单倍型相合造血干细胞移植后γδT细胞功能亚群重建与EBV感染的相关性及调控机制研究

Epstein–Barr virus (EBV) reactivation and its related diseases are the life-threatening clinical complications in patients who have undergone haploidentical hematopoietic stem cell transplantation (haploHSCT). It has been recognized that immune recovery after allogeneic HSCT is crucial for decreasing subsequent morbidity of virus infections. Delayed recovery of adaptive immunity was found to be associated with the increased risk of EBV infection. However, the link between immune recovery and EBV infection post-haploHSCT remains elusive. Our recent studies suggest that γδT cells could play a role in the control of EBV reactivation after haploHSCT. Human γδ T cells can recognize and respond to a wide variety of stress-induced antigens, thereby developing anti-tumor and anti-infective activity in a MHC-independent manner. The majority of γδ T cells in peripheral blood have the Vγ9Vδ2 T cell receptor. Although previous studies have demonstrated that Vγ9Vδ2 T cells are capable to eliminate the EBV-transformed B lymphocytes in vitro and in the mouse models, the reconstitution characteristic of Vδ2+ T cells and its functional involvement in anti-EBV infection after HSCT remain to be elucidated. In contrast, expansion of Vδ1+ or Vδ2-negative T cells in response to EBV reactivation has been reported in the individual patient following bone marrow or cord blood transplantation. To clarify the functional role of γδ-T subsets in this regard, in the current project, we are going to elucidate the recovery characteristics of γδT cell and its subpopulations after haploHSCT with "Beijing protocol". The mechanism of anti-EBV immunity and γδT-based therapeutic strategies in this context will be also explored. Our findings will benefit to control the serious complications due to EBV infection post-haploHSCT and improve the overall survival of haploHSCT recipients.

EB病毒(EBV) 感染是单倍型相合造血干细胞移植(haploHSCT)后常见的致死性并发症，严重影响患者生存。虽然免疫重建延迟已被认为是移植后发生感染的重要因素，但目前对haploHSCT后与EBV感染相关的免疫细胞亚群（特别是固有免疫细胞）缺乏明确阐述。申请人前期工作提示，γδT细胞可能与haploHSCT后发生EBV感染相关。虽然体外和动物模型实验已表明γδT细胞具有抗EBV功能，但γδT细胞在haploHSCT模式中的重建特征、重建的γδT细胞是否与移植后EBV感染及作用机制未见报道。本项目以原创性“北京方案”移植模式为平台，旨在阐明haploHSCT后γδT细胞及亚群的重建规律、其与发生EBV感染的确切关系，揭示调控重建γδT细胞亚群抗EBV感染的机制，进而探索移植后干预EBV感染的新策略。该研究结果将有助于解决haploHSCT所面临的严重感染问题，提高临床疗效、改善患者生存。

EB病毒（Epstein-Barr virus，EBV）再激活和EBV相关淋巴增殖性疾病（Epstein-Barr virus-related lymphoproliferative disorders，EBV-LPD）是异基因造血细胞移植（allogeneic hematopoietic cell transplantation，alloHCT）后常见的严重并发症，可显著降低患者生存。阐明移植受者体内参与抗EBV感染的免疫机制对改善预后、提高患者生存具有重要意义。本课题通过临床队列研究和体外实验首次明确了HLA单倍型相合HCT后早期外周血Vδ2+ T细胞亚群重建水平低与发生EBV再激活显著相关，对于全面揭示移植后抗EBV细胞免疫机制具有重要理论和实践意义。本课题进一步研究发现，单倍型相合HCT后重建的树突状细胞（DCs）亚群比例低与Vδ2+ T细胞重建延迟具有显著相关性，体外诱导的自体/异体DCs 均可促进移植后重建的Vδ2+ T 细胞扩增，且扩增后的Vδ2+ T 细胞具有较好的活性和功能。本课题通过体外和小鼠实验、以及临床队列研究首次阐明移植方案中常用免疫抑制剂酶酚酸酯对Vδ2+ T细胞功能活性及抗EBV效应具有抑制作用，揭示了免疫抑制剂的应用增加移植后EBV感染风险的免疫学机制，临床上缩短移植后使用酶酚酸酯的疗程可显著改善Vδ2+ T细胞重建、降低EBV再激活和EBV-LPD的发生率。尽管体外和小鼠实验都证实Vδ2+ T细胞具有杀伤EBV靶细胞的作用，但其杀伤效率仍有待提高。丙戊酸（valproic acid，VPA）是一种组蛋白去乙酰化酶抑制剂，既往研究显示其对化疗或免疫治疗具有协同抗肿瘤作用。双膦酸盐（如pamidronate，PAM）和/或VPA能否增强EBV靶细胞对Vδ2+ T细胞的免疫原性既往未见报道。本课题研究发现，VPA联合PAM通过甲羟戊酸途径提高了EBV感染的靶细胞和EBV相关肿瘤细胞的免疫原性，从而增强了Vδ2+ T细胞的杀伤作用。上述研究成果丰富了对造血细胞移植后抗病毒免疫机制的认识，为治疗致死性病毒感染并发症提供了新的靶点和选择策略，建立和发展以Vδ2+ T 细胞为基础的细胞免疫治疗手段有利于更好地发挥γδT 细胞抗感染效应，从而提高alloHCT受者总生存率。同时对研究γδT 细胞在抗肿瘤免疫的作用具有重要借鉴意义。