The Targeted Low-toxic Vector NNP Mediated MiR-34a Delivery And Induced Tumor Cell Apoptosis

Cancer is one of the world's five most incurable diseases.Breast cancer,the second leading cause of death among women,is on the rise.Among many cancer treatments,targeted gene delivery vector system is considered to have the most promising clinical application.Based on the above reasons,in this paper to design and build new targeted low toxicity nanometer gene carrier,will be targeted to tumor cells MCF-7 carcino embryonic antigen on the surface of six(CEACAM 6)single domain antibodies and lowers a cationic carrier of positive charge of N-Ac-Leu jointly modify genes in high loading capacity,high efficiency of gene delivery PEI 25 K,the formation of new targeted low carrier NNP,and characterization of its specific properties.We systematically studied the particle size potential,gene assembly ability,antiserum damage protection ability,targeting ability,biocompatibility ability and cell transfection ability of NNP.Through a series of experiments,we found that the new targeted low-toxicity carrier NNP has particle size that is easy to enter tumor cells,weak positive charge,good gene loading capacity,and effective protection of nucleic acid from serum damage,so as to effectively extend the half-life of nano-drugs in vivo.On the one hand,NNP vector conjugates Nb(mono domain antibody),giving it the ability to target tumor cells MCF-7.On the other hand,N-Ac-Leu,which can reduce the positive charge of cationic carriers,was coupled on NNP carriers,which greatly improved the biocompatibility of NNP carriers.Through cell transfection experiments,we found that NNP has a good ability of cell transfection.To further explore the anti-tumor efficacy of the new vector NNP-delivered small nucleic acid mi R-34 a,we tested the apoptosis of small bodies by cell proliferation inhibition test,staining test of living dead cells,DAPI staining test,and flow cytometry test.We found that NNP/ mi R-34 a nanocomposite can effectively induce early apoptosis of MCF-7 cells.Through cell cycle arrest experiments,we also found that NNP/ mi R-34 a nano-complex can enable MCF-7 cells to produce G1 phase cell arrest.By observing the changes of cell mitochondrial membrane potential,it can be proved that NNP/ mi R-34 a nanocomposite can guide cells to produce mitochondrial related endogenous apoptosis pathway mechanism.Western blot was used to explore the molecular mechanism of the protein,and it was found that NNP/ mi R-34 a nanocomposite had different degrees of influence on apoptosis-related proteins.Finally,we explored the influence of NNP/ mi R-34 a nano-complex on the migration and invasion of MCF-7 cells in tumor cells,through scratch test,Transwell test,colony formation test and protein imprinting test.We conclude that NNP/ mi R-34 a nano-complex can effectively inhibit the migration,invasion and colony formation of MCF-7 cells.To sum up,the specific characteristics of the new targeted low-toxicity nano-carrier NNP and the effects of NNP-loaded mi R-34 a forming nano-complexes on the proliferation,apoptosis ability,cycle arrest and apoptosis-related proteins of MCF-7 cells were explored in detail from the cytological and molecular levels.Moreover,NNP-loaded mi R-34 a can effectively inhibit the migration,invasion and colony formation of MCF-7 cells.The research of NNP/ mi R-34 a nanocomposite has contributed to the targeted therapy of tumor,nanomedicine,gene therapy and other aspects.