Immunogenicity Risk Assessment for an Engineered Human Cytokine Analogue Expressed in Different Cell Substrates

The purpose of this article is to illustrate how performance of an immunogenicity risk assessment at the earliest stage of product development can be instructive for critical early decision-making such as choice of host system for expression of a recombinant therapeutic protein and determining the extent of analytical characterization and control of heterogeneity in co- and post-translational modifications. Application of a risk-based approach for a hypothetical recombinant DNA analogue of a human endogenous cytokine with immunomodulatory functions is described. The manner in which both intrinsic and extrinsic factors could interact to influence the relative scale of risk associated with expression in alternative hosts, namely Chinese hamster ovary (CHO) cells, Pichia pastoris, Escherichia coli, or Nicotinia tabacum is considered in relation to the development of the investigational product to treat an autoimmune condition. The article discusses how particular product-related variants (primary amino acid sequence modifications and post-translational glycosylation or other modifications) and process-derived impurities (host cell proteins, endotoxins, beta-glucans) associated with the different expression systems might influence the impact of immunogenicity on overall clinical benefit versus risk for a therapeutic protein candidate that has intrinsic MHC Class II binding potential. The implications of the choice of expression system for relative risk are discussed in relation to specific actions for evaluation and measures for risk mitigation, including use of in silico and in vitro methods to understand intrinsic immunogenic potential relative to incremental risk associated with non-human glycan and protein impurities. Finally, practical guidance on presentation of this information in regulatory submissions to support clinical development is provided.