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

异基因造血干细胞移植(HSCT)是目前治疗化疗难治或复发的急性髓系白血病(AML)患者的标准疗法。HSCT的疗效是基于供者免疫系统的免疫反应，尤其是供者T细胞的免疫反应。供者T细胞通过组织相容性抗原识别患者的AML细胞，是移植物抗白血病(GVL)效应的主要介导者。然而，供者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将供者T细胞定向到AML细胞，并在不引发GvHD的情况下增强GVL效应。因此，鉴定仅在AML细胞上表达的T细胞定义的抗原是可取的。一个潜在的靶抗原是人类白细胞抗原-DP，它属于人类白细胞抗原II类分子，在稳定状态下选择性表达于造血细胞上。目前，在寻找供者时只考虑了人类白细胞抗原-A、-B、-C、-DR和-DQ。由于HLADP和其他HLAII类基因座的遗传连锁不平衡，80%的无血缘关系的供者HSC移植中至少有一个HLADPB1等位基因不匹配。因此，产生选择性识别患者造血细胞的HLA-DPB1不合反应性供者T细胞似乎是合理的。在之前的异种移植研究中，我们能够证明，在体外产生的allo-HLA-DPB1反应性人类T细胞可以消除植入免疫缺陷小鼠的原发的HLA-DPB1+AML细胞。然而，在炎症条件下，非造血细胞上的HLA-DPB1表达上调，因此allo-HLA-DPB1反应性T细胞也可能触发GvHD。在提议的项目中，我想确定allo-HLA-DPB1反应性CD4+T细胞专用于识别造血细胞。首先，通过对不同的造血细胞和非造血细胞进行序列分析，鉴定与人类白细胞抗原-DPB1结合的多肽。识别仅限于造血细胞的多肽表位将使我能够产生allo-HLA-DPB1反应性CD4+T细胞，在没有GvHD的情况下应该会介导选择性GVL效应。在第二部分中，我将从这些allo-HLA-DPB1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产生来自不同供者的allo-HLA-DPB1反应性T细胞，这将为该方法在异基因HSCT后过继T细胞免疫治疗中的临床应用铺平道路。

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.