A Paclitaxel-based Supramolecular Hydrogel Loaded With Mifepristone For The Effective Inhibition Of Breast Cancer Metastasis

Objectives: Breast cancer is the leading cause of cancer-related death in women,and almost all breast cancer deaths are related to metastasis.Previous studies have shown that the combination of paclitaxel and mifepristone can reduce the metastasis rate of breast cancer.However,due to the short half-life of blood circulation,strong hydrophobicity of paclitaxel and mifepristone,lack of targeting,drug bioavailability is low and a series of adverse reactions are occurred.Therefore,our aim is to design and synthesize a nano-drug that can significantly improve the bioavailability of free drugs and enhance the anti-metastasis effect of breast cancer by co-delivery of paclitaxel and mifepristone.Content: To synthesize a paclitaxel-based hydrogel loaded with mifepristone,and to study the effect on the metastasis of breast cancer and its mechanism.Methods: Co-loaded paclitaxel and mifepristone hydrogel(PM-nano)was prepared by self-assembly.The formation,stability and viscoelasticity of hydrogels were tested by rheological test.The microstructure and secondary structure were characterized by transmission electron microscope and circular dichroism.In vitro drug release was quantitatively determined by dialysis and high performance liquid chromatography(HPLC).Drug uptake was determined by inverted fluorescence microscope and HPLC.Hemolysis test,blood routine test and blood biochemical analysis were carried out to evaluate the biocompatibility.MTT test,scratch test and Transwell test were performed to evaluate the effects of drugs on proliferation,migration and invasion of breast cancer cells.Western blot analysis was used to detect the possible mechanism.The 4T1-luciferase cells were inoculated to establish the in-situ tumor metastasis model of breast cancer.The body weight and tumor volume were measured every other day,in vivo imaging of small animals was performed every week.Finally,the tumor and main organs were used to evaluate the anti-tumor and anti-metastasis effects by pathological analysis.Results: A PTX-conjugated and MIF-loaded supramolecular hydrogel was developed with favorable water solubility,stability and viscoelasticity,in which paclitaxel was both the drug and the carrier.The form of nanofibers and a β-sheet structure was shown by transmission electron microscope and circular dichroism,respectively.In a weak acidic tumor microenvironment,free drugs were rapidly released from PM-nano.The cell intake of PM-nano was higher than that of free drugs.Even at the highest concentration of paclitaxel(0.5 mg/m L),PM-nano did not show significant hemolysis.All indicators of blood routine and blood biochemical tests were within the normal range,and there was no significant difference before and after administration.MTT assay showed that the cell survival rate of PM-nano was lowest,suggesting PM-nano had the strongest inhibitory effect on the proliferation of breast cancer cells.The scratch test showed that PM-nano had the largest scratch area and the lowest wound healing rate.The transwell analysis showed that the number of cells that passed through the membrane to the lower chamber was least in PM-nano group,and the absorbance value was also the lowest.Therefore,PM-nano has the strongest anti-proliferation and anti-tumor metastasis effect,which proves that PM-nano improves the efficacy of free drugs.PM-nano might inhibit breast cancer metastasis by glucocorticoid receptor/receptor tyrosine kinase-like orphan receptor 1 and matrix metalloproteinases.The tumor inhibition experiments showed PM-nano had the slowest tumor growth rate and the smallest tumor volume in all groups,indicating PM-nano had the best anti-tumor effect in vivo.There was no significant difference in body weight among all the groups.In vivo imaging of small animals and pathological analysis showed that only mice in PM-nano group did not have tumor metastasis;lung and liver were the most common metastasis sites,but heart,spleen,kidney,and adrenal gland could also have breast cancer metastasis,which suggested that clinicians should pay attention to follow up some rare metastasis sites.Conclusion: PM-nano can co-delivery of paclitaxel and mifepristone in the form of "carrier-free",greatly improve the drug bioavailability,significantly enhance the anti-breast cancer metastasis effect,and have good biological safety.