ABSTRACT Liver cancer is the most rapidly increasing cancer in the United States. Hepatocellular carcinoma (HCC), the most common type of primary liver cancer with limited treatment options, accounts for about 75% of all liver cancer and 30,000 deaths per year. Despite recent FDA approval of several new drugs for the treatment of advanced liver cancer, five-year survival rate for HCC patients still remains to be around 20 percent, highlighting the need to develop better therapeutic options. Aberrant expression of Fibroblast Growth Factor 19 (FGF19) has been recently identified as an oncogenic driver in a subset of HCC patients. FGF19 is an ileum-derived enterokine that belongs to endocrine Fibroblast Growth Factor (FGF) family, which normally functions as an endocrine hormone and signals though FGF receptor (FGFR) pathway in the presence of obligate receptor, β- Klotho. Overexpression of FGF19 in liver, however, promotes uncontrolled proliferation through FGF19-FGFR4- β-Klotho pathway in an autocrine-paracrine manner. Our previous studies revealed that β-Klotho recognizes FGF19 in the same manner as it recognizes FGF21, another member of endocrine FGF family with no mitogenic properties, and that the molecular interactions between β-Klotho and FGF21 or FGF19 occur at the identical interface. Based on our mechanistic understanding of cellular signaling by FGF19 and FGF21, we hypothesize that FGF21 variants with enhanced binding affinities to β-Klotho will be able to effectively compete with FGF19 and consequently inhibit aberrant FGF19 signaling in cancers. The aims of this proposal seek to address this overall hypothesis by (1) performing in-depth analyses of FGF21 variants using biophysical, biochemical, and structural studies, (2) testing their capacity to inhibit FGF19 signaling and block proliferation of FGF19-driven cancers, and (3) examining the abilities of FGF21 variants to overcome/prevent the resistance to kinase inhibitors currently in clinical trials. The outcome of our studies will provide deeper mechanistic and biological understanding of FGF19 signaling in cancers and offer novel means to prevent and treat FGF19-driven cancers.