Mingming Han1, Jianchang Qian1, Zhize Ye1, Renai Xu2, Daoxing Chen1, Saili Xie3, Jianping Cai4, Guoxin Hu5. 1. School of Pharmaceutical Sciences, Wenzhou Medical University, Wenzhou, Zhejiang, PR China. 2. The First Affiliated Hospital of Wenzhou Medical University, Wenzhou, Zhejiang, PR China. 3. The First Affiliated Hospital of Wenzhou Medical University, Wenzhou, Zhejiang, PR China. Electronic address: xiesaili1988@163.com. 4. School of Pharmaceutical Sciences, Wenzhou Medical University, Wenzhou, Zhejiang, PR China; The Ministry of Health (MOH) Key Laboratory of Geriatrics, Beijing Hospital, National Center of Gerontology, Beijing, PR China. Electronic address: caijp61@vip.sina.com. 5. School of Pharmaceutical Sciences, Wenzhou Medical University, Wenzhou, Zhejiang, PR China. Electronic address: hgx@wmu.edu.cn.
Abstract
AIM: We aimed (i) to study the effects of genetic polymorphism of cytochrome P450 3A4 (CYP3A4) and drug interactions on acalabrutinib (ACA) metabolism and (ii) to investigate the mechanisms underlying the effects of CYP3A4 variants on the differential kinetic profiles of ACA and ibrutinib. METHOD: Recombinant human CYP3A4 and variants were expressed using a Bac-to-Bac baculovirus expression system. The cell microsome was prepared and subjected to kinetic study. The analyte concentrations were determined by UPLC-MS/MS. A molecular docking assay was employed to investigate the mechanisms leading to differences in kinetic profiles. RESULTS: The kinetic parameters of ACA, catalyzed by CYP3A4 and 28 of its variants, were determined, including Vmax, Km, and Ksi. CYP3A4.6-8, 12, 13, 17, 18, 20, and 30 lost their catalytic function. No significant differences were found for CYP3A4.4, 5, 10, 15, 31, and 34 compared with CYP3A4.1 with respect to intrinsic clearance (Vmax/Km, Clint). However, the Clint values of CYP3A4.9, 14, 16, 19, 23, 24, 28, 32 were obviously decreased, ranging from 0.02 to 0.05 μL/min/pmol. On the contrary, the catalytic activities of CYP3A4.2, 3, 11, 29, and 33 were increased dramatically. The Clint value of CYP3A4.11 was 5.95 times as high as that of CYP3A4.1. Subsequently, CYP3A4.1, 3, 11, 23, and 28 were chosen to study the kinetic changes in combination with ketoconazole. Interestingly, we found the inhibitory potency of ketoconazole varied in different variants. In addition, the kinetic parameters of ibrutinib and ACA were accordingly compared in different CYP3A4 variants. Significant differences in relative clearance were observed among variants, which would probably influence the distance between the redox site and the heme iron atom. CONCLUSION: Genetic polymorphism of CYP3A4 extensively changes its ACA-metabolizing enzymatic activity. In combination with a CYP inhibitor, its inhibitory potency also varied among different variants. Even the same variants exhibited different capabilities catalyzing ACA. Its enzymatic capabilities are probably determined by the distance between the substrate and the heme iron atom, which could be impacted by mutation.
AIM: We aimed (i) to study the effects of genetic polymorphism of cytochrome P450 3A4 (CYP3A4) and drug interactions on acalabrutinib (ACA) metabolism and (ii) to investigate the mechanisms underlying the effects of CYP3A4 variants on the differential kinetic profiles of ACA and ibrutinib. METHOD: Recombinant humanCYP3A4 and variants were expressed using a Bac-to-Bac baculovirus expression system. The cell microsome was prepared and subjected to kinetic study. The analyte concentrations were determined by UPLC-MS/MS. A molecular docking assay was employed to investigate the mechanisms leading to differences in kinetic profiles. RESULTS: The kinetic parameters of ACA, catalyzed by CYP3A4 and 28 of its variants, were determined, including Vmax, Km, and Ksi. CYP3A4.6-8, 12, 13, 17, 18, 20, and 30 lost their catalytic function. No significant differences were found for CYP3A4.4, 5, 10, 15, 31, and 34 compared with CYP3A4.1 with respect to intrinsic clearance (Vmax/Km, Clint). However, the Clint values of CYP3A4.9, 14, 16, 19, 23, 24, 28, 32 were obviously decreased, ranging from 0.02 to 0.05 μL/min/pmol. On the contrary, the catalytic activities of CYP3A4.2, 3, 11, 29, and 33 were increased dramatically. The Clint value of CYP3A4.11 was 5.95 times as high as that of CYP3A4.1. Subsequently, CYP3A4.1, 3, 11, 23, and 28 were chosen to study the kinetic changes in combination with ketoconazole. Interestingly, we found the inhibitory potency of ketoconazole varied in different variants. In addition, the kinetic parameters of ibrutinib and ACA were accordingly compared in different CYP3A4 variants. Significant differences in relative clearance were observed among variants, which would probably influence the distance between the redox site and the hemeiron atom. CONCLUSION: Genetic polymorphism of CYP3A4 extensively changes its ACA-metabolizing enzymatic activity. In combination with a CYP inhibitor, its inhibitory potency also varied among different variants. Even the same variants exhibited different capabilities catalyzing ACA. Its enzymatic capabilities are probably determined by the distance between the substrate and the hemeiron atom, which could be impacted by mutation.
Authors: Anssi J H Mykkänen; Suvi Taskinen; Mikko Neuvonen; Maria Paile-Hyvärinen; E Katriina Tarkiainen; Tuomas Lilius; Tuija Tapaninen; Janne T Backman; Aleksi Tornio; Mikko Niemi Journal: Clin Pharmacol Ther Date: 2022-06-24 Impact factor: 6.903