The Study Of Interactions Between Biflavonoids In Stellera Chamaejasme L And Transporters

Stellera chamaejasme L. (Thymelaeaceae) is ingrained in the Chinese Pharmacopoeia(2015). It is a toxic perennial herb possessing antivirus, antitumor, antibacterial, immunomodulatory activities. But its clinical application is limited because of its narrow safety scope and its obscure mechanism of metabolism and toxicity. Biflavonoid is the main active constituent in this herbal medicine. To our knowledge, ther were a few studies about the absorption and metabolism of biflavonoid isolated from Stellera chamaejasme L. Drug absorption, distribution, metabolism and excretion play an important role in efficacy and toxicity of medicines. Our studies focused on interactions between the biflavonoids such as chamaechromone(CMC), neochamaejasmin B(NCB), neochamaejasmin A(NCA) in Stellera chamaejasme L and transporters, using cells overexpressing drug transports, the molecular docking model and whole animal models, both in vitro and in vivo. In vitro, substrates and/or inhibitors of CMC, NCB, NCA over tranporters MRP2, BCRP, P-gp,OAT1 were evaluated through cell accumulation, uptake, cell kinetics studies. In vivo, pharmacokinetic studies in rats were performed to clarify the mechanism of drug transporter interactions and pharmacokinetic interactions between different components via transporters. The results were showed as following.1. The rapid and sensitive liquid chromatography-mass spectrometry methods were established and validated for the determination of biflavonoids (CMC, NCB, NCA) and lamivudine in biological specimens, respectively.2. Interactions between biflavonoids and transporters in vitroThe transport and cellular accumulation studies in Madin-Darby canine kidney (MDCK) cells overexpressing human multidrug resistance protein 2 (MRP2) or P-gp, LLC-PK1 cells overexpressing human breast cancer resistance protein (BCRP) were performed. The results indicated that NCB is a substrate of MRP2 and P-gp, repectively, NCA is a substrate of P-gp and BCRP, repectively, CMC is a substrate of MRP2 and BCRP, repectively. NCB inhibited the P-gp-mediated efflux in a concentration-dependent manner. The data from uptake study showed that CMC, NCB, NCA are a substrate of OAT1. And all of the three compounds were revealed to be the inhibitors of OAT 1. The inhibition of NCB over P-gp and OAT1 are the mixed type of competitive and non-competitive inhibition. Meanwhile, the inhibition of NCB over MRP2 is a competitive type inhibition.Binding selectivity based on molecular docking was studied. The molecular docking data indicated that NCB had higher affinity to P-gp than to single flavonoid structure Ligl. 3. Interactions between biflavonoids and transporters in vivoNCB increased the bioavailability of CMC via inhibition of MRP2 and BCRP in rats. The results showed that the area under the plasma concentration-time curve (AUC0-γ) and maximal plasma concentration (Cmax) of CMC were increased by 48.9% and 81.9%, respectively. The mechanism of improving the oral bioavailability of CMC was attributable to the inhibition of the intestinal BCRP-and MRP2-mediated efflux of CMC by NCB. Therefore, the efficiency and toxicity of an ingredient can be affected by co-existed other ingredients in an herbal medicine.The effects of Stellera chamaejasme L extracts (234.5 mg/kg, containing CMC, NCB and NCA) on the pharmacokinetics of lamivudine (15 mg/kg) were assessed. Both AUC0-∞ and Cmax of lamivudine were increased. The mechanism is that lamivudine is a substrate of OAT1 and CMC, NCB and NCA in Stellera chamaejasme L extracts can inhibit OAT1 in rat kidney which results in an increase of lamivudine blood concentration.Pharmacokinetics of CMC, NCB and NCA were measured after Stellera chamaejasme L extracts was administrated. The results showed that CMC absorption in rat was less than NCB although its contents are high than NCB in extracts. The absorption of NCB in rats was highest among the three components, while NCA was the least one.In summary, there are the drug transporters-based drug interaction among CMC, NCB and NCA in Stellera chamaejasme L extracts, which led to change their ADME properties, suggesting that the disposition of components in Stellera chamaejasme L extracts is a complex process compared to single compound.