T2 A novel drug target: IL-36 signalling drives inflammation and poor bacterial clearance in COPD

The inflammatory cytokine IL-36γ is increased in COPD lungs, whereas its endogenous inhibitor IL-36 receptor antagonist (IL-36RA) is reduced. IL-36 receptor signalling drives neutrophilic inflammation. Macrophages are critical in modulating lung inflammation and clearance of bacteria but the effect of IL-36γ on macrophages is not fully understood. To understand how IL-36γ affected macrophage function, non-smoker(NSm) and COPD monocyte-derived macrophages (MDM) were cultured with 100ng IL-36γ. IL-36γ reduced phagocytosis of H.influenzae by 23% in COPD MDM (p<0.01,n=13) but there was no effect on NSm MDM. Cytokine concentrations were measured in MDM supernatants by ELISA. IL-36γ did not affect IL-6, CXCL1 or CXCL8 secretion. Small airway fibroblasts (SAF) are a known effector cell of IL-36γ. To assess cross-talk between cell types, SAF supernatants were transferred onto MDM and cytokines measured in MDM supernatants. Untreated SAF did not stimulate MDM, however IL-36γ-treated SAF caused increased IL-6 (NT SAF vs IL36γ SAF: 2.8±0.4 vs 49.6±13.1ng/ml), CXCL1 (3.4±1.5 vs 14.1±4.9ng/ml) and CXCL8 (13.9±3.4 vs 53.3±11.0ng/ml) secretion from MDM. Lung macrophages are a major source of IL-36RA. IL36RN expression was measured in monocytes and MDM. COPD monocytes had lower IL36RN expression than NSm (NSm:1.52±0.38 vs COPD:0.32±0.12ng/ml, p<0.01, n=8), suggesting an innate pre-disposition towards low IL-36RA in COPD. IL36RN expression increased on monocyte differentiation to MDM, with NSm MDM having 5-fold higher IL36RN expression than COPD MDM (p<0.01). To investigate the mechanism regulating IL-36RA expression, MDM were incubated with PIK75 (PI3K inhibitor), UO0126 (ERK inhibitor), VX745 (p38 inhibitor), SP600125 (JNK inhibitor) and TPCA1 (NF-κB inhibitor) for 24h and IL36RN measured. Only PIK75 increased IL36RN expression (NT:0.63±0.33, PIK75:1.63±0.75, p<0.05, n=11), indicating that PI3K signalling mediates its reduced expression in COPD. This study demonstrates that IL-36γ causes a cascade of inflammation and impairs bacterial clearance in COPD. As IL-36γ is secreted in response to viral stimuli, the effects of IL-36γ described above could be important in pathogenesis of COPD exacerbations and secondary bacterial infection. IL36RN is reduced in COPD macrophages, which may be due to PI3K expression which is increased in COPD: Further work is needed to identify how this pathway regulates IL36RN to identify a potential drug target.