Assessing the Utility of Circular Dichroism and FTIR Spectroscopy in Monoclonal-Antibody Comparability Studies.
Protein characterization is a necessary activity during development, technical transfers, and licensure. One important aspect of protein characterization is higher order structure assessment, which can be accomplished in a variety of ways. Circular dichroism (CD) and Fourier transform infrared (FTIR) spectroscopies provide global higher order structure and are routinely used to measure the overall structure for product characterization; however, their use as comparability tools is uncertain because of their insensitivity to local or small structure changes. We use a monoclonal antibody (mAb) to explore the usefulness of CD and FTIR compared with other indirect methods of structure characterization such as size-exclusion and ion-exchange chromatographies (SEC and IEC). A panel of degraded samples of a mAb was generated; their higher order structure evaluated using CD and FTIR and was found to be largely unchanged. However, the SEC and IEC chromatograms of certain degraded samples were found to have measurable changes. Based on these studies, we conclude that the application of CD and FTIR should be reserved for global higher order structure identification or product characterization only. The use of CD or FTIR comparability of mAbs should be carefully evaluated, as comparability can be sensitively determined using indirect methods based on chromatography.
Late Stage Pharmaceutical Development, Genentech, South San Francisco, California 94080. Electronic address: lin.jasper@gene.com.
Jasper C
JC
Lin
Late Stage Pharmaceutical Development, Genentech, South San Francisco, California 94080.
Zephania Kwong
ZK
Glover
Late Stage Pharmaceutical Development, Genentech, South San Francisco, California 94080.
Alavattam
A
Sreedhara
D000911:Antibodies, Monoclonal / Q000737:chemistry*; D002850:Chromatography, Gel / Q000379:methods; D002852:Chromatography, Ion Exchange / Q000379:methods; D002942:Circular Dichroism / Q000379:methods; D011487:Protein Conformation; D011506:Proteins / Q000737:chemistry; D017550:Spectroscopy, Fourier Transform Infrared / Q000379:methods
D000911:Antibodies, Monoclonal; D011506:Proteins
FTIR; HPLC; biophysical characterization of proteins; circular dichroism; comparability; electronic circular dichroism; monoclonal antibodies; protein structure; proteins higher order structure