Division of Pharmaceutical Chemistry and Technology, and ‡Division of Pharmaceutical Biosciences, Faculty of Pharmacy, University of Helsinki , 00014 University of Helsinki, Finland.
Mol Pharm. 2017 Oct 2;14(10):3299-3311. doi: 10.1021/acs.molpharmaceut.7b00366. Epub 2017 Sep 13.
Xenobiotic and endobiotic glucuronides, which are generated in hepatic and intestinal epithelial cells, are excreted via efflux transporters. Multidrug resistance proteins 2-4 (MRP2-MRP4) and the breast cancer resistance protein (BCRP) are efflux transporters that are expressed in these polarized cells, on either the basolateral or apical membranes. Their localization, along with expression levels, affects the glucuronide excretion pathways. We have studied the transport of three planar cyclic glucuronides and glucuronides of the two propranolol enantiomers, by the vesicular transport assay, using vesicles from baculovirus-infected insect cells expressing human MRP2, MRP3, MRP4, or BCRP. The transport of estradiol-17β-glucuronide by recombinant MRP2-4 and BCRP, as demonstrated by kinetic values, were within the ranges previously reported. Our results revealed high transport rates and apparent affinity of MRP4 toward the glucuronides of 4-methylumbelliferone, 1-naphthol, and 1-hydroxypyrene (K values of 168, 13, and 3 μM, respectively) in comparison to MRP3 (K values of 278, 98, and 8 μM, respectively). MRP3 exhibited lower rates, but stereoselective transport of propranolol glucuronides, with higher affinity toward the R-enantiomer than the S-enantiomer (K values 154 vs 434 μM). The glucuronide of propranolol R-enantiomer was not significantly transported by either MRP2, MRP4, or BCRP. Of the tested small glucuronides in this study, BCRP transported only 1-hydroxypyrene glucuronide, at very high rates and high apparent affinity (V and K values of 4400 pmol/mg/min and 11 μM). The transport activity of MRP2 with all of the studied small glucuronides was relatively very low, even though it transported the reference compound, estradiol-17β-glucuronide, at a high rate (V = 3500 pmol/mg/min). Our results provide new information, at the molecular level, of efflux transport of the tested glucuronides, which could explain their disposition in vivo, as well as provide new tools for in vitro studies of MRP3, MRP4, and BCRP.
外源性和内源性葡萄糖醛酸苷在肝和肠上皮细胞中生成，然后通过外排转运体排泄。多药耐药相关蛋白 2-4（MRP2-MRP4）和乳腺癌耐药蛋白（BCRP）是在这些极化细胞中表达的外排转运体，位于基底外侧或顶端膜上。它们的定位和表达水平会影响葡萄糖醛酸苷的排泄途径。我们通过使用昆虫细胞来源的囊泡进行囊泡转运试验，研究了三种平面环状葡萄糖醛酸苷和两种普萘洛尔对映体葡萄糖醛酸苷的转运，该囊泡是由表达人 MRP2、MRP3、MRP4 或 BCRP 的杆状病毒感染的昆虫细胞制备的。从动力学值来看，重组 MRP2-4 和 BCRP 转运雌二醇-17β-葡萄糖醛酸苷的结果表明，其转运速率和对映体亲和力都在之前报道的范围内。与 MRP3（K 值分别为 278、98 和 8 μM）相比，我们的结果显示 MRP4 对 4-甲基伞形酮、1-萘酚和 1-羟基芘葡萄糖醛酸苷具有较高的转运速率和亲和力（K 值分别为 168、13 和 3 μM）。MRP3 对普萘洛尔葡萄糖醛酸苷的转运速率较低，但具有立体选择性，对 R-对映体的亲和力高于 S-对映体（K 值分别为 154 和 434 μM）。MRP2、MRP4 或 BCRP 均不能显著转运普萘洛尔 R-对映体葡萄糖醛酸苷。在本研究中测试的小分子葡萄糖醛酸苷中，BCRP 仅以非常高的速率和高表观亲和力转运 1-羟基芘葡萄糖醛酸苷（V 和 K 值分别为 4400 pmol/mg/min 和 11 μM）。尽管 MRP2 以高速率转运参比化合物雌二醇-17β-葡萄糖醛酸苷（V = 3500 pmol/mg/min），但它对所有测试的小分子葡萄糖醛酸苷的转运活性都非常低。我们的研究结果提供了分子水平上测试的葡萄糖醛酸苷外排转运的新信息，这可以解释它们在体内的分布情况，并且为 MRP3、MRP4 和 BCRP 的体外研究提供了新工具。
