Reformulation and Thermal Stability of a Therapeutic Anti-Cocaine mAb

Abstract

We concentrated and reformulated the anti-cocaine mAb, h2E2, to reduce the amount of sucrose and histidine buffer infused with the mAb, to satisfy FDA maximum exposure levels for those components for use in clinical trials. After concentration of the original 20 mg/ml mAb, 4 reformulation buffers were evaluated for suitability. The concentration of histidine was reduced from 10 mM to 3 or 0 mM, and the concentration of sucrose reduced from 10% to 2, 4, or 6%. The approximately 100 mg/ml reformulated mAb samples were analyzed for oligomer formation, aggregation, concentration of the emulsifier polysorbate 80, and thermal stability. These reformulated mAb samples were also assessed for their stability at 40°C from 1 day to 12 weeks. As expected, long term thermal resistance to oligomer formation increased as a function of increasing sucrose concentration. Interestingly, unbuffered reformulated mAb displayed a less than or equal to tendency to form oligomers and aggregates, compared to the histidine buffered samples. Importantly, even after 12 weeks at 40°C, all the reformulated samples displayed little aggregation, and bound their antigen (cocaine) with identical affinities and thermodynamics, as measured by isothermal titration calorimetry (ITC). These ITC thermodynamic binding parameters are consistent with recently published values for the original formulation of this mAb. In all reformulated samples there was a slight decrease in the number of cocaine binding sites after 12 weeks at 40°C, likely due to the parallel small increase in soluble oligomeric antibody, suggesting that soluble oligomeric mAb may no longer bind cocaine with high affinity.