a Department of Biological Engineering, Koch Institute for Integrative Cancer Research, Massachusetts Institute of Technology , Cambridge , MA , USA.
c Adimab LLC , Lebanon , NH , U.S.A.
MAbs. 2017 Oct;9(7):1036-1040. doi: 10.1080/19420862.2017.1356529. Epub 2017 Jul 26.
Early stage assays that evaluate monoclonal antibody drug-like properties serve as valuable tools for selection of lead candidates. One liability for clinical development, off-target reactivity, is often assessed by binding to a mixture or panel of noncognate proteins. While robust, these mixes are often ill-defined, and can suffer from issues such as lot-to-lot variability. In this study, we discovered in immunoprecipitation experiments that certain chaperones are present in one of these mixtures;we then explored the use of recombinant chaperone proteins as well-characterized agents to predict antibody nonspecificity. Antibody binding to the heat shock proteins HSP70, HSP90, or trigger factor all served as predictors of cross-interaction propensity, with HSP90 providing the greatest ability to predict antibody clearance rates in mouse. Individual chaperone binding correlates surprisingly closely with binding to complex cell extracts, with the exception of a few "false negatives" (assuming a complex cell extract as the "true" value). As defined reagents, these chaperone reagents present advantages for high throughput assays of nonspecificity.
早期阶段的评估单克隆抗体类药物特性的检测可作为候选药物先导物筛选的有价值的工具。非靶标反应性是临床开发的一个不利因素，通常通过与非同源蛋白混合物或面板的结合来评估。虽然这些混合物很强大，但往往定义不明确，并且可能存在批次间变异性等问题。在这项研究中，我们在免疫沉淀实验中发现某些伴侣蛋白存在于其中一种混合物中；然后我们探索了使用重组伴侣蛋白作为经过充分表征的试剂来预测抗体的非特异性。抗体与热休克蛋白 HSP70、HSP90 或触发因子的结合均可作为交叉相互作用倾向的预测因子，其中 HSP90 最能预测抗体在小鼠中的清除率。令人惊讶的是，单个伴侣蛋白的结合与与复杂细胞提取物的结合非常密切相关，除了少数“假阴性”（假设复杂细胞提取物为“真实”值）。作为定义明确的试剂，这些伴侣蛋白试剂为非特异性的高通量检测提供了优势。
