Most therapeutic proteins encounter stresses that could lead to denaturation during or after lyophilization. Adding appropriate excipients and the design of suitable drying cycles are crucial for ensuring the retention of native structure in lyophilized proteins of pharmaceutical significance. Biophysical techniques that can interrogate proteins in the solid state are necessary to optimize formulation and process development. Solid-state hydrogen deuterium exchange mass spectrometry (ssHDX-MS) is among the emerging techniques for the analysis of conformational stability of lyophilized protein therapeutics. Herein, we have analyzed the effect of different drying methods (freeze drying with uncontrolled and controlled ice nucleation and spray drying) on a fragment antigen-binding (Fab) protein formulated with sucrose using ssHDX-MS. The solid Fab formulations were exposed to D2O in the vapor phase at controlled 11% RH and at 23°C, deuterium incorporation at different time points was determined using LC-MS. The extent of deuterium uptake and the kinetic parameters showed that the spray drying of a Fab-sucrose spray dried formulation had less deuterium incorporation than in two other drying processes; in previous studies using myoglobin formulations, lower deuterium incorporation was correlated with better storage stability. The relationship between deuterium uptake and stability for the Fab formulations, that were prepared using different processing methods, will be discussed in this presentation.

Solid-state hydrogen-deuterium exchange mass spectrometry (ssHDX-MS) analysis of therapeutic fragment antigen-binding (Fab) protein formulations

253rd National Meeting (2017)

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