Article Review – Long-term stability of diluted solutions of the monoclonal antibody rituximab

Full Article Title – Muriel Paul, Victoire Vieillard, Emmanuel Jaccoulet, Alain Astier. Long-term stability of diluted solutions of the monoclonal antibody rituximab. International Journal of Pharmaceutics (2012) 436, pp 282– 290


This article represents a good piece of academic research to investigate changes to the chemical, physical and functional attributes of Rituximab upon long-term storage as a diluted solution. The approach taken in the article to identify changes upon storage is well considered, as the authors appreciate and explicitly state the fundamental requirements for the analytical protocols to evaluate primary, secondary, tertiary and quaternary structure for a comprehensive characterization of chemical/physical stability, as well the need to utilize cell-based methods for evaluating functional activity.

Perhaps most importantly though, the article represents an excellent example of when care and diligence must be exercised in ‘interpreting’ an academic study, particularly when one seeks to consider applying its conclusions to extending the shelf-life of a real product. Although this article stands as a good piece of academic research, it does not attempt to address several key regulatory requirements, nor would these ever be considered during the peer-review process for an academic article.

Here, we briefly describe just a couple of examples where results from this study would not be considered sufficient for their use in applying an extended shelf-life to Rituximab for clinical use.


The article conducts all stability testing onRituximab samples stored at only a single concentration (1 mg/mL). It is recommended (ICH Q5c) that stability testing must be carried out at concentrations representative of the final product, which in the case of Rituximab, would be over a wide range of concentrations as stated in the SPC (1-4 mg/mL).


Turbidity was the only technique used in the article to characterize larger aggregates and insoluble precipitates, so stands as the only method of characterizing and quantifying particulate levels. Strict limits for particle numbers with size ranges ≥10 micron and ≥25 micron are applied in the BP and EP to injectibles, including monoclonal antibodies.

Functional Activity

For the purpose of assigning an extended shelf life, a suitable assay to determine functional activity would need to demonstrate it was unlikely that any significant change in clinical efficacy has occurred as a result of prolonged storage. This article describes the use of a cell based assay for measuring direct cytotoxicity of Rituximab towards the Raji cell line, as its measure of functional activity.


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[2] Glennie MJ, French R, Cragg MS, and Taylor RP: Mechanisms of killing by anti-CD20 monoclonal antibodies. Mol. Immunol. (2007) 44, 3823-3837.