[關(guān)鍵詞]
[摘要]
目的 建立替諾福韋酯(TDF)的代謝產(chǎn)物替諾福韋(TFV)、替芬泰(Y101)及其代謝物M8的LC-MS/MS分析方法,基于大鼠體內(nèi)藥動學(xué)、腎排泄及體外腎切片攝取模型研究Y101與TDF的藥物-藥物相互作用(DDI)。方法 ①藥動學(xué)實驗:SD雄性大鼠隨機分為3組,分別單次ig給藥Y101(60 mg·kg-1)、TDF(30 mg·kg-1)及TDF(30 mg·kg-1)+Y101(60 mg·kg-1),通過LC-MS/MS方法測定給藥后血漿中Y101、M8、TFV濃度,并采用非房室模型統(tǒng)計矩法計算藥動學(xué)參數(shù)。②腎排泄實驗: SD雄性大鼠隨機分為3組,分別單次iv給藥Y101(25 mg·kg-1)、TDF(30 mg·kg-1)及TDF(30 mg·kg-1)+Y101(25 mg·kg-1)。利用LC-MS/MS方法測定尿樣中Y101、M8、TFV濃度,分析藥物及代謝物的累積排泄率;③腎切片實驗:大鼠腎切片分別在含有M8(5.0 μmol·L-1)、TFV(10 μmol·L-1)、TFV(10 μmol·L-1)+Y101(2.0 μmol·L-1)和TFV(10 μmol·L-1)+M8(5.0 μmol·L-1)的藥液中孵育一定時間后,收集樣品經(jīng)適當(dāng)處理后利用LC-MS/MS方法測定腎臟對TFV、Y101、M8的攝取量。結(jié)果 ①藥動學(xué)實驗:對Y101及M8開展部分方法學(xué)驗證,對TFV開展全面的方法學(xué)驗證,驗證結(jié)果表明LC-MS/MS方法專屬性強、靈敏度高。采用LC-MS/MS方法測定大鼠血漿中的TFV、Y101和M8,發(fā)現(xiàn)與單獨用藥組相比,Y101+TDF組大鼠血漿中TFV、Y101和M8藥時曲線下面積(AUC0-t和AUC0-∞)顯著性增加,血漿清除率(CLP)顯著性減小。②腎排泄實驗:Y101+TDF組的尿累積排泄分?jǐn)?shù)低于單獨用藥組,且TFV、M8均出現(xiàn)顯著性降低。③腎切片實驗:TFV+M8組與單獨孵育組相比,腎切片對TFV或M8的攝取量均顯著性下降。結(jié)論 TDF的代謝產(chǎn)物TFV與Y101的代謝產(chǎn)物M8可能通過競爭性抑制有機陰離子轉(zhuǎn)運體(3 OAT3),對TFV、M8的血藥濃度及腎排泄造成影響,提示臨床合用需要進(jìn)行劑量調(diào)整。
[Key word]
[Abstract]
Objective To establish LC-MS/MS methods for the analysis of tenofovir (TFV), bentysrepinine (Y101) and its metabolite M8. The drug-drug interactions between Y101 and tenofovir disoproxil fumarate (TDF) and its mechanism were studied by using the model of in vivo pharmacokinetics, renal excretion and in vitro uptake in kidney slices. Methods ① Pharmacokinetic experiments: SD male rats were randomly divided into three groups: rats were orally given Y101 (60 mg·kg-1), TDF (30 mg·kg-1) or TDF (30 mg·kg-1) + Y101 (60 mg·kg-1), respectively. Plasma concentrations of Y101, M8 and TFV were determined by LC-MS/MS methods, and pharmacokinetic parameters were calculated by Phenix WinNonlin with non-compartmental analysis. ② Renal excretion experiments: SD male rats were randomly divided into three groups: Rats via tail vein were injected with Y101(25 mg·kg-1), TDF (30 mg·kg-1) or TDF (30 mg·kg-1) + Y101 (25 mg·kg-1), respectively. The concentrations of Y101, M8 and TFV in urine samples were determined by LC-MS/MS methods, and the cumulative excretion rates of drugs and their metabolites were calculated. ③ Kidney slices experiments: Rat kidney slices were incubated in buffer containing M8 (5.0 μmol·L-1), TFV (10 μmol·L-1), TFV (10 μmol·L-1) + Y101 (2.0 μmol·L-1) or TFV (10 μmol·L-1) + M8 (5.0 μmol·L-1) for a certain period of time, respectively. The samples were collected and properly treated, and then the renal uptakes of TFV, Y101 and M8 were determined by LC-MS/MS methods. Results ① Pharmacokinetic experiments: A full validation of TFV and a partial validation of Y101 and M8 were carried out. These results showed that the LC-MS/MS methods were highly specific and sensitive. The concentrations of TFV, Y101 and M8 in rat plasma were determined by validated LC-MS/MS methods. It was found that compared with the Y101 or TDF treatment group, the area under the curve (AUC0-t and AUC0-∞) of TFV, Y101 and M8 in rat plasma increased significantly and the plasma clearance rate (CLP) decreased significantly in the group of TDF + Y101. ② Renal excretion studies: The results showed that the cumulative urinary excretion of TFV and M8 in the TDF+Y101 group was significantly lower than the Y101 or TDF alone group. ③ Kidney slices experiments: Compared with the TFV or M8 alone group, the uptakes of TFV or M8 decreased significantly in the TFV + M8 group. Conclusion TFV and M8 may have an impact on the blood concentrations and renal excretion of TFV and M8 through competitive inhibition of OAT3, suggesting that the dose needs to be adjusted in clinical combination.
[中圖分類號]
R969.2
[基金項目]
國家自然科學(xué)基金青年基金項目(82104284)