Mechanism Of Methotrexate Combined With Tripterygium Wilfordii In The Treatment Of Rheumatoid Arthritis And Study Of Tripterygium Wilfordii Prodru

Rheumatoid arthritis(RA)is a chronic andsystemic autoimmune inflammatory disorderthat affects about 0.5-1.0%of total adult population and brings heavy burden on healthcare systems.RA ischaracterized by systemic and synovial inflammation,which leads to permanent joint damageand disability if uncontrolled.Disease-modifying antirheumatic drugs(DMARDs)are recommended as thefirst-line treatment for RA early clinical intervention.In global,methotrexate(MTX)as a DMARDs,owing to its efficacy,acceptable,and low cost,has become(and remains)a mainstay in the treatment of RA.It is noteworthy that an increasing number of studies have found that RA is a heterogeneous disease with complex molecular network of pathophysiology,while MTX can only target a limited number of proteins.To gain a better therapeutic effect,combination regimens are recommended and lead to better outcomes than with either agent alone.Tripterygiumwilfordii Hook F(TwHF)is a traditional Chinese medicinal herb,and its extract,tripterygium glycosides(TGS),exhibit immunosuppressive and anti-inflammatory effects.Tripterygium glycosides tablets(TGTS)as a prescription medicine,mainly containing TGS,have been widely used in China treatment of RA for several decades.Actually,MTX plus TGTs has been empirically usedon treatment ofRA in China for decades,while the interferences of drug blood level and synergistic mechanism attributed to their joint treatment remains unknown.In the view of plasmapharmacochemistry,only when the drug is absorbed into the blood can exert its biological activity.To avoid the probable danger that drug combination can lead to a shift towards the producti on of another contri buti ng to the undesi rabl e side effects.It is essential to establish aquantitativeand comprehensive monitor strategy on the variation of bioactive migratory constituents and terminal metabolic phenotypes.The pseudotargeted metabolomics methodi ntegrated the advantages of both untargeted and targeted methods,is demonstrated to be a comprehensive strategy with high-quality and information-abundant data The ultra-high-performance liquid chromatography Q Exactive hybrid quadrupole-orbitrap high-resolution accurate MS(UHPLC-QE Orbitrap HRMS)instrument is very important for in vivo metabolomics studies regarding to the identification of untargeted metabolites in complex matrices,and is capable of analyzing a broad range of metabolites by database.For targeted metabolites’ quantification,multiple reaction monitoring(MRM)performed on a triple quadrupole tandem mass spectrometry(QqQ-MS)is recognized as the gold standard,attributed to its wide linear dynamic range,high sensitivity and high repeatability.As the research went on,we found that the most important active component of TwHF is triptolide,which has poor water solubility and low bioavailability.These shortcomingsseriously affected its clinical application.In recent years,the development of prodrug technology provides a new opportunity to improve the water-solubility of active compounds.This study proposes a strategy based on plasma pharmacochemistry and pseudotargeted metabolomics to explore the mechanism of drug combination.Finally,based on a clinical trial,14 drug prototype components and 6 biotransformed metabolites in plasma of patients with rheumatoid arthritis were quantitatively determined after MTX administration alone,TGTs administration alone and the combination of the two and 19 differential endogenous metabolites were quantitatively determined.The final results showed that TGTsgroup was as effective as MTX group and the MTX plus TGTS combination group was better than that of their alone groups in RA.The results of this study confirmed that the combination of MTX and TGTS did not significantly affect the absorption and metabolism of MTX plus TGTS compared with their monotherapy,respectively.MTX is intended to exert therapeutic effects by regulating DHFR,TS and purine DE novo synthesis,while TGTS is intended to exert therapeutic effects by regulating ADA and energy metabolism.In the end,the blood drug concentrations after the combined effects of MTX and TGTS were described from the perspective of quantitative plasma pharmacochemistry.Meanwhile,the metabolic disorders in RA and the combined mechanism of MTX and TGTS were systematically described from the perfective of metabolomics.In the further study,the chemical structure of triptolide was modified with the help of prodrug technology.Triptolide and succinic anhydride were combined by covalent bond to form a new compound with strong water solubility,which in order to enhance the water solubility and to improvethe bioavailability of triptolide.