| Summary: | <p>Interferon regulatory factor 5 (IRF5), is a multifunctional regulator of immune responses, with a critical role in controlling pro-inflammatory responses of macrophages. IRF5 contributes to the pathogenesis of several inflammatory and autoimmune diseases. Given the importance of IRF5 function in physiology and disease, IRF5 represents a therapeutic target. However, the mechanisms regulating IRF5 activation and the kinases responsible for its phosphorylation in vivo have been unclear. The purpose of this study was to identify novel IRF5 kinases as potential targets for specific control of IRF5-driven inflammatory conditions. Based on a kinase inhibitor library screen and rounds of validation PYK2 and MEKK3 have been prioritised as candidate IRF5 kinases.</p>
<p>Using the CRISPR-Cas9 system I have generated PYK2 and MEKK3 deficient macrophages. Deficiency in PYK2 had an effect on IRF5 activation, IRF5 recruitment to the genomic loci and transcription of IRF5 target inflammatory genes. Moreover, I have identified the PYK2 dependent tyrosine site within IRF5 and demonstrated its importance in IRF5 activation. PYK2 inhibition led to a comparable impact on macrophage transcriptomic signature as IRF5 deficiency, and suppressed cytokine production in biopsies taken from the inflamed gut mucosal tissue of patients with ulcerative colitis. The molecular dissection of the PYK2/IRF5 axis carried out in this thesis has a direct translational potential for inflammatory conditions such as ulcerative colitis, where both the PYK2 and IRF5 gene loci have been associated with increased genetic risk.</p>
<p>Deficiency in MEKK3 had an effect on IRF5 activity and inflammatory gene expression. In the absence of MEKK3 inhibitors, clinically validated kinase inhibitors with off-target MEKK3 potency, were used as initial leads to design MEKK3-specific inhibitors. Chemical modifications of the lead compound Neratinib resulted in the identification and synthesis of analogs with comparable biochemical potency on MEKK3, improved selectivity on a small panel of pathway relevant kinases, with reduced IRF5-driven TNF activation in an IRF5- and MEKK3- specific manner.</p>
|
|---|