Skin-Intrinsic Immunosuppressive Mesenchymal Stem Cells and aGVHD

项目总结/摘要 急性移植物抗宿主病（aGVHD）是异基因造血干细胞移植的主要限制性并发症 移植用于治疗白血病/淋巴瘤和某些实体瘤。aGVHD发生时，T 来自供体骨髓的淋巴细胞攻击受体体内对维持生命功能至关重要的细胞， 器官，即皮肤、肝脏和肠道。在过去三十年的研究中，墨菲博士的实验室 发现在皮肤和鳞状粘膜中， 次要组织相容性抗原的差异，涉及携带Vb特异性标记的T细胞， 选择性地作用于上皮网脊以及鼠毛囊的富含上皮干细胞的凸起区域。 这种关联导致表达干细胞标志物的相邻上皮细胞凋亡而死亡， 细胞角蛋白15。这些效应肢的系统性治疗抑制充满了问题，包括 对免疫重建和移植物抗肿瘤效应钝化的附带影响。然而，我们现在知道 通过与该提案的共同PI Markus和Natasha Frank博士合作发现， 皮肤和鳞状粘膜具有能够有效免疫抑制的内在细胞途径。 具体来说，我们已经发现了一种具有干细胞特征的真皮树突状细胞， 多药耐药转运蛋白ABCB 5，也表达PD-1。这种细胞有能力阻止T细胞 反应，包括同种异体反应性，因此具有重要的，但尚未探索的承诺，作为皮肤- 内源性途径，可用于抑制aGVHD中的组织损伤。在这份提案中，墨菲博士和 弗兰克博士已经联合力量，以解决利用皮肤的内在的长期目标， 免疫抑制能力来阻止aGVHD。该提案的具体目标是确定 表达PD-1的免疫抑制性真皮间充质干细胞在最早和最特异性 效应细胞-靶细胞相互作用导致aGVHD。为此，我们提出了三个具体目标： 1)将时空多重免疫谱与疾病发病率和恢复的所有方面相关联 在aGVHD中; 2）剖析ABCB 5真皮干细胞在aGVHD中的功能，并定义PD-1/PD-L1中心 皮肤内源性免疫抑制策略; 3）应用ABCB 5+真皮干细胞移植 利用时空多重免疫谱作为效应-靶点影响aGVHD严重程度的策略 生物测定这些方法涉及最先进的效应细胞-靶细胞的多重免疫表型分析 在aGVHD中敏感和特异地代表组织损伤的相互作用，并利用ABCB 5基因敲除 以及PD-L1 IG给药和ABCB 5+干细胞移植背景下的谱系追踪模型。的 基本原理是，一旦成功完成这项工作，我们将确定新的治疗方法， 通过皮肤操作治疗和预防aGVHD皮肤和鳞状粘膜病理， 内在免疫抑制途径。

Project Summary/Abstract Acute graft-versus-host disease (aGVHD) is a major limiting complication of allogeneic stem cell transplantation for the treatment of leukemia/lymphoma and certain solid tumors. aGVHD occurs when T lymphocytes from donor bone marrow attack cells within the recipient that are critical to the function vital organs, namely skin, liver and gut. During the past three decades of research, Dr. Murphy’s laboratory has found that in skin and squamous mucosae, the first sites of effector-target cell interaction provoked by differences in minor histocompatibility antigens, involve T cells bearing Vb-specific markers that home selectively to epithelial rete ridges as well as the epithelial stem cell-rich bulge regions of murine hair follicles. This association results in death by apoptosis of adjacent epithelial cells that express the stem cell marker, cytokeratin 15. Systemic therapeutic inhibition of these effector limbs is riddled with problems that include collateral impact on immune reconstitution and blunting of graft-versus-tumor effects. However, we now know through discoveries made in collaboration with co-PIs of this proposal, Drs. Markus and Natasha Frank, that the skin and squamous mucosae harbor intrinsic cellular pathways capable of potent immunosuppression. Specifically, we have discovered a dermal dendritic cell that has stem cell characteristics, expresses the multidrug resistance transporter ABCB5, and also expresses PD-1. This cell has the ability to thwart T cell responses, including alloreactivity, and therefore holds significant but as yet unexplored promise as a skin- intrinsic pathway that may be leveraged to inhibit the tissue injury in aGVHD. In this proposal, Dr. Murphy and the Drs. Frank have combined forces to address the long-term objective of harnessing the skin’s intrinsic immunosuppressive capabilities to thwart aGVHD. The proposal’s specific objective is to determine the impact of PD-1-expressing immunosuppressive dermal mesenchymal stem cells on the earliest and most specific effector-target cell interactions responsible for aGVHD. To this end, we have proposed three specific aims: 1) to correlate spatiotemporal multiplex immune profiling with all aspects of disease morbidity and recovery in aGVHD; 2) to dissect ABCB5 dermal stem cell function in aGVHD and define PD-1/PD-L1-centered strategies for skin-intrinsic immunosuppression; and 3) to apply ABCB5+ dermal stem cell transplant strategies to influence aGVHD severity using spatiotemporal multiplex immune profiling as effector-target bioassay. The approaches involve state-of-the-art multiplex immunophenotyping of effector-target cell interactions that sensitively and specifically typify tissue injury in aGVHD, and leverage ABCB5 gene knockout and lineage tracing models in context of PD-L1 Ig administration and ABCB5+ stem cell engraftment. The rationale is that upon successful completion of this work, we will identify novel therapeutic approaches to the treatment and prevention of aGVHD skin and squamous mucosal pathology through manipulation of skin- intrinsic immunosuppressive pathways.