Heterogeneous and evolving epitope landscape of clinical anti-drug antibodies against multidomain biotherapeutic: a case study of TAK-186

The emergence of anti-drug antibodies (ADAs) poses a major obstacle in the clinical development of therapeutic proteins (TPs) such as monoclonal antibodies and their derivatives. While standard multitiered ADA assays and neutralizing antibody assays offer valuable insights into the humoral immunogenicity risks of TPs, they are not sufficient to provide in-depth knowledge such as ADA epitope specificities. For complex multidomain biotherapeutics (MDBs), ADAs targeting individual domains can elicit distinct pharmacological effects. Therefore, it is crucial to implement straightforward and reliable methodologies to deconvolute ADA epitope profiles of MDBs. Herein, we report a case study using domain specificity analysis, linear peptide scanning and bioinformatic B cell epitope prediction to unveil the clinical ADA epitope landscape of TAK-186, a multidomain T cell engager that has been discontinued from clinical development. By applying this workflow, we observed strong domain specificity variability among patient samples. Furthermore, the data showed that many patients demonstrated evolved ADA epitope specificities throughout the course of the treatment. Several potential linear epitopes were identified subsequently through experimental and computational approaches. Overall, we presented in this study a practical strategy to elucidate and potentially mitigate the immunogenicity liabilities of complex biotherapeutics.