Biophysical Characterization of Binary Therapeutic Monoclonal Antibody Mixtures

Abstract

Coformulations containing two therapeutic monoclonal antibodies (mAbs) could offer various benefits like enhanced therapeutic efficacy and better patient compliance. However, there are very few published studies on coformulations and binary mixtures of mAbs. It remains unclear to what extent mAbs with different physicochemical properties can be combined in solution without detrimental effects on protein stability. Here, we present a study including six model mAbs of the IgG1 subclass that are commercially available. In silico and biophysical characterization shows that the proteins have different physicochemical properties. Thus, their combinations represent various scenarios for coformulation development. We prepared all possible binary mixtures of the six mAbs and determined several biophysical parameters that are assessed during early-stage protein drug product development. The measured biophysical parameters are indicative of the conformational protein stability (inflection points of the thermal protein unfolding transitions) and the colloidal protein stability (aggregation onset temperatures and interaction parameter k D from dynamic light scattering). Remarkably, all 15 binary mAb mixtures do not exhibit biophysical parameters that indicate inferior conformational or colloidal stability compared to the least stable mAb in the mixture. Our findings suggest that the coformulation of some therapeutic monoclonal antibodies of the IgG1 subclass could be possible in a straightforward way as severe detrimental effects on the stability of these proteins in binary mixtures were not observed.

Stability Convergence in Antibody Coformulations

“…Only a few previous studies have looked into the stability of the coformulation of antibodies. 6,8,9,32 This specific Fab interacted with Hub07 or Hub19 to increase their stability, as we observed previously for a more closely related IgG1. 30 Acting as an aggregation decelerator, the use of a bioactive Fab represents a promising coformulation component for antibody candidates.…”

“…The creation of antibody coformulations can be more challenging than the combination of small molecules. Stability of antibodies is susceptible to changes in temperature, mechanical force, pH, ionic strength, and protein concentration. − Mixtures of antibodies and other therapeutic or excipient proteins could also create the risk of heterogeneous aggregationan irreversible change of protein structure leading to immunogenic species and reduction of biological activity. ,,, Therefore, the impact of coformulation on the individual stability of the antibodies must be carefully investigated during the development of coformulated products.…”

“…Antibody-based therapies have become a major class of pharmaceutical products. The rapid growth of successful antibody therapeutic approvals is now increasingly leading toward their exploration for combined use to achieve a synergistic inhibition of therapeutic targets. − Clinical studies of antibody combinations have shown promising improvement in a number of diseases. , However, multiantibody therapies bring additional complexities and challenges to their innovation as products suitable for combined administration, particularly when coformulated into a single product dosage form. − …”

“… 4 , 5 However, multiantibody therapies bring additional complexities and challenges to their innovation as products suitable for combined administration, particularly when coformulated into a single product dosage form. 6 − 10 …”

Improving Biologics’ Effectiveness in Clinical Oncology: From the Combination of Two Monoclonal Antibodies to Oligoclonal Antibody Mixtures

“…Similarly, the single formulation of three antibodies (atoltivimab + maftivimab + odesivimab) has been approved by the FDA in 2020 for Ebola hemorrhagic fever. As demonstrated for the co-formulation of six approved antibodies [ 83 ], it seems that therapeutic monoclonal antibodies of the IgG1 subclass can be combined without severe detrimental effects to the stability of these proteins in binary mixtures.…”