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High-Throughput Screening of Excipients Intended to Prevent Antigen Aggregation at Air-Liquid Interface

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ABSTRACT

Purpose

The aim was to develop a high-throughput screening method compatible with low protein concentrations, as present in vaccines, in order to evaluate the performance of various excipients in preventing the aggregation at air-liquid interface of an experimental recombinant antigen called Antigen 18A.

Methods

Aggregation of Antigen 18A was triggered by shaking in a half-filled vial or by air bubbling in a microplate. Size-exclusion chromatography, turbidimetry, Nile Red fluorescence spectroscopy, and attenuated total reflection Fourier-transform infrared spectroscopy were used to assess Antigen 18A aggregation. A high-throughput method, based on tryptophan fluorescence spectroscopy, was set up to screen excipients for their capability to prevent Antigen 18A aggregation at air-liquid interface.

Results

While a similar aggregation profile was obtained with both stress tests when using size-exclusion chromatography, spectroscopic and turbidimetric methods showed an influence of the stress protocol on the nature of the aggregates. The high-throughput screening revealed that 7 out of 44 excipients significantly prevented Antigen 18A from aggregating. We confirmed the performance of hydroxypropyl-β-cyclodextrin and hydroxypropyl-γ-cyclodextrin, as well as poloxamers 188 and 407, in half-filled shaken vials.

Conclusions

A high-throughput screening approach can be followed for evaluating the performance of excipients against aggregation of a protein antigen at air-liquid interface.

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ACKNOWLEDGMENTS

This work was supported by GlaxoSmithKline Biologicals. The authors thank BASF, CyDex, Roquette and Sasol for providing gift samples of excipients, Laurent Bessemans and Caroline de Raikem for their support in programming the liquid handling robot, Carine Schroeders for technical assistance with ELISA analyses, Bénédicte Gbaguidi for advice with ATR-FTIR spectroscopy, the GlaxoSmithKline Biologicals R&D Media Preparation team for preparing excipient solutions, and Ulrike Krause and Pascal Cadot for their continuous support in reviewing the manuscript.

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Authors and Affiliations

  1. Louvain Drug Research Institute, Université catholique de Louvain, Unité de Pharmacie Galénique, Avenue E. Mounier 73, UCL 7320, 1200, Brussels, Belgium

    Sébastien Dasnoy & Véronique Préat

  2. GlaxoSmithKline Biologicals, Rue de l’Institut 89, 1330, Rixensart, Belgium

    Nancy Dezutter, Dominique Lemoine & Vivien Le Bras

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  1. Sébastien Dasnoy
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  2. Nancy Dezutter
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  3. Dominique Lemoine
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  5. Véronique Préat
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Correspondence to Véronique Préat.

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Dasnoy, S., Dezutter, N., Lemoine, D. et al. High-Throughput Screening of Excipients Intended to Prevent Antigen Aggregation at Air-Liquid Interface. Pharm Res 28, 1591–1605 (2011). https://doi.org/10.1007/s11095-011-0393-x

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  • DOI: https://doi.org/10.1007/s11095-011-0393-x

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