Influence of N-glycosylation in the A and C domains on the immunogenicity of factor VIII.
Influence of N-glycosylation in the A and C domains on the immunogenicity of factor VIII.
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DOI：
10.1182/bloodadvances.2021005758
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发表时间：
2022-07-26
期刊：
影响因子：
7.5
通讯作者：
Miao, Carol H.
中科院分区：
文献类型：
--
作者：
Vander Kooi, Amber;Wang, Shuaishuai;Fan, Meng-Ni;Chen, Alex;Zhang, Junping;Chen, Chun-Yu;Cai, Xiaohe;Konkle, Barbara A.;Xiao, Weidong;Li, Lei;Miao, Carol H.
消除FVIII A和C结构域位点的N-糖基化对其免疫原性的影响不同，但对特异性活性无影响。在位点N2118处的有效甘露糖基化糖肽表位被鉴定并表征为活化T细胞。血友病A治疗中最显著的并发症是形成针对因子VIII（FVIII）蛋白的抑制剂。聚糖和聚糖结合蛋白是正常运作的免疫系统的核心。本研究的重点是FVIII的糖基化是否在诱导和调节抗FVIII免疫应答中起重要作用。我们研究了4个N-糖基化位点，包括A1结构域中的N41和N239，A3结构域中的N1810和C1结构域中的N2118，在调节其免疫原性中的潜在作用。血浆源性FVIII的糖组学分析显示，位点N41、N239和N1810主要含有唾液酸化的复合糖型，而位点N2118主要含有高甘露糖聚糖。引入了一种将天冬酰胺（N）取代为谷氨酰胺（Q）的错义变体，以消除这些位点上的糖基化。在基因转移编码B结构域缺失FVIII（BDD-FVIII）和这4种FVIII变体的质粒后，发现血浆中FVIII的比活性在所有治疗组中保持相似。与BDD-FVIII质粒给药小鼠相比，在N41 Q、N239 Q和N1810 Q FVIII变体质粒给药小鼠中观察到免疫应答略微增加或相当，在N2118 Q FVIII质粒给药小鼠中观察到免疫应答显著降低。在AAV介导的基因转移实验中也证明了N2118 Q FVIII变体对抑制剂应答的降低。此外，N2118糖基化位点周围的特异性糖肽表位被鉴定并表征为在FVIII特异性增殖测定中激活T细胞。这些结果表明，FVIII的N-糖基化对其免疫原性具有显著影响。
Elimination of N-glycosylation at sites in the A and C domains of FVIII differentially affects its immunogenicity but not specific activity. A potent mannosylated glycopeptide epitope at site N2118 was identified and characterized to activate T cell's. The most significant complication in hemophilia A treatment is the formation of inhibitors against factor VIII (FVIII) protein. Glycans and glycan-binding proteins are central to a properly functioning immune system. This study focuses on whether glycosylation of FVIII plays an important role in induction and regulation of anti-FVIII immune responses. We investigated the potential roles of 4 N-glycosylation sites, including N41 and N239 in the A1 domain, N1810 in the A3 domain, and N2118 in the C1 domain of FVIII, in moderating its immunogenicity. Glycomics analysis of plasma-derived FVIII revealed that sites N41, N239, and N1810 contain mostly sialylated complex glycoforms, while high mannose glycans dominate at site N2118. A missense variant that substitutes asparagine (N) to glutamine (Q) was introduced to eliminate glycosylation on each of these sites. Following gene transfer of plasmids encoding B domain deleted FVIII (BDD-FVIII) and each of these 4 FVIII variants, it was found that specific activity of FVIII in plasma remained similar among all treatment groups. Slightly increased or comparable immune responses in N41Q, N239Q, and N1810Q FVIII variant plasmid-treated mice and significantly decreased immune responses in N2118Q FVIII plasmid-treated mice were observed when compared with BDD-FVIII plasmid-treated mice. The reduction of inhibitor response by N2118Q FVIII variant was also demonstrated in AAV-mediated gene transfer experiments. Furthermore, a specific glycopeptide epitope surrounding the N2118 glycosylation site was identified and characterized to activate T cells in an FVIII-specific proliferation assay. These results indicate that N-glycosylation of FVIII can have significant impact on its immunogenicity.
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影响因子：
7.5
作者：
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通讯作者：
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