Abstract
Many biopharmaceuticals (BPs) are known to be immunogenic in the clinic, which can result in modified pharmacokinetics, reduced efficacy, allergic reactions and anaphylaxis. During recent years, several technologies to predict immunogenicity have been introduced, but the predictive value is still considered low. Thus, there is an unmet medical need for optimization of such technologies. The generation of T cell dependent high affinity anti-drug antibodies plays a key role in clinical immunogenicity. This study aimed at developing and evaluating a novel in vitro T cell:PBMC assay for prediction of the immunogenicity potential of BPs. To this end, we assessed the ability of infliximab (anti-TNF-α), rituximab (anti-CD20), adalimumab (anti-TNF-α) and natalizumab (anti-α4-integrin), all showing immunogenicity in the clinic, to induce a CD4+ T cells response. Keyhole limpet hemocyanin (KLH) and cytomegalovirus pp65 protein (CMV) were included as neo-antigen and recall antigen positive controls, respectively. By analyzing 26 healthy donors having HLA-DRB1 alleles matching the European population, we calculated the frequency of responding donors, the magnitude of the response, and the frequency of BP-specific T cells, as measured by 3[H]-thymidine incorporation and ELISpot IL-2 secretion. KLH and CMV demonstrated a strong T cell response in all the donors analyzed. The frequency of responding donors to the BPs was 4% for infliximab, 8% for adalimumab, 19% for rituximab and 27% for natalizumab, which is compared to and discussed with their respective observed clinical immunogenicity. This study further complements predictive immunogenicity testing by quantifying the in vitro CD4+ T cell responses to different BPs. Even though the data generated using this modified method does not directly translate to the clinical situation, a high sensitivity and immunogenic potential of most BPs is demonstrated.
Highlights
Biopharmaceuticals (BPs), such as monoclonal antibodies are widely used for the treatment of autoimmune disease, and cancer
To develop a sensitive and high throughput T cell assay we combined and optimized the current assays used by contract research organizations (CRO), including peripheral blood mononuclear cell (PBMC) and dendritic cell (DC):T cell assays from Antitope Ltd (EpiScreenTM), Lonza (EpiBaseTM), ProImmuneLtd (REVEAL1), EpiVax Inc. and ImmunXperts
Comparing the optimized T cell:PBMC assay with a standard CD8+ T cell-depleted PBMC assay, an increased response to Keyhole limpet hemocyanin (KLH), cytomegalovirus pp65 protein (CMV) and Protective antigen (PA) for both IL-2 secretion and proliferation was observed (Fig 2)
Summary
Biopharmaceuticals (BPs), such as monoclonal antibodies (mAbs) are widely used for the treatment of autoimmune disease, and cancer. A major concern regarding treatment with therapeutic proteins is the risk of provoking an unwanted immune response, such as the development of anti-drug antibodies (ADAs). Many factors contribute to the immunogenicity of BPs, including product-, disease-, treatment- and patient-related factors [3]. Several methods to evaluate T cell responses have been published and applied during drug development to reduce the risk for immunogenicity in the clinic. These include peripheral blood mononuclear cell (PBMC)-based assays [14], dendritic cell (DC):T cell assays [15, 16] and more complex assays where naïve T cells are amplified polyclonally [17] or antigen- [18, 19]
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