This is an open access article under the terms of the Creative Commons Attribution-NonCommercial-NoDerivs License 4.0 (CC BY-NC-ND 4.0), which permits use and distribution in any medium, provided the original work is properly cited, the use is non-commercial and no modifications or adaptations are made.
Aggregation is a critical issue that hampers the development of monoclonal antibody therapeutics (Mabs). Traditionally, aggregation is considered a process in which native forms of proteins are transformed into an unstable highly associated form through an intermediate formation step. Here we describe the unfolding of an anti CD40 antibody using a folding model based on Lumry-Eyring nucleated polymerization (LENP) model. This model captures several experimental features of the thermal unfolding of this protein as studied by common in situ biophysical techniques such as circular dichroism, fluorescence spectroscopy and turbidity measurements. According to this model, the unfolding and aggregation of the anti CD40 antibody is determined by several distinct steps that include conformational change(s) to generate aggregation prone states, reversible oligomer formation, nucleation and growth as well as their kinetics and the formation of higher order assemblies/aggregates. Furthermore, the loss of monomer is controlled by both thermodynamic (equilibrium unfolding) and kinetic determinants of the unfolding process. This approach captures both of these rate-limiting steps. It can be concluded that this approach is sensitive to formulation conditions such as protein concentration, changes in buffer conditions, and temperature stress. The potential use of this approach in formulation development and stability testing of Mabs is discussed.
Rao, G., Iyer, V., Kosloski, M. P., Pisal, D. S., Shin, E., Middaugh, C. R., & Balu-Iyer, S. V. (2010). Use of a folding model and in situ spectroscopic techniques for rational formulation development and stability testing of Monoclonal antibody therapeutics. Journal of Pharmaceutical Sciences, 99(4), 1697–1706. http://doi.org/10.1002/jps.21938
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