7 research outputs found
Mechanistic Investigation of Factors Affecting Electrostatic and Hydrodynamic Behavior of Proteins in Formulations at Low Ionic Strength
Archival abstract submitte
Solubility Challenges in High Concentration Monoclonal Antibody Formulations: Relationship with Amino Acid Sequence and Intermolecular Interactions
The
purpose of this work was to elucidate the molecular interactions leading
to monoclonal antibody self-association and precipitation and utilize
biophysical measurements to predict solubility behavior at high protein
concentration. Two monoclonal antibodies (mAb-G and mAb-R) binding
to overlapping epitopes were investigated. Precipitation of mAb-G
solutions was most prominent at high ionic strength conditions and
demonstrated strong dependence on ionic strength, as well as slight
dependence on solution pH. At similar conditions no precipitation
was observed for mAb-R solutions. Intermolecular interactions (interaction
parameter, <i>k</i><sub>D</sub>) related well with high
concentration solubility behavior of both antibodies. Upon increasing
buffer ionic strength, interactions of mAb-R tended to weaken, while
those of mAb-G became more attractive. To investigate the role of
amino acid sequence on precipitation behavior, mutants were designed
by substituting the CDR of mAb-R into the mAb-G framework (GM-1) or
deleting two hydrophobic residues in the CDR of mAb-G (GM-2). No precipitation
was observed at high ionic strength for either mutant. The molecular
interactions of mutants were similar in magnitude to those of mAb-R.
The results suggest that presence of hydrophobic groups in the CDR
of mAb-G may be responsible for compromising its solubility at high
ionic strength conditions since deleting these residues mitigated
the solubility issue