Department of Drug Disposition, Development/Commercialization, Lilly Research Laboratories, Lilly Corporate Center, Indianapolis, Indiana, USA.
Department of Biotechnology Discovery Research, Lilly Research Laboratories, Lilly Corporate Center, Indianapolis, Indiana, USA.
AAPS J. 2018 Sep 25;20(6):103. doi: 10.1208/s12248-018-0263-0.
Among the numerous antibody-drug conjugate (ADC) clinical candidates, one of the most prevalent types utilizes the interchain cysteines in antibodies to conjugate auristatin via a maleimide-containing linker. In this class of ADCs, there are a paucity of systematic studies characterizing how IgG subclass influences the biophysical properties and in vivo pharmacokinetics of the ADC molecules. In the current investigation, we studied cysteine-conjugated ADCs using a model system consisting of human IgG1, IgG2, and IgG4 antibodies with the same variable region. Our findings identified some unforeseen differences among the three ADCs. Drug conjugation profiling by LC-MS revealed that 50% of inter heavy-light chain disulfide bonds are disrupted to conjugate drugs in IgG1 antibody while only 10% in IgG2 antibody and 20% in IgG4 antibody. The solution behavior of the ADCs was interrogated in concentrating experiments and diffusion interaction parameter measurements. We found that drug conjugation affected the solution property of the three antibodies differently, with the IgG2-based ADC having the most increased propensity to aggregate. Rat PK studies using a sensitive LC-MS-based bioanalytical method showed that the IgG1-based ADC has poor peripheral linker-payload stability while the IgG2- and IgG4-based ADCs are stable. The conjugate stability of the IgG2-based ADC was further confirmed in a cynomolgus monkey PK study. Overall, the IgG2-based ADC exhibited the best PK/conjugate stability but also the most deterioration in stability among the three ADCs. Our findings provide important information and present multifactorial considerations for the selection of IgG subclass during ADC drug discovery when employing stochastic cysteine conjugation.
在众多抗体药物偶联物 (ADC) 的临床候选物中，有一种最常见的类型是利用抗体的链间半胱氨酸通过含有马来酰亚胺的连接子将奥瑞他汀连接起来。在这类 ADC 中，很少有系统的研究来描述 IgG 亚类如何影响 ADC 分子的物理化学性质和体内药代动力学。在当前的研究中，我们使用由人 IgG1、IgG2 和 IgG4 抗体组成的模型系统研究了半胱氨酸偶联的 ADC。我们的研究结果发现这三种 ADC 之间存在一些意外的差异。通过 LC-MS 进行的药物偶联分析表明，50%的重链-轻链间二硫键被破坏以在 IgG1 抗体中结合药物，而在 IgG2 抗体中只有 10%，在 IgG4 抗体中只有 20%。通过浓缩实验和扩散相互作用参数测量来研究 ADC 的溶液行为。我们发现药物偶联以不同的方式影响三种抗体的溶液性质，以 IgG2 为基础的 ADC 具有最强的聚集倾向。使用基于灵敏 LC-MS 的生物分析方法的大鼠 PK 研究表明，基于 IgG1 的 ADC 在外周连接子-有效载荷稳定性差，而 IgG2 和 IgG4 为基础的 ADC 是稳定的。在食蟹猴 PK 研究中进一步证实了 IgG2 为基础的 ADC 的缀合物稳定性。总体而言，基于 IgG2 的 ADC 表现出最佳的 PK/缀合物稳定性，但在三种 ADC 中稳定性也最差。我们的研究结果提供了重要的信息，并为在采用随机半胱氨酸偶联进行 ADC 药物发现时选择 IgG 亚类提供了多方面的考虑。
