盐酸度洛西汀对P-糖蛋白功能影响的体内外结合研究
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摘要
一、目的
通过考察盐酸度洛西汀对Caco-2细胞上P-糖蛋白功能和表达的影响以及对P-糖蛋白底物他林洛尔人体药动学的影响,探讨盐酸度洛西汀对P-糖蛋白的作用。
二、方法
(一)盐酸度洛西汀对Caco-2细胞上P-糖蛋白功能和表达的影响
1.细胞培养以及形态学验证
用MEM培养基对Caco-2细胞进行常规培养,建立Caco-2细胞模型,流式细胞术进行P-糖蛋白表达验证。
2.细胞毒性试验
用MTT法考察盐酸度洛西汀对Caco-2细胞的毒性,选择细胞存活率大于90%的药物浓度作为最大无毒浓度进行下一步细胞试验。
3.罗丹明-123外排试验
用维拉帕米作为P-糖蛋白功能抑制的阳性对照,用流式细胞仪测定细胞内罗丹明-123的荧光强度,考察度洛西汀对P-糖蛋白介导的罗丹明-123外排作用的影响,从而判断药物对P-糖蛋白功能的影响。
4.P-糖蛋白表达的测定
药物与细胞作用72h后采用流式细胞术分析度洛西汀对Caco-2细胞上P-糖蛋白表达的影响。
(二)盐酸度洛西汀对他林洛尔人体药动学的影响
1.给药方法
采用随机开放,两周期自身前后对照试验设计,12名健康受试者第一周期单剂量口服他林洛尔1片(50mg),第二周期受试者服用度洛西汀(30mg/次,Bid),连续服用6天,于第七天晨加服他林洛尔1片(50mg)。洗脱期:1周。
2.血样采集方法
在他林洛尔服药前及服药后0.5,1,1.5,2,3,4,5,6,7,9,12,24,36,48和60h由肘静脉取血5mL置肝素化离心管中,分离出血浆,置-70℃冰箱中保存待测。
3.测定方法
采用HPLC-ESI-MS/MS法测定血浆中他林洛尔的浓度。
三、结果
(一)盐酸度洛西汀对P-糖蛋白作用的研究结果
1.细胞培养以及形态学验证
Caco-2形态正常;流式细胞仪测定显示Caco-2细胞高度表达P-糖蛋白,适合用于盐酸度洛西汀对P-糖蛋白功能和表达影响的研究。
2.细胞毒性试验
盐酸度洛西汀在≦10μmol/L时,与细胞培养72h后细胞的存活率大于90%,因此,盐酸度洛西汀的最大无毒浓度为10μmol/L。
3.盐酸度洛西汀对罗丹明-123外排的影响
与阴性对照组相比,低、中、高浓度(0.2、5、10μmol/L)的盐酸度洛西汀显著降低了罗丹明-123从Caco-2细胞的外排(p＜0.05),但不同药物处理组间差别不明显。
4.盐酸度洛西汀对P-糖蛋白表达的影响
Caco-2细胞与低、中、高浓度(0.2、5、10μmol/L)的盐酸度洛西汀培养72h后,其荧光强度与阴性对照组相比,没有显著性差异。说明试验浓度的盐酸度洛西汀对P-糖蛋白的表达无明显作用。
(二)盐酸度洛西汀对他林洛尔人体药动学的影响
1.他林洛尔的测定
他林洛尔在2.0～240.0ng/mL浓度范围内线性关系良好。萃取回收率均大于86.6%,方法回收率为94.3%～105.6%,日内、日间RSD均小于13%,稳定性试验RSD均小于10.5%。
2.盐酸度洛西汀对他林洛尔药动学的影响
受试者多剂量服用盐酸度洛西汀后,他林洛尔的AUC_(0-60)和C_(max)由单用时的1348.54ng·h/mL,91.90ng/mL分别增加至1498.30ng·h/mL和125.21ng/mL。AUC和C_(max)分别增加了11%和36%。等效性分析显示两周期AUC和C_(max)比值的90%置信区间分别为77%—106%和68%—112%,均落于等效范围之外。他林洛尔的其它药动学参数T_(max),t_(1/2),CL/F,V/F在两周期间并无显著性差异。
四、结论
本试验首次从体内、体外两方面考察了盐酸度洛西汀对P-糖蛋白功能的影响。体外试验显示:低、中、高浓度(0.2、5、10μmol/L)的盐酸度洛西汀均抑制了P-糖蛋白的功能,降低了罗丹明-123从Caco-2细胞中的外排,但是共同培养72h并不影响P-糖蛋白的表达。这说明盐酸度洛西汀对P-糖蛋白的影响仅为功能上的抑制,其抑制的可能原因为竞争性抑制,或者抑制ATP酶等,其具体机制还需进一步研究。体内试验显示:多剂量服用度洛西汀使得他林洛尔的AUC和C_(max)增加,提高其生物利用度,但并不影响其他的药动学参数。我们推测可能的原因是盐酸度洛西汀抑制了肠道P-糖蛋白的功能,增加了其吸收程度。试验结果提示我们盐酸度洛西汀和P-糖蛋白底物合用时可能通过抑制P-糖蛋白功能减少药物外排,增加其血药浓度,使得疗效增加,但同时应注意因药物浓度增加导致的药物不良反应。
Objective
To evaluate the effect of duloxetine on P- glycoprotein function by investigating the effects of duloxetine on the function and expression of P-glycoprotein in Caco-2 cells and quantitating the effect of orally administered duloxetine on the pharmacokinetics of talinolol.
Method
1.The effects of duloxetine on the function and expression of P-glycoprotein in Caco-2 cells
1.1 Cell culture and morphology validated of cells
Caco-2 cells were cultured with MEM(Minimal Essential Medium) which contains 10%FBS(Fetal Bovine Serum) in a humidified atmosphere of 95%air and 5%CO_2 at 37℃.The expression of p-glycoprotein in the cells was identified by flow cytometer.
1.2 Cytotoxicity
MTT(diphenyltetrazolium bromide) assay was used to determine the non-cytotoxicity dosage of duloxetine in Caco-2 cells.The dosage at which the survival rate of cells is above 90%was considered as the highest non-cytotoxic dosage.
1.3 Effects of duloxetine on the efflux of rhodamine-123 in Caco-2 cells
The effect of duloxetine on P-glycoprotein function was analyzed using rhodamine-123 assay.Flow cytometry was used to determine the intracellular rhodamine -123 concentration and verapamil was used as the positive control.
