The Preparation, Characterization And Antitumor Effect Of A Peptide Modified Polypyridyl Ruthenium Complex Liposome

Nowadays, cancer is still the enemy of human’s health and longevity. Among them, the cure rate of brain glioma has always been very low. Traditional surgical treatment cannot eradicate tumor tissue, and chemotherapy is a necessary auxiliary treatment. Because of the existence of Blood brain barrier (Blood brain barrier, BBB), it is very difficult for chemotherapy drugs to go into the brain. On the other hand, the drugs is easy to have side effects to normal brain tissue after they going into the brain. Therefore, how to break through the barrier of the blood brain and reduce drug toxicity, and improve the antitumor drug curative effect are the emergency problems to be solved.This thesis mainly focus on the above problems from two aspects:first, the synthesis of two kinds of polypyridyl ruthenium complex with the effect of broad-spectrum antitumor as precursor. It was encapsulated in liposome in order to reduce toxicity. Because the ruthenium coordination compounds with the advantages of low toxicity, easy absorption, easy excretion as well as easy modification and tracer. Second, build a kind of active targeting long circulation liposomes drug delivery system with pepteide modified, to make it carry themselves with fluorescent antitumor drugs of pyridine class ruthenium complexes targeting into the brain and breaking through the blood-brain barrier. Through the cell experiments in vitro and animal experiments in vivo to investigate its anti-tumor effect. Because the active targeting long circulation liposomes have some advantages such as active targeting, slow release, long cycle, low toxicity, high stability and so on.In the first part, the long cycle of polypyridyl ruthenium complex complective targeting liposomes with membrane peptide (RVG-derived peptide, RDP) modified was prepared, and its physical and chemical properties were evaluated. Firstly, two kinds of pyridine synthesis class ruthenium complexes which have the effect of antitumor were synthesised. They are 2.2’-polypyridyl ruthenium complex complexes and 1.10-phenanthroline ruthenium complexes. The MTT experiments was used to determine the half of inhibition rate of tumor cells (IC50) respectively. And then choose 2.2’-polypyridyl ruthenium complex complexes as a representative of the drug, to further study its antitumor properties.Secondly, the peptide RDP, with the function of brain targeting, was conjugated with two stearic acyl phosphatidyl (DSPE-PEG-NHS), and then DSPE-PEG-RDP was obtained. Thirdly, RDP modified active targeting of 2.2’-polypyridyl ruthenium complex complexes active targeting liposome (RDP-Ru-Lip) and ordinary passive targeting of 2.2’-polypyridyl ruthenium complex complexes liposome (Ru-Lip) were prepared by membrane dispersion. By using laser particle size and Zeta, transmissio potential analyzer, biological mass spectrometry and ultraviolet spectrophotometer were characterized, and its physical and chemical properties were also examined. Dialysis demulsification method was used to measure the entrapment efficiency of liposome. In vivo imaging, inverted microscope and immunofluorescence were used to test its targeting ability, and MTT experiments was used to verify its effect on tumor cells. The results showed that the average particle size of RDP-Ru-Lip was (94.56±5.97)nm, the entrapment efficiency was (78.86±2.78)%, and the average particle size of Ru-Lip was (90.20±2.23)nm, the entrapment efficiency was (79.62±3.62)%, and liposomes obtained are characterized with high reproducibility and stability.The second part focuses on the targeting property and effects on tumor cells of the liposome prepared before. The imaging experiments in vivo showed that RDP-Ru-Lip have good brain targeting ability. The fluorescent inverted microscope method and MTT experiments also showed that RDP-Ru-Lip has obvious addicted to neurotropic ability, to selectively into neurological cells, and IC5o was significantly lower than ordinary liposome group and polypyridyl ruthenium complex complexes group without liposome encapsulated. The DAPI staining results showed that liposomes carrying drugs into the cell, polypyridyl ruthenium complex most stay in the cytoplasm, rarely enter the nuclei, and speculated that its anti-tumor effect mainly play a role in the cytoplasm. In conclusion, RDP-Ru-Lip were stable and controllable, and have obvious targeting brain function, antitumor effect in vitro is expected to be targeted good carrier for the treatment of brain tumor diseases.The longitudinal results showed that the RDP-Ru-Lip prepared is of good physical and chemical properties, controllable stablity, and of high reproducibility. Through the cell experiments in vitro, it was obvious neurotropic to neurological cells and obvious effects of anti-tumor cells. The experiments in vivo showed that it can quickly break through the blood brain barrier, and carry united polypyridyl ruthenium complex drugs targeting into the brain, and improve the drug concentrations. These experimental studies provide a strong theoretical support for more active brain targeting long circulation liposomes and anti-tumor metal complexes of development.