Mechanistic model-based characterization of size-exclusion-mixed-mode resins for removal of monoclonal antibody fragments.

Autor: Altern SH; Department of Chemical and Biological Engineering, Rensselaer Polytechnic Institute, Troy, NY, USA; Center for Biotechnology and Interdisciplinary Studies, Rensselaer Polytechnic Institute, Troy, NY, USA., Kocot AJ; Department of Chemical and Biological Engineering, Rensselaer Polytechnic Institute, Troy, NY, USA; Center for Biotechnology and Interdisciplinary Studies, Rensselaer Polytechnic Institute, Troy, NY, USA., LeBarre JP; Department of Chemical and Biomolecular Engineering, North Carolina State University, Raleigh, NC, USA., Boi C; Department of Chemical and Biomolecular Engineering, North Carolina State University, Raleigh, NC, USA; Biomanufacturing Training and Education Center (BTEC), North Carolina State University, Raleigh, NC, USA; Department of Civil, Chemical, Environmental, and Materials Engineering, University of Bologna, Bologna, Italy., Phillips MW; Downstream Research and Development, EMD Millipore Corporation, Burlington, MA, USA., Roush DJ; Process Research and Development, Merck & Co., Inc., Rahway, NJ, USA., Menegatti S; Department of Chemical and Biomolecular Engineering, North Carolina State University, Raleigh, NC, USA; Biomanufacturing Training and Education Center (BTEC), North Carolina State University, Raleigh, NC, USA; North Carolina Viral Vector Initiative in Research and Learning (NC-VVIRAL), North Carolina State University, Raleigh, NC, USA., Cramer SM; Department of Chemical and Biological Engineering, Rensselaer Polytechnic Institute, Troy, NY, USA; Center for Biotechnology and Interdisciplinary Studies, Rensselaer Polytechnic Institute, Troy, NY, USA. Electronic address: crames@rpi.edu.
Jazyk: angličtina
Zdroj: Journal of chromatography. A [J Chromatogr A] 2024 Mar 15; Vol. 1718, pp. 464717. Date of Electronic Publication: 2024 Feb 08.
DOI: 10.1016/j.chroma.2024.464717
Abstrakt: Although antibody fragments are a critical impurity to remove from process streams, few platformable purification techniques have been developed to this end. In this work, a novel size-exclusion-mixed-mode (SEMM) resin was characterized with respect to its efficacy in mAb fragment removal. Inverse size-exclusion chromatography showed that the silica-based resin had a narrow pore size distribution and a median pore radius of roughly 6.2 nm. Model-based characterization was carried out with Chromatography Analysis and Design Toolkit (CADET), using the general rate model and the multicomponent Langmuir isotherm. Model parameters were obtained from fitting breakthrough curves, performed at multiple residence times, for a mixture of mAb, aggregates, and an array of fragments (varying in size). Accurate fits were obtained to the frontal chromatographic data across a range of residence times. Model validation was then performed with a scaled-up column, altering residence time and feed composition from the calibration run. Accurate predictions were obtained, thereby illustrating the model's interpolative and extrapolative capabilities. Additionally, the SEMM resin achieved 90% mAb yield, 37% aggregate removal, 29% [Formula: see text] removal, 54% Fab/Fc removal, 100% Fc fragments removal, and a productivity of 72.3 g mAbL×h. Model predictions for these statistics were all within 5%. Simulated batch uptake experiments showed that resin penetration depth was directly related to protein size, with the exception of the aggregate species, and that separation was governed by differential pore diffusion rates. Additional simulations were performed to characterize the dependence of fragment removal on column dimension, load density, and feed composition. Fragment removal was found to be highly dependent on column load density, where optimal purification was achieved below 100 mg protein/mL column. Furthermore, fragment removal was dependent on column volume (constant load mass), but agnostic to whether column length or diameter was changed. Lastly, the dependence on feed composition was shown to be complex. While fragment removal was inversely related to fragment mass fraction in the feed, the extent depended on fragment size. Overall, the results from this study illustrated the efficacy of the SEMM resin in fragment and aggregate removal and elucidated relationships with key operational parameters through model-based characterization.
Competing Interests: Declaration of Competing Interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Steven M. Cramer reports financial support was provided by National Institute for Innovation in Manufacturing Biopharmaceuticals. All other authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.
(Copyright © 2024 Elsevier B.V. All rights reserved.)
Databáze: MEDLINE
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