Research On The Interaction Between Three Furocoumarins And Organic Anion Transporters

Human organic anion transporters?hOATs?,which are expressed in many tissues,mediate the exchange of various small molecule compounds on the cell membrane to sustain the normal physiological function of the organism.hOATs are vital in the absorption,distribution,metabolism and excretion process of drugs or xenobiotics.Some clinic drugs are reported to be the substrates or the inhibitors of hOATs,and as for certain drugs with a narrow therapeutic window,the change of pharmacokinetics caused by the alteration of hOATs will cause toxic side effects.The application of traditional Chinese medicines has a long history in China.Nowadays traditional Chinese medicines are often administered concomitantly with conventional medications.However,due to the complex composition,many bioactive ingredients are reported to interact with conventional medications via effecting drug transporters,and it may change the therapeutic effects and induce toxic side effects.Therefore,the systematical studies on the interaction between bioactive ingredients and drug transporters will clarify the disposition of bioactive ingredients in vivo and help to improve the safety and effectiveness of the clinical application of traditional Chinese medicines.Furocoumarins exist in many traditional Chinese medicines such as Radix Angelica dahuricae and Angelica sinensis.They exhibit remarkable pharmacological activity such as anti-inflammatory and anti-tumor effects.It has been reported that imperatorin,isoimperatorin,and bergapten are highly exposed in the liver and kidney,and the hepatic uptake of these three furocoumarins involved both active transport and passive diffusion processes.However,it is still unknown which transporters are responsible for the transport of these furanocoumarins,and whether these furocoumarins have inhibitory effects on the transporters.In this study,three furocoumarins were used for investigating.Therefore the interactions between the furocoumarins and hOATs?hOAT1,hOAT2,hOAT3?were evaluated by using transgenic cell lines,including substrate/inhibitor verifications.This study will help researchers understand the disposal process of these three furocoumarins and will provide valuable data for the safe use of traditional Chinese medicines which contain furocoumarins in the clinic.The main conclusions are as follows:1.Establish and validate the analytical methods of LC-MS/MS of furocoumarinsTo study the interaction between three furocoumarins and hOATs,LC-MS/MS methods for the analysis of imperatorin,isoimperatorin,and bergapten in cell lysates were established separately and the method validations were carried out.The specificity,linear range,precision,accuracy,matrix effect,extraction recovery,and sample stability of analytical methods meet the guidelines requirements of quantitative analysis methods of biological samples.It indicated that the LC-MS/MS methods we established can provide reliable detection for further investigation.2.The interaction between three furocoumarins and hOATs in vitroThe transgenic cell models constructed previously were adopted for investigating the interaction between three furocoumarins and hOATs.All the data indicated that three furocoumarins were not the substrates of hOAT1-3.There was no difference in uptake between the transporter-overexpressed cells and the corresponding mock cells.Time-dependent and concentration-dependent experiments further confirmed that these three furanocoumarins entered the hOAT1-3 overexpressed cells via passive diffusion.Although they are not substrates,the three furanocoumarins show inhibitory effects on hOAT1-3.Imperatorin is the inhibitor of hOAT1-3 and it shows the strongest inhibitory effects on hOAT2 with the IC₅₀ value of 29.93±3.34?M.Isoimperatorin is also the inhibitor of hOAT1-3 and it shows the strongest inhibitory effects on hOAT3 with the IC₅₀value of 43.51±9.96?M.While bergapten shows a weaker inhibitory effect on hOAT1and hOAT2,and it has no inhibitory effect on hOAT3.All the vitro results indicate that the drug combination of certain traditional Chinese medicines which contain high amounts of these three furanocoumarins and clinic drugs which are substrates of hOAT1,hOAT2,and hOAT3 with narrow therapeutic windows should be avoided.