Manchester Pharmacy School , University of Manchester , Manchester M13 9PL , United Kingdom.
School of Chemical Engineering and Analytical Science , University of Manchester , Manchester M1 7DN , United Kingdom.
Mol Pharm. 2019 Dec 2;16(12):4775-4786. doi: 10.1021/acs.molpharmaceut.9b00430. Epub 2019 Nov 11.
The coformulation of monoclonal antibody (mAb) mixtures provides an attractive route to achieving therapeutic efficacy where the targeting of multiple epitopes is necessary. Controlling and predicting the behavior of such mixtures requires elucidating the molecular basis for the self- and cross-protein-protein interactions and how they depend on solution variables. While self-interactions are now beginning to be well understood, systematic studies of cross-interactions between mAbs in solution do not exist. Here, we have used static light scattering to measure the set of self- and cross-osmotic second virial coefficients in a solution containing a mixture of two mAbs, mAbA and mAbB, as a function of ionic strength and pH. mAbB exhibits strong association at a low ionic strength, which is attributed to an electrostatic attraction that is enhanced by the presence of a strong short-ranged attraction of nonelectrostatic origin. Under all solution conditions, the measured cross-interactions are intermediate self-interactions and follow similar patterns of behavior. There is a strong electrostatic attraction at higher pH values, reflecting the behavior of mAbB. Protein-protein interactions become more attractive with an increasing pH due to reducing the overall protein net charges, an effect that is attenuated with an increasing ionic strength due to the screening of electrostatic interactions. Under moderate ionic strength conditions, the reduced cross-virial coefficient, which reflects only the energetic contribution to protein-protein interactions, is given by a geometric average of the corresponding self-coefficients. We show the relationship can be rationalized using a patchy sphere model, where the interaction energy between sites and is given by the arithmetic mean of the and interactions. The geometric mean does not necessarily apply to all mAb mixtures and is expected to break down at a lower ionic strength due to the nonadditivity of electrostatic interactions.
单克隆抗体 (mAb) 混合物的共配方为实现治疗效果提供了一条有吸引力的途径，因为需要靶向多个表位。控制和预测此类混合物的行为需要阐明自蛋白-蛋白相互作用和它们如何依赖于溶液变量的分子基础。虽然自相互作用现在开始得到很好的理解，但在溶液中 mAbs 之间的交叉相互作用的系统研究尚不存在。在这里，我们使用静态光散射来测量包含两种 mAb（mAbA 和 mAbB）混合物的溶液中自和交叉渗透压第二维里系数的集合，作为离子强度和 pH 的函数。mAbB 在低离子强度下表现出强烈的缔合，这归因于静电吸引，而这种静电吸引通过存在强的短程非静电来源的吸引力而增强。在所有溶液条件下，测量的交叉相互作用处于中间自相互作用，并且表现出相似的行为模式。在较高 pH 值下存在强烈的静电吸引，这反映了 mAbB 的行为。由于减少蛋白质的总净电荷，蛋白质-蛋白质相互作用随着 pH 值的增加而变得更具吸引力，这种效应随着离子强度的增加而减弱，因为静电相互作用的屏蔽。在中等离子强度条件下，仅反映蛋白质-蛋白质相互作用的能量贡献的减小交叉维里系数由相应自系数的几何平均值给出。我们表明，该关系可以用带补丁的球体模型来合理化，其中位点 和 之间的相互作用能由 和 相互作用的算术平均值给出。几何平均值不一定适用于所有 mAb 混合物，并且由于静电相互作用的不可加性，预计在较低的离子强度下会失效。
