Patient-derived liver organoids as an in vitro model to study new personalized therapies targeting VDAC1 in intrahepatic cholangiocarcinoma

Introduction and Aim Intrahepatic cholangiocarcinoma (iCCA) is characterized by a very poor outcome, and reliable biomarkers as well as new therapeutic strategies are urgently needed. As a main actor in the regulation of mitochondria-mediated cell death and survival signaling pathways, Voltage Dependence Anion Selective Channel isoform 1 (VDAC1) became an attractive pharmacologic target. Many molecules have been conceived, however, due to promiscuity and side effects, none of them have been extensively used to treat patients. The aim of this study was to test a new class of small molecules targeting VDAC1 to induce activation of the apoptotic pathway in iCCA patient-derived liver cells and organoids. Methods To generate organoids, we minced tumor and paired non-tumor biopsies and shortly digested in small cell clusters that are seeded into Matrigel. After characterization using immunofluorescence and qPCR techniques, we treated primary cell cultures and organoids with different concentrations of small molecules targeting VDAC1, monitoring cell viability and ROS production, to verify the in vitro effects and the efficiency of these compounds on cells. Results We developed and established a biobank of human iCCA-derived organoids, evaluating the morphological characteristics and assessing a mathematical tool to model tumor growth. In addition, we analysed the presence of typical CCA markers (EpCAM, CK19, CK7, E-Cadherin, Ki67). We also investigated VDAC1 expression underlying higher levels in iCCA cells in comparison with non-tumor cells (p<0.005). We subsequently examined the efficiency of new small molecules targeting VDAC1, at different time points and concentrations, both in patient-derived cell cultures and organoids. In particular, we showed a significant decrease in viability in tumor cells only and a modulation in ROS production. Conclusion We developed and characterized a well-defined iCCA in vitro model that allowed us to investigate the effect of small molecules targeting VDAC1 as a new personalized therapy.