Assessing the immunogenicity of sub-visible aggregates
The presence of aggregates in biotherapeutics can have a critical effect on the biological activity of the molecule as well as immunogenicity in vivo. Manufacturing and product formulation processes, as well as handling of the product during shipment and storage pose certain stress factors for a protein, including freeze/thawing, pH shifts, heating and mechanical stress (stirring and shaking). This may result in the formation of protein aggregates with different properties, e.g. size, solubility, stability and chemical bonding. The presence of protein aggregates is known to facilitate potent immune responses via a variety of mechanisms. Abzena’s data from CD4+ T cell proliferation, MoDC phenotyping and cytokine analysis suggest that only a minor fraction of a protein therapeutic has to be aggregated to induce T cell activation, which in turn could lead to antidrug antibody formation via increased DC activation and consequently T cell responses.
Abzena can artificially stress proteins to induce the formation of sub-visible aggregates of different properties at quantities that may be present in clinical material (after storage and handling). The stressed antibodies are characterised with an orthogonal analytical approach and compared for their immunogenic potential in vitro.
The stress conditions are fine-tuned to ensure that for post-stress samples hardly any monomer is lost and the aggregates that are induced involve only a minor fraction of the total protein in the stressed samples. Therefore, these samples may reflect products that are administered to patients, e.g. after long-term or inadequate storage conditions or handling.
Abzena has developed a variety of ways to measure and characterise the potential immunogenicity of aggregates ex-vivo, these include:
- Measurement of CD4+ T cell proliferation in the presence of aggregated samples
- Measurement of cytokine secretion by cytokine bead array
- Measurement of aggregate uptake by MoDC
Working with Abzena
Abzena’s discovery services are tailored for each project to ensure that the objectives are met or exceeded. Experienced project teams are assigned to each study focusing on progressing projects through to results in the minimum amount of time. Our clients widely regard us as professional and attentive partners who deliver quality results.
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