Mechanism of inhibition of P-glycoprotein mediated efflux by vitamin E TPGS:: Influence on ATPase activity and membrane fluidity
Mechanism of inhibition of P-glycoprotein mediated efflux by vitamin E TPGS:: Influence on ATPase activity and membrane fluidity
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DOI：
10.1021/mp060121r
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发表时间：
2007-05-01
影响因子：
4.9
通讯作者：
Lehr, Claus-Michael
中科院分区：
文献类型：
--
作者：
Collnot, Eva-Maria;Baldes, Christiane;Lehr, Claus-Michael
外排泵(如P-gp、MRP1和BCRP)抑制已被认为是克服多药耐药和提高药物生物利用度的一种策略。除了小分子抑制剂，已知的表面活性剂如吐温80、Cremophor EL、几种Pluronics和维生素E TPGS(TPGS 1000)可以调节外排泵的活性。竞争抑制底物结合、改变膜流动性和抑制外排泵ATPase被认为是可能的机制。围绕TPGS 1000，我们的研究目的是通过比较Caco-2转运实验中的抑制实验结果与电子自旋共振(ESR)和ATPase活性研究的数据来揭示其抑制机制。使用5-十二烷基硬脂酸(5-SA)作为自旋探针的Caco-2细胞的ESR结果排除了细胞膜流态化的主要贡献；只有在表面活性剂浓度高于实现完全外流抑制所需浓度的100倍时，才能观察到膜流动性的变化。同时，TPGS 1000抑制底物诱导的ATPase活性，而不单独诱导显著的ATPase活性。通过研究不同聚乙二醇链长度的TPGS类似物和/或具有修饰的疏水核心的TPGS类似物，转运研究表明ATPase活性的调节与P-gp介导的外流的抑制潜力有关。因此，这些结果表明ATPase抑制是TPGS 1000抑制细胞外排泵机制中的一个重要因素。
Efflux pump (e.g., P-gp, MRP1, and BCRP) inhibition has been recognized as a strategy to overcome multi-drug resistance and improve drug bioavailability. Besides small-molecule inhibitors, surfactants such as Tween 80, Cremophor EL, several Pluronics, and Vitamin E TPGS (TPGS 1000) are known to modulate efflux pump activity. Competitive inhibition of substrate binding, alteration of membrane fluidity, and inhibition of efflux pump ATPase have been proposed as possible mechanisms. Focusing on TPGS 1000, the aim of our study was to unravel the inhibitory mechanism by comparing the results of inhibition experiments in a Caco-2 transport assay with data from electron spin resonance (ESR) and from ATPase activity studies. ESR results, on Caco-2 cells using 5-doxyl stearic acid (5-SA) as a spin probe, ruled out cell membrane fluidization as a major contributor; change of membrane fluidity was only observed at surfactant concentrations 100 times higher than those needed to achieve full efflux inhibition. Concurrently, TPGS 1000 inhibited substrate induced ATPase activity without inducing significant ATPase activity on its own. By investigating TPGS analogues that varied by their PEG chain length, and/or possessed a modified hydrophobic core, transport studies revealed that modulation of ATPase activity correlated with inhibitory potential for P-gp mediated efflux. Hence, these results indicate that ATPase inhibition is an essential factor in the inhibitory mechanism of TPGS 1000 on cellular efflux pumps.