Cholangiocyte Pathobiology in Cholestatic Liver Disease

PROJECT SUMMARY/ABSTRACT Primary Sclerosing Cholangitis (PSC) affects ~40,000 Americans, accounts for 6% of liver transplants, and has a median survival of ~15 years. Despite this clinical burden, there are currently no agency-approved pharma- cotherapies, underscoring a critical gap in our understanding of PSC pathogenesis and the lack of newly vali- dated, druggable targets. Thus, our OVERALL OBJECTIVE is to clarify the pathogenesis of PSC and identify new therapies. Cholangiocyte senescence is a feature of PSC, informing its pathogenesis and revealing new therapeutic targets. Experiments proposed here extend this observation and address the novel concept that PSC is a disease in which cholangiocytes display features of viral infection (i.e., an interferon response) in the absence of virus (viral mimicry), driving sterile fibroinflammation. Our preliminary data show that: (i) PSC cholangiocytes exhibit a Type-I interferon (IFN) response characterized by antiviral and proinflammatory gene expression; (ii) the IFN response correlates with derepression of human endogenous retroviruses (HERVs) [remnants of retro- viral infections comprising ~8% of the human genome]; experimentally induced expression of an HERV RNA element triggers cholangiocyte senescence and an IFN response; (iii) RIG-like receptors (RLRs, RIG-I and MDA5), known to bind viral and endogenous double-stranded (ds)RNA, are upregulated in PSC cholangiocytes, directly interact with HERV RNAs, and initiate antiviral signaling through mitochondrial antiviral signaling protein (MAVS); (iv) MAVS promotes both cholangiocyte proinflammatory gene expression and macrophage migra- tion/activation; MAVS knockout mice showed reduced hepatic IFN response gene expression in a model of PSC; and v) pharmacological inhibition of receptor-interacting serine/threonine protein kinase 1 (RIPK1), downstream of MAVS, suppresses the IFN response. The data support our CENTRAL HYPOTHESIS that in PSC, cholangi- ocyte HERV activation establishes persistent viral mimicry promoting a chronic antiviral response driving proin- flammatory gene expression and immune cell dysfunction. Our integrated approach tests 3 SPECIFIC AIMS: In Aim 1 we will determine the mechanisms of cholangiocyte immunostimulatory HERV expression. We will assess: i) how epigenetic derepression drives aberrant HERV expression, and ii) how HERVs interact with RIG- like receptors and drive MAVS aggregation. In Aim 2 we will assess the mechanisms and consequences of the cholangiocyte antiviral response. We will answer: i) how MAVS promotes RIPK1 activation and the antiviral IFN and proinflammatory response, and ii) how genetic targeting of RIPK1 influences the cholangiocyte antiviral/fi- broinflammatory response. In Aim 3 we will define the mechanisms and pathologic outcomes of cholangiocyte sterile inflammation and assess therapeutic approaches. We will determine: i) how the cholangiocyte RIPK1- mediated proinflammatory secretome drives macrophage activation and migration, and ii) how pharmacologic interventions targeting RIPK1 modify fibroinflammation in mouse models of PSC. The results of our experiments will provide novel pathophysiologic insights and potentially new druggable targets.