Design,Synthesis And Evaluation Of Anti-Inflammatory Activity Of Novel 8-Quinolinesulfonamide Derivatives

Inflammation underlies a variety of physiological and pathological processes and is the most basic form of defense against pathogens,damaged cells,and other stimuli that cause harm to the body.Inflammation is a complex set of interactions among soluble factors and cells that can arise in any tissue in response to traumatic,infectious,post-ischaemic,toxic or autoimmune injury.Now,we understand that inflammation is tightly linked to innate immunity and its molecular mechanisms,and that many inflammatory mediators are part of the innate immunity inherent in all multicellular organisms.It is generally believed that proper inflammatory feedback is beneficial to the body,but excessive inflammatory response can have adverse effects on the body.At present,studies have shown that the occurrence and development of various human diseases are closely related to body inflammation,including inflammatory bowel disease,asthma,rheumatoid arthritis,diabetes and Alzheimer’s disease.However,inflammation can be reduced by inhibiting the production of inflammatory factors such as TNF-α,IL-1β,IL-6 and NO.The classic drugs currently used to treat inflammation are non-steroidal anti-inflammatory drugs,such as aspirin,nimesulide,celecoxib,etc.,but these drugs may cause some adverse reactions.Therefore,it is still of great significance to continue research and development of new anti-inflammatory drugs.This study is based on the application of quinoline ring in anti-inflammatory drugs,and according to the sulfonamide structure contained in the drugs currently used in clinical（nimesulide,celecoxib and parecoxib,etc.）,a series of8-quinolinesulfonamide derivatives was synthesized.First,we preliminarily assessed the anti-inflammatory activity of all compounds in vitro by analyzing the production of NO,TNF-αand IL-1βin lipopolysaccharide（LPS）-induced RAW264.7 cells.Among them,the compound 3l with a single substituent CF₃introduced on the phenyl ring showed the best anti-inflammatory activity,with IC₅₀values of 2.61±0.39,9.74±0.85,and 12.71±1.34μM.In addition,western blotting experiments showed that 3l could reduce the expression of inflammatory mediators（i NOS and COX-2）in LPS-induced RAW264.7 cells in a concentration-dependent manner.Based on the above results,3l was considered to be the compound with the most anti-inflammatory potential,so we chose 3l as the selected compound for the further analysis.We first used the reverse virtual screening of Discovery Studio 2017 R2（DS 2017）software to obtain the potential target of the compound 3l to be toll-like receptor 4（TLR4）/MD-2,and then performed preliminary verification by molecular docking（CDOCKER）.The results showed that the compound could bind to the LPS binding site of TLR4/MD-2 and interact with amino acids on TLR4 and MD-2 receptor proteins,respectively.Next,we further assessed the effect of compound 3l on TLR4/MD-2 by co-immunoprecipitation（co-IP）to analyze the interaction of this compound with potential targets TLR4/MD-2.The results showed that compound 3l could significantly inhibit the heterodimerization of TLR4/MD-2 and the homodimerization of TLR4 to exert anti-inflammatory effect.Moreover,preliminary mechanism studies indicated that 3l could prevent TLR4 from being activated by preventing the binding of TLR4 and MD-2,thereby blocking the activation of the NF-κB/MAPK signaling pathway.In vivo experiment,it was proved that compound 3l had a significant therapeutic effect on adjuvant-induced arthritis（AIA）by observing and analyzing the swelling of the rat’s foot and the grade of arthritis,analyzing its joint pathology and detecting the levels of inflammatory factors（TNF-αand IL-1β）in the serum of the rat.These results indicated that compound 3l is a potential anti-inflammatory agent,from which more effective anti-inflammatory drugs could be developed.