High concentration formulations of recombinant human interleukin-1 receptor antagonist: I. Physical characterization

At relatively high protein concentrations (i.e., up to 100 mg/mL), recombinant human interleukin-1 receptor antagonist (rhIL-1ra) was found to exist in a monomer-dimer equilibrium controlled by solution ionic strength. Sedimentation equilibrium at 25 degrees C was used to measure the increase in the dimer dissociation constant (K-d) as a function of ionic strength. K-d increased from 2.0 to 12.6 mM as the solution ionic strength was increased from 0.011 to 0.184 molal. These K-d values were used with both static light scattering and membrane osmometry data collected over a protein concentration range of 1-100 mg/mL to determine second osmotic virial coefficients. Expanding the second osmotic virial coefficient model to account for separate monomer-monomer (B-22), monomer-dimer (B-23), and dimer-dimer (B-33) interactions reveals net monomer-dimer interactions are attractive, whereas the others are repulsive. Lastly, isothermal titration calorimetry dilution experiments showed that rhIL-1ra dimerization is enthalpically driven (Delta H-dimerization << 0), which is consistent with intermolecular cation-pi interactions previously proposed as the monomer-monomer contact sites in dimers. (C) 2007 Wiley-Liss, Inc. and the American Pharmacists Association.