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-?B. Both NOD2 and RIP2 are NF-?B 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-?B, 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.

Public Health Relevance

Faulty NOD2 activity is responsible for exacerbating inflammatory diseases. Diseases such as asthma, inflammatory bowel disease, sarcoidosis and arthritis can all be worsened by too much NOD2 activity. RIP2 is an obligate kinase in the NOD2 signaling pathway. In our prior granting period, we found FDA-approved drugs that could inhibit RIP2. This has attracted the interest of pharmaceutical companies as a number are now screening for new RIP2 inhibitors. Despite this, our knowledge of RIP2's role in the cell is murky. This grant application aims to identify RIP2 substrates, to identify biomarkers that can identify NOD2-driven inflammatory disease and predict which patient best will responds to RIP2 inhibitors, to identify additional pharmaceuticals to fine-tune the inhibition of this pathway and o determine efficacy of RIP2 inhibition in inflammatory disease.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
2R01GM086550-06A1
Application #
8693212
Study Section
Innate Immunity and Inflammation Study Section (III)
Program Officer
Dunsmore, Sarah
Project Start
2008-12-09
Project End
2018-02-28
Budget Start
2014-05-10
Budget End
2015-02-28
Support Year
6
Fiscal Year
2014
Total Cost
$324,925
Indirect Cost
$119,925
Name
Case Western Reserve University
Department
Pathology
Type
Schools of Medicine
DUNS #
077758407
City
Cleveland
State
OH
Country
United States
Zip Code
44106
Russo, Hana M; Rathkey, Joseph; Boyd-Tressler, Andrea et al. (2016) Active Caspase-1 Induces Plasma Membrane Pores That Precede Pyroptotic Lysis and Are Blocked by Lanthanides. J Immunol 197:1353-67
Dorand, R Dixon; Nthale, Joseph; Myers, Jay T et al. (2016) Cdk5 disruption attenuates tumor PD-L1 expression and promotes antitumor immunity. Science 353:399-403
Chirieleison, Steven M; Kertesy, Sylvia B; Abbott, Derek W (2016) Synthetic Biology Reveals the Uniqueness of the RIP Kinase Domain. J Immunol 196:4291-7
Perez, Jessica M; Chirieleison, Steven M; Abbott, Derek W (2015) An IκB Kinase-Regulated Feedforward Circuit Prolongs Inflammation. Cell Rep 12:537-44
Ryan, Sean O; Leal Jr, Sixto M; Abbott, Derek W et al. (2014) Mgat2 ablation in the myeloid lineage leads to defective glycoantigen T cell responses. Glycobiology 24:262-71
Tigno-Aranjuez, Justine T; Benderitter, Pascal; Rombouts, Frederik et al. (2014) In vivo inhibition of RIPK2 kinase alleviates inflammatory disease. J Biol Chem 289:29651-64
Ryan, Sean O; Abbott, Derek W; Cobb, Brian A (2014) Myeloid glycosylation defects lead to a spontaneous common variable immunodeficiency-like condition with associated hemolytic anemia and antilymphocyte autoimmunity. J Immunol 192:5561-70
Jun, Janice C; Kertesy, Sylvia; Jones, Mark B et al. (2013) Innate immune-directed NF-κB signaling requires site-specific NEMO ubiquitination. Cell Rep 4:352-61
Tigno-Aranjuez, Justine T; Bai, Xiaodong; Abbott, Derek W (2013) A discrete ubiquitin-mediated network regulates the strength of NOD2 signaling. Mol Cell Biol 33:146-58
Jun, Janice C; Cominelli, Fabio; Abbott, Derek W (2013) RIP2 activity in inflammatory disease and implications for novel therapeutics. J Leukoc Biol 94:927-32

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