Tissue specific HLA-DPB1 reactive T cells in allogeneic hematopoietic stem cell transplantation

异基因造血干细胞移植（HSCT）目前是化疗难治性或复发性急性髓性白血病（AML）患者的标准治疗。HSCT的疗效基于供体免疫系统的免疫反应，特别是供体T细胞的免疫反应。供体T细胞通过组织相容性抗原识别患者AML原始细胞，并且是这种移植物抗白血病（GvL）效应的主要介质。然而，供体T细胞也可以识别患者来源的非造血细胞，从而介导有时危及生命的移植物抗宿主病（GvHD）。HSCT研究的一个主要目的是特异性地重定向供体T细胞对抗AML细胞，并在不诱导GvHD的情况下加强GvL效应。因此，需要鉴定仅在AML细胞上表达的T细胞限定的抗原。潜在的靶抗原是HLA-DP，其属于HLA II类分子并且在稳态条件下选择性地在造血细胞上表达。目前，供体搜索仅考虑HLA-A、-B、-C、-DR和-DQ。由于HLA-DP和其他HLA-II类基因座的遗传连锁不平衡较低，80%的无关供体HSC移植在至少一个HLA-DPB 1等位基因上不匹配。因此，产生选择性识别患者造血细胞的HLA-DPB 1错配反应性供体T细胞似乎是合理的。在先前的异种移植研究中，我们能够证明体外产生的同种异体HLA-DPB 1反应性人T细胞根除移植到免疫缺陷小鼠中的原代HLA-DPB 1 + AML母细胞。然而，在炎症条件下，HLA-DPB 1在非造血细胞上显示上调表达，因此同种异体HLA-DPB 1反应性T细胞也可能触发GvHD。在拟议的项目中，我想确定allo-HLA-DPB 1反应性CD 4 + T细胞专门识别造血细胞。首先，我将通过质谱序列分析表征与不同造血和非造血细胞上的HLA-DPB 1结合的肽。限制于造血细胞的肽表位的鉴定将使我能够产生同种异体HLA-DPB 1反应性CD 4 + T细胞，其应该在GvHD不存在的情况下介导选择性GvL效应。在第二部分中，我将从这些异基因HLA-DPB 1反应性T细胞克隆中克隆T细胞受体（TCR）。TCR基因转移将用于快速生成来自不同供体的allo-HLA-DPB 1反应性T细胞，这将为该方法在同种异体HSCT后过继性T细胞免疫治疗背景下的未来临床应用铺平道路。

Allogeneic hematopoietic stem cell transplantation (HSCT) is currently the standard of care treatment for chemorefractory or relapsed acute myeloid leukemia (AML) patients. The curative effect of HSCT is based on an immune reaction of the donor immune system, especially of donor T cells. Donor T cells recognize patients AML blasts through histocompatibility antigens and are the main mediators of this graft-versus-leukemia (GvL) effect. However, donor T cells can also recognize non-hematopoietic cells of patient origin thereby mediating the sometimes life-threatening graft-versus-host disease (GvHD). One major aim of HSCT research is to specifically redirect donor T cells against AML cells and strengthen the GvL effect without inducing GvHD. Therefore, the identification of T-cell defined antigens which are exclusively expressed on AML cells would be desirable. A potential target antigen is HLA-DP, which belongs to HLA class II molecules and is under steady state conditions selectively expressed on hematopoietic cells. Currently, only HLA-A, -B, -C, -DR, and -DQ is taken into account for donor search. Because of low genetic linkage disequilibrium of HLA-DP and the other HLA class II loci, 80 % of unrelated donor HSC transplantations are mismatched in at least one HLA-DPB1 allele. Therefore, it appears reasonable to generate HLA-DPB1 mismatch reactive donor T cells selectively recognizing patient hematopoietic cells. In previous xenotransplantation studies, we were able to show that in vitro generated allo-HLA-DPB1 reactive human T cells eradicate primary HLA-DPB1+ AML blasts engrafted in immunodeficient mice. However, under inflammatory conditions HLA-DPB1 shows upregulated expression on non-hematopoietic cells and therefore allo-HLA-DPB1 reactive T cells might also trigger GvHD. In the proposed project, I would like to identify allo-HLA-DPB1 reactive CD4+ T cells exclusively recognizing hematopoietic cells. First of all, I will characterize peptides binding to HLA-DPB1 on different hematopoietic and non-hematopoietic cells by mass spectrometry sequence analysis. The identification of peptide epitopes restricted to hematopoietic cells would allow me to generate allo-HLA-DPB1 reactive CD4+ T cells which should mediate a selective GvL effect in the absence of GvHD. In the second part, I will clone the T cell receptor (TCR) from those allo-HLA-DPB1 reactive T cell clones. TCR gene transfer will be used for the rapid generation of allo-HLA-DPB1 reactive T cells from different donors, which should pave the way for future clinical application of this approach in the context of adoptive T-cell immunotherapy after allogeneic HSCT.