Design,Synthesis And Anti-hepatic Fibrotic Activity Evaluation Of Oxymatrine Derivatives

Oxymatrine has been applied as an anti-hepatitis B drug in China for decades.In recent years,its anti-hepatic fibrosis effect has been documented and further confirmed in clinic.In this study,taking oxymatrine as the hit,an in vitro anti-hepatic fibrosis high throughput screening was carried out among our innovative tricyclic matrine derivative library by a luciferase model based on COL1A1 promoter.Fortunately,12N-p-methylbenzenesulfonyl matrinic butyric acid(NS-1)was found to exert an inhibition effect against COL1A1 promoter with three times higher than the hit oxymatrine and was considered to be an ideal lead compound.Figure 1.Structure of matrine and discovery of its lead compound NS-1Therefore,in the present thesis,taking NS-1 as the lead,oriented by the down-regulation activity on COL1A1 promoter,a systematic structural modification was carried out.The structure-activity relationship(SAR)of down-regulating COL1A1 promoter was systematically summarized by synthesizing 107 new target matrine compounds,as shown in Figure 2.Figure 2.SAR of the tricyclic matrinic on inhibiting COL1A1 promoterButane was the most favorable 11-side chain.The introduction of 12N-substituent was beneficial for the activity;among all the substituents,aniline formyl was the most favorable;position,electrical property and number of substituents on benzene affected the activity;the matrinic nucleus with the 5S configuration was a druglike pharmacophore.Meanwhile,the IC50 values of twelve highly active 12n-aniline formyl matrinic butane derivatives with the inhibition rate of over 75%at 40 ?M were determined to further verify the activity and SAR.Their IC50 values ranged from 10 to 74 ?M.The activity order was basically consistent with the initial order in the primary screening.Three key matrine thiadiazole compounds(ND-2,NE-2 and NE-4)and 12N-anilineformyl matrinic butane compounds(FAM8c,FAM10c,FAM13c,FAM22c,FAM23c,FAM24c and FAM25c)could effectively reverse the increase of COL1A1,alpha smooth muscle actin(a-SMA)and other liver fibrosis-related factors induced by transforming growth factor beta 1(TGF-?1)on both mRNA level and protein levels.The preliminary safety evaluation showed that the oral LD50 values ofthem were over 500 mg/kg and 800 mg/kg,respectively,indicating good in vivo safety profiles.The in vivo pharmacokinetic behaviors of FAM8c,FAM10c,FAM13c and FAM22c in SD rat model at the dosage of 25 mg/kg via oral route were then investigated.It was revealed that most compounds showed poor metabolism behaviors,which might result from their poor absorption in vivo.Therefore,a high dosage of 200 mg/kg/day was chosen in the preliminary in vivo anti-hepatic fibrosis activity assay of key compound FAM22c in a biliary duct ligation(BDL)SD rat model.After a consecutive administration for 14 days,the liver volumes of the FAM22c-treated rats were about 2/3 of that of BDL model rats,and comparable to that of sham-operated rats.Fibrogenetic proteins such as COL1A1,?-SMA and TGF-?1 in the livers of FAM22c-treated rats were significantly reduced,representing the significant anti-hepatic fibrosis activity in vivo.Besides,the levels of ALT,AST,CHO and LDL in serum were significantly reduced,revealing the obvious hepato-protective effects.Therefore,FAM22c was promising for further study.The preliminary mechanism of matrine derivatives disclosed that they might exert the anti-hepatic fibrosis effect through repressing TGF?/Smad signaling pathway.Totally 107 new target matrine compounds were synthesized in this thesis,and their structures were confirmed by 1H NMR,13C NMR and HRMS.The anti-COL1A1 SAR was systematically summarize for the very first time and provided early data for development of anti-hepatic fibrosis agents targeting COL1A1.Among all the derivatives,the representative compound FAM22c displayed good anti-fibrotic effect in both in vitro and in vivo,and a high safety profile in vivo as well,which made it a potential candidate for the therapy of anti-hepatic fibrosis.The work has been partially published in Molecules(2018),and two Chinese patents have been applied based on it.In addition,the anti-influenza activity several compounds with bulky substituents at the terminal of 11-side chain were tested.Among them,ADCA3 displayed an IC50 value of 7.2 ?M against Type A influenza virus H3N2 A95-359.Meanwhile,it displayed a good stability to esterase.Preliminary mechanism showed that it might affect the earlt stage of viral replication.Therefore,it was a good anti-influenza lead compound,and worth further study.This part of work has been published in Molecules(2019)and Die Pharmazie(2019).