Modulation of Immune Reaction in Hydrodynamic Gene Therapy for Hemophilia A
Modulation of Immune Reaction in Hydrodynamic Gene Therapy for Hemophilia A
复制标题
A 型血友病水动力基因治疗中免疫反应的调节
DOI：
10.1089/hum.2021.145
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发表时间：
2022
影响因子：
4.2
通讯作者：
Xiao-Bing Zhang
中科院分区：
文献类型：
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作者：
Mei Zhao;Yi-Dan Sun;Meng-Di Yin;Juan-Juan Zhao;Si-Ang Li;Guo-Hua Li;Feng Zhang;Jing Xu;Fei-Ying Meng;Beldon Zhang;Xin-Yu Sun;Jian-Ping Zhang;Tao Cheng;Xiao-Bing Zhang
血友病A（Hemophilia A，HA）是一种单基因疾病，其特征是由于F8突变导致的血浆凝血因子8（Plasma clotting factor 8，F8）缺陷。我们一直在尝试使用CRISPR-Cas9基因编辑策略永久治愈HA。在这项研究中，我们诱导BDDF 8（B-domain-deleted F8）基因到HA小鼠的白蛋白基因座的靶向整合，通过流体动力学尾静脉注射编辑质粒载体。治疗后一周，在所有治疗的HA小鼠中观察到正常血清水平的70%至280%范围内的高F8活性，但在3-5周后降至背景水平。我们发现，针对F8的体液免疫反应是F8活性降低的主要原因。我们假设，流体动力注射诱导的肝损伤引发了大量的炎性细胞因子的释放。然而，共注射表达十几种免疫调节因子的质粒未能有效地抑制免疫反应和稳定F8活性。携带miR-142- 3 p靶序列的spCas 9质粒减轻了细胞免疫应答，但不能提供治疗功效。引人注目的是，免疫抑制剂环磷酰胺几乎消除了免疫反应，导致长达一年的稳定F8水平。我们的研究结果应该在使用流体动力学基因递送方法在小鼠模型中开发疗法方面具有重要意义，突出了调节由肝损伤引发的先天免疫反应的必要性。
Hemophilia A (HA) is a monogenic disease characterized by plasma clotting factor 8 (F8) deficiency due to F8 mutation. We have been attempting to cure HA permanently using a CRISPR-Cas9 gene-editing strategy. In this study, we induced targeted integration of BDDF8 (B-domain-deleted F8) gene into the albumin locus of HA mice by hydrodynamic tail vein injection of editing plasmid vectors. One week after treatment, a high F8 activity ranging from 70% to 280% of normal serum levels was observed in all treated HA mice but dropped to background levels 3-5 weeks later. We found that the humoral immune reaction targeting F8 is the predominant cause of the decreased F8 activity. We hypothesized that hydrodynamic injection-induced liver damage triggered the release of large quantities of inflammatory cytokines. However, coinjection of plasmids expressing a dozen immunomodulatory factors failed to curtail the immune reaction and stabilize F8 activity effectively. The spCas9 plasmid carrying a miR-142-3p target sequence alleviated the cellular immune response but could not deliver therapeutic efficacy. Strikingly, immunosuppressant cyclophosphamide virtually abolished the immune response, leading to a year-long stable F8 level. Our findings should have important implications in developing therapies in mouse models using the hydrodynamic gene delivery approach, highlighting the necessity of modulating the innate immune response triggered by liver damage.