Abstract 5588: Efficacy of TGR-5 inhibition in cholangiocarcinoma using the novel small molecule inhibitor SBI-364

Abstract Background: Cholangiocarcinoma (CCA) is a malignancy of the biliary epithelium with increasing incidence and poor survival. Despite the advances in targeted therapies, treatment success has been hindered by primary or acquired resistance. New therapeutic strategies are necessary to improve outcomes in this patient population. This study investigates the anti-tumor efficacy of TGR-5 inhibition in CCA using a novel small molecular inhibitor, SBI-364. Methods: Cell viability was assessed using a CellTiter-Glo assay in four unique CCA cell lines treated with SBI-364 at concentrations ranging from 1 nM to 100 µM for 48 hours. Viability was expressed as a percentage relative to untreated controls. Cell migration was assessed using a scratch wound healing assay at 0, 12, and 24 hours following SBI-364 treatment. Migration was reported as the percentage of wound closure relative to the initial scratch width. Apoptosis was assessed using the Caspase-Glo® 3/7 Assay (Promega) after 12 hours of treatment with SBI-364. Caspase activity was normalized to untreated controls and expressed as relative luminescence units. Results: SBI-364 reduced cell viability in a dose-dependent manner across all CCA cell lines with IC50 values ranging from 4.1-15.8 µM. Higher concentrations of SBI-364 induced increased cytotoxicity. In the scratch assay, untreated CCA cells achieved 100 % wound closure within 24 hours. Treatment with SBI-364 at 10 µM significantly inhibited cellular migration, with reduced wound closure compared to controls (p < 0.05). Apoptosis analysis revealed a significant increase in caspase-3/7 activity in three CCA cell lines following treatment with 10 µM SBI-364 for 12 hours (p < 0.01). Conclusion: TGR-5 inhibition using novel small molecule inhibitor SBI-364 demonstrates potent anti-tumor effects in CCA by reducing viability, inhibiting migration, and promoting apoptosis. These findings support further investigation of SBI-364 as a promising therapeutic strategy in CCA. Citation Format: Enis Ozmert, Jennifer L. Tomlinson, Danielle M. Carlson, Rory L. Smoot. Efficacy of TGR-5 inhibition in cholangiocarcinoma using the novel small molecule inhibitor SBI-364 [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 5588.