| Background:Malignant tumor has been a threat to human health all the time.As one of the common female malignant tumors, breast cancer, its incidence increased dramatically in recent years, and has a trend of getting younger over. Along with the biomedical progress, people have done deep research on the mechanism of breast cancer pathogenesis, treatment strategy change rapidly. Eukaryotic translation initiation factors, as important regulators of translation, played significant roles in solid tumor transformation, evolution and resistance to drugs, and this kind of gene has been found in various human cancers including breast cancer. eIF4E is a crucial factor of eIF4F complex, which possess the ability of recognizing mRNA5’cap and start the5’cap translation process. It is reported that the normal phosphorylation level of4EBP family could inhibit the binding of eIF4E and eIF4G, and then block the assembly of eIF4F complex. On the contrary, the over phosphorylation of4EBP could loss the capability of binding with eIF4E and then promote the formation of eIF4F complex. The phosphorylation of4EBP family was dependent on the recognition of mTOR to RAIP motif and TOS motif. So, we speculated that the delete of RAIP motif and TOS motif could decrease the phosphorylation level inhibit eIF4F assembly and affect tumor development.However, How to make4EBP1D go through cell membrane and treat cancers? Vascular endothelial growth factor was regarded as the most significant peptide and was essential to cancer growth. Regular VEGF-A isomers, which were always called tumor angiogenesis factors, were selectively clipped as VEGF-A isoforms, called VEGFxxxb isomers and they still possessed the function of anti-tumor angiogenesis. In the former researches, we have detected the effects of eIF4E targeted4EBP expression vector on mice breast cancer model. Nevertheless, gene targeted therapy is more worthy for our further exploration.Research objectivesOn the premise of eIF4F assembly and cancer development were all broken by4EBP expression vector. VEGF111b vector was constructed to disrupt angiogenesis. Using mice breast cancer model, the effects of two kinds of gene therapy on mice volume, weight, heart, liver, lung, spleen and kidney were explored during this research. Immunohistochemical study was carried out to assess the influence on ki-67index of argans for evaluating the function of combination therapy synstematically. Meanwhile, the next mechanisms were studied primarily:1. The molecular mechanism of4EBP1D,VEGF gene monotherapy2. The targeted mechanism of4EBP1D-VEGF combination therapy.Research methods and results1) The total RNA extracted from well-grown cells was used to construct expression vector by RT-PCR and molecular cloning technologies. Double digestion and sequencing were used to detect their accuracy.2) The4T1breast cancer mice model was successfully established at first and then InvivojetPEI mediated pSecX and pSecX-4EBP1D pVEGF111b、 pVEGF11b-BP1were used to assess the targeted anti-cancer efficiency of monotherapy and combination therapy of4EBP1D and VEGF111b. Intravenous injection of plasmid was carried out once every three days until five times gene therapy finished. The results told us that the cancer volume of truncated4EBP1D group and VEGF111b gene therapy group was reduced41.5%and42.8%while gene combination therapy group reduced cancer volume by57.1%.3) The tissues of experiment group and control group were fixated, dehydrated, transparented, wax dipped and sliced for HE staining to know the pathological changes of cancer tissues. VEGF111b targeted4EBP1D displayed better therapy efficacy than the others. Conclusion4EBP1D-VEGF111b gene combination therapy, compared with single drug treatment, could relieve the development of mouse breast cancer animal model and reduce the side effects on heart, liver, spleen, lung and kidney better. The study provided a basis of animal experiments for using gene-targeted drugs to treat breast cancer. |
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