Life Sciences Division, Human Health Therapeutics Research Centre, National Research Council Canada, Ottawa, Canada.
Department of Biochemistry, Microbiology and Immunology, Faculty of Medicine, University of Ottawa, Canada.
FEBS J. 2022 Jul;289(14):4304-4327. doi: 10.1111/febs.15809. Epub 2021 Mar 25.
Single-domain antibodies (sdAbs), the autonomous variable domains of camelid and shark heavy-chain antibodies, have many desirable properties as components of biologic drugs. However, their sequences may increase the risk of immunogenicity and antidrug antibody (ADA) development in humans, and thus, sdAbs are routinely humanized during development. Here, we review and summarize the available evidence regarding the factors governing immunogenicity of sdAbs and our current state of knowledge of strategies to mitigate immunogenicity risks by humanization. While several sdAb properties, including high homology of camelid V Hs with human IGHV3 gene products, favor low immunogenicity in humans, epitopes absent in the human repertoire including the exposed V :V interface may be intrinsically immunogenic. While most clinical trials have demonstrated minimal sdAb immunogenicity, two notable exceptions (the tetrameric DR5-specific V H TAS266 and the TNFR1-specific V GSK1995057) illustrate that special caution must be taken in identifying preexisting ADAs against highly potent sdAbs. Nonhuman sequence alone does not adequately explain sdAb immunogenicity, as some camelid V Hs are nonimmunogenic while some fully human V s elicit ADAs. The presence of preexisting ADAs directed against the exposed C-termini of some sdAbs in a significant proportion of individuals awaits a molecular explanation. Whether sdAb humanization reduces or promotes immunogenicity remains unclear: reduction of nonhuman sequence content at the expense of introducing low-level aggregation in humanized variants may be counterproductive. Further work will establish thresholds for V H and V humanization to maximize human sequence content while avoiding loss of binding affinity and/or immunogenicity resulting from aggregation or decreased stability.
单域抗体（sdAb）是骆驼科和鲨鱼重链抗体的独立可变结构域，作为生物药物的组成部分具有许多理想的特性。然而，它们的序列可能会增加在人类中产生免疫原性和抗药物抗体（ADA）的风险，因此，sdAb 在开发过程中通常会被人源化。在这里，我们回顾和总结了关于 sdAb 免疫原性的决定因素的现有证据，以及我们目前对通过人源化减轻免疫原性风险的策略的了解。虽然 sdAb 的一些特性，包括骆驼科 V Hs 与人 IGHV3 基因产物的高度同源性，有利于在人类中产生低免疫原性，但在人类中不存在的表位，包括暴露的 V : V 界面，可能具有内在的免疫原性。虽然大多数临床试验表明 sdAb 的免疫原性很低，但有两个值得注意的例外（四聚体 DR5 特异性 V H TAS266 和 TNFR1 特异性 V GSK1995057）表明，在鉴定针对高活性 sdAb 的预先存在的 ADA 时必须特别小心。非人类序列本身并不能充分解释 sdAb 的免疫原性，因为一些骆驼科 V Hs 是非免疫原性的，而一些完全人源化的 V s 则会引发 ADA。在相当一部分个体中，一些 sdAb 的暴露 C 末端存在预先存在的针对 ADA 的情况，这有待分子解释。sdAb 人源化是否会降低或促进免疫原性仍不清楚：以牺牲人源化变体中低水平聚集为代价，降低非人类序列含量可能适得其反。进一步的研究将确定 V H 和 V 人源化的阈值，以最大限度地提高人源序列含量，同时避免由于聚集或稳定性降低而导致的结合亲和力和/或免疫原性丧失。