1.4 Effect of duloxetine on P-glycoprotein expression
The expression of P-glycoprotein in Caco-2 cells was measured by flow cytometry after incubation with duloxetine for 72h.
2.The effect of orally administered duloxetine on the pharmacokinetics of talinolol
2.1 Dose Administration
The study was conducted in a self-controlled two-period experiment in a randomized,open-labeled design in 12 healthy Chinese volunteers.In the 1~(st) period,the volunteers received a single oral dose of 50 mg talinolol. After 1 week washout,the volunteers received 30 mg duloxetine twice daily for 6 consecutive days.On the 7~(th) day,they received 30 mg duloxetine and 50 mg talinlolol concomitantly.
2.2 Blood samples collection
Serial blood samples were collected from an in-dwelling venous catheter(anti-coagulated with sodium heparin) at 0、0.5、1、1.5、2、3、4、5、6、7、9、12、24、36、48、60 h after talinolol administration.The plasma was separated immediately and frozen at-70℃until analysis.
2.3 Analytic Method
Concentration of talinolol in plasma was determined by HPLC-ESI-MS/MS.
Results
1.Effects of duloxetine on the function and expression of P-glycoprotein in Caco-2 cells.
1.1 Cell Culture and Morphology Validated
The expression of P-glycoprotein is plentiful in Caco-2 cell and Caco-2 cell is suitable for evaluating the effect of duloxetine on P-glycoprotein function and expression.
1.2 Cytotoxicity
The 90%cells survived after incubation with duloxetine when the concentration of duloxetine was no more than 10μmol/L.The max concentration without cytotoxicity for duloxetine is 10μmol/L.
1.3 Effects of duloxetine on the efflux of rhodamine-123 in Caco-2 cells
Various concentrations of duloxetine decreased the efflux of rhodamine-123 significantly compared with negative control,but the differences between dose groups were not significant.
1.4 The effects of duloxetine on the expression of P-glycoprotein
The P-glycoprotein expression in Caco-2 cells after incubation with various concentrations of duloxetine(0.2、5、10μmol/L) was not significant different with the negative control.So,duloxetine in these concentrations doesn't modulate the expression of P-glycoprotein in Caco-2 cells.
2.The effect of duloxetine on the pharmacokinetics of talinolol
2.1 Determination of talinolol
The calibration curve was linear in the range of 2.0～240.0 ng/mL for talinolol.The average extraction recoveries for talinolol were above 86.6%.The methodology recoveries were between 94.3%and 105.6%. The intra-day and inter-day RSD were less than 13%and the RSD of stability test were no more than 10.5%.
2.2 The effect of duloxetine on the pharmacokinetics of talinolol
After subjects treated with duloxetine,the AUC_(0-60) and C_(max) of talinolol increased from 1348.54 ng·h/mL to 1498.30 ng·h/mL and 91.90 ng/mL to 125.21 ng/mL,respectively.The AUC_(0-60) and C_(max) of talinolol were increased by 11%and 36%,respectively.The 90%CIs for the ratios of AUC(0.77-1.06) and C_(max)(0.68-1.12) were not fell within the accepted range for lack of interaction(0.80-1.25).No significant difference in any other pharmacokinetic parameters was observed between the two periods(T_(max),t_(1/2),CL/F,V/F).
Discussion
The study for the first time evaluated the effect of duloxetine on the P-glycoprotein function in vitro and in vivo.In vitro study showed that various concentrations of duloxetine inhibit the function of P-glycoprotein and decrease the effiux of rhodamine-123,but doesn't modulate the expression of P-glycoprotein in Caco-2 cell.This suggested that duloxetine just influence the function of P-glycoprotein.The possible reason is competitive inhibition or inhibition of ATP enzyme.In vivo study showed that a 6 -day treatment period with duloxetine increased the oral bioavailability of talinolol,but other pharmacokinetic parameters were not changed.Because talinolol is not metabolized to a relevant extent,we speculate that the increase in oral bioavailability of talinolol by co-administration of duloxetine can be attributed with high certainty to an inhibition of P-glycoprotein in the small intestine.
Our results suggest that duloxetine could inhibit the function of P-gp in vitro and in vivo,and caution should be exercised when duloxetine is to be co- administered with drugs that are P-gp substrate.
通过考察盐酸度洛西汀对Caco-2细胞上P-糖蛋白功能和表达的影响以及对P-糖蛋白底物他林洛尔人体药动学的影响,探讨盐酸度洛西汀对P-糖蛋白的作用。
二、方法
(一)盐酸度洛西汀对Caco-2细胞上P-糖蛋白功能和表达的影响
1.细胞培养以及形态学验证
用MEM培养基对Caco-2细胞进行常规培养,建立Caco-2细胞模型,流式细胞术进行P-糖蛋白表达验证。
2.细胞毒性试验
用MTT法考察盐酸度洛西汀对Caco-2细胞的毒性,选择细胞存活率大于90%的药物浓度作为最大无毒浓度进行下一步细胞试验。
3.罗丹明-123外排试验
用维拉帕米作为P-糖蛋白功能抑制的阳性对照,用流式细胞仪测定细胞内罗丹明-123的荧光强度,考察度洛西汀对P-糖蛋白介导的罗丹明-123外排作用的影响,从而判断药物对P-糖蛋白功能的影响。
