Insights from native mass spectrometry approaches for top- and middle- level characterization of site-specific antibody-drug conjugates
Insights from native mass spectrometry approaches for top- and middle- level characterization of site-specific antibody-drug conjugates
复制标题
DOI：
10.1080/19420862.2017.1316914
复制
发表时间：
2017-01-01
期刊：
影响因子：
5.3
通讯作者：
Cianferani, Sarah
中科院分区：
文献类型：
--
作者：
Botzanowski, Thomas;Erb, Stephane;Cianferani, Sarah
抗体-药物偶联物（ADC）已经作为一类有望成为有效免疫疗法的化合物而出现。第一代ADC主要通过在链间二硫键还原后在赖氨酸侧链胺或半胱氨酸硫醇基团上的反应产生，从而产生每个抗体具有可变数量的药物负载的异质群体。为了控制药物负载的位置和数量，已经开发了旨在产生更均匀的位点特异性缀合物的新缀合策略。我们在这里报告的第一个多层次的表征位点特异性ADC的国家的最先进的质谱（MS）方法，包括天然MS和其联用离子迁移率（IM-MS）。我们证明了用于位点特异性ADC分析的原生MS方法的多功能性，具有在一次运行中提供几个关键质量属性的独特能力，沿着ADC同质性/异质性的直接快照，而无需大量的数据解释。还强调了天然IM-MS直接访问位点特异性ADC构象信息的能力。最后，通过比较位点特异性DAR 4 ADC与第一代ADC的分析表征来说明这些技术用于评估ADC的异质性/同质性的潜力。总之，我们的结果突出了天然MS方法用于分析表征所有类型ADC（包括位点特异性缀合物）的兼容性、多功能性和优势。因此，我们设想将天然MS和IM-MS方法，即使是最新的最先进形式，整合到基准生物缀合策略的工作流程中。
Antibody-drug conjugates (ADCs) have emerged as a family of compounds with promise as efficient immunotherapies. First-generation ADCs were generated mostly via reactions on either lysine side-chain amines or cysteine thiol groups after reduction of the interchain disulfide bonds, resulting in heterogeneous populations with a variable number of drug loads per antibody. To control the position and the number of drug loads, new conjugation strategies aiming at the generation of more homogeneous site-specific conjugates have been developed. We report here the first multi-level characterization of a site-specific ADC by state-of-the-art mass spectrometry (MS) methods, including native MS and its hyphenation to ion mobility (IM-MS). We demonstrate the versatility of native MS methodologies for site-specific ADC analysis, with the unique ability to provide several critical quality attributes within one single run, along with a direct snapshot of ADC homogeneity/heterogeneity without extensive data interpretation. The capabilities of native IM-MS to directly access site-specific ADC conformational information are also highlighted. Finally, the potential of these techniques for assessing an ADC's heterogeneity/homogeneity is illustrated by comparing the analytical characterization of a site-specific DAR4 ADC to that of first-generation ADCs. Altogether, our results highlight the compatibility, versatility, and benefits of native MS approaches for the analytical characterization of all types of ADCs, including site-specific conjugates. Thus, we envision integrating native MS and IM-MS approaches, even in their latest state-of-the-art forms, into workflows that benchmark bioconjugation strategies.