Investigation On 2-methoxyestradiol Albumin Nanoparticles Based On The Specificity Of The Monoclonal Antibody

Targeted drug delivery system has been one of hot research fields in tumor therapy. The objective of the study was to prepare a new targeting drug delivery system of HER2-2-ME-BSANPs which include the carrier of bovine serum albumin(BSA), the target agents of anti-HER2 monoclonal antibody and the model drug of2-ME. The drug delivery has the abilities to target tumor cells, induce more drugs into tumor cells, improve the antitumor effect of 2-ME and reduce the toxicity of drugs to normal tissues. The contents of the study were mainly divided into three aspects:Firstly, the preparation and characterization of HER2-2-ME-BSANPs.In this study, HER2-2-ME-BSANPs were prepared by N-succinimidyl-3-(2-Pyridyl dithio) propionate(SPDP) conjuncted anti-HER2 monoclonal antibody and2-ME-BSANPs which were prepared successfully by desolvation method in our prior research. HER2-2-ME-BSANPs were prepared successfully and had been proved to keep high immunocompetence and specificity by methods of SDS-Polyacrylamide gel electrophoresis(SDS-PAGE), agglutination test and immune-fluorescent assay.The conjugation ratio of anti-HER2 monoclonal antibody to 2-ME-BSANPs was36.28% detected by DTT method, indicating the system could be used to targeted cancer therapy. The nanoparticles obtained were negatively charged with a zeta potential of(-25.59 ± 0.69) mV and characterized(221.5 ± 2.1) nm with a narrow size distribution and better stability. 2-Methoxyestradiol loading efficiency and loading amount of the nanoparticles were(89.15±3.80)% and(8.31±2.50)%, respectively,improved greatly the solubility and bioavailability of 2-ME. In vitro, the release study of HER2-2-ME-BSANPs indicated 2-ME solution released quickly, while the release of the targeting drug delivery system was releasing slowly. The results showed that the targeted drug delivery system has a certain slow-release effect and improve the the half-life of 2-ME.Secondly, investigation on the antitumor activity and tumor-targeting of HER2-2-ME-BSANPs in vitro.In this study, human breast cancer SK-BR-3(HER2-positive expression, HER2+)and MCF-7(HER2-negtive expression, HER2-) cells were chosen as model cells. Theresults of MTT showed that the inhibitory effect of HER2-2-ME-BSANPs in SK-BR-3 cells was stronger than that in MCF-7 cells. HER2-2-ME-BSANPs can inhibit the two cells proliferation both with time and concentration dependent manner and HER2-2-ME-BSANPs had higher inhibition efficiency on these two cells than2-ME at all the time point. Flow cytometry were adopted to investigate the uptake ability in SK-BR-3 and MCF-7 cells. The results showed that HER2-2-ME-BSANPs were transferred into the SK-BR-3 cells(HER2+) more than MCF-7 cells(HER2-)and much faster than 2-ME because of anti-HER2 monoclonal antibody mediated.Flow cytometry revealed that HER2-2-ME-BSANPs could markedly arrest cell cycle in G2/M phase and did not change the mechanism of 2-ME. The results of cell apoptosis and cell autophagy showed that HER2-2-ME-BSANPs could induce the cell apoptosis and cell autophagy of SK-BR-3 cells and MCF-7 cells. In vitro, the targeted drug delivery system of HER2-2-ME-BSANPs which has good antineoplastic activity could transport the drugs into cancer cells accurately and effectively.Thirdly, investigating on the pharmacokinetics, pharmacodynamics and targeting of HER2-2-ME-BSANPs in vivo.Sprague-Dawle(SD) rats were applied to investigate the metabolism of HER2-2-ME-BSANPs. The results showed that the elimination half-life of emulsion was prolonged, AUC of HER2-2-ME-BSANPs was increased, the clearance rate decreased significantly. Therefore, HER2-2-ME-BSANPs was successfully prepared and with remarkable sustained release property, increased drug concentration in tumor cells and extended the time of effective drug concentration, improved the biology utilization, enhanced the therapeutic effects of 2-ME and played a very important role in targeting therapy of cancer long-lasting.In this study, the pharmacodynamics and tissue distribution of HER2-2-MEBSANPs in rats were explored by the classic method of pharmacology. The resluts showed that HER2-2-ME-BSANPs had higher anti-tumor activity than that of2-ME-BSANPs and 2-ME. The tissue distribution of the delivery system which marked by DIR fluorochrome was observed in vivo imaging technology. The results indicated that the fluorescence intensity of the system was tronger in HER2-positivecancer tissue than that in HER2-negtive cancer and had lower accumulation in the liver of the rats. The results indicated that the drug delivery system can prolong and improve phrmacodynamic due to the high concentration of drugs in tumor.