4.P-糖蛋白表达的测定
药物与细胞作用72h后采用流式细胞术分析度洛西汀对Caco-2细胞上P-糖蛋白表达的影响。
(二)盐酸度洛西汀对他林洛尔人体药动学的影响
1.给药方法
采用随机开放,两周期自身前后对照试验设计,12名健康受试者第一周期单剂量口服他林洛尔1片(50mg),第二周期受试者服用度洛西汀(30mg/次,Bid),连续服用6天,于第七天晨加服他林洛尔1片(50mg)。洗脱期:1周。
2.血样采集方法
在他林洛尔服药前及服药后0.5,1,1.5,2,3,4,5,6,7,9,12,24,36,48和60h由肘静脉取血5mL置肝素化离心管中,分离出血浆,置-70℃冰箱中保存待测。
3.测定方法
采用HPLC-ESI-MS/MS法测定血浆中他林洛尔的浓度。
三、结果
(一)盐酸度洛西汀对P-糖蛋白作用的研究结果
1.细胞培养以及形态学验证
Caco-2形态正常;流式细胞仪测定显示Caco-2细胞高度表达P-糖蛋白,适合用于盐酸度洛西汀对P-糖蛋白功能和表达影响的研究。
2.细胞毒性试验
盐酸度洛西汀在≦10μmol/L时,与细胞培养72h后细胞的存活率大于90%,因此,盐酸度洛西汀的最大无毒浓度为10μmol/L。
3.盐酸度洛西汀对罗丹明-123外排的影响
与阴性对照组相比,低、中、高浓度(0.2、5、10μmol/L)的盐酸度洛西汀显著降低了罗丹明-123从Caco-2细胞的外排(p＜0.05),但不同药物处理组间差别不明显。
4.盐酸度洛西汀对P-糖蛋白表达的影响
Caco-2细胞与低、中、高浓度(0.2、5、10μmol/L)的盐酸度洛西汀培养72h后,其荧光强度与阴性对照组相比,没有显著性差异。说明试验浓度的盐酸度洛西汀对P-糖蛋白的表达无明显作用。
(二)盐酸度洛西汀对他林洛尔人体药动学的影响
1.他林洛尔的测定
他林洛尔在2.0～240.0ng/mL浓度范围内线性关系良好。萃取回收率均大于86.6%,方法回收率为94.3%～105.6%,日内、日间RSD均小于13%,稳定性试验RSD均小于10.5%。
2.盐酸度洛西汀对他林洛尔药动学的影响
受试者多剂量服用盐酸度洛西汀后,他林洛尔的AUC_(0-60)和C_(max)由单用时的1348.54ng·h/mL,91.90ng/mL分别增加至1498.30ng·h/mL和125.21ng/mL。AUC和C_(max)分别增加了11%和36%。等效性分析显示两周期AUC和C_(max)比值的90%置信区间分别为77%—106%和68%—112%,均落于等效范围之外。他林洛尔的其它药动学参数T_(max),t_(1/2),CL/F,V/F在两周期间并无显著性差异。
四、结论
本试验首次从体内、体外两方面考察了盐酸度洛西汀对P-糖蛋白功能的影响。体外试验显示:低、中、高浓度(0.2、5、10μmol/L)的盐酸度洛西汀均抑制了P-糖蛋白的功能,降低了罗丹明-123从Caco-2细胞中的外排,但是共同培养72h并不影响P-糖蛋白的表达。这说明盐酸度洛西汀对P-糖蛋白的影响仅为功能上的抑制,其抑制的可能原因为竞争性抑制,或者抑制ATP酶等,其具体机制还需进一步研究。体内试验显示:多剂量服用度洛西汀使得他林洛尔的AUC和C_(max)增加,提高其生物利用度,但并不影响其他的药动学参数。我们推测可能的原因是盐酸度洛西汀抑制了肠道P-糖蛋白的功能,增加了其吸收程度。试验结果提示我们盐酸度洛西汀和P-糖蛋白底物合用时可能通过抑制P-糖蛋白功能减少药物外排,增加其血药浓度,使得疗效增加,但同时应注意因药物浓度增加导致的药物不良反应。
Objective
To evaluate the effect of duloxetine on P- glycoprotein function by investigating the effects of duloxetine on the function and expression of P-glycoprotein in Caco-2 cells and quantitating the effect of orally administered duloxetine on the pharmacokinetics of talinolol.
Method
1.The effects of duloxetine on the function and expression of P-glycoprotein in Caco-2 cells
1.1 Cell culture and morphology validated of cells
Caco-2 cells were cultured with MEM(Minimal Essential Medium) which contains 10%FBS(Fetal Bovine Serum) in a humidified atmosphere of 95%air and 5%CO_2 at 37℃.The expression of p-glycoprotein in the cells was identified by flow cytometer.
1.2 Cytotoxicity
MTT(diphenyltetrazolium bromide) assay was used to determine the non-cytotoxicity dosage of duloxetine in Caco-2 cells.The dosage at which the survival rate of cells is above 90%was considered as the highest non-cytotoxic dosage.
1.3 Effects of duloxetine on the efflux of rhodamine-123 in Caco-2 cells
The effect of duloxetine on P-glycoprotein function was analyzed using rhodamine-123 assay.Flow cytometry was used to determine the intracellular rhodamine -123 concentration and verapamil was used as the positive control.
1.4 Effect of duloxetine on P-glycoprotein expression
The expression of P-glycoprotein in Caco-2 cells was measured by flow cytometry after incubation with duloxetine for 72h.
2.The effect of orally administered duloxetine on the pharmacokinetics of talinolol
2.1 Dose Administration
The study was conducted in a self-controlled two-period experiment in a randomized,open-labeled design in 12 healthy Chinese volunteers.In the 1~(st) period,the volunteers received a single oral dose of 50 mg talinolol. After 1 week washout,the volunteers received 30 mg duloxetine twice daily for 6 consecutive days.On the 7~(th) day,they received 30 mg duloxetine and 50 mg talinlolol concomitantly.
2.2 Blood samples collection
Serial blood samples were collected from an in-dwelling venous catheter(anti-coagulated with sodium heparin) at 0、0.5、1、1.5、2、3、4、5、6、7、9、12、24、36、48、60 h after talinolol administration.The plasma was separated immediately and frozen at-70℃until analysis.
2.3 Analytic Method
Concentration of talinolol in plasma was determined by HPLC-ESI-MS/MS.
Results
1.Effects of duloxetine on the function and expression of P-glycoprotein in Caco-2 cells.
1.1 Cell Culture and Morphology Validated
The expression of P-glycoprotein is plentiful in Caco-2 cell and Caco-2 cell is suitable for evaluating the effect of duloxetine on P-glycoprotein function and expression.
1.2 Cytotoxicity
The 90%cells survived after incubation with duloxetine when the concentration of duloxetine was no more than 10μmol/L.The max concentration without cytotoxicity for duloxetine is 10μmol/L.
1.3 Effects of duloxetine on the efflux of rhodamine-123 in Caco-2 cells
Various concentrations of duloxetine decreased the efflux of rhodamine-123 significantly compared with negative control,but the differences between dose groups were not significant.
