Novel GvHD-Specific Gene Identification for Optimal Control of GvHD

摘要 移植物抗宿主病(GvHD)是异基因造血成功的重要障碍之一。 细胞移植(allo-HCT)。该项目的目标是识别其遗传/药理学特性的新基因。 阻断可以选择性地预防GvHD，同时保持或增强抗白血病活性(又名移植物- 抗白血病或GVL效应)。异基因血细胞移植仍然是血液病患者最有效的治疗方法 恶性肿瘤，作为allo-HCT的治疗益处，主要来自GVL。然而，allo-HCT也 存在这样的风险，即移植(移植物)中的捐赠者T细胞会变得过于狂热，并开始攻击 不仅是白血病，还有患者的皮肤、肠道和肝脏，导致移植物抗宿主病。因为一个强大的 两种供者T细胞介导的免疫反应之间的联系，特别是限制GvHD仍然是 Allo-HCT的目标。控制GvHD的威胁将扩大allo-HCT的范围和好处。几个 包括我们在内的组织已经提出了减少GvHD的治疗策略，而不是取消有益的 GVL在动物模型和人类患者中的作用。然而，异基因供者T细胞的机制 GvHD和GVL的差异调制在很大程度上仍然是未知的。我们机械论理解中的这个鸿沟 妨碍我们具体预防/治疗GvHD的能力。我们假设我们识别的基因是 与GvHD器官相比，与浸润性肿瘤的供者T细胞不同相关的T细胞将是 在不对GVL产生负面影响的情况下预防GvHD。为此，我们在全基因组范围内进行了无偏见的 CRISPR/CAS9库屏幕。因此，我们首次确定了487个与GvHD相关的新基因 这可能成为GvHD的分子靶点，因为这些基因的引导RNA被丰富了 仅表达于肿瘤结节中的T细胞，而不存在于GvHD靶器官中。我们发现有五套 参与已知途径的基因在这487个基因中最为显著地丰富。这五个都是 通路与RELN-RAP1-FGFR3轴紧密相连，但在异源基因中尚未被积极研究。 Hct字段。因此，我们将确定RELN-RAP1-FGFR3轴的遗传/药物抑制是否会导致 在我们的小鼠模型中维持或增强GVL的GVHD特异性免疫调节 GVHD/GVL和人PBMC/T细胞介导的异种GvHD/GVL。如果成功，我们的新疗法的发现 有选择地以最佳方式控制GvHD的目标将是潜在的游戏规则改变者。我们建议的研究将 对GvHD及其预防提供重要的机械性见解，这可能具有变革性 对治疗发展的影响不仅对实体器官移植排斥反应，而且对炎症性 炎症性肠病、牛皮癣和类风湿性关节炎等疾病。此外，我们的研究可能 也有助于提高抗癌治疗和免疫治疗方法的有效性 。

ABSTRACT Graft-versus-host disease (GvHD) is one of the most significant barriers to success for allogeneic hematopoietic cell transplantation (allo-HCT). The goal of this project is to identify novel genes whose genetic/pharmacologic blockade may selectively prevent GvHD while maintaining or enhancing anti-leukemia activities (aka graft- versus-leukemia or GvL effect). Allo-HCT remains the most effective treatment for patients with hematologic malignancies, as the therapeutic benefit of allo-HCT is primarily derived from GvL. However, allo-HCT also carries the risk that the donor T cells in the transplant (graft) will become overzealous and begin to attack not only the leukemia but also the patient's skin, intestines and liver, resulting in GvHD. Because of a strong association between the two donor T cell-mediated immune responses, specifically limiting GvHD remains the goal of allo-HCT. Managing the threat of GvHD would broaden the scope and benefit of allo-HCT. Several groups, including ours, have proposed therapeutic strategies to reduce GvHD without abrogating the beneficial GvL effect in animal models and human patients. However, the mechanisms by which allogeneic donor T cells differentially modulate GvHD and GvL remain largely unknown. This gap in our mechanistic understanding hinders our ability to specifically prevent/treat GvHD. We hypothesize that the genes we identify as being differentially associated with donor T cells that infiltrate tumors versus GvHD organs will be critical targets for the prevention of GvHD without negatively affecting GvL. To this end, we performed an unbiased genome-wide CRISPR/Cas9 library screen. As a result, we have for the first time identified 487 novel GvHD-associated genes that could potentially serve as molecular targets in GvHD, as the guide RNAs for these genes were enriched only in T cells obtained from tumor nodules but not from any GvHD target organs. We found that five sets of genes that are involved in known pathways are most significantly enriched in these 487 genes. All of these five pathways are closely linked to the RELN-RAP1-FGFR3 axis that has not been actively investigated in the allo- HCT field. Thus, we will determine if genetic/pharmacologic inhibition of the RELN-RAP1-FGFR3 axis results in GvHD-specific immune modulation while maintaining or enhancing GvL in our mouse models of murine GvHD/GvL and human PBMC/T cell-mediated xeno-GvHD/GvL. If successful, our findings of novel therapeutic targets to selectively and optimally control GvHD will be potential game-changers. Our proposed studies will provide significant mechanistic insights into GvHD and its prevention, which may have transformative implications for therapeutic development not only for solid organ transplant rejection but also inflammatory diseases such as inflammatory bowel disease, psoriasis and rheumatoid arthritis. In addition, our studies may also help improve the efficacies of anti-cancer therapies and immunotherapeutic approaches .