Synthesis, Characterization And PTP1B Inhibitory Activity Of The Metal Complexes With Salicyladehyde Schiff Base Ligands

The number of patients suffering from diabetes mellitus(DM) is increasing throughout the world,with this disease becoming most significant disease in the century.Up to now,there are not still potent methods to cure the disease thoroughly.DM is a disease that results in chronic hyperglycemia due to an absolute or relative lack of insulin and/or insulin resistance.It is proved that protein-tyrosine phosphatase 1B(PTP1B) is an important negative regulator of insulin and leptin signaling in vivo.PTP1B can associate with and dephosphorylate activated insulin receptor(IR) or insulin receptor substrates.Overexpression of PTP1B in cell cultures decreases insulin-stimulated phosphorylation of nsulin receptor(IR) or insulin receptor substrates.So it is very important to study the inhibitors'interaction with PTP1B and their mechanism.Furthermore,the potent and specificity PTP1B inhibitors may be promising candidates for novel anti-diabetic drug development.Currently,a large number of the studies on PTP1B inhibitors have been reported.While most of the inhibitors are organic moleculars,the inorganic metal-complex PTP1B inhibitors have been few reported.Many experiments in vivo have shown the close relation of the trace metal elements to diabetes precaution and treatment.However,the mechanism for the insulin-sensitizing of metal effects has not been well understood by far,though the activities of decreasing the blood sugar by metal compounds had been associated with protein tyrosine phosphatase inhibition.Therefore,how metal compounds directly inhibit PTP1B and then influence metabolism need more investigation.In this thesis,we design and synthesis a series novel metal compounds employing the ligands with wide biological activities,and their inhibition effect and mechanism against PTP1B were investigated.The main results are as follows:1.A new series of oxovanadium complexes with mixed ligands,a tridentate ONO donor Schiff base ligand[viz.,salicylidene anthranilic acid(SAA)]and a bidentate polypyridyle ligand,have been synthesized and characterized by elemental analysis,FT-IR,EPR,DFT calculations,X-ray crystallography, ESI-MS,UV-vis,Molar conductivity measurment and potentiometric pH titrations.The results show that this kind of oxovanadium complex with a distorted octahedral geometry is stable and neutral,and the center vanadium atom is the oxidation state ofâ…¤(â…£) with d¹ configuration.The complexes have been found to be potent inhibitors against PTP1B(IC₅₀ approximate 10^-8M) in vitro.This kind of complex selectively inhibits PTP1B over the other two phosphatases(approximate 4～9-fold selectivity against SHP-1 and about 2-fold selectivity against TCPTP).Kinetics assays suggest that these complexes inhibit PTP1B in a competitive manner.The binding model of the kind of complex with PTP1B is predicted by using molecular docking techniques.The putative mode of interaction of the kind of complex with PTP1B suggests that the complex is capable of binding to the catalytic site of PTP1B.In the model,the vanadyl oxygen is close to the sulphur atom of the active site cysteine(Cys 215).In order to test the substitution of ligands effects on the PTP1B-inhibition activities,we replaced "salicylaldehyde" with "5-bromo-salicylaldehyde" and "2-hydroxy naphthalene-1-carbaldehyde",respectively,then synthesize and characterized a series of new similar mixed oxovanadium complexes.The results of inhibition assays show the PTP1B-inhibition activities of these oxovanadium complexes of derivatives are slightly lower than the corresponding complexes.It appears that the spatial bulk of substitute of Schiff base weakly diminishes the PTP1B-inhibition activities in these complexes.2.Based on the good activities of decreasing blood sugar by the oxovanadium complex with N₂O₂ coordination form.A tetradentate donor Schiff base ligand and derivatives with N₂O₂ coordination form were chosen to synthesize a series of oxovanadium complexes.These complexes were characterized by elemental analysis,FT-IR,EPR,DFT,UV-vis,conductivity measurment.The results show this kind of oxovanadium complex is five-coordinated neutral complex with distorted tetragonal pyramid geometry. The center vanadium atom is V(IV) with d¹ configuration.The complexes have been shown to be potent inhibitors of PTP1B.Kinetics assays suggest that the complex VO(sal-BDA) inhibits PTP1B in a competitive manner and inhibition constant of 31nM,was determined for the inhibition of PTP1B. The binding model of the complex VO(sal-BDA) with PTP1B is studied by using molecular docking techniques.The putative mode of interaction of the complex with PTP1B suggests that the vanadyl oxygen of the complex is close to the sulphur atom of cysteine(Cys 215) at the active site.3.Copper and zinc are essential trace elements that play some central roles in the biochemistry of every living organism.The antitumor activity of copper complexes based on DNA cleavage has been studied extensively. However,the investigation of copper complexes interaction with PTP1B, which is regared as the target of antitumor drugs,is not reported.Our group first explores PTP1B-inhibitory activities of copper complexes and zinc complexes with Schiff base ligands.In this thesis,the copper complexes with Schiff base.or 1,10-phenanthroline ligands are synthesized and nsulin receptor(IR) or insulin receptor substratesthen characterized by elemental analysis,FT-IR,X-ray crystallography,UV-vis,conductivity measurement. The results of inhibition assays show these copper complexes are potent PTP1B inhibitors.IC₅₀ values ranged from 10^-6 to 10^-7M.More studies show that PTP1B inhibitory activities of copper complexes with Schiff base ligands decrease in the presence of GSH and PTP1B inhibitory activities of the copper complex[Cu(phen)₃]²⁺ increase in the presence of oxygen,implying the existence of oxidation mechanism during the PTP1B-inhibition process by copper complexes.Noted that the binuclear copper complex [Cu(phen)₂(μ-IDA)Cu(phen)(NO₃)]NO₃ inhibits PTP1B in a uncompetitive manner.And the complex selectively inhibits TCPTP over the other two phosphatases SHP-1 and TCPTP.4.Using oxovanadium complex VO(SAA)(phen) and copper complex [Cu(phen)₂(μ-IDA)Cu(phen)(NO₃)]NO₃ as the representatives,we attempt to cultivate and PTP1B/inhibitor compound crystals by the soaking method and obtain two X-ray crystal structures of PTP1B soaked in the copper complex and oxovanadium complex.In comparison with the native PTP1B,the structures in PTP1B soaked in the copper complex and oxovanadium complex have some difference,and it is mainly obvious for the SH group of Cys 215,which indicates the ever happened interactions between the inhibitors and active region of PTP1B.Remarkably,the SH of catalytic cysteine of PTP1B is partially oxidized in the PTP1B/(copper complex) compound crystal,and the phenomenon is similar to that reported by the reference published in Nature.