Nuclease Activity And Antitumor Activity Of Targeted Copper, Platinum, Nickel Complexes

As a kind of potential artificial nucleases, Cu complexes have been widely studied in the field of biochemistry and molecular biology, a large number of copper complexes with novel structure and excellent nuclease activity have potential application values in DNA sequence determination, footprint reagents, abnormal DNA identification and cleavage aspects. In the artificial nuclease research field, cleavage efficiency is one of the focus to study, and adding auxiliary ligand to improve the original property of the complex is commonly used method. Meanwhile, due to their low toxicity and suitable redox potential, copper complexes have also made significant progress in the antitumor activity study, while the study of copper complexes with targeting groups to some organelles of the cell is rare. Therefore, targeting copper complexes as antitumor drugs have been studied more and more. In addition, the biodistribution, cellular accumulation, and cytostatic process of copper complexes seem to be different from those of platinum complexes, which create an opportunity to overcome the tumor resistance to conventional platinum drugs. Since the antitumor activity of cisplatin was widely concerned and accepted, the study on the antitumor activity of platinum complexes has become to the hot spot. The later designed platinum complexes such as carboplatin and oxaliplatin also have good antitumor effect. However, due to a variety of side effects and drug resistance, the applications of the above platinum complexes have been limited a lot. Since then, original designed platinum complexes to overcome drug resistance and having targeting ability to reduce side efffects have started to be studied. The main contents of this thesis are designing of copper and platinum complexes, the improvement of their properties, artificial nuclease activity and antitumor activity.First of all, a ternary copper-terpyridine complex [Cu(ttpy)(Gly)(NO3)](NO3)-H2O (2-1)(ttpy=4’-p-tolyl-2,2’:6,2"-terpyridine) is synthesized and characterized by X-ray crystallography and ESI-MS as an artificial nuclease. Glycine (Gly) is introduced into the complex to enhance the water-solubility and electrostatic affinity for the nucleic acid target. The DNA binding ability of complex2-1has been studied by UV-vis spectroscopy, fluorescence spectra and CD spectra, and the DNA cleavage activity of complex2-1has been studied by gel electrophoresis, using a structural analog [Cu(ttpy)(NO3)2](2-2) as the reference. The results indicate that carefully positioned auxiliary groups in a copper complex can significantly affect the substrate binding or activation ability and consequently the nuclease efficiency of the complex.Secondly, three Cu(Ⅱ), Pt(Ⅱ), Ni(Ⅱ)-ttpy-tpp complexes, where [ttpy-tpp=4’-p-tolyl-(2,2’:6’,2"-terpyridyl)triphenylphosphonium bromide], were synthesized and characterized by X-ray crystallography and ESI-MS as a targeted anticancer agent. Triphenylphosphine (TPP) is introduced into the complex for its mitochondrion-targeting ability and lipophilic character to obtain an original targeting complex, being able to pass through tumor cell membrane. The DNA binding ability of the complex has been studied by UV-vis spectroscopy, fluorescence spectra and CD spectra, and the DNA cleavage activity of the complex has been studied by gel electrophoresis. Results indicate that the complex bind with DNA stably through insert mode and electrostatic interaction, and could cleave pBR322DNA and DNA extracted from cells through oxidation cleavage, which lay the foundation of the further study on its antitumor activity.Thirdly, the cellular level mechanism of ttpy-tpp complexes have been studied. Flow cytometry, ICP and MTT method are used to detect its reactive oxygen species, the accumulation of copper in mitochondria and in cytoplasm as a whole, mitochondrial membrane potential, in vitro cytotoxity, cell cycle and cell membrane phospholipids asymmetry and integrity, so its reactive oxygen species level, mitochondrial targeting property, the damage degree to mitochondria, cytotoxicity and cell apoptosis mode have been precisely studied. The results indicate that the complex has had ideal targeting ability, high cytotoxicity and low drug resistance, it could be one of the potential antitumor drugs.Lastly, platinum complexes [Pt(BPAN)C1](5-1), Pt(BPANF)C1](5-2)(BPAN=N1,N1-bis((pyridin-2-yl)methyl)ethane-1,2-diamine, BPANF=N1,N1-bis((pyridin-2-yl)methyl)ethane-N2-folacin-1,2-diamine) are synthesized and characterized by ESI-MS, NMR and IR. On the structure basis of complex5-1,a tumor targeting group folic acid (FA) is added to complex5-2to get a new type of targeting platinum complex. The in vitro cytotoxicity of complex5-1and5-2has been studied to lay the foundation of further cellular level mechanism.