1.4 The effects of duloxetine on the expression of P-glycoprotein
The P-glycoprotein expression in Caco-2 cells after incubation with various concentrations of duloxetine(0.2、5、10μmol/L) was not significant different with the negative control.So,duloxetine in these concentrations doesn't modulate the expression of P-glycoprotein in Caco-2 cells.
2.The effect of duloxetine on the pharmacokinetics of talinolol
2.1 Determination of talinolol
The calibration curve was linear in the range of 2.0～240.0 ng/mL for talinolol.The average extraction recoveries for talinolol were above 86.6%.The methodology recoveries were between 94.3%and 105.6%. The intra-day and inter-day RSD were less than 13%and the RSD of stability test were no more than 10.5%.
2.2 The effect of duloxetine on the pharmacokinetics of talinolol
After subjects treated with duloxetine,the AUC_(0-60) and C_(max) of talinolol increased from 1348.54 ng·h/mL to 1498.30 ng·h/mL and 91.90 ng/mL to 125.21 ng/mL,respectively.The AUC_(0-60) and C_(max) of talinolol were increased by 11%and 36%,respectively.The 90%CIs for the ratios of AUC(0.77-1.06) and C_(max)(0.68-1.12) were not fell within the accepted range for lack of interaction(0.80-1.25).No significant difference in any other pharmacokinetic parameters was observed between the two periods(T_(max),t_(1/2),CL/F,V/F).
Discussion
The study for the first time evaluated the effect of duloxetine on the P-glycoprotein function in vitro and in vivo.In vitro study showed that various concentrations of duloxetine inhibit the function of P-glycoprotein and decrease the effiux of rhodamine-123,but doesn't modulate the expression of P-glycoprotein in Caco-2 cell.This suggested that duloxetine just influence the function of P-glycoprotein.The possible reason is competitive inhibition or inhibition of ATP enzyme.In vivo study showed that a 6 -day treatment period with duloxetine increased the oral bioavailability of talinolol,but other pharmacokinetic parameters were not changed.Because talinolol is not metabolized to a relevant extent,we speculate that the increase in oral bioavailability of talinolol by co-administration of duloxetine can be attributed with high certainty to an inhibition of P-glycoprotein in the small intestine.
Our results suggest that duloxetine could inhibit the function of P-gp in vitro and in vivo,and caution should be exercised when duloxetine is to be co- administered with drugs that are P-gp substrate.
引文
[1].Gottesman.M.M.Pastan,I.Biochemistry of multidrug resistance mediated by the multidrug transporter.[J].Annu.Rev,.Biochem,1993,62:385-427.
[2].Barrand MA,Bagrij T,Neo SY.Multidrug resistance-associated protein:a protein distinct from P-glycoprotein involved in cytoxic drug expulsion[J].Gen Pharma,1997,28:629-651.
[3].Hennessy M,Spiers JP.A primer on the mechanics of P-glycoprotein the multidrug transporter[J].Pharmacol Res,2007,(1):1 - 15.
[4].Schinkel AH.P-Glycoprotein,a gatekeeper in the blood-brain barrier[J].Adv Drug Deliv Rev,1999,36:179-194.
[5].Jin Sun,Zhong-Gui He,Gang Cheng,et al.Multidrug resistance P-glycoprotein:crucial significance in drug disposition and interaction.Med Sci Monit,2004,10:5-14.
[6].Scheffer GL,Pijnenborg AC,Smit EF,et al.Multidrug resistance related molecules in human and murine lung[J].J Clin Pathol,2002,55:332-339.
[7].Dietrich CG,Geier A,Oude Elferink RP.ABC of oral bioavailability:transporters as gatekeepers in the gut[J].Gut,2003,52:1788-1795.
[8].Chandra P,Brouwer KL.The complexities of hepatic drug transport:current knowledge and emerging concepts[J].Pharm Res,2004,21(5):719-735.
[9].Graft CL,Pollack GM.Drug transport at the blood-brain barrier and the choroid plexus[J].Curt Drug Metab,2004,5:95-108.
[10].Leslie EM,Deeley RG,Cole SP.Multidrug resistance proteins:role of P-glycoprotein,MRP1,MRP2,and BCRP(ABCG2) in tissue defense[J].Toxicol Appl Pharmacol,2005,204:216-237.
[11].Akiyama SI,Comwell MM,Kuwano M,et al.Most drugs that reverse multidrug resistance also inhibit photoaffinity labeling of p-glycoprotein by a vinblastine analog[J].Mol Pharmacol,1988,33:144-147.
[12].Richard B Kin.Drugs as P-glycoprotein substrates,inhibitors,and inducers [J].Drug Metab Rev,2002,34:47-54
[13].Seelig A.A general pattern for substrate recognition by P-glycoprotein[J].Eur J Biochem,1998,251:252-261.
[14].Anathea BW,Peter K.Duloxetine Hydrochloride[J].Nature Reviews,2004,3:907-908.
[15].Wimett L,Laustsen G.Duloxetine fights major depressive disorder[J].Nurse Pract,2005,30:6-7.
[16].Cruz MP,Gonzales ME,Jacobs J,et al.Duloxetine HCl(Cymbalta) for the treatment ofdepression,neuropathic pain,fibromyalgia and stress urinary incontinence [J].Drug Forecast,2006,31:84-97.
[17].Dan P,Yaron D,Dina M,Rimona M(2004).Fluoxetine inhibits multidrug resistance extrusion pumps and enhances responses to chemotherapy in syngeneic and in human xenograft mouse tumor models.Cancer Res,64,7562-7569.
[18].Johanna W,Dormann SM,Martin-Facklam M,Kerpen CJ,Ketabi-Kiyanvash N,Haefeli WE(2003).Inhibition of P-glycoprotein by newer antidepressants.J.Pharmaacol.Exp.Ther,305,197-204.
[19].刘志伟.Caco-2细胞单层模型及其在毒理学中的应用.卫生研究,2004,33:756-759.
[20].关溯,赵立子,陈杰,等.Caco-2细胞模型的建立及意义.山东医药,2005,45:1-3.
[21].Maurer H,H.Tenberken O,Kratzsch C,et al.Screening for library-assisted identification and fully validated quantification of 22 beta-blockers in blood plasma by liquid chromatography-mass spectrometry with atmospheric pressure chemical ionization.J.Chromatogr.A,2004,1058:169-181.
[22].Beate T,R Oertel,K Richter,et al.Disposition and bioavailability of the β1-adrenoceptor antagonist talinolol in man.Biopharm dru dispo,1995,16:403-414.
