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Asthma is a highly rhythmic disease with increased airway sputum eosinophilia and decline in lung function (FEV1) observed early morning. 1 We have previously shown that the time of day of allergen challenge in mice governs the amplitude of the resulting airway hyperresponsiveness (AHR). Furthermore, the clock gene, NR1D1, regulates time of day changes in AHR.2 Since the molecular clock in the airway epithelium is also the pacemaker for the lung, we are interested to discover how allergen challenge regulates rhythmic downstream inflammatory processes in asthma (AHR). However, whether targeting this molecular clock with its agonist (SR9009) can attenuate House Dust Mite (HDM)-induced inflammation and pathophysiological changes remain unknow.We employed a conditional NR1D1 reporter BAC construct with luciferase under the NR1D1 gene’s regulatory control, crossed with CCSP-iCre mice to generate NR1D1-STOPluc+/− (Cre+/−) animals (figure 1A). Mice were exposed to HDM allergen intranasally three times weekly for five weeks while a subset received the NR1D1/Rev-erba agonist SR9009 (i.p, at ZT6/1 PM) every day in the last 2 weeks of the HDM administration while another subset received vehicle alone. These mice were subjected to bioluminescence imaging performed every four hours post-luciferin injection, utilizing an IVIS imaging system. Precision-cut lung slices were also prepared, imaged continuously over 72 hours to assess bioluminescence in response to allergen exposure.Abstract P245 Figure 1The protective effect of Rev-erba agonist SR9009 against allergic airways disease. (A) schematic diagram of the generation of NR1D1/Rev-erba StopLuc in CCSP- expressing airway epithelial cells hence, NR1D1-STOPluc:CCSP-iCre±(Cre±) reporter mice. (B) bioluminescence imaging of NR1D1- STOPluc:CCSP-iCre±mice demonstrating dampened expression of NR1D1 with HDM allergen challenge in comparison to vehicle PBS but prevented with SR9009. (C) SR9009 prevents eosinophilia and influx of inflammatory cells in the lungs. (D) SR9009 prevented airway remodelling associated with T2 asthmaWe propose that the Rev-erba agonist SR9009, will protect against HDM induced airway inflammation and associated pathologyThe results indicate a diurnal pattern in NR1D1 expression in NR1D1-STOPluc mice, peaking at ZT12 in non-allergic animals and showing significant reductions in HDM-treated mice (p<0.001 at ZT12, figure 1B). Administration of SR9009 to HDM challenged animals recovered NR1D1 expression and concomitantly reduced HDM-associated eosinophilia (figure 1C). SR9009 also reduced pathological changes induced by HDM such as epithelial thickness and subepithelial collagen (figure 1D,E). The result demonstrates the therapeutic potential of SR9009 and its protective role in T2 asthma.References Durrington HJ, et al. AJRCCM. 2018;198(12):1578–1581.Chakraborty A, et al. European Respiratory Journal 2023;62(suppl 67):OA4208.