Targeting dendritic cells for selective modulation of GVHD

在AIM 1下进行的实验表明，即使相对轻微的GVHD也会降低对疫苗接种的数量T细胞免疫反应和对表达疫苗靶向抗原的肿瘤的功能性免疫反应。这项工作已经发表(Capitini等人，布拉德，2009)，并证明了如果骨髓移植要优化为针对恶性肿瘤的免疫治疗方法的平台，预防移植物抗宿主病的重要性。在目标2下，我们已经确定抑制干扰素-γ信号可以阻止GVHD的发展。重要的是，接受供者骨髓和缺乏干扰素γ受体的T细胞可以防止移植物抗宿主病的发生。然而，这些小鼠对疫苗接种没有反应，也不能防止肿瘤的发展。有趣的是，干扰素γ受体缺陷的骨髓受者可以被注入大量表达干扰素伽马受体的完全免疫活性T细胞，而不会引发移植物抗宿主病。此外，这些小鼠对树突状细胞疫苗产生了强烈的反应，从而对肿瘤具有保护作用。接下来，我们研究了干扰素-伽马途径的其他成分是否可以在捐赠者的骨髓中靶向，以预防移植物抗宿主病。利用干扰素-γ信号转导所必需的转录因子STAT1的骨髓缺陷，我们已经证实，在骨髓来源的细胞中干扰这一途径可以在保留免疫功能的情况下预防GVHD。为了确定相关的骨髓源性细胞群，我们选择性地以STAT1为靶标，方法是生成带有STAT1基因的小鼠(从Lothar Hennighausen博士那里获得)，该基因在非T细胞启动子(CD11c(DC表达)、溶菌酶(在所有吞噬细胞上)和CD19(B细胞表达)下表达Cre重组酶)。在这些捐献者的骨髓接受者中，STAT1基因(因此，干扰素伽马信号)将在选择性细胞群中被去除。使用这些小鼠作为骨髓捐赠者的异基因移植实验正在进行中。目标3还处于初级阶段。我们已经获得了选择性的STAT1抑制剂，并开始在体外测试这些药物。

The experiments conducted under aim 1 have demonstrated that even relatively mild GVHD can diminish quantitative T cell immune responses to vaccination and functional immune responses to tumors expressing vaccine-targeted antigens. This work has been published (Capitini et al, Blood, 2009) and demonstrated the importance of preventing GVHD if BMT is to be optimized as a platform for immunotherapeutic approaches targeting malignancy. Under aim 2, we have established that inhibition of interferon gamma signaling can prevent the development of GVHD. Importantly, recipients of donor bone marrow and T cells deficient in interferon gamma receptor prevented the development of GVHD. However, these mice were unable to response to vaccination and could not be protected against tumor development. Interestingly, recipients of bone marrow deficient in interferon gamma receptor could be infused with large number of fully immune competent T cells expressing the interferon gamma receptor did not without inducing GVHD. In addition, these mice generated robust responses to dendritic cell vaccination resulting in protection against tumor. We next studied whether other components of the interferon gamma pathway could be targeted in donor bone marrow to prevent GVHD. Using bone marrow deficient in STAT1, a transcription factor necessary for interferon gamma signaling, we have confirmed that interference with this pathway in bone marrow-derived cells can prevent GVHD with preserved immune competence. To identify the relevant bone-marrow-derived cell population, we have selectively targeted STAT1 by generating mice with a floxed STAT1 gene (obtained from Dr. Lothar Hennighausen) that express the Cre recombinase under non-T cell promoters (CD11c (DC expression), lysozyme (on all phagocytic cells), and CD19 (B cell expression). In recipients of bone marrow from these donors, the STAT1 gene (and, thus, interferon gamma signaling) will be ablated in selective cell populations. Allogeneic transplant experiments using these mice as bone marrow donors are ongoing. Aim 3 is in the initial stage. We have obtained selective Stat1 inhibitors and are beginning to test these in vitro.