Submicron Aggregation of Chemically Denatured Monoclonal Antibody

Isothermal chemical denaturation (ICD) has been widely used to evaluate the conformational stability of therapeutic proteins such as monoclonal antibodies. However, the chemical unfolding pathway and the subsequent aggregation of antibodies are not yet well-understood. In the present work, we conduc...

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Bibliographic Details
Published in:	Molecular pharmaceutics Vol. 15; no. 10; pp. 4710 - 4721
Main Authors:	Rowe, Jacob B, Flynn, Rhiannon P, Wooten, Harrison R, Noufer, Hailey A, Cancel, Rachel A, Zhang, Jifeng, Subramony, J. Anand, Pechenov, Sergei, Wang, Ying
Format:	Journal Article
Language:	English
Published:	United States American Chemical Society 01-10-2018
Subjects:	differential scanning calorimetry (DSC) protein unfolding dynamic light scattering (DLS) isothermal chemical denaturation (ICD) protein aggregation submicron particles
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Summary:	Isothermal chemical denaturation (ICD) has been widely used to evaluate the conformational stability of therapeutic proteins such as monoclonal antibodies. However, the chemical unfolding pathway and the subsequent aggregation of antibodies are not yet well-understood. In the present work, we conducted a systematic study on an ICD-induced aggregation of a pharmaceutical monoclonal antibody. Using dynamic light scattering, we monitored formation and growth of submicron aggregates in various buffers. Our experiments revealed a nucleation-controlled submicron aggregation of the antibody in the presence of chemical denaturant. After the unfolded protein reached a steady state, we reduced the denaturant concentration by dilution or dialysis to trigger further aggregation after ICD. In this way, we studied the pH effect on aggregation of the stressed protein after removal of denaturant. The ICD-dilution experiment provides a practical means for studying the propensity of unfolded proteins to form aggregates under various formulation conditions. This unique method allows us to control the degree of protein unfolding and the initiation of post-ICD aggregation.
Bibliography:	ObjectType-Article-1 SourceType-Scholarly Journals-1 ObjectType-Feature-2 content type line 23
ISSN:	1543-8384 1543-8392
DOI:	10.1021/acs.molpharmaceut.8b00690