7 and MDA-MB-231 cells,with inhibition rates of 62.94%and 55.62%,respectively.Compared with single gene silencing or co-silencing of any two genes,co-silencing of three genes inhibited the proliferation of two types of breast cancer cells more effectively.The results indicated that simultaneous silencing of BIRC5,NAMPT and TOP1 had a synergistic inhibitory effect on the proliferation of MCF7 and MDA-MB-231 cells.Similarly,co-silencing of HSP72,NAMPT and m TOR had significant inhibitory effects on the proliferation of MCF7 and MDA-MB-231 cells,with inhibitory rates of 46.22%and 49.05%,respectively.Compared with single gene silencing or co-silencing of any two genes,co-silencing of three genes inhibited the proliferation of two types of breast cancer cells more effectively,showing a synergistic inhibitory effect.In addition,by comparing the inhibitory effects of the two target combinations on the proliferation of MCF7 and MDA-MB-231 cells,it was found that the inhibitory effect of the combination of BIRC5,NAMPT and TOP1 targets was superior to that of the combination of HSP72,NAMPT and m TOR targets.Moreover,the combination of HSP72,NAMPT and m TOR targets showed better inhibitory effect on the proliferation of MDA-MB-231 cells than MCF7 cells.The experimental results were consistent with the prediction results of the model,which verified the reliability of the prediction results.Then,LQZ-7I,FK866 and topotecan,which are specific small molecule inhibitors of BIRC5,NAMPT and TOP1,were used as the therapeutic drug combination of this target combination.In vitro anti-tumor activity evaluation results showed that the drug combination had strong proliferation inhibition and synergistic anti-tumor effects on MCF7 and MDA-MB-231 cells under different combination ratios,with IC₅₀less than 20μM and combination index（CI）less than 1.Among them,the drug combination had the best synergistic anti-tumor effect on both MCF7 and MD-MB-231 cells when the molar ratio of2.5:10:1,with IC₅₀of 7.04μM and 13.16μM,and CI of 0.26 and 0.19,respectively.In vivo anti-tumor activity evaluation results showed that the drug combination had the strongest inhibitory effect on tumor growth,with tumor inhibition rate of 62.05%.In vivo safety evaluation results showed that compared with blank control group,the drug combination had no significant effects on body weight,liver and kidney function of mice.In addition,there was no significant difference in hematological toxicity between the same dose of topotecan alone and in combination with LQZ-7I and FK866.These results indicated that the combination of LQZ-7I,FK866 and topotecan could enhance the anti-breast cancer effect,but did not increase the poisonous side effects.Conclusion:In this study,the combination of BIRC5,NAMPT and TOP1 targets was successfully mined based on the anti-tumor synergism prediction model,showing good anti-tumor effects on MCF7 and MDA-MB-231 cells.LQZ-7I,FK866 and topotecan,the specific small molecule inhibitors of BIRC5,NAMPT and TOP1 respectively,were used as the therapeutic drug combination of this target combination,showing strong synergistic anti-tumor effect and good safety.The research results of this study are expected to provide a new combination therapy scheme for breast cancer treatment,and provide theoretical basis and methodological reference for the research and development of synergistic anti-tumor targets and drug combinations.