Developing a Panel of Recombinant Antibody Controls for Cytokine Release Assays

The National Institute for Biological Standards and Control (NIBSC), a UK-based organization with a leading role in creating international standards and reference materials, recently developed the first antibody reference panel for the qualification and validation of cytokine release assays, which are critical tools for the preclinical safety assessment of novel therapeutic antibodies.

Absolute Antibody was chosen to recombinantly produce the reference antibodies, manufacturing gram quantities of two research-grade biosimilar positive controls and three isotype-matched negative controls using our proprietary transient expression system.

Antibody Panel for Cytokine Release Assays

It is important to ensure that new antibody therapeutics do not cause undesired immune reactions in humans before clinical trials begin. One concern is cytokine release syndrome (CRS), a rapid systemic inflammatory response characterized by the secretion of pro-inflammatory cytokines from immune cells. A variety of in vitro cytokine release assays (CRAs) are used to evaluate the CRS risk of new antibody drugs in preclinical safety studies. To date, however, a key challenge has been the lack of available standardized positive and negative control antibodies for use in assay qualification. Labs might struggle to source clinically relevant controls due to high cost or restricted availability, and each company uses their own control set, which makes it difficult to compare CRA performance.

The NIBSC therefore led an international effort to generate and validate the first antibody reference panel for cytokine release assays. The panel includes three antibodies shown to induce CRS in the clinic at varying levels, as well as three isotype-matched negative controls. It was evaluated by 11 laboratories from leading pharmaceutical companies worldwide, showing that cytokine release was induced at expected levels in a variety of assay platforms. The international collaborative study was published in the journal Cytokine X.

“The use of this common set of positive and negative control monoclonal antibodies will provide information on the reproducibility, robustness and potential limitations of a CRA platform,” said Dr. Sandrine Vessillier, Head of Immunotoxicology at NIBSC and lead author on the paper. “It will establish confidence in the ability of any novel CRA to identify potential CRS risk.”

Positive and Negative Control Antibodies

Absolute Antibody was selected to manufacture the antibodies for the CRA reference panel after responding to a NIBSC proposal request, as we were able to supply one gram of each antibody at the desired quality specifications. We used our transient antibody expression platform to recombinantly manufacture aggregate-free antibodies with high purity (≥98%), no albumin contamination, and low endotoxin levels compatible with all in vitro and in vivo applications.

We provided the following positive control antibodies for the CRA reference panel: Anti-CD52 [Campath-1H or Alemtuzumab] in human IgG1 format and Anti-CD3 epsilon [OKT-3 or Muromonab] in mouse IgG2a format. We also provided three isotype-matched negative controls from our anti-NP panel of recombinant isotype controls: human IgG1, human IgG4 and mouse IgG2a.

The entire panel of recombinant antibody controls for cytokine release assays is now available for researchers worldwide via the NIBSC catalog here. For more information, read the full text of the new paper or contact us with any questions.

 

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