Protein solubilization: A novel approach

Abstract

Formulation development presents significant challenges with respect to protein therapeutics. One component of these challenges is to attain high protein solubility (> 50 mg/ml for immunoglobulins) with minimal aggregation. Protein-protein interactions contribute to aggregation and the integral sum of these interactions can be quantified by a thermodynamic parameter known as the osmotic second virial coefficient (B-value). The method presented here utilizes high-throughput measurement of B-values to identify the influence of additives on protein-protein interactions. The experiment design uses three tiers of screens to arrive at final solution conditions that improve protein solubility. The first screen identifies individual additives that reduce protein interactions. A second set of B-values are then measured for different combinations of these additives via an incomplete factorial screen. Results from the incomplete factorial screen are used to train an artificial neural network (ANN). The “trained” ANN enables predictions of B-values for more than 4,000 formulations that include additive combinations not previously experimentally measured. Validation steps are incorporated throughout the screening process to ensure that 1) the protein’s thermal and aggregation stability characteristics are not reduced and 2) the artificial neural network predictive model is accurate. The ability of this approach to reduce aggregation and increase solubility is demonstrated using an IgG protein supplied by Minerva Biotechnologies, Inc.

Feasibility of Using Gluconolactone, Trehalose and Hydroxy-Propyl Gamma Cyclodextrin to Enhance Bendroflumethiazide Dissolution Using Lyophilisation and Physical Mixing Techniques

“…Trehalose is a disaccharide formed by a 1,1-glucoside bond between two α-glucose units, freely soluble in water, and used as a carrier to increase protein stability, trehalose is used with protein formulations to prevent degradation and to enhance proteins’ stability [ 30 ] and it was used in this study to investigate its effect on drug dissolution.…”

Low-volume solubility assessment during high-concentration protein formulation development

“…Comparable effects to previous studies were observed, where chaotropic salts and increasing ionic strength decreased predominant attractive interactions and enhanced protein solubility. In contrast, very little data are available for protein solubility dependent on other additives . Besides the effect of pH and buffer salts, valuable data are presented in this study about the influence of sugars, amino acids or surfactants on protein solubility without the influence of pH and ionic strength.…”

“…A widely recognized approach uses protein–protein interactions (PPI) to predict solubility behaviour of proteins. While attractive interactions are unfavourable, repulsive interactions are preferred during HCF development, as they increase protein solubility . Most investigations were performed under dilute conditions utilizing the second virial coefficient, B 22 .…”

Structure, heterogeneity and developability assessment of therapeutic antibodies

“…224,225 Among many different ways to obtain B 22 values, such as selfinteraction chromatography (SIC), 226 membrane osmometry (MO) 227 and analytical ultracentrifugation (AUC), 228 the most common method is through static light scattering (SLS). 225,229,230 Recently this value was used to determine a universal solubility line, the "liquidus" line, as part of a phase diagram for a mAb. 231,232 Cross-interaction chromatography (CIC) assay, introduced by Jacob et al 233 takes advantage of the accumulative effect on a column to capture weak binding between testing mAb and a large quantity (30 mgs) of immobilized human serum IgGs.…”