Systematic errors in detecting biased agonism: Analysis of current methods and development of a new model-free approach
Systematic errors in detecting biased agonism: Analysis of current methods and development of a new model-free approach
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DOI：
10.1038/srep44247
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发表时间：
2017-03-14
影响因子：
4.6
通讯作者：
Costa, Tommaso
中科院分区：
文献类型：
--
作者：
Onaran, H. Ongun;Ambrosio, Caterina;Costa, Tommaso
发现偏向激动剂需要一种方法，该方法可以可靠地区分由于不同转导蛋白的不平衡激活引起的信号传导中的偏向与所研究的系统固有的差异扩增的偏向，所研究的系统固有的差异扩增总是由生物信号传导网络及其测量的非线性性质引起。我们系统地比较了七种偏倚诊断方法的性能，所有这些方法都是基于根据经典受体理论分析配体的浓度-响应曲线。我们通过比较受体-转导子与Gs、Gi和抑制蛋白相互作用的BRET测定，计算了一些β-肾上腺素能激动剂的偏倚因子。使用相同的配体，我们还比较了源自相同受体-换能器相互作用的信号步骤的响应，其中理论上不可能有偏倚的功效。在这两种情况下，我们发现了高水平的假阳性结果和方法之间普遍缺乏相关性。总之，该分析表明，所有测试的方法，包括文献中最广泛使用的一些方法，都不能有把握地区分真正的配体偏倚和“系统偏倚”。我们还提出了两种新的半定量方法的偏见诊断，似乎是更强大和可靠的比目前可用的策略。
Discovering biased agonists requires a method that can reliably distinguish the bias in signalling due to unbalanced activation of diverse transduction proteins from that of differential amplification inherent to the system being studied, which invariably results from the non-linear nature of biological signalling networks and their measurement. We have systematically compared the performance of seven methods of bias diagnostics, all of which are based on the analysis of concentration-response curves of ligands according to classical receptor theory. We computed bias factors for a number of beta-adrenergic agonists by comparing BRET assays of receptor-transducer interactions with Gs, Gi and arrestin. Using the same ligands, we also compared responses at signalling steps originated from the same receptor-transducer interaction, among which no biased efficacy is theoretically possible. In either case, we found a high level of false positive results and a general lack of correlation among methods. Altogether this analysis shows that all tested methods, including some of the most widely used in the literature, fail to distinguish true ligand bias from "system bias" with confidence. We also propose two novel semi quantitative methods of bias diagnostics that appear to be more robust and reliable than currently available strategies.