[23].Gramatte T,Oertel R,Terhaag B.Direct demonstration of small intestinal secretion and site-dependent absorption of the beta-blocker talinolol in humans.Clin Pharmacol Ther,1996,59:541-549.
[24].Krueger M,H,chenbach A,Terhaag B.Pharmacokinetics of oral talinolol following a single dose and during steady state in patients with chronic renal failure and healthy volunteers.In J Clin Pharm and Ther,2001,39:61-66.
[25].Ulrike W,Hildegard S.L,Ernst M.Evidence for intestinal secretion as an additional clearance pathway of talinolol enatiomers:concentration- and dose-dependent absorption in vitro and in vivo.Pharmaceutic Res,1996,13:514-522.
[26].Schwarz UI,Dresser GK,Oertel R,Kim RB.Talinolol-verapamil interaction is not solely due to P-glycoprotein inhibition[abstract].Clin Pharmacol Ther,2001,69,P85.
[27].Shirasaka Y,Kawasaki M,Sakane T,et al.Induction of human P-glycoprotein in Caco-2 cells:development of a highly sensitive assay system for P-glycoprotein-mediated drug transport[J].Drug Metab Pharmacokinet,2006,21:414-423.
[28].Han Y,Tan TM,Lim LY.Effects of capsaicin on P-gp function and expression in Caco-2 cells[J].Biochem Pharmacol,2006,71:1727-1734.
[29].张向荣,张逸凡,钟大放.体外Caco-2细胞模型在药物吸收中的应用进展[J].中国医院药学杂志,2004,24:773-775.
[30].Mosmann T.Rapid colorimetric assay for cellular growth and survival:application to proliferation and cytotoxicity assays.J Immunol Methods,1983,65:55-63.
[31].DuBuske LM.The role of P-glycoprotein and organic anion-transporting polypeptides in drug interactions[J].Drug Saf,2005,28:789-801.
[32].Couture L,Nash JA,Turgeon J.The ATP-binding cassette transporters and their implication in drug disposition:a special look at the heart[J].Pharmacol Rev,2006,58:244-258.
[33].Breier A,Barancik M,Sulova Z,et al.P-glycoprotein-implications of metabolism of neoplastic cells and cancer therapy[J].Curr Cancer Drug Targets,2005,5:457-468.
[34].Thomas H,Coley HM.Overcoming multidrug resistance in cancer:an update on the clinical strategy of inhibiting P-glycoprotein[J].Cancer Contro,2003,10:159-165.
[35].Petriz J,O'Connor JE,Carmona MIs,et al.rhodamine123 an appropriate fluorescent probe to assess P-glycoprotein mediated multidrug resistance in vinblastine-resistant CHO cells.Anal Cell Pathol,1997,14:129-140.
[36].Ryoko Yumoto,Teruo Murakami,Yuko Nakamoto,et al.Transport of Rhodamine 123,a P-Glycoprotein Substrate,across Rat Intestine and Caco-2 Cell Monolayers in the Presence of Cytochrome P-450 3A-Related Compounds.J Pharmacol Exp Ther,1999,289:149-155.
[37].He L,Liu GQ.Effects of various principles from Chinese herbal medicine on rhodamine 123 accumulation in brain capillary endothelial cells.Acta Pharm Sin,2002,23:591-596.
[38].Anuchapreeda S,Leechanachai P,Smith M,et al.Modulation of P-glycoprotein expression and function by curcumin in multidrug-resistant human KB cells[J].Biochem Pharmacol,2002,64:573.
[39].Durr,D,Stieger B,Kullak-Ublick,GA,et al.St.John's wort induces intestinal P-glycoprotein/MDR1 and intestinal and hepatic CYP3A4[J].Clin Pharmacol Ther,2000,68:598-604.
[40].Hennessy M,Kelleher D,Spiers JP,et al.St.John's wort increases expression of P-glycoprotein:implications for drug interactions[J].Br J Clin Pharmacol,2002,53:75.
[41].Matsuyama N,Kojima.Simultaneous five cell-lineage flow cytometric analysis system for detection of leucocyte antibodies[J].Transfusion Medicine,2006,16:111-118.
[42].Katrien S,Barbara DM,Catharina D,et al.Comparison of two functional flow cytometric assays to assess P-gp activity in acute leukemia[J].Leukemia and Lymphoma,2004,11:2221-2228.
[43].Han Y,Tan TM,Lim LY.Effects of capsaicin on P-gp function and expression in Caco-2 cells[J].Biochem Pharmacol,2006,71:1727-1734.
[44].U.I.Schwarz,T.Gramatte,J.Krappweis.P-glycoprotein inhibitor erythromycin increases oral bioavailability of talinolol in humans[J].In d Clin Pharm and Ther,2000,38:161-167
[45].Thomas grate,reinhard ortel.Intestinal secretion of intravenous talinolol is inhibited by luminal R-verapamil[J].Clin Pharmacol Ther,1999;66:239-245
[46].Weber CC,Kressmann S,Ott M,Fricker G,Muller WE.Inhibition of P-glycoprotein function by several antidepressants may not contribute to clinical efficacy[J].Pharmacopsychiatry,2005,38:293-300.
[47].Leibovitz A,Bilchinsky T,Gil I,Habot B.Elevated serum digoxin level associated with coadministered fluoxetine[J].Arch Intern Med,1998,158:1152-1153.
[48].Yasui-Furukori N,Kaneko S.Digitalis intoxication induced by paroxetine co-administration[J].Lancet.2006,367:788.
[49].Juurlink DN.Mamdani MM,Kopp A,et al.A population-based assessment of the potential interaction between serotonin specific reuptake inhibitors and digoxin [J].Br J Clin Pharmacol,2005;59:102-107.
[50].S Anuchapreeda,P Leechanachai,M Smith,et al.Modulation of P-glycoprotein expression and function by curcumin in multidrug-resistant human KB cells.Biochemical pharmacology[J].2002,64:573-582.
[1]Mickisch GH,Kossig J,Keilhauer G,et al.Efects of calcium antagonists in multidrug resistant primary human renal cell carcinomas[J].Cancer,1990,50:3670.
[2]Pol MA,Broxterman HJ,Paler JM,et al.Function of the ABC transporters,P-glycoprotein,multidrug resistance protein and breastcancer resistance protein,minimal residual disease in acute myeloid leukemia[J].Haematologica,2003,88:134.
