In situ precipitation and vacuum drying of interferon alpha-2a: Development of a single-step process for obtaining dry, stable protein formulation

Abstract

Feasibility studies were performed to develop a process for obtaining stable dry protein formulations based on in situ polyethylene glycol (PEG)-induced precipitation and vacuum drying of interferon alpha-2a (IFNα2a) solution in a vial. Using a laboratory scale freeze dryer, the process was carried out in two phases: first, protein solution containing PEG was concentrated to achieve protein precipitation, and second, remaining water was removed by further reducing the chamber pressure. Drying conditions, i.e. temperature and pressure, and solution composition were selected to ensure maximal precipitation (solubility of IFNα2a), to achieve precipitation without boiling, and to ensure stability. Dried formulations were subjected to stability studies (40 °C). Concentration and precipitation could be achieved at a fast rate by utilizing pressures slightly above the vapor pressure of water. Fluorescence and circular dichroism (CD) studies showed that precipitated IFNα2a maintained its native structure. Fourier transform infrared spectroscopy (FTIR) studies showed that IFNα2a when dried in the presence of trehalose, maintained its secondary structure. Trehalose also prevented formation of aggregates during drying. Moisture contents of 1% (w/w) were achieved within 48 h of drying. Dry formulation containing 1:20:100 (w/w) IFNα2a:trehalose:mannitol was stable against aggregation and oxidation (6% oxidized at 40 °C, 6 months). Stability profile was comparable to a similar lyophilized formulation.