Multiomic Characterization of Treatment Response to JAK Inhibition in Patients with APECED

Background Autoimmune polyendocrinopathy-candidiasis-ectodermal dystrophy (APECED) is an inborn error of immunity caused by loss-of-function of the AIRE gene. Lack of AIRE impairs thymic negative selection of T cells, causing multiorgan autoimmunity, making APECED a prototypical disease for failed central tolerance. IFNγ secreted by T cells drives inflammation in APECED, and targeting IFNγ downstream signaling with JAK1/2 inhibitor ruxolitinib remits autoimmunity in the patients. How JAK inhibition affects tissue-specific responses at the single-cell level remains unclear. Methods In longitudinal samples from four APECED patients treated with ruxolitinib on a phase-II clinical trial at National Institutes of Health, we combined single-cell RNA sequencing (scRNAseq) from blood, enteritis-affected duodenum, and scalp skin affected by alopecia with serum proteome, autoantibody microarray, and metabolome to characterize tissue-specific autoimmune responses and their normalization upon JAK inhibition. Results In tissues affected by autoimmunity, we detected multifaceted cross talk between immune and stromal cells, ushering IFNγ-driven inflammation. Ruxolitinib downregulated expression of tissue-homing chemokines and antigen presentation by stromal cells and resulted in decreased accumulation of immune cells to the tissues, as well as dampening of pro-inflammatory responses. Normalization of tissue homeostasis ensued; for example, hair keratinocytes became detectable, aligning with clinical hair regrowth. Many of the transcriptional tissue changes were discernible by the serum proteome, and the treatment response was apparent in rejuvenation of proteome-based organ age. However, treatment was also associated with subclinical proteomic changes reflecting potential side effects of JAK inhibitors, including anemia, weight gain, and hypercoagulability. We expanded the proteome evaluation to 89 additional APECED patients affected by autoimmunity, identifying a patient cluster with the most severe clinical phenotype, highest autoantibody burden, and proteome-based premature aging of multiple organs. These patients had a distinct inflammatory profile, which, in addition to excess IFNγ, was characterized by heightened IL6 responses and TGFβ resistance, partly driven by alterations in sex hormone production. Conclusions Multiomic characterization of treatment response to JAK inhibition uncovers pathogenesis and normalization of tissue-specific autoimmunity. JAK inhibitors are increasingly used to target various immune pathways in inborn errors of immunity, and understanding their complex effects systemically and in the tissues is a prerequisite for safe and successful therapy.