肠壁中的药物代谢影响口服药物的生物利用度,并受年龄的影响。因此,充分了解肠道的药物代谢能力对预测全身暴露非常重要.这项研究的目的是研究肠样物质作为研究儿童和成人CYP3A4/5介导的代谢的工具的潜力。咪达唑仑的生物转化,CYP3A4/5型号基材,在Ussing室中使用肠样单层和组织外植体进行了研究,两者均来自儿科[中位数(范围年龄):54周(2天-13岁),n=21]和成人(n=5)组织。采用Caco-2细胞单层作为对照。此外,CYP3A4的mRNA表达在肠样单层(n=11)中测定,组织(n=23)和Caco-2使用RT-qPCR。咪达唑仑代谢在所有的肠样膜中成功检测到,以及在Ussing室研究的所有组织外植体中,而Caco-2显示没有明显的代谢物形成。在肠样单层和组织之间,咪达唑仑的提取部分相似。在来自0-70周龄供体的肠样单层中,提取的咪达唑仑的比例随着年龄的增长而增加。在组织中未观察到统计学上显著的相关性,这可能是由于观察到的高变异性和研究中包括的较小供体数量。在基因表达水平,组织中CYP3A4随年龄增加(n=32),而这没有反映在肠样结构单层中(n=16)。值得注意的是,在类肠和组织中观察到不对称代谢物形成,在屏障的腔侧形成较高的代谢物。总之,我们证明肠样物质可用于测量CYP3A4/5咪达唑仑代谢,我们显示与在新鲜分离的组织中观察到的相似。儿童和成人都是如此,表明肠样物质预测肠道代谢的潜力。这项研究为进一步开发肠样物质以研究体外药物代谢提供了有希望的数据,并有可能预测特殊人群的口服吸收,以替代使用新鲜组织。
Drug metabolism in the intestinal wall affects bioavailability of orally administered drugs and is influenced by age. Hence, it is important to fully understand the drug metabolizing capacity of the gut to predict systemic exposure. The aim of this study was to investigate the potential of
enteroids as a tool to study CYP3A4/5 -mediated metabolism in both children and adults. Bioconversion of midazolam, a CYP3A4/5 model substrate, was studied using enteroid monolayers as well as tissue explants in the Ussing chamber, both derived from pediatric [median (range age): 54 weeks (2 days - 13 years), n = 21] and adult (n = 5) tissue. Caco-2 cellular monolayers were employed as controls. In addition, mRNA expression of CYP3A4 was determined in enteroid monolayers (n = 11), tissue (n = 23) and Caco-2 using RT-qPCR. Midazolam metabolism was successfully detected in all enteroid monolayers, as well as in all tissue explants studied in the Ussing chamber, whereas Caco-2 showed no significant metabolite formation. The extracted fraction of midazolam was similar between enteroid monolayers and tissue. The fraction of midazolam extracted increased with age in enteroid monolayers derived from 0 to 70 week old donors. No statistically significant correlation was observed in tissue likely due to high variability observed and the smaller donor numbers included in the study. At the level of gene expression, CYP3A4 increased with age in tissues (n = 32), while this was not reflected in enteroid monolayers (n = 16). Notably, asymmetric metabolite formation was observed in
enteroids and tissue, with higher metabolite formation on the luminal side of the barrier. In summary, we demonstrated that
enteroids can be used to measure CYP3A4/5 midazolam metabolism, which we show is similar as observed in fresh isolated tissue. This was the case both in children and adults, indicating the potential of
enteroids to predict intestinal metabolism. This study provides promising data to further develop
enteroids to study drug metabolism in vitro and potentially predict oral absorption for special populations as an alternative to using fresh tissue.