Valliere-Douglass John, Wallace Alison, Balland Alain
Analytical and Formulation Sciences, Amgen Inc., Seattle, WA 98119, USA.
J Chromatogr A. 2008 Dec 19;1214(1-2):81-9. doi: 10.1016/j.chroma.2008.10.078. Epub 2008 Oct 25.
The following report describes the use of hydrophobic-interaction chromatography (HIC) to separate and characterize populations of monoclonal antibodies resulting from variable N- and C-terminal processing, stressed-induced covalent modifications and conformationally altered populations present in the drug product. We investigated the use of HIC to characterize heterogeneity in the intact molecule and the Fab and Fc sub-domains resulting from papain cleavage. We found that certain classes of covalent modifications to antibodies are highly amenable to HIC separation. Specific covalent modifications occurring on antibodies could be separated into pure fractions which contained unmodified, singly modified (on 1 heavy or light chain) and doubly modified (on both heavy or light chains) molecules. This report demonstrates the utility of HIC for assessing the heterogeneity, stability and, in some cases, potency of monoclonal antibodies.
以下报告描述了使用疏水相互作用色谱法（HIC）来分离和表征药物产品中因可变的N端和C端加工、应激诱导的共价修饰以及构象改变而产生的单克隆抗体群体。我们研究了使用HIC来表征木瓜蛋白酶裂解产生的完整分子以及Fab和Fc亚结构域中的异质性。我们发现，某些类型的抗体共价修饰非常适合HIC分离。抗体上发生的特定共价修饰可以分离成纯组分，其中包含未修饰的、单修饰的（在1条重链或轻链上）和双修饰的（在重链或轻链上均有修饰）分子。本报告证明了HIC在评估单克隆抗体的异质性、稳定性以及在某些情况下的效价方面的实用性。
