Profiling antibody drug conjugate positional isomers: a system-of-equations approach.

Antibody drug conjugates enable the targeted delivery of potent chemotherapeutic agents directly to cancerous cells. They are made by the chemical conjugation of cytotoxins to monoclonal antibodies, which can be achieved by first reducing interchain disulfide bonds followed by conjugation of the resulting free thiols with drugs. This process yields a controlled, but heterogeneous, population of conjugated products that contains species with various numbers of drugs linked to different former interchain disulfide cysteine residues on the antibodies. We have developed a mathematical approach using inputs from capillary electrophoresis and hydrophobic interaction chromatography to determine the positional isomer distribution within a population of antibody drug conjugates. The results are confirmed by analyzing isolated samples of specific drug-to-antibody ratio species. The procedure is amenable to rapid determination of positional isomer distributions and features low material requirements. A survey of several antibody drug conjugates based on the same IgG framework and small molecule drug combination has shown a very similar distribution of isomers among all of the molecules using this technique, suggesting a robust conjugation process.

Viscosity behavior of high-concentration protein mixtures.

Highly concentrated protein solutions are becoming increasingly commonplace within the biopharmaceutical industry as more products are developed that feature high doses of drug intended for subcutaneous administration. An as-yet undeveloped subclass of these products feature multiple proteins coformulated together in high-concentration protein mixtures. Previous work has illustrated that the viscosity of aqueous solutions of various proteins at high concentrations can be remarkably different across otherwise similar molecules. This work characterizes the viscosity behavior of mixtures of such proteins, primarily monoclonal antibodies, and shows that a simple mixing rule first proposed by Arrhenius predicts the viscosity of an arbitrary mixture. This approach is shown to successfully calculate the viscosity of mixtures of proteins ranging up to 250 mg/mL total protein concentration and approximately 1700 cP at different ionic strengths and with accuracy errors of less than 10%. Only information about the viscosity of the isolated protein components of the mixture is required for the calculations.