The Fate of Bacterial Cocaine Esterase (CocE): An In Vivo Study of CocE-Mediated Cocaine Hydrolysis, CocE Pharmacokinetics, and CocE Elimination
The Fate of Bacterial Cocaine Esterase (CocE): An In Vivo Study of CocE-Mediated Cocaine Hydrolysis, CocE Pharmacokinetics, and CocE Elimination
复制标题
DOI：
10.1124/jpet.111.186049
复制
发表时间：
2012-01-01
影响因子：
3.5
通讯作者：
Sunahara, Roger K.
中科院分区：
文献类型：
--
作者：
Brim, Remy L.;Noon, Kathleen R.;Sunahara, Roger K.
可卡因滥用和毒性问题在美国仍然普遍存在。目前，由于没有食品和药物管理局批准的药物治疗这种适应症，可卡因毒性只能对症治疗。为了解决未满足的需求，研究人员开发了一种稳定的细菌可卡因酯酶突变体[T172R/G173Q-CocE (DM-CocE)]，该突变体可将可卡因水解为无活性代谢物，具有较低的免疫原性潜力，并已在可卡因毒性动物模型中进行了测试。在这里，我们记录了低剂量DM-CocE在体外和体内对可卡因的快速水解，以及DM-CocE蛋白在大鼠体内的药代动力学和分布。质谱法测定，50.5 μ g/kg的DM-CocE能在2 min内有效消除雄性和雌性大鼠体内4 mg/kg的可卡因。我们通过在大鼠和猴子中使用药理学相关剂量的DM-CocE （0.32 mg/kg）来水解惊厥剂量的可卡因来扩展这些发现。DM-CocE在注射后迅速将可卡因降至检出限以下；然而，DM-CocE的消除导致外周可卡因再分布30至60分钟。采用[S-35]标记法定量测定DM-CocE在大鼠体内的半衰期为2.1 h。少量尿排出DM-CocE也被观察到。免疫组织化学、Western blotting和x线摄影均用于阐明DM-CocE消除、快速蛋白水解和氨基酸再循环到所有组织的机制。这种快速消除DM-CocE是治疗可卡因毒性的理想特性，并且随着DM-CocE走向临床应用，应该减少免疫原性反应或不良反应的可能性。
Cocaine abuse and toxicity remain widespread problems in the United States. Currently cocaine toxicity is treated only symptomatically, because there is no Food and Drug Administration-approved pharmacotherapy for this indication. To address the unmet need, a stabilized mutant of bacterial cocaine esterase [T172R/G173Q-CocE (DM-CocE)], which hydrolyzes cocaine into inactive metabolites and has low immunogenic potential, has been developed and previously tested in animal models of cocaine toxicity. Here, we document the rapid cocaine hydrolysis by low doses of DM-CocE in vitro and in vivo, as well as the pharmacokinetics and distribution of the DM-CocE protein in rats. DM-CocE at 50.5 mu g/kg effectively eliminated 4 mg/kg cocaine within 2 min in both male and female rats as measured by mass spectrometry. We expanded on these findings by using a pharmacologically relevant dose of DM-CocE (0.32 mg/kg) in rats and monkeys to hydrolyze convulsant doses of cocaine. DM-CocE reduced cocaine to below detection limits rapidly after injection; however, elimination of DM-CocE resulted in peripheral cocaine redistribution by 30 to 60 min. Elimination of DM-CocE was quantified by using [S-35] labeling of the enzyme and was found to have a half-life of 2.1 h in rats. Minor urinary output of DM-CocE was also observed. Immunohistochemistry, Western blotting, and radiography all were used to elucidate the mechanism of DM-CocE elimination, rapid proteolysis, and recycling of amino acids into all tissues. This rapid elimination of DM-CocE is a desirable property of a therapeutic for cocaine toxicity and should reduce the likelihood of immunogenic or adverse reactions as DM-CocE moves toward clinical use.