A New Pyrazolo[4,3-d]pyrimidine Derivative:Synthesis And Anti-inflammatory Activity

Inflammation is a defense response when the body responds to stimuli.It is a double-edged sword.Moderate inflammation can limit the spread of inflammation,clear necrotic tissue,and restore organ function.However,the persistent undiminished inflammatory response will cause damage to the body,cause loss of body function,cause a series of inflammatory diseases,and even cause tumors and death.Inflammasome are multi-protein complexes that are assembled by cytoplasmic pattern recognition receptors.The most thoroughly studied is the NLRP3 inflammasome,which is an important component of innate immunity and can be activated by a variety of pathogenic microorganisms and danger signals,has played a huge role in a variety of human diseases.Therefore,the NLRP3 inflammasome is the core of the inflammatory response,and its research may provide new therapeutic targets for various inflammation-related diseases.NF-κB is a transcription regulator that has a wide range of effects in the body,and the activation of the classic NLRP3 inflammasome also depends on the role of NF-κB.The structure of pyrazolo [4,3-d] pyrimidine is very similar to that of purine.It has various pharmacological activities,and plays a very important role on energy supply,coenzyme formation and regulating metabolism.Based on the virtual screening of anti-inflammatory activities,through computer molecular docking,we have discovered and synthesized a novel pyrazolo [4,3-d] pyrimidine derivative,which can be used as a new lead compound to improve inflammation.After the compound was screened out through preliminary virtual screening,a more systematic pharmacological and anti-inflammatory activity study was conducted,and the possible anti-inflammatory mechanism of the compound was initially revealed.RAW264.7 cells were stimulated with LPS,simulating an in vitro cellular inflammation model.First,observing the morphological changes of RAW264.7 cells under a microscope,and comparing the experimental group,which the compound is added,model group and normal group to determine whether the compound has an anti-inflammatory effect.The effect of this compound on NO production in LPS-stimulated RAW264.7 cells was determined by the Griess method using a nitric oxide kit.Quantitative changes of TNF-α,IL-6,IL-1β,and COX-2 m RNA were measured by real-time PCR.Expression of COX-2 protein,NF-κB signaling pathway protein,and NLRP3 inflammasome associated protein were measured by Western blot.Furthermore,it was confirmed that this compound exerts a preliminary mechanism of anti-inflammatory activity through the NF-κB / NLRP3 inflammasome signaling pathway.