Abstract
An In Vitro Comparative Immunogenicity Assessment (IVCIA) assay was evaluated as a tool for predicting the potential relative immunogenicity of biotherapeutic attributes. Peripheral blood mononuclear cells from up to 50 healthy naïve human donors were monitored up to 8 days for T-cell proliferation, the number of IL-2 or IFN-γ secreting cells, and the concentration of a panel of secreted cytokines. The response in the assay to 10 monoclonal antibodies was found to be in agreement with the clinical immunogenicity, suggesting that the assay might be applied to immunogenicity risk assessment of antibody biotherapeutic attributes. However, the response in the assay is a measure of T-cell functional activity and the alignment with clinical immunogenicity depends on several other factors. The assay was sensitive to sequence variants and could differentiate single point mutations of the same biotherapeutic. Nine mAbs that were highly aggregated by stirring induced a higher response in the assay than the original mAbs before stirring stress, in a manner that did not match the relative T-cell response of the original mAbs. In contrast, mAbs that were glycated by different sugars (galactose, glucose, and mannose) showed little to no increase in response in the assay above the response to the original mAbs before glycation treatment. The assay was also used successfully to assess similarity between multiple lots of the same mAb, both from the same manufacturer and from different manufacturers (biosimilars). A strategy for using the IVCIA assay for immunogenicity risk assessment during the entire lifespan development of biopharmaceuticals is proposed.
Highlights
Immunogenicity to protein based biotherapeutics is a complex process that involves numerous patient and product specific factors [1,2]
In order to appraise the In Vitro Comparative Immunogenicity Assessment (IVCIA) assay as an effective in vitro tool for assessing the relative risk of immunogenicity in the clinic, ten IgG1 and IgG2 biotherapeutic monoclonal antibodies, with known rates of clinical immunogenicity (Table 1) [26,27], were tested in the IVCIA assay for their ability to elicit an immune response in peripheral blood mononuclear cells (PBMC)
An additional Monoclonal antibodies (mAbs), which has not been tested in the clinic, was tested and is predicted to have a rate of clinical immunogenicity that is intermediate among the selected mAbs based on in silico methods
Summary
Immunogenicity to protein based biotherapeutics is a complex process that involves numerous patient and product specific factors [1,2]. Monoclonal antibodies (mAbs) are a major class of protein biotherapeutics that have many product specific factors that are critical for the quality of the drug product. Several formats of cell-based assay platforms have been explored to assess the risk of immunogenicity These include assay systems using: whole blood, peripheral blood mononuclear cells (PBMC), CD8+-depleted PBMC, immortalized cell lines, dendritic cells/monocytes/macrophages co-cultured with autologous CD4+ Tcells, and artificial lymph node systems, to name a few [5,6,7,8,9,10,11,12]. Various biological outcomes can be measured at different stages of immune cell activation in these in vitro assays including: cytokine secretion, expression of cell surface markers of activation, identification of HLA-DR bound peptides, signal transduction events, phagocytosis by antigen presenting cells (APC), and proliferation
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