Culot Maxime, Fabulas-da Costa Anaëlle, Sevin Emmanuel, Szorath Erica, Martinsson Stefan, Renftel Mila, Hongmei Yan, Cecchelli Romeo, Lundquist Stefan
BBB Laboratory EA 2465, IMPRT: IFR114 Université Lille Nord de France, UArtois, Lens, France.
PLoS One. 2013 Dec 3;8(12):e80634. doi: 10.1371/journal.pone.0080634. eCollection 2013.
Historically, the focus has been to use in vitro BBB models to optimize rate of drug delivery to the CNS, whereas total in vivo brain/plasma ratios have been used for optimizing extent. However, these two parameters do not necessarily show good correlations with receptor occupancy data or other pharmacological readouts. In line with the free drug hypothesis, the use of unbound brain concentrations (Cu,br) has been shown to provide the best correlations with pharmacological data. However, typically the determination of this parameter requires microdialysis, a technique not ideally suited for screening in early drug development. Alternative, and less resource-demanding methodologies to determine Cu,br employ either equilibrium dialysis of brain homogenates or incubations of brain slices in buffer to determine fraction unbound brain (fu,br), which is subsequently multiplied by the total brain concentration to yield Cu,br. To determine Cu,br/Cu,pl ratios this way, still requires both in vitro and in vivo experiments that are quite time consuming. The main objective of this study was to explore the possibility to directly generate Cu,br/Cu,pl ratios in a single in vitro model of the BBB, using a co-culture of brain capillary endothelial and glial cells in an attempt to mimick the in vivo situation, thereby greatly simplifying existing experimental procedures. Comparison to microdialysis brain concentration profiles demonstrates the possibility to estimate brain exposure over time in the BBB model. A stronger correlation was found between in vitro Cu,br/Cu,pl ratios and in vivo Cu,br/Cu,pl obtained using fu,br from brain slice than with fu,br from brain homogenate for a set of 30 drugs. Overall, Cu,br/Cu,pl ratios were successfully predicted in vitro for 88% of the 92 studied compounds. This result supports the possibility to use this methodology for identifying compounds with a desirable in vivo response in the CNS early on in the drug discovery process.
从历史上看，重点一直是使用体外血脑屏障模型来优化药物向中枢神经系统的递送速率，而体内脑/血浆总比率则用于优化药物递送程度。然而，这两个参数不一定与受体占有率数据或其他药理学指标显示出良好的相关性。根据游离药物假说，已证明使用未结合的脑浓度（Cu,br）能与药理学数据提供最佳相关性。然而，通常该参数的测定需要微透析，这是一种不太适合早期药物开发筛选的技术。用于测定Cu,br的替代且资源需求较少的方法，要么采用脑匀浆的平衡透析，要么将脑片在缓冲液中孵育以测定未结合脑分数（fu,br），随后将其乘以总脑浓度以得出Cu,br。要以此方式测定Cu,br/Cu,pl比率，仍需要耗时的体外和体内实验。本研究的主要目的是探索在单一的血脑屏障体外模型中直接生成Cu,br/Cu,pl比率的可能性，使用脑毛细血管内皮细胞和神经胶质细胞的共培养来模拟体内情况，从而极大地简化现有实验程序。与微透析脑浓度曲线的比较表明，在血脑屏障模型中有可能估计随时间的脑暴露情况。对于一组30种药物，发现体外Cu,br/Cu,pl比率与使用脑片的fu,br获得的体内Cu,br/Cu,pl之间的相关性，比与使用脑匀浆的fu,br之间的相关性更强。总体而言，在体外成功预测了92种研究化合物中88%的Cu,br/Cu,pl比率。这一结果支持了在药物发现过程早期使用该方法来识别在中枢神经系统具有理想体内反应的化合物的可能性。
