树突状细胞在干扰素联合供体淋巴细胞输注治疗移植后白血病复发中增强抗白血病效应的机制研究

Acute leukemia (AL) relapse is the leading cause of death following allogeneic stem cell transplantation (allo-HSCT) and the overall survival is poor due to lack of effective treatment. Our group pioneered a novel strategy using donor lymphocyte infusion (DLI) in combination with interferon-α (IFN-α).We demonstrated that the combination of IFN-α with DLI (aDLI) significantly improved the therapeutic efficacy compared to traditional DLI (tDLI). Our data showed that in the group of aDLI,the complete remission (CR) rate and 3-year leukemia free survival (LFS) was 81.3% and 48.6% respectively.Furthermore, our preliminary results showed that IFN-α could enhance the anti-leukemia effects of donor lymphocyte via the activation of dendritic cell (DC) and subsequent enhancement of antigen- presentation and cytotoxity of T cells. However, the detailed mechanisms remain unknown. In this study, we will investigate the association between the activation and maturation of IFN-α-induced DC (IFN-DC) and the enhanced anti-leukemia effects in aDLI treatment using both in vitro and in vivo models as well as clinical specimen. Moreover, we will also elucidate the molecular mechanisms underlying the induction of IFN-DC activation and maturation triggered by IFN-α.We believe that the results of this research will provide valuable theoretical information for designing more optimal and effective adoptive immunotherapy in the future.

异基因造血干细胞移植术后复发的急性白血病（AL）患者缺乏有效治疗手段，预后恶劣，是临床急需解决的一大难题。近来本课题组在国际上创新性的应用干扰素-α（IFN-α）联合供体淋巴细胞输注（aDLI）治疗移植后复发的对化疗或常规DLI无效的高危AL，81.3%的患者获得完全缓解，3年无白血病生存率达48.6%，疗效远优于常规DLI。但确切机制尚待研究。预试验结果显示IFN-α可能通过有效活化树突状细胞（DC），达到增强抗原提呈，增强效应细胞的活化和杀伤效应。本课题拟用病患标本，离体细胞培养和小鼠模型，从IFN-α诱导的DC(IFN-DC)的表型特征，功能（抗原提呈，促Th1极化，促抗原特异性CD8+ CTL活化增殖，细胞毒性，移行能力）及其成熟活化的相关分子机制，研究IFN-DC在aDLI治疗中增强抗白血病效应的可能机制，为设计更优化、更有效的新型过继免疫治疗方案提供理论依据。

急性白血病（AL）患者异基因造血干细胞移植（allo-HSCT）后复发是导致移植后死亡的首要原因，是造血干细胞移植领域急需解决的一大难题。本课题组在国际上创新性的应用干扰素-α（IFN-α）联合供体淋巴细胞输注（aDLI）治疗移植后复发的对化疗或常规DLI无效的高危AL，疗效远优于常规DLI，确切机制不明。本课题通过临床治疗（包括病患标本检测），体外树突状细胞（DC）培养体系和小鼠模型三个层面进一步从临床疗效，细胞体外功能实验及动物模型证实：1. aDLI组无论在缓解率和5年OS和DFS均明显优于tDLI组; 2. aDLI组患者和aDLI治疗有效组患者的DC尤其是IFN-α诱导产生的IFN-DC的数量及活化水平以及T细胞及其活化亚群（尤其是效应细胞CD8+CTL, CD3+TCRγδ+）较tDLI组和无效组患者明显增高，与杀伤效应相关的穿孔素、颗粒酶B的mRNA转录水平，TH1方向相关的细胞因子水平（IL-12，IFN-γ）在aDLI后+2-+4周达到高峰，而且aDLI有效的HLA-A*0201阳性的患者白血病特异性CTL（WT-1+CTL）细胞水平增高，且在aDLI后+3-+4周达到高峰；2.建立的小鼠移植后复发模型和aDLI模型以及tDLI模型亦证实上述结果；3. 体外细胞培养：① 建立了与aDLI和tDLI相对应的体外培养体系即 IFN-DC与IL-4-DC，并从生物学特征及细胞功能方面证实IFN-DC与IL-4-DC为不同的DC群体； ②体外3天诱导分化的IFN-DC能在抗原提呈及迁移趋化能力方面优于传统的体外7天诱导并促成熟的IL-4-DC；③经IFN-α诱导产生的高度活化的IFN-DC刺激淋巴细胞增殖的能力较传统的IL-4-DC更强；④经抗原肽负载后的IFN-DC激活T细胞产生的特异性的细胞杀伤效应较IL-4-DC更强；⑤IFN-α促进单核细胞分化产生高度活化的IFN-DC部分是JAK-STAT信号通路激活的结果；上述研究结果证实了我们的科学假说：即新型DLI中的IFN-α是通过有效激活DC，尤其是IFN-DC，达到增强白血病抗原提呈，增强效应T细胞的活化和对白血病细胞的杀伤效应，从而达到快速和超越常规DLI疗效的优势。本课题为设计更优化、更有效的新型过继免疫治疗方案提供了理论依据。