Flow Activation\nEnergy of High-Concentration Monoclonal\nAntibody Solutions and Protein–Protein Interactions Influenced\nby NaCl and Sucrose

The solution viscosity and protein–protein interactions\n(PPIs) as a function of temperature (4–40 °C) were measured\nat a series of protein concentrations for a monoclonal antibody (mAb)\nwith different formulation conditions, which include NaCl and sucrose.\nThe flow activation energy (<i>E</i><sub>η</sub>)\nwas extracted from the temperature dependence of solution viscosity\nusing the Arrhenius equation. PPIs were quantified via the protein\ndiffusion interaction parameter (<i>k</i><sub>D</sub>) measured\nby dynamic light scattering, together with the osmotic second virial\ncoefficient and the structure factor obtained through small-angle\nX-ray scattering. Both viscosity and PPIs were found to vary with\nthe formulation conditions. Adding NaCl introduces an attractive interaction\nbut leads to a significant reduction in the viscosity. However, adding\nsucrose enhances an overall repulsive effect and leads to a slight\ndecrease in viscosity. Thus, the averaged (attractive or repulsive)\nPPI information is not a good indicator of viscosity at high protein\nconcentrations for the mAb studied here. Instead, a correlation based\non the temperature dependence of viscosity (i.e., <i>E</i><sub>η</sub>) and the temperature sensitivity in PPIs was observed\nfor this specific mAb. When <i>k</i><sub>D</sub> is more\nsensitive to the temperature variation, it corresponds to a larger\nvalue of <i>E</i><sub>η</sub> and thus a higher viscosity\nin concentrated protein solutions. When <i>k</i><sub>D</sub> is less sensitive to temperature change, it corresponds to a smaller\nvalue of <i>E</i><sub>η</sub> and thus a lower viscosity\nat high protein concentrations. Rather than the absolute value of\nPPIs at a given temperature, our results show that the temperature\nsensitivity of PPIs may be a more useful metric for predicting issues\nwith high viscosity of concentrated solutions. In addition, we also\ndemonstrate that caution is required in choosing a proper protein\nconcentration range to extract <i>k</i><sub>D</sub>. In\nsome excipient conditions studied here, the appropriate protein concentration\nrange needs to be less than 4 mg/mL, remarkably lower than the typical\nconcentration range used in the literature.