title: Intermolecular interactions of IgG1 monoclonal antibodies at high concentrations characterized by light scattering.
abstract: Light scattering intensity measurements of solutions of two purified monoclonal antibodies were performed over a wide range of concentrations (0.5-275 mg/mL) and ionic strengths (0.02 to 0.6 M). Despite extensive sequence homology between these mAbs, alteration of ∼20 amino acids in the complementarity determining regions resulted in different net intermolecular interactions and responses to solution ionic strength. The concentration dependence of scattering was analyzed by comparison with the predictions of three models, allowing for intermolecular interaction of various types. In order of increasing complexity, the three models account for: (1) steric repulsions (simple hard-sphere model), (2) steric repulsion with short-ranged attractive interactions of varying magnitude (adhesive hard-sphere model), and (3) steric and nonsteric repulsive interactions between several species whose relative concentrations may change as a function of total protein concentration as dictated by equilibrium self-association (effective hard-sphere mixture model). Simple scattering models of noninteracting and adhesive hard-sphere species permitted qualitative interpretation of contributions from excluded volume, electrostatic, and van der Waals interactions on net mAb interactions at high concentration as a function of ionic strength. mAb2 electrostatic interactions were repulsive, whereas mAb1 interactions were net attractive at low ionic strengths, attributed to an anisotropic distribution of molecular charge. The effective hard-sphere mixture model can account quantitatively for the dependence of scattering for both antibodies over the entire concentration range and at salt concentrations exceeding 40 mM. This analysis showed that at high ionic strength both mAbs self-associate weakly to form dimer with an affinity that varies little with salt concentration at concentrations exceeding 75 mM. In addition, mAb1 appears to self-associate further to form oligomers with stoichiometry of 4-6 and an affinity that declines substantially with increasing ionic strength. All three models lead to the conclusion that at high concentrations repulsive interactions are predominantly due to excluded volume, whereas additional features are salt-dependent and reflect a substantial electrostatic contribution to intermolecular interactions of both mAbs.
authors.affiliation: Genentech Incorporated, Late Stage Pharmaceutical and Processing Development, 1 DNA Way, South San Francisco, California 94080, USA. tscherer@gene.com
authors.forename: Thomas M
authors.initials: TM
authors.lastname: Scherer
authors.forename: Jun
authors.initials: J
authors.lastname: Liu
authors.forename: Steven J
authors.initials: SJ
authors.lastname: Shire
authors.forename: Allen P
authors.initials: AP
authors.lastname: Minton
mesh_terms: D000911:Antibodies, Monoclonal / Q000737:chemistry*; D007074:Immunoglobulin G / Q000737:chemistry*; D008027:Light; D009994:Osmolar Concentration; D012542:Scattering, Radiation; D055672:Static Electricity
chemical_list: D000911:Antibodies, Monoclonal; D007074:Immunoglobulin G