Decoupling Protein Concentration and Aggregate Content Using Diffusion and Water NMR

Overview

Journal Anal Chem

Specialty Chemistry

Date 2024 Jun 29

PMID 38943616

Authors

Mark I Grimes

Matthew Cheeks

Jennifer Smith

Fabio Zurlo

Mick D Mantle

Affiliations

Soon will be listed here.

Abstract

Protein-based biopharmaceutical drugs, such as monoclonal antibodies, account for the majority of the best-selling drugs globally in recent years. For bioprocesses, key performance indicators are the concentration and aggregate level for the product being produced. In water NMR (NMR), the use of the water transverse relaxation rate [(HO)] has been previously used to determine protein concentration and aggregate level; however, it cannot be used to separate between them without using an additional technique. This work shows that it is possible to "decouple" these two key characteristics by recording the water diffusion coefficient [(HO)] in conjunction with (HO), even in the event of overlap in either (HO) or (HO). This method is demonstrated on three different systems, following appropriate (HO) or (HO) calibration data acquisition for a protein of interest. Our method highlights the potential use of benchtop NMR as an at-line process analytical technique.

References

Foster R, Damion R, Baboolal T, Smye S, Ries M . A nuclear magnetic resonance study of water in aggrecan solutions. R Soc Open Sci. 2016; 3(3):150705. PMC: 4821274. DOI: 10.1098/rsos.150705. View

Feng Y, Taraban M, Yu Y . Linear dependence of the water proton transverse relaxation rate on the shear modulus of hydrogels. Chem Commun (Camb). 2014; 50(81):12120-2. PMC: 4176820. DOI: 10.1039/c4cc04717f. View

Taraban M, Briggs K, Merkel P, Yu Y . Flow Water Proton NMR: In-Line Process Analytical Technology for Continuous Biomanufacturing. Anal Chem. 2019; 91(21):13538-13546. DOI: 10.1021/acs.analchem.9b02622. View

Chen J, Sambol E, Kennealey P, OConnor R, DeCarolis P, Cory D . Water suppression without signal loss in HR-MAS 1H NMR of cells and tissues. J Magn Reson. 2004; 171(1):143-50. DOI: 10.1016/j.jmr.2004.08.009. View

Colsenet R, Mariette F, Cambert M . NMR relaxation and water self-diffusion studies in whey protein solutions and gels. J Agric Food Chem. 2005; 53(17):6784-90. DOI: 10.1021/jf050162k. View

Zalesskiy S, Danieli E, Blumich B, Ananikov V . Miniaturization of NMR systems: desktop spectrometers, microcoil spectroscopy, and "NMR on a chip" for chemistry, biochemistry, and industry. Chem Rev. 2014; 114(11):5641-94. DOI: 10.1021/cr400063g. View

Hermeling S, Crommelin D, Schellekens H, Jiskoot W . Structure-immunogenicity relationships of therapeutic proteins. Pharm Res. 2004; 21(6):897-903. DOI: 10.1023/b:pham.0000029275.41323.a6. View

Bielser J, Wolf M, Souquet J, Broly H, Morbidelli M . Perfusion mammalian cell culture for recombinant protein manufacturing - A critical review. Biotechnol Adv. 2018; 36(4):1328-1340. DOI: 10.1016/j.biotechadv.2018.04.011. View

Sinharoy P, Aziz A, Majewska N, Ahuja S, Handlogten M . Perfusion reduces bispecific antibody aggregation via mitigating mitochondrial dysfunction-induced glutathione oxidation and ER stress in CHO cells. Sci Rep. 2020; 10(1):16620. PMC: 7538420. DOI: 10.1038/s41598-020-73573-4. View

10.

Hass R, Munzke D, Vargas Ruiz S, Tippmann J, Reich O . Optical monitoring of chemical processes in turbid biogenic liquid dispersions by Photon Density Wave spectroscopy. Anal Bioanal Chem. 2015; 407(10):2791-802. DOI: 10.1007/s00216-015-8513-9. View

11.

Capito F, Skudas R, Kolmar H, Hunzinger C . Mid-infrared spectroscopy-based antibody aggregate quantification in cell culture fluids. Biotechnol J. 2013; 8(8):912-7. DOI: 10.1002/biot.201300164. View

12.

Marchand A, Mishra R, Bernard A, Dumez J . Online Reaction Monitoring with Fast and Flow-Compatible Diffusion NMR Spectroscopy. Chemistry. 2022; 28(52):e202201175. DOI: 10.1002/chem.202201175. View

13.

Parr M, Montacir O, Montacir H . Physicochemical characterization of biopharmaceuticals. J Pharm Biomed Anal. 2016; 130:366-389. DOI: 10.1016/j.jpba.2016.05.028. View

14.

Song J, Taraban M, Yu Y, Lu L, Biswas P, Xu W . biophysical characterization of high-concentration protein formulations using NMR. MAbs. 2024; 16(1):2304624. PMC: 10841025. DOI: 10.1080/19420862.2024.2304624. View

15.

Wang Q, Chen Y, Park J, Liu X, Hu Y, Wang T . Design and Production of Bispecific Antibodies. Antibodies (Basel). 2019; 8(3). PMC: 6783844. DOI: 10.3390/antib8030043. View

16.

Taraban M, Jones M, Yu Y . Rapid and Noninvasive Quantification of Capsid Gene Filling Level Using Water Proton Nuclear Magnetic Resonance. Anal Chem. 2021; 93(48):15816-15820. DOI: 10.1021/acs.analchem.1c04088. View

17.

Urquhart L . Top companies and drugs by sales in 2020. Nat Rev Drug Discov. 2021; 20(4):253. DOI: 10.1038/d41573-021-00050-6. View

18.

Carter P . Potent antibody therapeutics by design. Nat Rev Immunol. 2006; 6(5):343-57. DOI: 10.1038/nri1837. View

19.

Bryant R, Mendelson D, Lester C . The magnetic field dependence of proton spin relaxation in tissues. Magn Reson Med. 1991; 21(1):117-26. DOI: 10.1002/mrm.1910210114. View

20.

Briggs K, Taraban M, Yu Y . Quality assurance at the point-of-care: Noninvasively detecting vaccine freezing variability using water proton NMR. Vaccine. 2020; 38(31):4853-4860. DOI: 10.1016/j.vaccine.2020.05.049. View