Identification of in vitro protein biomarkers of idiosyncratic liver toxicity
Identification of in vitro protein biomarkers of idiosyncratic liver toxicity
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DOI：
10.1016/j.tiv.2004.01.012
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发表时间：
2004-08-01
影响因子：
3.2
通讯作者：
Dambach, DM
中科院分区：
文献类型：
--
作者：
Gao, J;Garulacan, LA;Dambach, DM
药物诱导的特异质肝毒性仍然是制药行业的重要安全性问题。这种毒性部分是由于目前临床前研究系统的预测性有限。形成了一个假设，即用临床上与特异质肝毒性相关的药物处理现有的体外肝细胞培养物将导致释放指示肝毒性的细胞外蛋白生物标志物。为了检验这一假设，蛋白质组学和免疫学技术的组合被用来首先识别，并随后验证，从永生化的人肝细胞过表达细胞色素P450 3A 4的蛋白负载的条件培养基的组件。这些细胞分别用由噻唑烷二酮和L-酪氨酸PPARgamma激动剂和HIV蛋白酶抑制剂的组合组成的七种单独的化合物加上载体对照（二甲基亚砜）处理。对于每类药物，比较了临床确定的肝毒性和非肝毒性化合物。两种蛋白质BMS-PTX-265和BMS-PTX-837在来自用每种毒性化合物处理的细胞的条件培养基中与来自用无毒化合物（和媒介物）处理的细胞的培养基相比可再现地且显著地增加。该结果支持该假设，因此使用一系列连续测定（蛋白质印迹和酶联免疫吸附测定）来测量这两种蛋白质作为20种化合物的扩展组的函数的响应。对于所有20种药物，BMS-PTX-265的升高与已知的安全性特征完全相关;而BMS-PTX-837的变化正确预测了20种药物中19种的安全性特征（1个假阴性）。总之，数据支持临床前体外方法作为鉴定肝毒性新生物标志物的手段，以及生物标志物本身的有效性。(C)2004爱思唯尔有限公司保留所有权利。
Drug-induced idiosyncratic hepatotoxicity continues to be an important safety issue for the pharmaceutical industry. This toxicity is due, in part, to the limited predictive nature of current pre-clinical study systems. A hypothesis was formed that treatment of existing in vitro hepatocyte cultures with drugs clinically linked to idiosyncratic hepatotoxicity would result in the release of extracellular protein biomarkers indicative of liver toxicity. To test this hypothesis, a combination of proteomic and immunological techniques were used to first identify, and subsequently verify, components of the protein-laden conditioned culture media from immortalized human hepatocytes which overexpressed cytochrome P450 3A4. These cells were treated separately with seven individual compounds made up of a combination of thiazolidinedione and L-tyrosine PPARgamma agonists and HIV protease inhibitors, plus a vehicle control (dimethyl sulfoxide). For each drug class, clinically determined hepatotoxic and non-hepatotoxic compounds were compared. Two proteins, BMS-PTX-265 and BMS-PTX-837, were reproducibly and significantly increased in the conditioned media from cells treated with each of the toxic compounds as compared to media from cells treated with the non-toxic compounds (and vehicle). This result supported the hypothesis, and so a series of successive assays (western blots and enzyme linked immunosorbent assays) were used to measure the response of these two proteins as a function of an expanded set of 20 compounds. For all 20 drugs, elevations of BMS-PTX-265 correlated exactly with the known safety profile; whereas changes in BMS-PTX-837 correctly predicted the safety profile in 19 of 20 drugs (one false negative). In summary, the data supports both the pre-clinical in vitro method as a means to identify new biomarkers of liver toxicity, as well as the validity of the biomarkers themselves. (C) 2004 Elsevier Ltd. All rights reserved.