The Synthesis Of Pyrimido[4,5-b]indole Derivtives And Their Effects On UrotensinⅡ

UrotensinⅡ is a kind of cyclic peptide which was isolated from fish urophysis, and it is a kind of nerve functional peptide. It makes vascular smooth muscle being vasoconstrictor and vasodilatation.As one of the strongest vascular contractility peptide,UⅡhas strong effect on promoting vascular smooth muscle cells division, plays an important role in the cardiovascular system, participates in the formation of atherosclerosis,hypertension and cardiomyopathy.Therefore, studying of UⅡinhibitor is necessary to treatment disease.The peptide ligands of UⅡbecome a hot target to researchers these years. Designing new chemical compounds and making them be real will be a new research direction. Our research takes o-Fluoronitrobenzene and ethyl cyanoacetate as the main raw materials to compound four new molecules, which were named PI-01, PI-02, PI-03, PI-04, belonging to 9H-pyrimido[4,5-b]indole.Through spectrum analysis solution, the molecular structure of the four kinds of compounds can be determined, ensuring the physical and chemical properties, anticipating phamacological effects.Studying four new molecules is in order to ensure their stimulating effect on VSMC. To ensure the cytotoxicity of the four compounds, we use MTT method to detect PI-01, PI-02, PI-03,PI-04 with the same concentration gradient, calculating the inhibition rate and IC50 respectively.Results show that PI-01 and PI-03 have capacity to inhibit growing cells,whereas PI-02 and PI-04 express they can promote cells. So we only calculate PI-01 and PI-03. Their IC50 are 1.803×10-5mol/L and 2.231×10-7mol/L. The inhibition of SMC by PI-01 is dose depended, while PI-03 is not. The conclusion is we can chose PI-01 to be de main compound to research.Determined by MTT method,PI-01 inhibit UⅡ in SMC. Let set up the research: The blank group, the negative group, model group( give 10-8mol/L UⅡ), the medicine group( give UⅡfor40min, and then give PI-01 10-7mol/L, 10-8mol/L, 10-9mol/L, totally three groups). Each group has eight data. Then the results comes out. The various of concentrations can inhibit UⅡ. The higher concentration take stronger inhibition. We prefer to choose 10-8mol/L UⅡ to take cell signaling study. But it is not sure and we should make more research to improve it.We have to use FCM to analyzing the cell cycle. There are four groups in our research: the blank group, the model group( give 10-8mol/L UⅡ) the medicine group( give UⅡfor 40 min, and then give PI-01 10-8mol/L), the drug control group( only put PI-01 for, 10-8mol/L). The result shows that the medicine group in S period rise obviously, while G2/M is reduced, Shows that cell cycle is inhibited by G2 period. And the result is the same as the result in MTT. It means that, we can study signals with PI-01 in 10-8mol/L.Analyzing weather the PI-01 take Notch signaling pathway to inhibit UⅡin SMC by RT-PCR.Notch signaling pathway has four subunits named Notch1,Notch2, Notch3, Notch4. We promote UⅡcan inhibit Notch signal, while PI-01 do the opposite function to UⅡ.PI-01 can do the same action to the four subunits, showing that PI-01 inhibiting U Ⅱ associated with Notch signaling pathway.