Safety Assessment UK, AstraZeneca R&D Alderley Park, Macclesfield, SK10 4TG, UK.
Toxicol Appl Pharmacol. 2012 Apr 15;260(2):162-72. doi: 10.1016/j.taap.2012.02.007. Epub 2012 Feb 21.
Measurement of cardiac contractility is a logical part of pre-clinical safety assessment in a drug discovery project, particularly if a risk has been identified or is suspected based on the primary- or non-target pharmacology. However, there are limited validated assays available that can be used to screen several compounds in order to identify and eliminate inotropic liability from a chemical series. We have therefore sought to develop an in vitro model with sufficient throughput for this purpose. Dog ventricular myocytes were isolated using a collagenase perfusion technique and placed in a perfused recording chamber on the stage of a microscope at ~36 °C. Myocytes were stimulated to contract at a pacing frequency of 1 Hz and a digital, cell geometry measurement system (IonOptix™) was used to measure sarcomere shortening in single myocytes. After perfusion with vehicle (0.1% DMSO), concentration-effect curves were constructed for each compound in 4-30 myocytes taken from 1 or 2 dog hearts. The validation test-set was 22 negative and 8 positive inotropes, and 21 inactive compounds, as defined by their effect in dog, cynolomolgous monkey or humans. By comparing the outcome of the assay to the known in vivo contractility effects, the assay sensitivity was 81%, specificity was 75%, and accuracy was 78%. With a throughput of 6-8 compounds/week from 1 cell isolation, this assay may be of value to drug discovery projects to screen for direct contractility effects and, if a hazard is identified, help identify inactive compounds.
在药物发现项目中，测量心脏收缩力是临床前安全性评估的一个合乎逻辑的部分，特别是如果基于主要或非靶标药理学已经确定或怀疑存在风险。然而，可用的经过验证的测定方法有限，这些方法可以用于筛选几种化合物，以确定并消除化学系列中的变力性毒性。因此，我们一直在寻求为此目的开发一种具有足够通量的体外模型。使用胶原酶灌注技术从狗心室肌细胞中分离出来，并在显微镜台上的灌注记录室中以约 36°C 的温度放置。以 1 Hz 的起搏频率刺激肌细胞收缩，并使用数字细胞几何测量系统（IonOptix™）测量单个肌细胞中的肌节缩短。在施用载体（0.1%DMSO）后，从 1 或 2 只狗的心脏中取出的 4-30 个肌细胞中构建每个化合物的浓度-效应曲线。验证测试集由 22 个阴性变力药和 8 个阳性变力药以及 21 个无活性化合物组成，这些化合物的作用在狗、食蟹猴或人类中被定义。通过将测定结果与已知的体内收缩力效应进行比较，该测定的灵敏度为 81%，特异性为 75%，准确性为 78%。从 1 个细胞分离物中每周进行 6-8 种化合物的检测，该测定方法可能对药物发现项目有价值，可以筛选直接的收缩力效应，如果发现危险，则有助于识别无活性的化合物。
