Wsól Vladimír, Szotáková Barbora, Skálová Lenka, Maser Edmund
Department of Biochemical Sciences, Faculty of Pharmacy, Charles University, Heyrovského 1203, CZ-500 05, Hradec Králové, Czech Republic.
Chem Biol Interact. 2003 Feb 1;143-144:459-68. doi: 10.1016/s0009-2797(02)00221-1.
Oracin, 6-[2-(2-hydroxyethyl)aminoethyl]-5,11-dioxo-5,6-dihydro-11H-indeno[1,2-c] isoquinoline, is a potential cytostatic drug for oral use and presently in phase II of clinical trials. Major advantages of this novel chemotherapeutic are the possibility of oral administration, its negative results in the Ames test on mutagenicity, and the lack of cardiotoxicity. Metabolic studies on oracin have revealed that the principal metabolite in all laboratory animals is 11-dihydrooracin (DHO), which is produced by carbonyl reduction of the parent compound. Since the carbonyl moiety of oracin is a pro-chiral centre, reduction may lead to the two stereoisomer forms (+)-DHO and (-)-DHO. The aim of the present study was to infer if 11beta-hydroxysteroid dehydrogenase type 1 (11beta-HSD 1) is responsible for carbonyl reduction of oracin in mouse liver and if this enzyme exhibits stereospecificity in DHO formation. 11beta-HSD 1 was purified from mouse liver microsomes, and the kinetics and stereospecificity regarding DHO formation were determined and compared to values obtained from the whole microsomal fraction. We could show that purified mouse liver 11beta-HSD 1 catalyzes the stereospecific carbonyl reduction of oracin, thereby following a sigmoidal dose-response kinetics. Due to a different ratio of (+)-DHO and (-)-DHO (93:7) formed by purified 11beta-HSD 1 compared to that produced in whole microsomes (70:30), the existence of at least one other oracin carbonyl reducing enzyme can be expected in mouse liver microsomes. This suggestion is further supported by the fact that the Hill coefficient of 2 for purified 11beta-HSD 1 (which is supporting earlier data on the cooperativity of this dimeric enzyme) changes to a Hill coefficient of 3 in whole microsomes (which is indicative for another enzyme participating in oracin carbonyl reduction).
奥拉辛，即6-[2-(2-羟乙基)氨基乙基]-5,11-二氧代-5,6-二氢-11H-茚并[1,2-c]异喹啉，是一种有潜力的口服细胞生长抑制剂，目前正处于临床试验的II期。这种新型化疗药物的主要优点包括可口服给药、在艾姆斯诱变试验中呈阴性结果以及无心脏毒性。对奥拉辛的代谢研究表明，所有实验动物体内的主要代谢产物是11-二氢奥拉辛（DHO），它是由母体化合物的羰基还原产生的。由于奥拉辛的羰基部分是一个前手性中心，还原可能会产生两种立体异构体形式：(+)-DHO和(-)-DHO。本研究的目的是推断11β-羟基类固醇脱氢酶1型（11β-HSD 1）是否负责小鼠肝脏中奥拉辛的羰基还原，以及该酶在DHO形成过程中是否表现出立体特异性。从小鼠肝脏微粒体中纯化出11β-HSD 1，并测定了其与DHO形成相关的动力学和立体特异性，并与从整个微粒体部分获得的值进行比较。我们能够证明，纯化后的小鼠肝脏11β-HSD 1催化奥拉辛的立体特异性羰基还原，从而呈现出S形剂量反应动力学。与整个微粒体中产生的(+)-DHO和(-)-DHO比例（70:30）相比，纯化后的11β-HSD 1形成的比例不同（93:7），因此可以预期小鼠肝脏微粒体中至少存在另一种奥拉辛羰基还原酶。纯化后的11β-HSD 1的希尔系数为2（这支持了关于这种二聚体酶协同性的早期数据），而在整个微粒体中变为3（这表明有另一种酶参与奥拉辛羰基还原），这一事实进一步支持了这一推测。
