Crosstalk of Immune Dysregulation and Impaired Wound Healing in inherited STAT4-mediated Autoinflammatory Disease - Abstract Disabling pansclerotic morphea (DPM) is a severe, systemic disorder of childhood with debilitating skin, joint, and mucosal symptoms, immune dysregulation and malignancy. DPM is known to be severely recalcitrant to therapy, leading to death within 10 years of diagnosis. Our investigation of the specific contribution of STAT4 to the immunodysregulatory phenotype seen in DPM patients revealed the following key observations: 1) The STAT4 variant is a gain-of-function mutation that leads to impaired wound healing; 2) STAT4 variation leads to an enhanced immunodysregulatory phenotype; 3) persistent phosphorylation of STAT4 results in IL-6 mediated autoinflammation; 4) JAK inhibition upstream can reduce IL-6 and improve wound healing in vitro and in vivo. Based on our preliminary data we propose the CENTRAL HYPOTHESIS that gain-of-function mutations in STAT4 lead to persistent JAK-STAT signaling due to steady STAT4 phosphorylation, erroneous gene transcription, increased cytokine production, persistent systemic inflammation, and subsequently the failure to regulate normal wound healing. To investigate our hypothesis, our proposal has the following SPECIFIC AIMS. FIRST, we will determine the mechanism of STAT4 gain-of-function mediated immune dysregulation in DPM. We will test the hypothesis that enhanced STAT4 activity leads to a failure of normal NK and T cell development and cellular activation due to intrinsic defects in hematopoietic precursors leading to altered differentiation and subsequent lymphocyte exhaustion from systemic inflammation using novel patient- derived induced pluripotent stem cells (iPSCs) to examine development, transcriptomics, proteomics and immune cell function. Human studies will be complemented by in vivo assays using knockin murine progenitor cells, bone marrow cells, and peripheral cells at progressive stages of disease. SECOND, we will dissect the role of enhanced STAT4 signaling in neovascularization and immune cell crosstalk. We will test the hypothesis that gain-of-function mutations in STAT4 lead to upregulation of angiogenic factors that drive neovascularization, life-threatening bleeds, and contribute to poor wound healing. We will characterize the role of STAT4 signaling in endothelial cells and vessel formation by differentiation of patient-derived iPSCs to endothelial cells. We will assess the competency of vessels in vitro, and the role of oxidative stress in perpetuating poor wound healing. We will investigate translational mechanisms targeting JAK/STAT and angiogenic pathways in DPM disease using transcriptomics of iPSC generated vessels. Finally, we will investigate translational mechanisms targeting the JAK/STAT pathway in DPM.
