Subclass Effects on Self-Association and Viscosity of Monoclonal Antibodies at High Concentrations

AA Chowdhury and N Manohar and MA Witek and MA Woldeyes and R Majumdar and KK Qian and WD Kimball and SF Xu and A Lanzaro and TM Truskett and KP Johnston, MOLECULAR PHARMACEUTICS, 20, 2991-3008 (2023).

DOI: 10.1021/acs.molpharmaceut.3c00023

The effects of a subclass of monoclonal antibodies (mAbs)on protein- proteininteractions, formation of reversible oligomers (clusters), and viscosity(eta) are not well understood at high concentrations. Herein,we quantify a short-range anisotropic attraction between the complementarity-determiningregion (CDR) and CH3 domains (KCDR-CH3) for vedolizumabIgG1, IgG2, or IgG4 subclasses by fitting small-angle X-ray scattering(SAXS) structure factor S (eff)(q) data with an extensive library of 12-bead coarse-grained (CG) moleculardynamics simulations. The KCDR-CH3 bead attractionstrength was isolated from the strength of long-range electrostaticrepulsion for the full mAb, which was determined from the theoreticalnet charge and a scaling parameter psi to account for solventaccessibility and ion pairing. At low ionic strength (IS), the strongestshort-range attraction (KCDR-CH3) and consequentlythe largest clusters and highest eta were observed with IgG1,the subclass with the most positively charged CH3 domain. Furthermore,the trend in KCDR-CH3 with the subclass followedthe electrostatic interaction energy between the CDR and CH3 regionscalculated with the BioLuminate software using the 3D mAb structureand molecular interaction potentials. Whereas the equilibrium clustersize distributions and fractal dimensions were determined from fitsof SAXS with the MD simulations, the degree of cluster rigidity underflow was estimated from the experimental eta with a phenomenologicalmodel. For the systems with the largest clusters, especially IgG1,the inefficient packing of mAbs in the clusters played the largestrole in increasing eta, whereas for other systems, the relativecontribution from stress produced by the clusters was more significant.The ability to relate eta to short-range attraction from SAXSmeasurements at high concentrations and to theoretical characterizationof electrostatic patches on the 3D surface is not only of fundamentalinterest but also of practical value for mAb discovery, processing,formulation, and subcutaneous delivery.

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