Human Health Therapeutics Research Centre, National Research Council Canada, 100 Sussex Drive, Ottawa, ON, K1A 0R6, Canada.
Department of Biochemistry, Microbiology and Immunology, University of Ottawa, 451 Smyth Road, Ottawa, ON, K1H 8M5, Canada.
Immunol Cell Biol. 2019 Jul;97(6):526-537. doi: 10.1111/imcb.12238. Epub 2019 Mar 3.
Human IgG1 and IgG3 antibodies (Abs) can mediate Ab-dependent cellular cytotoxicity (ADCC), and engineering of the Ab Fc (point mutation; defucosylation) has been shown to affect ADCC by modulating affinity for FcRγIIIa. In the absence of a C 1 domain, many camelid heavy-chain Abs (HCAbs) naturally bear very long and flexible hinge regions connecting their V H and C 2 domains. To better understand the influence of hinge length and structure on HCAb ADCC, we produced a series of hinge-engineered epidermal growth factor receptor (EGFR)-specific chimeric camelid V H-human Fc Abs and characterized their affinities for recombinant EGFR and FcRγIIIa, their binding to EGFR-positive tumor cells, and their ability to elicit ADCC. In the case of one chimeric HCAb (EG2-hFc), we found that variants bearing longer hinges (IgG3 or camelid hinge regions) showed dramatically improved ADCC in comparison with a variant bearing the human IgG1 hinge, in similar fashion to a variant with reduced C 2 fucosylation. Conversely, an EG2-hFc variant bearing a truncated human IgG1 upper hinge region failed to elicit ADCC. However, there was no consistent association between hinge length and ADCC for four similarly engineered chimeric HCAbs directed against distinct EGFR epitopes. These findings demonstrate that the ADCC of some HCAbs can be modulated simply by varying the length of the Ab hinge. Although this effect appears to be heavily epitope-dependent, this strategy may be useful to consider during the design of V H-based therapeutic Abs for cancer.
人 IgG1 和 IgG3 抗体（Abs）可介导抗体依赖的细胞细胞毒性（ADCC），并且已经表明通过调节与 FcRγIIIa 的亲和力来修饰 Ab Fc（点突变；去岩藻糖基化）可以影响 ADCC。在没有 C1 结构域的情况下，许多骆驼科重链 Abs（HCAbs）天然地具有非常长而灵活的铰链区，将它们的 V H 和 C2 结构域连接起来。为了更好地了解铰链长度和结构对 HCAb ADCC 的影响，我们生产了一系列铰链工程化的表皮生长因子受体（EGFR）特异性嵌合骆驼科 V H-人 Fc Abs，并对它们与重组 EGFR 和 FcRγIIIa 的亲和力、与 EGFR 阳性肿瘤细胞的结合能力以及引发 ADCC 的能力进行了表征。在一种嵌合 HCAb（EG2-hFc）的情况下，我们发现，与具有人 IgG1 铰链的变体相比，具有较长铰链（IgG3 或骆驼科铰链区）的变体显示出明显改善的 ADCC，与具有降低的 C2 岩藻糖基化的变体类似。相反，具有截断的人 IgG1 上铰链区的 EG2-hFc 变体未能引发 ADCC。然而，针对不同 EGFR 表位的四个类似工程化的嵌合 HCAbs 的铰链长度与 ADCC 之间没有一致的关联。这些发现表明，通过改变 Ab 铰链的长度，可以调节一些 HCAbs 的 ADCC。尽管这种效应似乎严重依赖于表位，但在设计基于 V H 的治疗性 Abs 用于癌症时，这种策略可能值得考虑。
