血小板靶向因子 VIII 基因治疗诱导的血友病 A 小鼠免疫耐受与 CD4 T 细胞有关。
Immune tolerance induced by platelet-targeted factor VIII gene therapy in hemophilia A mice is CD4 T cell mediated.
机构信息
Blood Research Institute, Blood Center of Wisconsin, Milwaukee, WI, USA.
Fujian Institute of Hematology, Fujian Medical University Union Hospital, Fuzhou, China.
出版信息
J Thromb Haemost. 2017 Oct;15(10):1994-2004. doi: 10.1111/jth.13800. Epub 2017 Sep 11.
UNLABELLED
Essentials The immune response is a significant concern in gene therapy. Platelet-targeted gene therapy can restore hemostasis and induce immune tolerance. CD4 T cell compartment is tolerized after platelet gene therapy. Preconditioning regimen affects immune tolerance induction in platelet gene therapy.
SUMMARY
Background Immune responses are a major concern in gene therapy. Our previous studies demonstrated that platelet-targeted factor VIII (FVIII) (2bF8) gene therapy together with in vivo drug selection of transduced cells can rescue the bleeding diathesis and induce immune tolerance in FVIII mice. Objective To investigate whether non-selectable 2bF8 lentiviral vector (LV) for the induction of platelet-FVIII expression is sufficient to induce immune tolerance and how immune tolerance is induced after 2bF8LV gene therapy. Methods Platelet-FVIII expression was introduced by 2bF8LV transduction and transplantation. FVIII assays and tail bleeding tests were used to confirm the success of platelet gene therapy. Animals were challenged with rhF8 to explore if immune tolerance was induced after gene therapy. Treg cell analysis, T-cell proliferation assay and memory B-cell-mediated ELISPOT assay were used to investigate the potential mechanisms of immune tolerance. Results We showed that platelet-FVIII expression was sustained and the bleeding diathesis was restored in FVIII mice after 2bF8LV gene therapy. None of the transduced recipients developed anti-FVIII inhibitory antibodies in the groups preconditioned with 660 cGy irradiation or busulfan plus ATG treatment even after rhF8 challenge. Treg cells significantly increased in 2bF8LV-transduced recipients and the immune tolerance developed was transferable. CD4 T cells from treated animals failed to proliferate in response to rhF8 re-stimulation, but memory B cells could differentiate into antibody secreting cells in 2bF8LV-transduced recipients. Conclusion 2bF8LV gene transfer without in vivo selection of manipulated cells can introduce immune tolerance in hemophilia A mice and this immune tolerance is CD4 T cell mediated.
目的
免疫反应是基因治疗中的一个重要关注点。血小板靶向基因治疗可以恢复止血功能并诱导免疫耐受。CD4 T 细胞在血小板基因治疗后被耐受。预处理方案影响血小板基因治疗中的免疫耐受诱导。
摘要
背景免疫反应是基因治疗中的一个主要关注点。我们之前的研究表明，血小板靶向因子 VIII（FVIII）（2bF8）基因治疗结合转导细胞的体内药物选择可以挽救 FVIII 小鼠的出血倾向并诱导免疫耐受。目的研究非选择性 2bF8 慢病毒载体（LV）诱导血小板-FVIII 表达是否足以诱导免疫耐受，以及 2bF8LV 基因治疗后如何诱导免疫耐受。方法通过 2bF8LV 转导和移植引入血小板-FVIII 表达。FVIII 测定和尾巴出血试验用于确认血小板基因治疗的成功。用 rhF8 对动物进行攻击，以探讨基因治疗后是否诱导免疫耐受。使用 Treg 细胞分析、T 细胞增殖测定和记忆 B 细胞介导的 ELISPOT 测定来研究免疫耐受的潜在机制。结果我们表明，2bF8LV 基因治疗后，FVIII 小鼠的血小板-FVIII 表达持续存在，出血倾向得到恢复。在接受 660 cGy 照射或白消安加 ATG 预处理的组中，没有转导的受者在 rhF8 挑战后产生抗 FVIII 抑制性抗体。在 2bF8LV 转导的受者中，Treg 细胞显著增加，并且所开发的免疫耐受具有可转移性。来自治疗动物的 CD4 T 细胞在对 rhF8 再刺激的反应中无法增殖，但记忆 B 细胞可以在 2bF8LV 转导的受者中分化为产生抗体的细胞。结论在没有体内选择操作细胞的情况下，2bF8LV 基因转移可以在血友病 A 小鼠中引入免疫耐受，并且这种免疫耐受是由 CD4 T 细胞介导的。
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