Design And Synthesis Of A Small Peptide RWR Containing RGD And Initial Exploration Of Function

Objectives1. To Design and synthesize the small peptide RWR containing RGD2. To measure the effect of RWR on platelet aggregation.3. To observe the effect of RWR on thrombosis.4. Verify RWR antithrombotic effect,to provide a theoretical basis for the researchand development of antithrombotic drug containing RGD.Methods1. The design and synthesis of the small peptide RWRUsing Fmoc solid-phase synthesis of chemical method synthesized RWR.On thebasis of RGD, adding a hydrophobic amino acid and a basic amino acid.The sequencecharacteristics is RGDWR,the research group named this new compound RWR.Thesmall peptide possesses the RGD structure,in theory the small peptide has the abilityof specific binding with the integrin αⅡbβ3receptor on the surface of the platelet.2. RWR affected the platelet aggregation rate in vitro dose-dependent relationshipDetecting the effect of different concentrations of RWR on platelet aggregation rateinduced by ADPNa2by platelet instrument。The whole blood of rabbits was separatedinto PPP and PRP.After PPP was compared to zero,adding different concentrations ofRWR to PRP tube,then adding the same dose of platelet aggregation inducerADPNa2,started to detect the maximum platelet aggregation rate within5min.Usingthe same method to detect the effect of RWR on human platelet aggregation rate.3. RWR compared with traditional medicine aspirin and tirofiban on the platelet aggre- gation rateDifferent concentrations of aspirin and tirofiban affected the maximum plateletaggregation rate,calculating the IC50of both,Compared the IC50of RWR with the IC50of aspirin and tirofiban.4. RWR effect platelet aggregation in vivo agingBefore administration,measured the heart blood platelet aggregation rate,then earvein injection of RWR,0h,0.25h,0.75h,1h,2h respectively measured the heart bloodplatelet aggregation rate,observed RWR in vivo metabolism and its role in aging.5. The effect of RWR on thrombosisThe effect of RWR on thrombus was researched through the thrombosis models ofthe rabbit arteriovenous bypass loop and thrombosis models of SD rats FeCl3.Thethrombosis models of the rabbit arteriovenous bypass loop illustrated the effect ofRWR from the thrombus weight. The thrombosis models of SD rats FeCl3explainedantithrombotic function from the change of vascular internal morphology by HEstaing.6. The influence of RWR on the content of cGMP in the plasmaBy ELISA kits of cGMP to determine the content of cGMP in the blood,understanding the changes of thrombosis, reflecting the RWR effect.Results1. Mass spectrometry analysis confirmed that the RWR sequence was correct, Thepurity was97.06%detected by HPLC.2. Platelet aggregation experiments demonstrated the platelet aggregation rate gradu-ally decreased with the concentration of RWR increasing.The effect of RWR wasmore notable on the role of human platelet aggregation.The anti-platelet aggrega-tion activity of RWR was higher than aspirin and tirofiban.The role of RWR onhuman platelet aggregation rate was more pronounced compared with the platelet of rabbits.3. With the concentrations of RWR increasing,the rabbit arteriovenous bypass loopexperiments showed that the thrombus weight gradually decreased,the effect ofRWR on thrombus weight was stronger than tirofiban;Ferric chloride thrombosismodels in rats showed the intravascular lacunar smaller.4. The content of cGMP in the plasma increased with the concentration of RWRincreasing.Conclusions1. Synthesis of new small peptide RWR,structure accorded with the design.2. Platelet aggregation experiments confirmed that the RWR had obviouscharacteristics of the inhibition of platelet aggregation in vitro and invivo.Compared with the traditional anti-thrombotic drugs aspirin (aspirin) andtirofiban, RWR was stronger on platelet aggregation.3. Two thrombosis model experiments confirmed RWR could inhibit thrombosis,theeffect of RWR on thrombus weight was stronger than tirofiban,Plasma cGMPincreased verified the RWR antithrombotic effect.