University of Coimbra, Faculty of Pharmacy, Coimbra, Portugal; University of Coimbra, Coimbra Institute for Biomedical Imaging and Translational Research, Coimbra, Portugal.
CICS-UBI - Health Sciences Research Centre, University of Beira Interior, Covilhã, Portugal.
Int J Pharm. 2021 Jan 25;593:120161. doi: 10.1016/j.ijpharm.2020.120161. Epub 2020 Dec 9.
The breast cancer resistance protein (BCRP) is an efflux transporter expressed at the apical surface of human brain endothelial cells of the blood-brain barrier (BBB). It was proposed as one of the transporters responsible for the development of drug resistance to several central nervous system (CNS) drugs, including antiepileptic drugs (AEDs). In this context, the present work aimed to characterize the interaction between new-generation AEDs, lacosamide, levetiracetam and zonisamide, and BCRP, in order to investigate whether intranasal administration can successfully avoid the impact of BCRP on brain drug distribution, preventing the development of refractory epilepsy. Firstly, BCRP substrates and/or inhibitors were identified resorting to intracellular accumulation and bidirectional transport assays on Madin-Darby canine kidney (MDCK) cells and the transfected cell line with human ABCG2 (MDCK-BCRP). Furthermore, in vivo pharmacokinetic studies were carried out for BCRP substrates with and without elacridar, a well-known P-gp and BCRP modulator, to assess the impact of efflux inhibition on brain drug distribution. The extent of drug equilibration between plasma and brain was compared after intravenous (IV) and intranasal administration to mice. Among the three tested AEDs, zonisamide was the only AED identified as BCRP substrate in vitro, as demonstrated by the net flux ratio of 2.73, which decreased 53.85 % in the presence of a BCRP inhibitor, Ko143. Lacosamide revealed to inhibit BCRP in all tested concentrations (2.5-75 µM), exhibiting a significant increase (p < 0.001) of the intracellular accumulation of a BCRP substrate (Hoechst 33342) in MDCK-BCRP cells. Levetiracetam did not behave as a BCRP substrate nor inhibitor. After IV administration, the plasma concentrations of zonisamide were unaffected by elacridar, but its extent of brain exposure increased three-fold (as assessed by AUC, 674.12 vs 284.47 µg.min/mL). These results corroborate the previous in vitro findings, suggesting that BCRP is involved in the transport of zonisamide through the BBB. In opposition, no significant changes were found in plasma or brain concentrations after the administration of zonisamide by intranasal route, indicating that the influence of BCRP is less relevant than for IV route. In addition, direct nose-to-brain delivery of zonisamide, given by the direct transport percentage, was approximately 49 %. Altogether, these assays demonstrated that the impact of BCRP on the delivery of zonisamide to the brain is lower after intranasal administration, probably due to direct nose-to-brain transport. Therefore, the intranasal administration of AEDs may be a relevant strategy to avoid the impact of efflux transporters at the BBB and the development of drug resistance.
乳腺癌耐药蛋白（BCRP）是一种位于人血脑屏障（BBB）脑内皮细胞顶表面的外排转运体。它被认为是导致几种中枢神经系统（CNS）药物耐药的转运体之一，包括抗癫痫药物（AEDs）。在这种情况下，本工作旨在研究新一代 AED 拉考沙胺、左乙拉西坦和唑尼沙胺与 BCRP 之间的相互作用，以研究经鼻给药是否能成功避免 BCRP 对脑内药物分布的影响，从而预防难治性癫痫的发生。首先，通过在 Madin-Darby 犬肾（MDCK）细胞和转染有人类 ABCG2（MDCK-BCRP）的细胞系上进行细胞内积累和双向转运试验，确定了 BCRP 的底物和/或抑制剂。此外，还进行了体内药代动力学研究，以评估外排抑制对脑内药物分布的影响，研究对象为有和没有已知 P-糖蛋白和 BCRP 调节剂埃拉西达的 BCRP 底物。比较了静脉（IV）和鼻内给药后小鼠血浆和脑内药物平衡程度。在三种测试的 AED 中，唑尼沙胺是唯一一种在体外被鉴定为 BCRP 底物的 AED，净通量比为 2.73，当存在 BCRP 抑制剂 Ko143 时，该值降低了 53.85%。拉考沙胺在所有测试浓度（2.5-75μM）下均显示出对 BCRP 的抑制作用，在 MDCK-BCRP 细胞中，对 BCRP 底物（Hoechst 33342）的细胞内积累有显著增加（p<0.001）。左乙拉西坦既不是 BCRP 底物也不是抑制剂。静脉给药后，埃拉西达对唑尼沙胺的血浆浓度没有影响，但脑内暴露量增加了三倍（AUC 评估，674.12 与 284.47μg·min·mL-1）。这些结果与之前的体外研究结果一致，表明 BCRP 参与了唑尼沙胺通过 BBB 的转运。相反，经鼻给予唑尼沙胺后，血浆或脑内浓度无明显变化，表明 BCRP 的影响小于 IV 途径。此外，直接通过鼻内途径给予的唑尼沙胺的直接脑内输送百分比约为 49%。总之，这些试验表明，经鼻给予唑尼沙胺后，BCRP 对唑尼沙胺向脑内输送的影响降低，这可能是由于直接从鼻到脑的输送。因此，AED 的经鼻给药可能是一种相关策略，可以避免 BBB 中流出转运体的影响和药物耐药性的发展。
