Raymond G. Perelman Center for Cellular and Molecular Therapeutics, the Children's Hospital of Philadelphia, Philadelphia, PA, USA.
Division of Hematology, the Children's Hospital of Philadelphia, Philadelphia, PA, USA.
Nat Commun. 2024 Aug 21;15(1):7193. doi: 10.1038/s41467-024-51296-8.
Durable factor VIII expression that normalizes hemostasis is an unrealized goal of hemophilia A adeno-associated virus-mediated gene therapy. Trials with initially normal factor VIII activity observed unexplained year-over-year declines in expression while others reported low-level, stable expression inadequate to restore normal hemostasis. Here we demonstrate that male mice recapitulate expression-level-dependent loss of factor VIII levels due to declines in vector copy number. We show that an enhanced function factor VIII variant (factor VIII-R336Q/R562Q), resistant to activated protein C-mediated inactivation, normalizes hemostasis at below-normal expression without evidence of prothrombotic risk in male hemophilia A mice. These data support that factor VIII-R336Q/R562Q may restore normal factor VIII function at low levels of expression to permit durability using low vector doses to minimize dose-dependent adeno-associated virus toxicities. This work informs the mechanism of factor VIII durability after gene transfer and supports that factor VIII-R336Q/R562Q may safely overcome current hemophilia A gene therapy limitations.
持久的 VIII 因子表达，使其正常止血，是血友病 A 腺相关病毒介导的基因治疗的一个未实现的目标。最初观察到因子 VIII 活性正常的试验中，发现表达水平每年都在不明原因地下降，而其他试验则报告了低水平、不稳定的表达，不足以恢复正常止血。在这里，我们证明雄性小鼠由于载体拷贝数下降而重现了VIII 因子水平的表达水平依赖性丧失。我们表明，一种增强功能的 VIII 因子变体（VIII-R336Q/R562Q），对激活蛋白 C 介导的失活有抗性，可在低于正常表达水平下使止血正常化，而在男性血友病 A 小鼠中没有血栓形成风险的证据。这些数据支持 VIII-R336Q/R562Q 可能以低表达水平恢复正常 VIII 因子功能，从而允许使用低载体剂量实现持久性，以最小化剂量依赖性腺相关病毒毒性。这项工作阐明了基因转移后 VIII 因子持久性的机制，并表明 VIII-R336Q/R562Q 可能安全地克服当前血友病 A 基因治疗的限制。
