Integrated Drug Discovery, Sanofi, Waltham, MA.
Department of Biochemistry and Molecular Biology, University of Chicago, Chicago, IL; and.
Blood. 2021 May 27;137(21):2970-2980. doi: 10.1182/blood.2020009197.
Interaction of factor VIII (FVIII) with von Willebrand factor (VWF) is mediated by the VWF D'D3 domains and thrombin-mediated release is essential for hemostasis after vascular injury. VWF-D'D3 mutations resulting in loss of FVIII binding are the underlying cause of von Willebrand disease (VWD) type 2N. Furthermore, the FVIII-VWF interaction has significant implications for the development of therapeutics for bleeding disorders, particularly hemophilia A, in which endogenous VWF clearance imposes a half-life ceiling on replacement FVIII therapy. To understand the structural basis of FVIII engagement by VWF, we solved the structure of BIVV001 by cryo-electron microscopy to 2.9 Å resolution. BIVV001 is a bioengineered clinical-stage FVIII molecule for the treatment of hemophilia A. In BIVV001, VWF-D'D3 is covalently linked to an Fc domain of a B domain-deleted recombinant FVIII (rFVIII) Fc fusion protein, resulting in a stabilized rFVIII/VWF-D'D3 complex. Our rFVIII/VWF structure resolves BIVV001 architecture and provides a detailed spatial understanding of previous biochemical and clinical observations related to FVIII-VWF engagement. Notably, the FVIII acidic a3 peptide region (FVIII-a3), established as a critical determinant of FVIII/VWF complex formation, inserts into a basic groove formed at the VWF-D'/rFVIII interface. Our structure shows direct interaction of sulfated Y1680 in FVIII-a3 and VWF-R816 that, when mutated, leads to severe hemophilia A or VWD type 2N, respectively. These results provide insight on this key coagulation complex, explain the structural basis of many hemophilia A and VWD type 2N mutations, and inform studies to further elucidate how VWF dissociates rapidly from FVIII upon activation.
FVIII 与 VWF 的相互作用由 VWF D'D3 结构域介导，而凝血酶介导的释放对于血管损伤后的止血至关重要。导致 FVIII 结合丧失的 VWF-D'D3 突变是 2N 型血管性血友病（VWD）的根本原因。此外，FVIII-VWF 相互作用对出血性疾病治疗药物的开发具有重要意义，特别是在血友病 A 中，内源性 VWF 清除对替代 FVIII 治疗施加半衰期上限。为了了解 VWF 与 FVIII 结合的结构基础，我们通过冷冻电镜解析了 BIVV001 的结构，分辨率为 2.9 Å。BIVV001 是一种用于治疗血友病 A 的生物工程临床阶段 FVIII 分子。在 BIVV001 中，VWF-D'D3 通过共价键与 B 结构域缺失的重组 FVIII（rFVIII）Fc 融合蛋白的 Fc 结构域连接，导致 rFVIII/VWF-D'D3 复合物稳定化。我们的 rFVIII/VWF 结构解析了 BIVV001 的结构，并提供了对先前与 FVIII-VWF 结合相关的生化和临床观察的详细空间理解。值得注意的是，FVIII 酸性 a3 肽区（FVIII-a3）被确定为 FVIII/VWF 复合物形成的关键决定因素，插入到 VWF-D'/rFVIII 界面形成的碱性凹槽中。我们的结构显示了 FVIII-a3 中带负电荷的 Y1680 与 VWF-R816 的直接相互作用，当突变时，分别导致严重的血友病 A 或 2N 型 VWD。这些结果提供了对该关键凝血复合物的深入了解，解释了许多血友病 A 和 2N 型 VWD 突变的结构基础，并为进一步阐明 VWF 在激活后如何快速从 FVIII 解离的研究提供了信息。
