Raman spectroscopic characterization of drying-induced structural changes in a therapeutic antibody: correlating structural changes with long-term stability

• Published in: Journal of pharmaceutical sciences Vol. 93; no. 4; p. 1005
• Main Authors: Sane, Samir U, Wong, Rita, Hsu, Chung C
• Format: Journal Article
• Language: English
• Published: United States 01-04-2004
• Subjects: Antibodies, Monoclonal - chemistry; Chemistry, Pharmaceutical; Chromatography, Gel; Drug Stability; Freeze Drying; Histidine - chemistry; Humans; Pharmaceutical Solutions; Protein Conformation; Protein Structure, Secondary; Spectrum Analysis, Raman; Sucrose; Trehalose - chemistry
• ISSN: 0022-3549
• DOI: 10.1002/jps.20014

We characterized the secondary structure of a therapeutic recombinant humanized monoclonal antibody (rhuMAb), formulated with different concentrations of sucrose, trehalose, and histidine and in solution, lyophilized, and spray-dried states. Quantitative secondary structure estimates were obtained using amide I band Raman spectroscopy and a previously developed spectral deconvolution procedure. On lyophilization or spray drying in the absence of sugar, the antibody underwent significant structural perturbation. The beta-sheet content decreased with corresponding gain in the turn and unordered content. With increasing amount of sucrose or trehalose, the extent of structural perturbation decreased. Eventually, at sugar-to-protein molar ratios of > or =360, almost complete structural preservation was observed. Histidine also protected the antibody against lyophilization-induced structural changes. The extent of structural perturbation immediately after lyophilization or spray drying exhibited good correlation with the rate of aggregation for the antibody during long-term storage under accelerated conditions. The results demonstrate that amide I band Raman spectroscopy could be a quick and reliable way to screen excipients and their concentrations during lyophilized or spray dried formulation development.