Role of F-Box Proteins in Lung Transplantation

摘要：肺移植是终末期肺部疾病的唯一治疗选择； 然而，存活率落后于其他器官移植，主要是由于慢性同种异体肺移植功能障碍。 (衣着)，最常见的是闭塞性细支气管炎综合征(BOS)。急性细胞排斥反应(ACR)的发作有 尽管以钙调神经磷酸酶抑制剂为基础(如环孢素A；CsA)，但CLAD的常见和主要危险因素 常规免疫抑制疗法(IST)。实验性移植模型和人体研究表明 1型T细胞免疫，以干扰素-γ的产生为标志，由转录因子T-bet驱动，发挥着 在同种异体移植排斥反应中的关键作用。因此，新的有效的治疗策略是一个尚未得到满足的主要需求。 它是防止肺排斥反应和改善预后的迫切需要。我们发现了一部小说， 促炎症泛素，E3连接酶亚单位，Fbxo3，它降解抗炎钙调素(CaM)- 相关蛋白质称为Fbxl2。我们的初步数据显示，Fbxo3在小鼠ACR期间被诱导 在小鼠原位肺移植(OLT)模型上，随着T-bet的增加，Fbxl2明显降低。此外， 使用一流的新型Fbxo3小分子抑制剂BC-1261，降低T-bet/1型免疫和 ACR的严重性，并在我们的早期研究中保留了Fbxl2。我们对肺移植受者(Ltrs)的研究表明 肺和血液中1型同种异体免疫反应的优势，Fbxo3上调和抑制 BC-1261的免疫应答。因此，我们假设Fbxl2是T-bet/Type-1的关键调节因子 T细胞免疫，FbxO_3：Fb_x12平衡和T-bet/干扰素-γ诱导是肺的关键决定因素 同种异体移植排斥与接受。为了检验这一点，我们提出了三个目标。在目标1中，我们将检验假设 在活化的T细胞中，Fbxo3的诱导降解了Fbxl2并增加了T-bet。我们将确定Fbxl2是否 泛素化T-bet，CaM是否作为连接分子调节T-bet，并检测Fbxo3的调节 在T细胞激活过程中。在目标2中，使用小鼠原位移植模型，我们将检验以下假设： 功能性Fbxo3：Fbxl2和T-bet/Type-1免疫是同种异体肺移植结果的关键决定因素。在这里，我们 将评估我们的新型Fbxo3抑制剂BC-1261与CsA对ACR或闭塞性呼吸道疾病(OAD； 建立BOS模型)，并测试新型Fbxo3缺陷小鼠的肺排斥反应是否减少。在目标3中，我们将测试 假设Fbxo3和T-bet/1型免疫通路是人类LT和 BC-1261是否抑制同种异体免疫反应。利用我们不断扩展的Ltr注册/生物信息库 支气管肺泡灌洗(BAL)和PBMC样本，我们将确定F-box蛋白是否发挥重要作用 在肺移植和全身同种异体免疫中的作用。私家侦探麦克代尔博士和陈博士带来了多学科 结合泛素生物学、小鼠和人类肺移植免疫学的专业知识，并将使用多重 PI格式。R01的成功将对预防同种异体肺移植排斥反应的科学产生革命性的影响。 为测试针对促炎泛素、E3连接酶亚单位Fbxo3的一流疗法奠定了基础。

Abstract: Lung transplantation (LT) is the only therapeutic option for patients with end-stage lung disease; however, survival lags behind other organ transplants, and is primarily due to chronic lung allograft dysfunction (CLAD), most often the bronchiolitis obliterans syndrome (BOS). Episodes of acute cellular rejection (ACR) are common and the major risk factor for CLAD, despite calcineurin inhibitor-based (e.g., cyclosporine; CSA) conventional immunosuppression therapy (IST). Experimental transplant models and humans studies indicate that Type-1 T cell immunity, marked by IFN-γ production, and driven by the transcription factor, T-bet, play a key role in allograft rejection. Thus, new and effective therapeutic strategies represent a major unmet need in LT and are desperately needed to prevent lung rejection and improve outcomes. We have discovered a novel, pro-inflammatory ubiquitin, E3 ligase subunit, Fbxo3, which degrades an anti-inflammatory calmodulin (CaM)- associated protein called Fbxl2. Our preliminary data show Fbxo3 is induced during ACR in the mouse orthotopic lung transplant (mouse OLT) model, along with T-bet, while Fbxl2 is markedly reduced. Further, using a first-in-class novel small molecule inhibitor of Fbxo3, BC-1261, reduces T-bet/Type-1 immunity and ACR severity, and preserves Fbxl2 in our early studies. Our studies in lung transplant recipients (LTRs) show a predominance of Type-1 alloimmune responses in the lung and blood, up-regulation of Fbxo3, and inhibition of immune responses with BC-1261. Therefore, we hypothesize that Fbxl2 is a critical regulator of T-bet/Type-1 immunity in T cells, and that Fbxo3: Fbxl2 balance and T-bet/IFN-γ induction are key determinants of lung allograft rejection versus acceptance. To test this, we propose 3 aims. In Aim 1, we will test the hypothesis that induction of Fbxo3 degrades Fbxl2 and increases T-bet in activated T cells. We will determine whether Fbxl2 ubiquitinates T-bet, whether CaM acts as a linker molecule in regulating T-bet, and test the regulation of Fbxo3 during T cell activation. In Aim 2, using the mouse OLT model, we will test the hypothesis that the balance of functional Fbxo3: Fbxl2 and T-bet/Type-1 immunity are key determinants of lung allograft outcomes. Here, we will assess our novel Fbxo3 inhibitor, BC-1261 versus CSA on ACR or obliterative airways disease (OAD; modeling BOS), and test whether lung rejection is reduced in novel Fbxo3-deficient mice. In Aim 3, we will test the hypothesis that the Fbxo3 and T-bet/Type-1 immunity pathways are putative targets in human LT and whether BC-1261 inhibits alloimmune responses. Making use of our expanding LTR registry/biorepository of bronchoalveolar lavage (BAL) and PBMC samples, we will determine whether F-box proteins play an important role in lung allograft and systemic alloimmunity. The PIs, Drs. McDyer and Chen, bring multidisciplinary expertise combining ubiquitin biology, mouse and human lung transplant immunology, and will use the multiple PI format. Success in this R01 will be transformative in the science to prevent lung allograft rejection, and lay the foundation to test a first-in-class therapy targeting the pro-inflammatory ubiquitin, E3 ligase subunit, Fbxo3.