Beyond glass transitions: studying the highly viscous and elastic behavior of frozen protein formulations using low temperature rheology and its potential implications on protein stability.
OBJECTIVE
A novel application of oscillatory shear rheology was used to directly monitor global phase behavior of protein formulations in the frozen state and study its correlation with physical instability of frozen protein formulations.


METHODS
Oscillatory rheology was used to measure changes in rheological parameters and to identify mechanical softening temperature (Ts*) and related properties of an IgG2 mAb formulation. Rheological measurements were compared to DSC/MDSC. Physical stability of IgG2 formulations was monitored by SE-HPLC.


RESULTS
Rheological parameters and Ts* of an IgG2 formulation were sensitive to physical/morphological phase changes during freezing and thawing. Ts* of the frozen formulation was a function of concentration of protein and excipient. Complex modulus, G*, and phase angle, δ, for IgG2 at 70 mg/mL in a sucrose-containing formulation showed the system was not completely frozen at -10°C, which correlated to stability data consistent with ice-induced protein aggregation.


CONCLUSIONS
We report the first application of oscillatory shear rheology to study phase behavior of IgG2 in a sucrose-containing formulation and its correspondence with physical stability not explained by glass transition (Tg'). We provide a mechanism and data suggesting that protein instability occurs at the ice/water interface.
Drug Product Development Process and Product Development, Amgen Inc., MS 8-1-C, One Amgen Center Drive, Thousand Oaks, California, 91320, USA.
Jian Hua
JH
Gu
Alice
A
Beekman
Tian
T
Wu
Deirdre Murphy
DM
Piedmonte
Priti
P
Baker
Michael
M
Eschenberg
Michael
M
Hale
Merrill
M
Goldenberg
D004548:Elasticity; D005079:Excipients / Q000737:chemistry*; D005615:Freezing; D007074:Immunoglobulin G / Q000737:chemistry*; D044367:Phase Transition; D055550:Protein Stability; D012212:Rheology; D013395:Sucrose / Q000737:chemistry*; D055682:Viscoelastic Substances / Q000737:chemistry*; D014783:Viscosity
D005079:Excipients; D007074:Immunoglobulin G; D055682:Viscoelastic Substances; D013395:Sucrose