Liver gene therapy with adeno-associated viral (AAV) vectors is under clinical investigation for haemophilia A (HemA), the most common inherited X-linked bleeding disorder. Major limitations are the large size of the F8 transgene, which makes packaging in a single AAV vector a challenge, as well as the development of circulating anti-F8 antibodies which neutralise F8 activity. Taking advantage of split-intein-mediated protein trans-splicing, we divided the coding sequence of the large and highly secreted F8-N6 variant in two separate AAV-intein vectors whose co-administration to HemA mice results in the expression of therapeutic levels of F8 over time. This occurred without eliciting circulating anti-F8 antibodies unlike animals treated with the single oversized AAV-F8 vector under clinical development. Therefore, liver gene therapy with AAV-F8-N6 intein should be considered as a potential therapeutic strategy for HemA.

中文翻译：

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内含肽介导的 F8 反式剪接的肝基因治疗可纠正小鼠血友病 A

腺相关病毒 (AAV) 载体的肝基因治疗正在临床研究 A 型血友病 (HemA)，这是最常见的遗传性 X 连锁出血性疾病。主要限制是F8转基因的大尺寸，这使得在单个 AAV 载体中包装成为一项挑战，以及开发中和 F8 活性的循环抗 F8 抗体。利用分裂内含肽介导的蛋白质反式剪接，我们将大且高度分泌的 F8-N6 变体的编码序列划分为两个单独的 AAV-内含肽载体，其共同给药于 HemA 小鼠导致治疗水平的表达F8 随着时间的推移。与用单个超大 AAV- F8治疗的动物不同，这在没有引发循环抗 F8 抗体的情况下发生临床开发中的载体。因此，用 AAV- F8 -N6 内含肽进行肝脏基因治疗应被视为 HemA 的潜在治疗策略。