[3]GraberA,Bjorkho|m M,Brineh R,et al.A phase Ⅰ/Ⅱ study of the MDR modulator PSC833combined with daunorubicin and cytarabine in patients with relapsed and primary refractory acute myeloid leukemia[J].Leuk Res,2003,27:323.
[4]Cruz F,Wolf A.Effects of the novel cyclosporine derivative PSC833 on glucose metabolism in rat primary cultures of neuronaland filial cells[J].Biochem Phannacol,2001,62:129.
[5]KalraR,Jones AM,Kirk J,et al.The effect of hypoxia on acquired drug resistance and response to epidermal growth factor in Chinese hamster lung fibroblasts and human breast-cancer cells in vitro[J].Cancer,1993,54:650.
[6]Dan P,Yaron D,Dina M,Rimona M(2004).Fluoxetine inhibits multidrug resistance extrusion pumps and enhances responses to chemotherapy in syngeneic and in human xenograft mouse tumor models.Cancer Re s.64,7562.
[7]Johanna W,Dormann SM,Martin-Facklam M,Kerpen CJ,Ketabi-Kiyanvash N,Haefeli WE (2003).Inhibition of P-glycoprotein by newer antidepressants.J.Pharmaacol.Exp.Ther.305,197.
[8]Limtrakul P,Anuchapreeda S,Buddhasukh D.Modulation of human multidrug-resistance MDR- 1 gene by natural curcuminoids[J].BMC Cancer,2004,17:4.
[9]Anuchapreeda S,Leechanachai P,Smith M,et al.Modulation of P-glycoprotein expression and function by curcumin in multidrug-resistant human KB cells[J].Biochem Pharmacol,2002,64:573.
[10]Fu L W,Deng Z A,Fan W,et al.Screening and discovery of novel MDR modifiers from naturally occurring bisbenzylisoquinoline alkaloids[J].Anticancer Res,2001,21:2273.
[11]陈宝安,杜鹃,张春秀,等.应用蛋白质芯片对汉防己甲素单用及与屈洛昔芬伍用逆转白血病细胞耐药机理的研究[J].中国实验血液学杂志,2005,13:999.
[12]Bae EA,Han MJ,Choo MK,et al.Metabolism of 20(S)- and 20(R)-ginsenoside Rg3 by human intestinal bacteria and its relation to in vitro biological activities[J].Biol Pharm Bull,2002,25:58.
[13]Kim S W,Kwon H Y,Chi D W,et al.Reversal of P-glycoprotein-mediated multidrug resistance by ginsenoside Rg(3)[J].Biochem Pharmacol,2003,65:75.
[14]Hasegawa H,Sung J H,Matsumiya S,et al.Reversal of daunomycin and vinblastine resistance in multidrug-resistant P388 leukemia in vitro through enhanced cytotoxicity by triterpenoids[J].Planta Med,1995,61:409.
[15]Choi C H,Kang G,Min Y D,et al.Reversal of P-glycoprotein-mediated multidrug resistance by protopanaxatriol ginsenosides from Korean red ginseng[J].Planta Med,2003,69:235.
[16]Jodoin J,Demeule M,Beliveau R.Inhibition of the multidrug resistance P-glycoprotein activity by green tea polyphenols[J].Biochim Biophys Acta,2002,1542:149.
[17]梁钢,张肃,黄志明,等.两种儿茶素成分体外逆转人肝癌细胞BEL-7404/ADR多药耐药性[J].癌症,2004,23:401.
[18]Sun M,Xu X,Lu Q,et al.Schisandrin B:A dual inhibitor of P-glycopmtein and multidrug resistance-associated protein 1[J].Cancer Lett,2007,246:300.
[19]Shen X,Chen G,Zhu G,et al.(+/-)-3'-0,4'-O-dicynnamoyl-cis-khellactone,a derivative of (+/-)-praeruptorin A,reverses P-glycoprotein mediated multidrug resistance in cancer cells[J].Bioorg Med Chem,2006,14:7138.
[20]韩艳秋,石永进,袁家颖,等.小檗胺逆转MCF7/ADR细胞耐药性及其机制探讨[J].解剖学报,2004,35:161.
[21]Fong WF,Wang C,Zhu GY,et al.Reversal of multidrug resistance in cancer cells by Rhizoma Alismatis extract[J].Phytomedicine,2007,14:160.
[2].Barrand MA,Bagrij T,Neo SY.Multidrug resistance-associated protein:a protein distinct from P-glycoprotein involved in cytoxic drug expulsion[J].Gen Pharma,1997,28:629-651.
[3].Hennessy M,Spiers JP.A primer on the mechanics of P-glycoprotein the multidrug transporter[J].Pharmacol Res,2007,(1):1 - 15.
[4].Schinkel AH.P-Glycoprotein,a gatekeeper in the blood-brain barrier[J].Adv Drug Deliv Rev,1999,36:179-194.
[5].Jin Sun,Zhong-Gui He,Gang Cheng,et al.Multidrug resistance P-glycoprotein:crucial significance in drug disposition and interaction.Med Sci Monit,2004,10:5-14.
[6].Scheffer GL,Pijnenborg AC,Smit EF,et al.Multidrug resistance related molecules in human and murine lung[J].J Clin Pathol,2002,55:332-339.
[7].Dietrich CG,Geier A,Oude Elferink RP.ABC of oral bioavailability:transporters as gatekeepers in the gut[J].Gut,2003,52:1788-1795.
[8].Chandra P,Brouwer KL.The complexities of hepatic drug transport:current knowledge and emerging concepts[J].Pharm Res,2004,21(5):719-735.
[9].Graft CL,Pollack GM.Drug transport at the blood-brain barrier and the choroid plexus[J].Curt Drug Metab,2004,5:95-108.
[10].Leslie EM,Deeley RG,Cole SP.Multidrug resistance proteins:role of P-glycoprotein,MRP1,MRP2,and BCRP(ABCG2) in tissue defense[J].Toxicol Appl Pharmacol,2005,204:216-237.
[11].Akiyama SI,Comwell MM,Kuwano M,et al.Most drugs that reverse multidrug resistance also inhibit photoaffinity labeling of p-glycoprotein by a vinblastine analog[J].Mol Pharmacol,1988,33:144-147.
