Reduction of the antigenicity of factor VIII toward complex inhibitory antibody plasmas using multiply-substituted hybrid human/porcine factor VIII molecules
Reduction of the antigenicity of factor VIII toward complex inhibitory antibody plasmas using multiply-substituted hybrid human/porcine factor VIII molecules
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DOI：
10.1182/blood.v95.2.564
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发表时间：
2000-01-15
期刊：
影响因子：
20.3
通讯作者：
Lollar, P
中科院分区：
文献类型：
--
作者：
Barrow, RT;Healey, JF;Lollar, P
因子VIII（fVIII）作为重链/轻链（A1-A2-B/apA 3-C1-C2）异二聚体循环。41个残基的轻链活化肽ap在凝血酶或因子Xa的蛋白水解活化期间从FVIII裂解。我们构建了7个活性重组杂合无B结构域的人/猪FVIII分子，其在A2、ap-A3和C2结构域内含有猪序列替换的组合。23种高滴度抑制性抗体在人FVIII和杂交体之间的交叉反应性与猪的置换程度呈负相关。在所有血浆中，所有3个区域的置换产生的交叉反应性与猪FVIII的交叉反应性没有显著差异。为了区分抑制剂与ap区和A3结构域的结合，我们构建了2个另外的杂交体，其含有猪A2和C2结构域取代以及猪A3或猪ap取代。在几种具有抗ap-A3区域活性的血浆中，猪ap片段的抗原性低于人ap片段。这表明，除了在先前研究中鉴定的A2、A3和C2结构域表位之外，一些抑制剂血浆还含有针对fVIII ap片段的抗体。人fVIII的A2、A3、C2和ap区内的猪序列的取代是必要的并且足以实现相对于猪fVIII的抗原性相对于大多数抑制性抗体血浆的最大降低。
Factor VIII (fVIII) circulates as a heavy chain/light chain (A1-A2-B/apA3-C1-C2) heterodimer. The 41-residue light chain activation peptide, ap, is cleaved from fVIII during proteolytic activation by thrombin or factor Xa, We constructed 7 active recombinant hybrid B-domainless human/porcine fVIII molecules that contained combinations of porcine sequence replacements within the A2, ap-A3, and C2 domains. The cross-reactivity of 23 high-titer inhibitory antibodies between human fVIII and the hybrids was inversely related to the degree of porcine substitution, In all plasmas, the substitution of all 3 regions yielded cross-reactivities that were not significantly different from those of porcine fVIII. To differentiate between inhibitor binding to the ap region and the A3 domain, we constructed 2 additional hybrids that contained porcine A2 and C2 domain substitutions and either porcine A3 or porcine ap substitutions. The porcine ap segment was less antigenic than the human ap segment in several plasmas that had activity against the ap-A3 region. This indicates that some inhibitor plasmas contain antibodies directed against the fVIII ap segment in addition to A2, A3, and C2 domain epitopes identified In previous studies. Substitution of porcine sequences within the A2, A3, C2, and ap regions of human fVIII is necessary and sufficient to achieve a maximal reduction in antigenicity relative to porcine fVIII with respect to most inhibitory antibody plasmas.