Palaschak Brett, Marsic Damien, Herzog Roland W, Zolotukhin Sergei, Markusic David M
Department of Pediatrics, University of Florida, Gainesville, FL 32610, USA.
Mol Ther Methods Clin Dev. 2017 Apr 19;5:142-152. doi: 10.1016/j.omtm.2017.04.004. eCollection 2017 Jun 16.
Multiple independent adeno-associated virus (AAV) gene therapy clinical trials for hemophilia B, utilizing different AAV serotypes, have reported a vector dose-dependent loss of circulating factor IX (FIX) protein associated with capsid-specific CD8 T cell (Cap-CD8) elimination of transduced hepatocytes. Hemophilia B patients who develop transient transaminitis and loss of FIX protein may be stabilized with the immune-suppressive (IS) drug prednisolone, but do not all recover lost FIX expression, whereas some patients fail to respond to IS. We developed the first animal model demonstrating Cap-CD8 infiltration and elimination of AAV-transduced hepatocytes of immune-deficient mice. Here, we extend this model to an immune-competent host where Cap-CD8 transfer to AAV2--treated mice significantly reduced circulating and hepatocyte FIX expression. Further, we studied two high-expressing liver tropic AAV2 variants, AAV2-LiA and AAV2-LiC, obtained from a rationally designed capsid library. Unlike AAV2, Cap-CD8 did not initially reduce circulating FIX levels for either variant. However, FIX levels were significantly reduced in AAV2-LiC--treated, but not AAV2-LiA--treated, mice at the study endpoint. Going forward, the immune-competent model may provide an opportunity to induce immunological memory directed against a surrogate AAV capsid antigen and study recall responses following AAV gene transfer.
多项针对B型血友病的独立腺相关病毒（AAV）基因治疗临床试验，使用了不同的AAV血清型，报告了与衣壳特异性CD8 T细胞（Cap-CD8）清除转导的肝细胞相关的循环凝血因子IX（FIX）蛋白的载体剂量依赖性损失。出现短暂性转氨酶升高和FIX蛋白损失的B型血友病患者，可能会通过免疫抑制（IS）药物泼尼松龙得以稳定，但并非所有患者都能恢复已损失的FIX表达，而一些患者对IS无反应。我们开发了首个动物模型，证明免疫缺陷小鼠的Cap-CD8浸润并清除了AAV转导的肝细胞。在此，我们将该模型扩展至免疫健全宿主，其中将Cap-CD8转移至接受AAV2治疗的小鼠显著降低了循环和肝细胞中的FIX表达。此外，我们研究了从合理设计的衣壳文库中获得的两种高表达肝脏嗜性AAV2变体，即AAV2-LiA和AAV2-LiC。与AAV2不同，Cap-CD8最初并未降低这两种变体的循环FIX水平。然而，在研究终点时，接受AAV2-LiC治疗的小鼠而非接受AAV2-LiA治疗的小鼠，FIX水平显著降低。展望未来，免疫健全模型可能提供一个机会，诱导针对替代AAV衣壳抗原的免疫记忆，并研究AAV基因转移后的回忆反应。
