Innate Immune Signal Transduction Specificity in Inflammatory Disease

描述（由申请人提供）：异常NOD2信号引起肉芽肿性炎症性疾病。NOD2等位基因功能缺失的患者易患克罗恩病，这是一种胃肠道炎症性疾病。相反，功能获得性NOD2突变的患者会发展为早发性结节病（EOS），这是一种以非干酪化肉芽肿为特征的炎症性疾病，可导致肺、肝和眼睛损伤。功能缺失多态性和功能获得突变都会引起炎症性疾病，这可能是由于NOD2作为一种变阻器帮助维持正常的免疫稳态。这种变阻器功能始于细菌侵入细胞，然后NOD2与细菌肽聚糖的分解产物结合。这激活了NOD2，这样它就可以调节先天免疫系统，帮助定制适应性免疫反应，以根除有害的病原体。NOD2激活过多或过少都可能是有害的，这种不平衡是炎症性疾病发展的核心。NOD2及其专性激酶RIP2是细胞内细菌识别导致NOD2:RIP2复合物被激活的正调控回路的一部分。除了刺激自噬、杀菌活性、MHC II类呈递和MAPK激活外，NOD2:RIP2复合体还激活NF-κB。NOD2和RIP2都是NF-κB调控的基因，因此，它们的激活会导致一个正反馈循环，其中NOD2:RIP2的激活会刺激进一步的激活和进一步的炎症。此外，NOD2和RIP2的表达受到多种炎症介质的刺激，包括TNF和IFN。鉴于此，在之前的批准期内，我们假设抑制这种正调节回路可能对治疗炎症性疾病有效。我们成功地鉴定了RIP2激酶活性的纳米摩尔抑制剂。尽管如此，一个令人不安的问题

DESCRIPTION (provided by applicant): Aberrant NOD2 signaling causes granulomatous inflammatory disease. Patients with loss-of-function NOD2 alleles are prone to the development of Crohn's disease, an inflammatory disorder of the gastrointestinal tract. In contrast, patients with gain-of-function NOD2 mutations develop Early Onset Sarcoidosis (EOS), an inflammatory disorder characterized by noncaseating granulomas that cause lung, liver and eye damage. The fact that both loss-of-function polymorphisms and gain-of-function mutations both cause inflammatory diseases is likely due to the fact that NOD2 functions as a rheostat to help maintain normal immunologic homeostasis. This rheostat function begins upon bacterial invasion of the cell whereupon NOD2 binds to a breakdown product of bacterial peptidoglycan. This activates NOD2 such that it can modulate the innate immune system to help tailor the adaptive immune response to eradicate the offending pathogen. Either too much or too little NOD2 activation can be deleterious, and this imbalance is central to the development of inflammatory disease. NOD2 and its obligate kinase RIP2 are part of a positive regulatory circuit in which intracellular bacterial recognition causes the NOD2:RIP2 complex to be activated. In addition to stimulating autophagy, bacteriocidal activity, MHC Class II presentation and MAPK activation, the NOD2:RIP2 complex activates NF-κB. Both NOD2 and RIP2 are NF-κB regulated genes, and as such, their activation causes a positive feedback loop in which activation of NOD2:RIP2 stimulates further activation and further inflammation. Additionally, NOD2 and RIP2 expression are stimulated by a variety of mediators of inflammation, including TNF and IFN. Given this, in the prior granting period, we hypothesized that inhibiting this positive regulatory circuit might be efficacious in treating inflammatory disease. We were successful in identifying nanomolar inhibitors of RIP2's kinase activity. Despite this, a troubling fact remains: We still don't know what the kinase activity of RIP2 is doing in the cell. Some studies have shown that the kinase activity is dispensable for NOD2 activity while others have shown that it's essential. Our work has helped clarify this as we showed that RIP2 was misclassified as a serine-threonine kinase. It is actually a dual specificity kinase, meaning that t phosphorylates serines, threonines and tyrosines. Our own work has shown that inhibition of RIP2 attenuates the acute NOD2 inflammatory response. While my lab has found that RIP2's kinase activity helps regulate NF-κB, we don't know its role in regulating other NOD2-driven responses like autophagy or MAPK signaling. The uncertainty regarding RIP2's kinase activity takes on added importance given the interest of pharmaceutical companies in inhibiting RIP2 in inflammatory diseases like sarcoidosis, asthma, IBD and inflammatory arthritis. Understanding the kinase activity is essential if the goal is to inhibit RIP2 in inflammatory disease and to then determine efficacy and response in those diseases. This knowledge is also essential to predict outcomes of RIP2 inhibition in inflammatory disease. This grant application aims to answer these key questions.