[12].Richard B Kin.Drugs as P-glycoprotein substrates,inhibitors,and inducers [J].Drug Metab Rev,2002,34:47-54
[13].Seelig A.A general pattern for substrate recognition by P-glycoprotein[J].Eur J Biochem,1998,251:252-261.
[14].Anathea BW,Peter K.Duloxetine Hydrochloride[J].Nature Reviews,2004,3:907-908.
[15].Wimett L,Laustsen G.Duloxetine fights major depressive disorder[J].Nurse Pract,2005,30:6-7.
[16].Cruz MP,Gonzales ME,Jacobs J,et al.Duloxetine HCl(Cymbalta) for the treatment ofdepression,neuropathic pain,fibromyalgia and stress urinary incontinence [J].Drug Forecast,2006,31:84-97.
[17].Dan P,Yaron D,Dina M,Rimona M(2004).Fluoxetine inhibits multidrug resistance extrusion pumps and enhances responses to chemotherapy in syngeneic and in human xenograft mouse tumor models.Cancer Res,64,7562-7569.
[18].Johanna W,Dormann SM,Martin-Facklam M,Kerpen CJ,Ketabi-Kiyanvash N,Haefeli WE(2003).Inhibition of P-glycoprotein by newer antidepressants.J.Pharmaacol.Exp.Ther,305,197-204.
[19].刘志伟.Caco-2细胞单层模型及其在毒理学中的应用.卫生研究,2004,33:756-759.
[20].关溯,赵立子,陈杰,等.Caco-2细胞模型的建立及意义.山东医药,2005,45:1-3.
[21].Maurer H,H.Tenberken O,Kratzsch C,et al.Screening for library-assisted identification and fully validated quantification of 22 beta-blockers in blood plasma by liquid chromatography-mass spectrometry with atmospheric pressure chemical ionization.J.Chromatogr.A,2004,1058:169-181.
[22].Beate T,R Oertel,K Richter,et al.Disposition and bioavailability of the β1-adrenoceptor antagonist talinolol in man.Biopharm dru dispo,1995,16:403-414.
[23].Gramatte T,Oertel R,Terhaag B.Direct demonstration of small intestinal secretion and site-dependent absorption of the beta-blocker talinolol in humans.Clin Pharmacol Ther,1996,59:541-549.
[24].Krueger M,H,chenbach A,Terhaag B.Pharmacokinetics of oral talinolol following a single dose and during steady state in patients with chronic renal failure and healthy volunteers.In J Clin Pharm and Ther,2001,39:61-66.
[25].Ulrike W,Hildegard S.L,Ernst M.Evidence for intestinal secretion as an additional clearance pathway of talinolol enatiomers:concentration- and dose-dependent absorption in vitro and in vivo.Pharmaceutic Res,1996,13:514-522.
[26].Schwarz UI,Dresser GK,Oertel R,Kim RB.Talinolol-verapamil interaction is not solely due to P-glycoprotein inhibition[abstract].Clin Pharmacol Ther,2001,69,P85.
[27].Shirasaka Y,Kawasaki M,Sakane T,et al.Induction of human P-glycoprotein in Caco-2 cells:development of a highly sensitive assay system for P-glycoprotein-mediated drug transport[J].Drug Metab Pharmacokinet,2006,21:414-423.
[28].Han Y,Tan TM,Lim LY.Effects of capsaicin on P-gp function and expression in Caco-2 cells[J].Biochem Pharmacol,2006,71:1727-1734.
[29].张向荣,张逸凡,钟大放.体外Caco-2细胞模型在药物吸收中的应用进展[J].中国医院药学杂志,2004,24:773-775.
[30].Mosmann T.Rapid colorimetric assay for cellular growth and survival:application to proliferation and cytotoxicity assays.J Immunol Methods,1983,65:55-63.
[31].DuBuske LM.The role of P-glycoprotein and organic anion-transporting polypeptides in drug interactions[J].Drug Saf,2005,28:789-801.
[32].Couture L,Nash JA,Turgeon J.The ATP-binding cassette transporters and their implication in drug disposition:a special look at the heart[J].Pharmacol Rev,2006,58:244-258.
[33].Breier A,Barancik M,Sulova Z,et al.P-glycoprotein-implications of metabolism of neoplastic cells and cancer therapy[J].Curr Cancer Drug Targets,2005,5:457-468.
[34].Thomas H,Coley HM.Overcoming multidrug resistance in cancer:an update on the clinical strategy of inhibiting P-glycoprotein[J].Cancer Contro,2003,10:159-165.
[35].Petriz J,O'Connor JE,Carmona MIs,et al.rhodamine123 an appropriate fluorescent probe to assess P-glycoprotein mediated multidrug resistance in vinblastine-resistant CHO cells.Anal Cell Pathol,1997,14:129-140.
[36].Ryoko Yumoto,Teruo Murakami,Yuko Nakamoto,et al.Transport of Rhodamine 123,a P-Glycoprotein Substrate,across Rat Intestine and Caco-2 Cell Monolayers in the Presence of Cytochrome P-450 3A-Related Compounds.J Pharmacol Exp Ther,1999,289:149-155.
[37].He L,Liu GQ.Effects of various principles from Chinese herbal medicine on rhodamine 123 accumulation in brain capillary endothelial cells.Acta Pharm Sin,2002,23:591-596.
[38].Anuchapreeda S,Leechanachai P,Smith M,et al.Modulation of P-glycoprotein expression and function by curcumin in multidrug-resistant human KB cells[J].Biochem Pharmacol,2002,64:573.
[39].Durr,D,Stieger B,Kullak-Ublick,GA,et al.St.John's wort induces intestinal P-glycoprotein/MDR1 and intestinal and hepatic CYP3A4[J].Clin Pharmacol Ther,2000,68:598-604.
[40].Hennessy M,Kelleher D,Spiers JP,et al.St.John's wort increases expression of P-glycoprotein:implications for drug interactions[J].Br J Clin Pharmacol,2002,53:75.
[41].Matsuyama N,Kojima.Simultaneous five cell-lineage flow cytometric analysis system for detection of leucocyte antibodies[J].Transfusion Medicine,2006,16:111-118.
[42].Katrien S,Barbara DM,Catharina D,et al.Comparison of two functional flow cytometric assays to assess P-gp activity in acute leukemia[J].Leukemia and Lymphoma,2004,11:2221-2228.
