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Experimental Model System to Study pH Shift-Induced Aggregation of Monoclonal Antibodies Under Controlled Conditions

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Abstract

Purpose

To present a convenient screening method for evaluating additive effects on the renaturation of an acid-exposed monoclonal antibody (mAb).

Methods

The assay involves brief incubation of a mAb at acidic pH and subsequent neutralization in the absence or presence of additive to induce mainly aggregation. An increase in absorbance depicted aggregation. The recorded aggregation data traces were fitted with a nucleation-autocatalytic growth model for the extraction of kinetic parameters.

Results

All kinetic data traces were fitted successfully with the selected model and the adjusted R square values were greater than 0.99. Trehalose had strongly stabilizing, proline mildly stabilizing and trimethylamine oxide had destabilizing effects on both the nucleation and growth phase of the reaction. Histidine was strongly stabilizing but was limited by its poor solubility.

Conclusion

The results demonstrate the suitability of the experimental mAb aggregation system and the nucleation-autocatalytic growth fit in the screening and quantification of additive effects on the renaturation of an acid-exposed mAb respectively. This will aid the investigation and derivation of quantitative structure-activity relationships of additive effects on mAb solubility.

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Abbreviations

Bis-ANS:

4, 4′-Bis (1-anilinonaphthalene 8-sulfonate)

ca.:

Circa

DMSO:

Dimethyl sulfoxide

DSF:

Differential Scanning Fluorimetry

Eq.:

Equation

HCl:

Hydrochloric acid

HPLC:

High Pressure Liquid Chromatography

HP-SEC:

High Performance–Size Exclusion Chromatography

mAb:

Monoclonal antibody

min:

Minutes

NaOH:

Sodium hydroxide

QSAR:

Quantitative Structure Activity Relationship

s:

Seconds

Tm :

(Apparent) Melting temperatures

TMAO:

Trimethylamine N-oxide

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ACKNOWLEDGMENTS AND DISCLOSURES

This research was funded by the Federal Ministry of Education and Research, Germany (BMBF), grant number 0315338A. The authors are grateful to Bernd Burghardt for preliminary data exploration and programming sessions in R statistical software. Thanks are due to René Handrick, Karen Schwab, Fabian Bickel, Albrecht Weber and Alba Signes Marrahi for gifts of mAb1 at various time points during the execution of this work.

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  1. Institute of Applied Biotechnology, Biberach University of Applied Sciences, Hubertus Liebrecht 35, 88400, Biberach Riss, Baden-Wurttemberg, Germany

    Olubukayo-Opeyemi Oyetayo & Hans Kiefer

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  1. Olubukayo-Opeyemi Oyetayo
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Correspondence to Olubukayo-Opeyemi Oyetayo.

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Oyetayo, OO., Kiefer, H. Experimental Model System to Study pH Shift-Induced Aggregation of Monoclonal Antibodies Under Controlled Conditions. Pharm Res 33, 1359–1369 (2016). https://doi.org/10.1007/s11095-016-1878-4

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  • DOI: https://doi.org/10.1007/s11095-016-1878-4

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