Chronic liver diseases (CLDs) including cirrhosis of any etiology are the major risk factors for the development of hepatocellular carcinoma (HCC). The national prevalence and mortality due to HCC in the US VA healthcare systems have increased by 3-fold during the last decade. Despite HBV vaccination, improved screening methods, and successful antiviral treatment for HCV, the incidence of CLD and cirrhosis has substantially (13%) increased since 2000. The existing treatment options (radiation, chemotherapy, and immunotherapy) of unresectable and metastatic HCC have not been successful in the majority of cases. Thus, HCC remains a significant healthcare challenge. The fundamental goal of this proposal is to determine mechanisms driving the transformation process and tumor growth in the liver, and develop strategies to improve clinical outcomes for these patients. In this quest, we found that doublecortin-like kinase 1 (DCLK1) protein is highly expressed in chronic viral hepatitis, cirrhosis, and HCC but not in normal liver. Elevated DCLK1 is associated with decreased survival of HCC patients. Recently, we demonstrated that DCLK1 activates an atypical β-catenin(48 kDa)/TCF4 signaling in hepatoma cells. However, its significance in the CLDs and HCC and DCLK1/β-catenin signaling axis in hepatic cell stemness is completely unknown. Using a mouse model of HCC, we demonstrated that DCLK1 is heavily induced in hepatocytes at sites of injury and likely contributes to the HCC-like tumor growth. Our preliminary data revealed that DCLK1-expressing hepatoma cells can polarize Kupffer cells (KCs) into immunosuppressive M2-like macrophages. IL-10 secreted by M2-like KCs has been reported to promote the selective apoptosis of M1 pro-inflammatory macrophages. Thus, DCLK1-dependent M2 polarization has the potential to promote immunosuppression in the hepatic tumor microenvironment (TME). Furthermore, inhibition of DCLK1 kinase activity resulted in a dramatic reduction in key pro-inflammatory (TNFα, IL-1β) and immunosuppressive IL-10. These observations suggested that DCLK1 is a central regulator of the complex interconnection of fibrogenic, immunoregulatory, and tumorigenic processes in CLDs. Our central hypothesis is that DCLK1 is a key mediator of oncogenic signaling and immune dysregulation in the tissue microenvironment in CLDs, which collectively stimulates the transformation of hepatocytes. We will test this hypothesis with the following interrelated yet independent specific aims. Aim 1 will determine the mechanisms by which DCLK1 mediates immunoregulatory and tumorigenic processes in the liver by introducing deletion and point mutation in DCLK1 and assay for the functional impacts. We will use inhibitors to DCLK1 kinase (DCLK1-IN- 1) and β-catenin/TCF interaction (FH535) to verify DCLK1-regulated clonogenicity, CSC properties, and monocyte/KCs polarization. Aim 2 will determine the effects of DCLK1 kinase inhibition on the DCLK1/β-catenin regulated signalin...