Kato Kota, Ohbuchi Masato, Hamamura Satoko, Ohshita Hiroki, Kazuki Yasuhiro, Oshimura Mitsuo, Sato Koya, Nakada Naoyuki, Kawamura Akio, Usui Takashi, Kamimura Hidetaka, Tateno Chise
Drug Metabolism Research Laboratories, Astellas Pharma Inc., Osaka, Japan (K.K., Ma.O., K.S., N.N., A.K., T.U.); PhoenixBio Co., Ltd., Hiroshima, Japan (S.H., H.O., C.T.); Liver Research Project Center, Hiroshima University, Hiroshima, Japan (C.T.); Department of Biomedical Science, Institute of Regenerative Medicine and Biofunction, Graduate School of Medical Science (Y.K., Mi.O.), Chromosome Engineering Research Center (Y.K., Mi.O.), Tottori University, Tottori, Japan; ADME & Tox Research Institute, Sekisui Medical Co., Ltd., Tokyo, Japan (H.K.).
Drug Metabolism Research Laboratories, Astellas Pharma Inc., Osaka, Japan (K.K., Ma.O., K.S., N.N., A.K., T.U.); PhoenixBio Co., Ltd., Hiroshima, Japan (S.H., H.O., C.T.); Liver Research Project Center, Hiroshima University, Hiroshima, Japan (C.T.); Department of Biomedical Science, Institute of Regenerative Medicine and Biofunction, Graduate School of Medical Science (Y.K., Mi.O.), Chromosome Engineering Research Center (Y.K., Mi.O.), Tottori University, Tottori, Japan; ADME & Tox Research Institute, Sekisui Medical Co., Ltd., Tokyo, Japan (H.K.)
Drug Metab Dispos. 2015 Aug;43(8):1208-17. doi: 10.1124/dmd.115.063479. Epub 2015 May 15.
We developed murine CYP3A knockout ko chimeric mice with humanized liver expressing human P450S similar to those in humans and whose livers and small intestines do not express murine CYP3A this: approach may overcome effects of residual mouse metabolic enzymes like Cyp3a in conventional chimeric mice with humanized liver, such as PXB-mice [urokinase plasminogen activator/severe combined immunodeficiency (uPA/SCID) mice repopulated with over 70% human hepatocytes] to improve the prediction of drug metabolism and pharmacokinetics in humans. After human hepatocytes were transplanted into Cyp3a KO/uPA/SCID host mice, human albumin levels logarithmically increased until approximately 60 days after transplantation, findings similar to those in PXB-mice. Quantitative real-time-polymerase chain reaction analyses showed that hepatic human P450s, UGTs, SULTs, and transporters mRNA expression levels in Cyp3a KO chimeric mice were also similar to those in PXB-mice and confirmed the absence of Cyp3a11 mRNA expression in mouse liver and intestine. Findings for midazolam and triazolam metabolic activities in liver microsomes were comparable between Cyp3a KO chimeric mice and PXB-mice. In contrast, these activities in the intestine of Cyp3a KO chimeric mice were attenuated compared with PXB-mice. Owing to the knockout of murine Cyp3a, hepatic Cyp2b10 and 2c55 mRNA levels in Cyp3a KO/uPA/SCID mice (without hepatocyte transplants) were 8.4- and 61-fold upregulated compared with PXB-mice, respectively. However, human hepatocyte transplantation successfully restored Cyp2b10 level nearly fully and Cyp2c55 level partly (still 13-fold upregulated) compared with those in PXB-mice. Intestinal Cyp2b10 and 2c55 were also repressed by human hepatocyte transplantation in Cyp3a KO chimeric mice.
我们培育出了小鼠CYP3A基因敲除（ko）嵌合小鼠，其肝脏人源化，表达与人类相似的人P450S，且肝脏和小肠不表达小鼠CYP3A。这种方法可能克服传统肝脏人源化嵌合小鼠（如PXB小鼠[用超过70%的人肝细胞重新填充的尿激酶型纤溶酶原激活剂/重症联合免疫缺陷（uPA/SCID）小鼠]）中残留小鼠代谢酶（如Cyp3a）的影响，以改善对人类药物代谢和药代动力学的预测。将人肝细胞移植到Cyp3a KO/uPA/SCID宿主小鼠后，人白蛋白水平呈对数增加，直至移植后约60天，这一结果与PXB小鼠相似。定量实时聚合酶链反应分析表明，Cyp3a KO嵌合小鼠肝脏中的人P450、UGT、SULT和转运蛋白mRNA表达水平也与PXB小鼠相似，并证实小鼠肝脏和肠道中不存在Cyp3a11 mRNA表达。Cyp3a KO嵌合小鼠和PXB小鼠肝脏微粒体中咪达唑仑和三唑仑的代谢活性结果相当。相比之下，与PXB小鼠相比，Cyp3a KO嵌合小鼠肠道中的这些活性减弱。由于小鼠Cyp3a基因的敲除，与PXB小鼠相比，Cyp3a KO/uPA/SCID小鼠（未进行肝细胞移植）肝脏中的Cyp2b10和2c55 mRNA水平分别上调了8.4倍和61倍。然而，与PXB小鼠相比，人肝细胞移植成功地几乎完全恢复了Cyp2b10水平，部分恢复了Cyp2c55水平（仍上调13倍）。在Cyp3a KO嵌合小鼠中，人肝细胞移植也抑制了肠道中的Cyp2b10和2c55。
