Ranking mAb-excipient interactions in biologics formulations by NMR spectroscopy and computational approaches

Area of Science:

• Biopharmaceutical formulation development
• Protein stability and characterization
• Molecular interactions

Background:

• Excipients are vital for biopharmaceutical formulations, enhancing protein stability and physicochemical properties.
• The precise mechanisms by which excipients confer stability to biologics, particularly monoclonal antibodies (mAbs), remain incompletely understood.
• Direct experimental evidence of excipient-protein interactions is needed to optimize formulation strategies.

Purpose of the Study:

• To elucidate the mechanism of excipient-mediated protein stabilization by directly measuring binding affinities.
• To rank excipients based on their binding affinity (K_D) and nonspecific binding (N_s) to a monoclonal antibody (mAb).
• To correlate excipient binding and proximity with mAb conformational and colloidal stability.

Main Methods:

• Utilized saturation transfer difference (STD) nuclear magnetic resonance (NMR) spectroscopy to determine mAb-excipient binding affinities.
• Employed molecular dynamics and SILCS-Monte Carlo simulations to assess excipient proximity to the protein.
• Correlated NMR-derived binding data with measurements of mAb conformational and colloidal stability.

Main Results:

• Successfully ranked a series of excipients based on their dissociation constants (K_D) and nonspecific binding constants (N_s) to a mAb.
• Simulation methods corroborated the STD NMR ranking of excipient proximity to the protein.
• Established a correlation between the determined mAb-excipient affinities and the overall stability of the mAb formulation.

Conclusions:

• Direct measurement of mAb-excipient binding affinity provides mechanistic insights into formulation stability.
• The combined NMR and simulation approach offers a powerful tool for ranking excipients.
• This methodology can accelerate and improve excipient selection for biologic formulations, reducing reliance on lengthy screening processes.