Shi Q, Schroeder J A, Kuether E L, Montgomery R R
Department of Pediatrics, Medical College of Wisconsin, Milwaukee, WI, USA.
Blood Research Institute, Blood Center of Wisconsin, Milwaukee, WI, USA.
J Thromb Haemost. 2015 Jul;13(7):1301-9. doi: 10.1111/jth.13001. Epub 2015 Jun 11.
Our previous studies have demonstrated that targeting FVIII expression to platelets results in FVIII storage together with von Willebrand factor (VWF) in platelet α-granules and that platelet-derived FVIII (2bF8) corrects the murine hemophilia A phenotype even in the presence of high-titer anti-FVIII inhibitory antibodies (inhibitors).
To explore how VWF has an impact on platelet gene therapy for hemophilia A with inhibitors.
2bF8 transgenic mice in the FVIII(-/-) background (2bF8(tg+/-) F8(-/-) ) with varying VWF phenotypes were used in this study. Animals were analyzed by VWF ELISA, FVIII activity assay, Bethesda assay and tail clip survival test.
Only 18% of 2bF8(tg+/-) F8(-/-) VWF(-/-) animals, in which VWF was deficient, survived the tail clip challenge with inhibitor titers of 3-8000 BU mL(-1) . In contrast, 82% of 2bF8(tg+/-) F8(-/-) VWF(+/+) mice, which had normal VWF levels, survived tail clipping with inhibitor titers of 10-50,000 BU mL(-1) . All 2bF8(tg+/-) F8(-/-) VWF(-/-) mice without inhibitors survived tail clipping and no VWF(-/-) F8(-/-) mice survived this challenge. Because VWF is synthesized by endothelial cells and megakaryocytes and is distributed in both plasma and platelets in peripheral blood, we further investigated the effect of each compartment of VWF on platelet-FVIII gene therapy for hemophilia A with inhibitors. In the presence of inhibitors, 42% of animals survived tail clipping in the group with plasma-VWF and 50% survived in the platelet-VWF group.
VWF is essential for platelet gene therapy for hemophilia A with inhibitors. Both platelet-VWF and plasma-VWF are required for optimal platelet-derived FVIII gene therapy for hemophilia A in the presence of inhibitors.
我们之前的研究表明，将FVIII表达靶向至血小板会导致FVIII与血管性血友病因子（VWF）一起储存在血小板α颗粒中，并且即使存在高滴度的抗FVIII抑制性抗体（抑制剂），血小板衍生的FVIII（2bF8）也能纠正小鼠血友病A的表型。
探讨VWF如何影响血友病A伴抑制剂的血小板基因治疗。
本研究使用了具有不同VWF表型的FVIII基因敲除背景下的2bF8转基因小鼠（2bF8(tg+/-)F8(-/-)）。通过VWF ELISA、FVIII活性测定、贝塞斯达测定和剪尾存活试验对动物进行分析。
在2bF8(tg+/-)F8(-/-)VWF(-/-)动物中，即VWF缺乏的动物，只有18%在抑制剂滴度为3 - 8000 BU/mL(-1)的剪尾挑战中存活。相比之下，2bF8(tg+/-)F8(-/-)VWF(+/+)小鼠，其VWF水平正常，82%在抑制剂滴度为10 - 50,000 BU/mL(-1)的剪尾试验中存活。所有无抑制剂的2bF8(tg+/-)F8(-/-)VWF(-/-)小鼠在剪尾试验中存活，而无VWF(-/-)F8(-/-)小鼠在此挑战中无一存活。由于VWF由内皮细胞和巨核细胞合成，并分布于外周血的血浆和血小板中，我们进一步研究了VWF的每个组分对血友病A伴抑制剂的血小板 - FVIII基因治疗的影响。在存在抑制剂的情况下，血浆VWF组中42%的动物在剪尾试验中存活，血小板VWF组中50%存活。
VWF对血友病A伴抑制剂的血小板基因治疗至关重要。在存在抑制剂的情况下，最佳的血友病A血小板衍生FVIII基因治疗需要血小板VWF和血浆VWF两者。
