Despite many advancements in the treatment of severe asthma (SA) with the advent of biologics, there still remain challenges because of incomplete understanding of the underlying immune and molecular aberrations. Towards this end, we analyzed the bronchoalveolar lavage (BAL) cells of patients enrolled in the severe asthma research program (SARP) and also of those in the P01-funded IMSA (Immune Mechanisms in Severe Asthma) program. In addition, we studied the transcriptome of these cells using bulk RNA-seq methods. Initial studies of the SARP cohort revealed that the BAL cells in 35-40% of SA patients harbor a high Type 1 (T1)/IFN-g immune signature and implicated IFN-g-activated STAT1 in steroid insensitivity due to co-operation with the glucocorticoid receptor (GR). Subjecting the BAL cells of 41 subjects in the IMSA cohort to mass cytometry/CyTOF and downstream bioinformatic analysis allowed clustering of the SA patients into two groups (PGs), PG2 and PG3, displaying distinct immune profiles. The PG2 group showed a heightened innate immune response dominated by FceRI+IL-4+ cells while PG3 was a lymphocyte-dominated group characterized by increased numbers of IFN-g+ CD4+ and CD8+ T cells, which included tissue-resident memory (TRM) cells. The immune cells were associated with gene modules by deconvolution of their transcriptomic data using our novel algorithm, ICLite. Module-associated gene expression suggests importance of FceRI- and IL-7-signaling in cytokine production by innate cells in PG2 and of co-stimulatory molecules in positive regulation of IFN-g production from TRMs in PG3. In the clinic, response to treatment of SA patients with the biologic, dupilumab, also highlights heterogeneity despite similar biomarker profiles of the treated patients. Thus, given the novel insight gained about distinct immune phenotypes of SA patients with specific transcriptomic signatures, we now have the opportunity to determine in fine resolution, immunologically and molecularly, how dupilumab differentially impacts the local immune (Proj. 1) and epithelial (Proj. 2) phenotypes in patients who show similar biomarker profiles and yet do not respond similarly to treatment. Collectively, these data allow us to hypothesize that two distinct immune mechanisms, one regulated by FceRI+ innate immune cells in conjunction with airway epithelial cells, and the second by T cells, with their impact on epithelial cells, collectively determines SA. To address this hypothesis, the specific aims of Project 1 are to: Aim 1. Characterize airway immune cells in high-resolution by CITE-seq and TCR-seq and determine impact of dupilumab on immune phenotype. Aim 2. Determine mechanisms that induce innate immune cell-driven SA phenotype using a novel fungal protease-driven model. Aim 3. Determine mechanisms that regulate a T1high adaptive immune response using an established HDM+cyclic dinucleotide-driven model. Collectively, the findings will qualify heterogeneity in immune res...