Abstract：
Coproporphyrin (CP) I and III are byproducts of haem synthesis currently investigated as biomarkers for drug-drug interactions involving hepatic organic anion transporting polypeptide (OATP) 1B transporters. Another hepatically expressed OATP-member is OATP2B1. The aim of this study was to test the impact of OATP2B1, which specifically transports CPIII, on CP serum levels, applying novel rat models.
CPIII transport kinetics and the interplay between OATP2B1 and multidrug resistance-associated proteins (MRPs) were determined in vitro using the vTF7 expression system. Novel rSlco2b1-/- and SLCO2B1+/+ rat models were characterized for physiological parameters and for CP serum levels. Hepatic and renal expression of transporters involved in CP disposition were determined by real-time qPCR, Western blot analysis, and immunohistochemistry.
In vitro experiments revealed differences in transport kinetics comparing human and rat OATP2B1 and showed a consistent, species-specific interplay with hMRP3/rMRP3. Deletion of rOATP2B1 was associated with a trend towards lower CPI serum levels compared with wildtype rats, while CPIII remained unchanged. Comparing SLCO2B1+/+ with knockout rats revealed an effect of sex: only in females the genetic modification influenced CP serum levels. Analysis of hepatic and renal transporters revealed marginal, but in part, statistically significant differences in rMRP2 abundance, which may contribute to the observed changes in CP serum levels.
Our findings support that factors other than OATP1B transporters are of relevance for basal CP levels. Only in female rats, humanization of SLCO2B1 affects basal CPI and CPIII serum levels, despite isomer selectivity of OATP2B1.
CPIII transport kinetics and the interplay between OATP2B1 and multidrug resistance-associated proteins (MRPs) were determined in vitro using the vTF7 expression system. Novel rSlco2b1-/- and SLCO2B1+/+ rat models were characterized for physiological parameters and for CP serum levels. Hepatic and renal expression of transporters involved in CP disposition were determined by real-time qPCR, Western blot analysis, and immunohistochemistry.
In vitro experiments revealed differences in transport kinetics comparing human and rat OATP2B1 and showed a consistent, species-specific interplay with hMRP3/rMRP3. Deletion of rOATP2B1 was associated with a trend towards lower CPI serum levels compared with wildtype rats, while CPIII remained unchanged. Comparing SLCO2B1+/+ with knockout rats revealed an effect of sex: only in females the genetic modification influenced CP serum levels. Analysis of hepatic and renal transporters revealed marginal, but in part, statistically significant differences in rMRP2 abundance, which may contribute to the observed changes in CP serum levels.
Our findings support that factors other than OATP1B transporters are of relevance for basal CP levels. Only in female rats, humanization of SLCO2B1 affects basal CPI and CPIII serum levels, despite isomer selectivity of OATP2B1.
摘要：
目的：卟啉（CP）I和CPIII是血红素合成的副产物，目前作为涉及肝脏有机阴离子转运多肽（OATP）1B转运蛋白的药物-药物相互作用的生物标志物进行研究。OATP家族的另一个肝表达成员是OATP2B1。这项研究的目的是测试OATP2B1的影响，它特别运输CPIII，应用新型大鼠模型对CP血清水平的影响。
方法：使用vTF7表达系统在体外测定了CPIII转运动力学以及OATP2B1和MRP之间的相互作用。对新型rSlco2b1-/-和SLCO2B1+/+大鼠模型的生理参数进行了表征，以及共产卟啉血清水平。通过实时qPCR确定参与CP处置的转运体的肝和肾表达，蛋白质印迹分析和免疫组织化学。
结果：体外实验揭示了人和大鼠OATP2B1的转运动力学差异，与hMRP3/rMRP3的物种特异性相互作用。与野生型大鼠相比，rOATP2B1的缺失与CPI血清水平降低的趋势有关，而CPIII保持不变。比较SLCO2B1+/+与基因敲除大鼠显示性别的影响：仅在雌性中，遗传修饰影响CP血清水平。肝肾转运体的分析显示边缘，但在某种程度上，rMRP2丰度的统计学差异，这可能有助于观察到CP血清水平的变化。
结论：我们的研究结果支持OATP1B转运蛋白以外的其他因素与基础CP水平相关。只有雌性老鼠，SLCO2B1的人源化影响基础CPI和CPIII血清水平，尽管OATP2B1的异构体选择性。
方法：使用vTF7表达系统在体外测定了CPIII转运动力学以及OATP2B1和MRP之间的相互作用。对新型rSlco2b1-/-和SLCO2B1+/+大鼠模型的生理参数进行了表征，以及共产卟啉血清水平。通过实时qPCR确定参与CP处置的转运体的肝和肾表达，蛋白质印迹分析和免疫组织化学。
结果：体外实验揭示了人和大鼠OATP2B1的转运动力学差异，与hMRP3/rMRP3的物种特异性相互作用。与野生型大鼠相比，rOATP2B1的缺失与CPI血清水平降低的趋势有关，而CPIII保持不变。比较SLCO2B1+/+与基因敲除大鼠显示性别的影响：仅在雌性中，遗传修饰影响CP血清水平。肝肾转运体的分析显示边缘，但在某种程度上，rMRP2丰度的统计学差异，这可能有助于观察到CP血清水平的变化。
结论：我们的研究结果支持OATP1B转运蛋白以外的其他因素与基础CP水平相关。只有雌性老鼠，SLCO2B1的人源化影响基础CPI和CPIII血清水平，尽管OATP2B1的异构体选择性。
