Biomimetic Synthesis And Anti-tumor Mechanism Of Cytisine-pterocarpan Derived Compounds

Sophora tonkinensis is a kind of traditional Chinese medicine originally recorded in“Kai Bao Ben Cao”.Previous studies showed that there are Cytisine-Pterocarpan derived compounds in S.tonkinensis,which had good cytotoxicity against cancer cell.However,the content of Cytisine-Pterocarpan derived compounds is low in S.tonkinensis.How to obtain a large number of target compounds has become an urgent matter.The biomimetic synthesis of Cytisine-Pterocarpan derived compounds were studied in the research,which laid a material foundation for the further study of this kind of compounds.Then the cytotoxic effect against different cancer cells were evaluated and the anti-tumor mechanism was studied.We are expecting to find new precursor compound or candidate drug.Objective and significance:According to the previous work,Cytisine-Pterocarpan derived compound had a certain cytotoxic effect against breast cancer cell.The short and high yield synthesis route was established using biomimetic synthesis method,the anti-tumor activities and the anti-tumor mechanism were studied in vitro.Research methods:Firstly,the biomimetic synthesis of Cytisine-Pterocarpan derived compound was carried out and the synthetic route conditions were optimized.The inhibitory activities of Tonkinensine B against different tumor cells were determined by MTT analysis.The value of IC₅₀0 was calculated and the effective concentration of Tonkinensine B was determined.The apoptosis morphology of MDA-MB-231 cells with the treatment of Tonkinensine B was detected by hoechst33342 staining.The apoptosis of MDA-MB-231 cells with the treatment of Tonkinensine B was studied by flow cytometry.The expression of Bcl-2,Bax and Cytochrome C protein in mitochondrion-mediated apoptotic pathway was determined by Western Blot.The data can determine the effect of Tonkinensine B on the expression of key proteins in MDA-MB-231 cells,which is triple-negative breast cancer cell.Research results:Cytisine-pterocarpan derived compounds were biomimetic synthesizedwith（-）-Cytisineand（-）-Maackiainviaa N,N-4-dimethyl-4-aminopyridine（DMAP）-mediated mild synthetic strategy in the presence of HCHO.The better synthetic route was established via optimizing the reaction conditions.The cytotoxicities of（-）-Cytisine,（-）-Maackiain and Tonkinensine B against MCF-7,MDA-MB-231,3LL,9L,Raw264.7,and BV2 cell lines were determined in vitro.The results showed that Cytisine-Pterocarpan derived compounds had obvious cytotoxicity against breast cancer cells,especially for MDA-MB-231 cell(IC₅₀=19.2μM).According to the research of cytotoxicities of Cytisine-pterocarpan derived compounds against Raw 264.7 and BV2 cells,it was suggested that this kind of compounds produced the lower cytotoxity on the central nervous system.The results of flow cytometry indicated that the cytotoxic activity of Cytisine-Pterocarpan derived compounds against MDA-MB-231 cell was induced by apoptosis.The results of western blot implied that this kind of compounds can regulate the expression of Bcl-2,Bax and Cytochrome C proteins in MDA-MB-231 cell.Conclusion:Cytisine-Pterocarpan derived compounds were synthesized with（-）-Cytisine and（-）-Maackiain using biomimetic synthesis method.It can effectively against MDA-MB-231 cell and accelerate cell apoptosis.The apoptosis is induced according to the mitochondria-mediated apoptotic pathway,which increasing the ratio of Bax/Bcl-2 protein and promoting the release of Cytochrome C from mitochondria to cytoplasm,finally leading to the cellular apoptosis.Research significance and application prospects:Breast cancer is a common malignant tumor among female group.Its incidence has occupied the first place in urban women’s tumors in China.With the rapid development of immunology and molecular biology,people have a better understanding of signal transduction in the apoptotic pathway.The human body can effectively prevent the rapid proliferation of tumor cells through apoptosis.Nowadays,the common methods of cancer treatment are preventing the proliferation of tumors according to inducing apoptosis of tumors cell.Cytisine-Pterocarpan derived compounds were synthesized using biomimetic synthesis method,and the cytotoxic effect of the novel compounds was screened.The results showed that this kind of compounds had obvious cytotoxity against MDA-MB-231 cell.With the modern biological and molecular biological methods,the cytotoxity of Cytisine-Pterocarpan derived compounds against MDA-MB-231 cell and the molecular mechanism of inducing cell apoptosis pathway were investigated.The results will provide more theoretical basis for the development of therapeutic drugs for breast cancer.