伊曲康唑是一种三唑类抗感染药物,已被证明可预防和治疗多种真菌和病毒感染,被认为是治疗COVID-19的潜在治疗方法。在这项研究中,我们旨在全面评估细胞色素P4503A4(CYP3A4)变异蛋白和药物相互作用对伊曲康唑在重组昆虫微粒体中代谢的影响,并表征了底物选择性的潜在机理。通过CYP3A4变体评估在有/没有洛匹那韦或达鲁那韦的情况下与伊曲康唑(0.2-15μM)的孵育,用UPLC-MS/MS测定代谢物羟基伊曲康唑的浓度。我们的数据显示,与CYP3A4.1相比,4种变体(CYP3A4.9、0.10、0.28和0.34)没有显着差异,和3个变体(CYP3A4.14,0.15和0.19)表现出增加的内在清除率(CLint),而其余17种变体蛋白显示出伊曲康唑催化作用的酶活性降低。此外,洛匹那韦和地瑞那韦对伊曲康唑代谢的抑制作用有不同程度的差异。此外,观察到10个变异体(CYP3A4.5、0.9、0.10、0.16、0.19、0.24、0.28、0.29、0.31和0.33)的动力学参数的不同变化趋势,尤其是CYP3A4.5和CYP3A4.16,这可能与代谢位点-血红素铁原子距离有关。在本研究中,我们首次功能分析了25种CYP3A4蛋白变体对伊曲康唑代谢的影响,并提供了伊曲康唑体外代谢的全面数据。这可能有助于更好地评估伊曲康唑在临床中的代谢和消除情况,以提高其临床治疗的安全性和有效性,也为COVID-19的治疗提供新的可能性。
Itraconazole is a triazole anti-infective drug that has been proven to prevent and treat a variety of fungal and viral infections and has been considered to be a potential therapeutic remedy for COVID-19 treatment. In this study, we aimed to completely evaluate the impacts of Cytochrome P450 3A4 (CYP3A4) variant proteins and drug interactions on the metabolism of itraconazole in recombinant insect microsomes, and to characterize the potential mechanism of substrate selectivity. Incubations with itraconazole (0.2-15 μM) in the presence/absence of lopinavir or darunavir were assessed by CYP3A4 variants, and the metabolite hydroxyitraconazole concentrations were measured by UPLC-MS/MS. Our data showed that when compared with CYP3A4.1, 4 variants (CYP3A4.9, .10, .28 and .34) displayed no significant differences, and 3 variants (CYP3A4.14, .15 and .19) exhibited increased intrinsic clearance (CLint), whereas the remaining 17 variant proteins showed decreased enzyme activities for the catalysis of itraconazole. Moreover, the inhibitory effects of lopinavir and darunavir on itraconazole metabolism varied in different degrees. Furthermore, different changed trend of the kinetic parameters in ten variants (CYP3A4.5, .9, .10, .16, .19, .24, .28, .29, .31, and .33) were observed, especially CYP3A4.5 and CYP3A4.16, and this may be related to the metabolic site-heme iron atom distance. In the present study, we functionally analyzed the effects of 25 CYP3A4 protein variants on itraconazole metabolism for the first time, and provided comprehensive data on itraconazole metabolism in vitro. This may help to better assess the metabolism and elimination of itraconazole in clinic to improve the safety and efficacy of its clinical treatment and also provide new possibilities for the treatment of COVID-19.