Ferguson Daniel C, Cheng Qiuying, Blanco Javier G
Department of Pharmaceutical Sciences, School of Pharmacy and Pharmaceutical Sciences, The State University of New York at Buffalo, Buffalo, New York.
Department of Pharmaceutical Sciences, School of Pharmacy and Pharmaceutical Sciences, The State University of New York at Buffalo, Buffalo, New York
Drug Metab Dispos. 2015 Jul;43(7):922-7. doi: 10.1124/dmd.115.064295. Epub 2015 Apr 27.
The anthracyclines doxorubicin and daunorubicin are used in the treatment of various human and canine cancers, but anthracycline-related cardiotoxicity limits their clinical utility. The formation of anthracycline C-13 alcohol metabolites (e.g., doxorubicinol and daunorubicinol) contributes to the development of anthracycline-related cardiotoxicity. The enzymes responsible for the synthesis of anthracycline C-13 alcohol metabolites in canines remain to be elucidated. We hypothesized that canine carbonyl reductase 1 (cbr1), the homolog of the prominent anthracycline reductase human CBR1, would have anthracycline reductase activity. Recombinant canine cbr1 (molecular weight: 32.8 kDa) was purified from Escherichia coli. The enzyme kinetics of "wild-type" canine cbr1 (cbr1 D218) and a variant isoform (cbr1 V218) were characterized with the substrates daunorubicin and menadione, as well as the flavonoid inhibitor rutin. Canine cbr1 catalyzes the reduction of daunorubicin to daunorubicinol, with cbr1 D218 and cbr1 V218 displaying different kinetic parameters (cbr1 D218 Km: 188 ± 144 μM versus cbr1 V218 Km: 527 ± 136 μM, P < 0.05, and cbr1 D218 Vmax: 6446 ± 3615 nmol/min per milligram versus cbr1 V218 Vmax: 15539 ± 2623 nmol/min per milligram, P < 0.01). Canine cbr1 also metabolized menadione (cbr1 D218 Km: 104 ± 50 μM, Vmax: 2034 ± 307 nmol/min per milligram). Rutin acted as a competitive inhibitor for the reduction of daunorubicin (cbr1 D218 Ki: 1.84 ± 1.02 μM, cbr1 V218 Ki: 1.38 ± 0.47 μM). These studies show that canine cbr1 metabolizes daunorubicin and provide the necessary foundation to characterize the role of cbr1 in the variable pharmacodynamics of anthracyclines in canine cancer patients.
蒽环类药物阿霉素和柔红霉素被用于治疗多种人类和犬类癌症，但蒽环类药物相关的心脏毒性限制了它们的临床应用。蒽环类药物C-13醇代谢物（如阿霉素醇和柔红霉素醇）的形成会导致蒽环类药物相关心脏毒性的发展。犬类中负责合成蒽环类药物C-13醇代谢物的酶仍有待阐明。我们推测犬羰基还原酶1（cbr1），即主要的蒽环类药物还原酶人类CBR1的同源物，会具有蒽环类药物还原酶活性。从大肠杆菌中纯化出重组犬cbr1（分子量：32.8 kDa）。用底物柔红霉素、甲萘醌以及类黄酮抑制剂芦丁对“野生型”犬cbr1（cbr1 D218）和一种变体同工型（cbr1 V218）的酶动力学进行了表征。犬cbr1催化柔红霉素还原为柔红霉素醇，cbr1 D218和cbr1 V218表现出不同的动力学参数（cbr1 D218 Km：188±144μM，而cbr1 V218 Km：527±136μM，P<0.05；cbr1 D218 Vmax：6446±3615 nmol/分钟每毫克，而cbr1 V218 Vmax：15539±2623 nmol/分钟每毫克，P<0.01）。犬cbr1也能代谢甲萘醌（cbr1 D218 Km：104±50μM，Vmax：2034±307 nmol/分钟每毫克）。芦丁对柔红霉素的还原起竞争性抑制作用（cbr1 D218 Ki：1.84±1.02μM，cbr1 V218 Ki：1.38±0.47μM）。这些研究表明犬cbr1能代谢柔红霉素，并为表征cbr1在犬类癌症患者蒽环类药物可变药效学中的作用提供了必要的基础。
