Project Summary Intrahepatic cholangiocarcinoma (ICC) ranks among the deadliest cancers, with a 5-year survival rate of only 5-15% due to late diagnosis and lack of effective treatment. The discovery that receptor tyrosine kinase Fibroblast Growth Factor Receptor 2 (FGFR2) is genetically activated in >20% of ICCs, most commonly via FGFR2 gene fusions, is leading to a new therapeutic paradigm for a subset of patients with this disease. Indeed, early clinical trials with FGFR inhibitors (FGFRi) show significant anti-tumor efficacy, demonstrating true `oncogene addiction' to FGFR2, analogous to the role of EGFR mutations or ALK fusions in lung cancer. However, as in lung cancer, drug resistance is a vexing challenge: patients treated with BGJ398, the most clinically advanced FGFRi, usually progress within 6 months. In this Project, we aim to address key barriers to improving care of patients with FGFR2+ ICC. We will define mechanisms of resistance to FGFRi's, pr will provide evidence to support both combination strategies as well as the testing of a novel class of FGFR2 inhibitors in the clinic. We propose a multi-pronged strategy for impactful translational research that leverages serial tumor and liquid biopsies from three FGFRi trials, a series of patient-derived xenografts (PDX) and organoids, a genetically engineered mouse model, and novel FGFR2 inhibitors. We will systematically address the key barriers to more effective targeted therapy against FGFR2 in ICC: 1) the mechanism of FGFRi resistance, either due to acquisition of secondary FGFR2 mutations or activation of `bypass' signaling pathways, 2) identification of more effective FGFR2 inhibitors, including those that can target the kinase after it acquires resistance mutations, and 3) defining novel combination therapies to target bypass pathways to counter resistance and prolong initial response. We propose the following Specific Aims: Aim 1. To identify and credential mutations causing FGFRi resistance in ICC. Aim 2. To identify FGFR inhibitors that more effectively target FGFR2. Aim 3. To develop combination strategies to overcome FGFR2-independent resistance mechanisms.