P-cresol sulfate (PCS) is a microbial metabolite derived from L-tyrosine and was recently discovered to have immunoregulatory influences on allergic airway inflammation. Administering PCS to mice reduced house dust mite-induced CCL20 production, a chemokine that recruits lymphocytes and dendritic cells (Wypych et al. Nature Immunology 2021). We are using PCS as a chemical template to develop novel therapeutics against allergic asthma. Our aim is to determine the molecular mechanism of action of PCS and its chemical derivatives in alleviating allergic airway inflammation. We isolated and stimulated primary mouse lung cells with lipopolysaccharide, a strong inducer of CCL20, identifying airway epithelial cells as the main cell type affected by PCS. RNA sequencing of ex vivo mouse lung epithelial cells revealed that PCS influenced heat shock protein 90 (HSP90) gene expression, and indeed, blockade of HSP90 reduced CCL20 production, suggesting it may be involved in the mechanism of action. In silico molecular modelling indicated a shared putative binding site for PCS and two chemical derivatives in the epidermal growth factor receptor (EGFR). Additionally, RNA-sequencing of the A549 human alveolar epithelial cell line identified an increase in genes regulated by the aryl hydrocarbon receptor (AHR) in PCS and chemical derivative treated cells. These results implicate EGFR and AHR in mediating the effects of PCS and its chemical derivatives. Overall, PCS acts on lung epithelial cells to reduce CCL20 production and consequently allergic airway inflammation. Elucidation of the molecule?s mechanism of action could lead to development of novel therapeutics against allergic asthma and other atopic diseases.