[43].Han Y,Tan TM,Lim LY.Effects of capsaicin on P-gp function and expression in Caco-2 cells[J].Biochem Pharmacol,2006,71:1727-1734.
[44].U.I.Schwarz,T.Gramatte,J.Krappweis.P-glycoprotein inhibitor erythromycin increases oral bioavailability of talinolol in humans[J].In d Clin Pharm and Ther,2000,38:161-167
[45].Thomas grate,reinhard ortel.Intestinal secretion of intravenous talinolol is inhibited by luminal R-verapamil[J].Clin Pharmacol Ther,1999;66:239-245
[46].Weber CC,Kressmann S,Ott M,Fricker G,Muller WE.Inhibition of P-glycoprotein function by several antidepressants may not contribute to clinical efficacy[J].Pharmacopsychiatry,2005,38:293-300.
[47].Leibovitz A,Bilchinsky T,Gil I,Habot B.Elevated serum digoxin level associated with coadministered fluoxetine[J].Arch Intern Med,1998,158:1152-1153.
[48].Yasui-Furukori N,Kaneko S.Digitalis intoxication induced by paroxetine co-administration[J].Lancet.2006,367:788.
[49].Juurlink DN.Mamdani MM,Kopp A,et al.A population-based assessment of the potential interaction between serotonin specific reuptake inhibitors and digoxin [J].Br J Clin Pharmacol,2005;59:102-107.
[50].S Anuchapreeda,P Leechanachai,M Smith,et al.Modulation of P-glycoprotein expression and function by curcumin in multidrug-resistant human KB cells.Biochemical pharmacology[J].2002,64:573-582.
[1]Mickisch GH,Kossig J,Keilhauer G,et al.Efects of calcium antagonists in multidrug resistant primary human renal cell carcinomas[J].Cancer,1990,50:3670.
[2]Pol MA,Broxterman HJ,Paler JM,et al.Function of the ABC transporters,P-glycoprotein,multidrug resistance protein and breastcancer resistance protein,minimal residual disease in acute myeloid leukemia[J].Haematologica,2003,88:134.
[3]GraberA,Bjorkho|m M,Brineh R,et al.A phase Ⅰ/Ⅱ study of the MDR modulator PSC833combined with daunorubicin and cytarabine in patients with relapsed and primary refractory acute myeloid leukemia[J].Leuk Res,2003,27:323.
[4]Cruz F,Wolf A.Effects of the novel cyclosporine derivative PSC833 on glucose metabolism in rat primary cultures of neuronaland filial cells[J].Biochem Phannacol,2001,62:129.
[5]KalraR,Jones AM,Kirk J,et al.The effect of hypoxia on acquired drug resistance and response to epidermal growth factor in Chinese hamster lung fibroblasts and human breast-cancer cells in vitro[J].Cancer,1993,54:650.
[6]Dan P,Yaron D,Dina M,Rimona M(2004).Fluoxetine inhibits multidrug resistance extrusion pumps and enhances responses to chemotherapy in syngeneic and in human xenograft mouse tumor models.Cancer Re s.64,7562.
[7]Johanna W,Dormann SM,Martin-Facklam M,Kerpen CJ,Ketabi-Kiyanvash N,Haefeli WE (2003).Inhibition of P-glycoprotein by newer antidepressants.J.Pharmaacol.Exp.Ther.305,197.
[8]Limtrakul P,Anuchapreeda S,Buddhasukh D.Modulation of human multidrug-resistance MDR- 1 gene by natural curcuminoids[J].BMC Cancer,2004,17:4.
[9]Anuchapreeda S,Leechanachai P,Smith M,et al.Modulation of P-glycoprotein expression and function by curcumin in multidrug-resistant human KB cells[J].Biochem Pharmacol,2002,64:573.
[10]Fu L W,Deng Z A,Fan W,et al.Screening and discovery of novel MDR modifiers from naturally occurring bisbenzylisoquinoline alkaloids[J].Anticancer Res,2001,21:2273.
[11]陈宝安,杜鹃,张春秀,等.应用蛋白质芯片对汉防己甲素单用及与屈洛昔芬伍用逆转白血病细胞耐药机理的研究[J].中国实验血液学杂志,2005,13:999.
[12]Bae EA,Han MJ,Choo MK,et al.Metabolism of 20(S)- and 20(R)-ginsenoside Rg3 by human intestinal bacteria and its relation to in vitro biological activities[J].Biol Pharm Bull,2002,25:58.
[13]Kim S W,Kwon H Y,Chi D W,et al.Reversal of P-glycoprotein-mediated multidrug resistance by ginsenoside Rg(3)[J].Biochem Pharmacol,2003,65:75.
[14]Hasegawa H,Sung J H,Matsumiya S,et al.Reversal of daunomycin and vinblastine resistance in multidrug-resistant P388 leukemia in vitro through enhanced cytotoxicity by triterpenoids[J].Planta Med,1995,61:409.
[15]Choi C H,Kang G,Min Y D,et al.Reversal of P-glycoprotein-mediated multidrug resistance by protopanaxatriol ginsenosides from Korean red ginseng[J].Planta Med,2003,69:235.
[16]Jodoin J,Demeule M,Beliveau R.Inhibition of the multidrug resistance P-glycoprotein activity by green tea polyphenols[J].Biochim Biophys Acta,2002,1542:149.
[17]梁钢,张肃,黄志明,等.两种儿茶素成分体外逆转人肝癌细胞BEL-7404/ADR多药耐药性[J].癌症,2004,23:401.
[18]Sun M,Xu X,Lu Q,et al.Schisandrin B:A dual inhibitor of P-glycopmtein and multidrug resistance-associated protein 1[J].Cancer Lett,2007,246:300.
[19]Shen X,Chen G,Zhu G,et al.(+/-)-3'-0,4'-O-dicynnamoyl-cis-khellactone,a derivative of (+/-)-praeruptorin A,reverses P-glycoprotein mediated multidrug resistance in cancer cells[J].Bioorg Med Chem,2006,14:7138.
[20]韩艳秋,石永进,袁家颖,等.小檗胺逆转MCF7/ADR细胞耐药性及其机制探讨[J].解剖学报,2004,35:161.
[21]Fong WF,Wang C,Zhu GY,et al.Reversal of multidrug resistance in cancer cells by Rhizoma Alismatis extract[J].Phytomedicine,2007,14:160.