Pihlaja Tea, Oksanen Timo, Vinkvist Netta, Sikanen Tiina
Faculty of Pharmacy, Drug Research Program, University of Helsinki, Helsinki, Finland.
Helsinki Institute of Sustainability Science, University of Helsinki, Helsinki, Finland.
Front Toxicol. 2024 Jul 15;6:1406942. doi: 10.3389/ftox.2024.1406942. eCollection 2024.
Pharmaceutical residues are widely detected in aquatic environment and can be taken up by nontarget species such as fish. The cytochromes P450 (CYP) represent an important detoxification mechanism in fish, like in humans. In the present study, we assessed the correlation of the substrate selectivities of rainbow trout CYP1A and CYP3A homologues with those of human, through determination of the half-maximal inhibitory concentrations (IC) of a total sixteen human pharmaceuticals toward CYP1A-like ethoxyresorufin O-deethylase (EROD) and CYP3A-like 7-benzyloxy-4-trifluoromethylcoumarin O-debenzylase (BFCOD) in rainbow trout () liver S9 fractions (RT-S9).
The inhibitory impacts (IC) of atomoxetine, atorvastatin, azelastine, bimatoprost, clomethiazole, clozapine, desloratadine, disulfiram, esomeprazole, felbinac, flecainide, orphenadrine, prazosin, quetiapine, sulpiride, and zolmitriptan toward the EROD and BFCOD activities in RT-S9 were determined using the IC shift assay, capable of identifying time-dependent inhibitors (TDI). Additionally, the nonspecific binding of the test pharmaceuticals to RT-S9 was assessed using equilibrium dialysis.
Most test pharmaceuticals were moderate to weak inhibitors of both EROD and BFCOD activity in RT-S9, even if most are noninhibitors of human CYP1A or CYP3A. Only bimatoprost, clomethiazole, felbinac, sulpiride, and zolmitriptan did not inhibit either activity in RT-S9. EROD inhibition was generally stronger than that of BFCOD and some substances (atomoxetine, flecainide, and prazosin) inhibited selectively only EROD activity. The strongest EROD inhibition was detected with azelastine and esomeprazole (unbound IC of 3.8 ± 0.5 µM and 3.0 ± 0.8 µM, respectively). None of the test substances were TDIs of BFCOD, but esomeprazole was a TDI of EROD. Apart from clomethiazole and disulfiram, the nonspecific binding of the test pharmaceuticals to the RT-S9 was extensive (unbound fractions <0.5) and correlated well ( = 0.7135) with their water-octanol distribution coefficients.
The results indicate that the P450 interactions in RT-S9 cannot be explicitly predicted based on human data, but the data reported herein can shed light on the substrate selectivity of rainbow trout CYP1A1 and CYP3A27 in comparison to their human homologues. The IC concentrations are however many orders of magnitude higher than average environmental concentrations of pharmaceuticals. The time-dependent EROD inhibition by esomeprazole could warrant further research to evaluate its possible interlinkages with hepatotoxic impacts on fish.
药物残留广泛存在于水生环境中，并可被鱼类等非靶标物种摄取。细胞色素P450（CYP）在鱼类中代表着一种重要的解毒机制，与人类相似。在本研究中，我们通过测定16种人类药物对虹鳟鱼肝S9组分（RT-S9）中CYP1A样乙氧异吩唑酮O-脱乙基酶（EROD）和CYP3A样7-苄氧基-4-三氟甲基香豆素O-脱苄基酶（BFCOD）的半数抑制浓度（IC），评估了虹鳟CYP1A和CYP3A同系物与人类底物选择性的相关性。
使用IC位移测定法测定了托莫西汀、阿托伐他汀、氮卓斯汀、比马前列素、氯美噻唑、氯氮平、地氯雷他定、双硫仑、埃索美拉唑、非布索坦、氟卡尼、奥芬那君、哌唑嗪、喹硫平、舒必利和佐米曲普坦对RT-S9中EROD和BFCOD活性的抑制作用（IC），该方法能够识别时间依赖性抑制剂（TDI）。此外，使用平衡透析评估了受试药物与RT-S9的非特异性结合。
大多数受试药物对RT-S9中EROD和BFCOD活性的抑制作用为中度至弱抑制，即使大多数对人类CYP1A或CYP3A无抑制作用。只有比马前列素、氯美噻唑、非布索坦、舒必利和佐米曲普坦对RT-S9中的任何一种活性均无抑制作用。EROD抑制作用通常强于BFCOD，一些物质（托莫西汀、氟卡尼和哌唑嗪）仅选择性抑制EROD活性。氮卓斯汀和埃索美拉唑对EROD的抑制作用最强（未结合IC分别为3.8±0.5μM和3.0±0.8μM）。受试物质均不是BFCOD的TDI，但埃索美拉唑是EROD的TDI。除氯美噻唑和双硫仑外，受试药物与RT-S9的非特异性结合广泛（未结合分数<0.5），且与其水-辛醇分配系数相关性良好（r = 0.7135）。
结果表明，不能根据人类数据明确预测RT-S9中的P450相互作用，但本文报道的IC数据可以揭示虹鳟CYP1A1和CYP3A27与其人类同系物相比的底物选择性。然而，IC浓度比药物的平均环境浓度高几个数量级。埃索美拉唑对EROD的时间依赖性抑制作用值得进一步研究，以评估其与对鱼类肝毒性影响的可能联系。
