Investigation of GLI1 selective inhibitor for targeted therapy in Intrahepatic Cholangiocarcinoma: a step towards precision medicine

Background and Aims: Intrahepatic cholangiocarcinoma (iCCA) is a biliary tract cancer, carrying aggressive nature and poor prognosis, with a 5-year survival rate <20%. The incidence and mortality of iCCA have escalated globally, due to late diagnoses, suboptimal chemotherapy responses, and a scarcity of targeted treatment options. A critical challenge in managing iCCA is its high inter- and intratumoral heterogeneity, which complicates biomarker discovery and the development of effective therapeutic strategies, impacting patient outcomes. World Health Organization (ICD-O-3.2) has endorsed the differentiation of iCCA into small and large bile duct types based on its molecular, histological, and clinical features. However, the relationship between molecular characteristics and specific iCCA subtypes remains unexplored, representing a significant gap in translational research. The Hedgehog (Hh) pathway plays a fundamental role in the onset and advancement of iCCA, influencing tumor growth, viability, cancer stem cells activation, and epithelial-mesenchymal transition. This research project seeks to assess the effects of a new natural compound known as Glabrescione B (GlaB) on Gli1 inhibition in primary and established cell lines in vitro, targeting the most important transcription factor and final effector in the Hh pathway. Method: The response of GlaB has been assessed by Trypan Blue Exclusion test. MTS assay was conducted to ascertain the IC50 value. Western blot has been utilized for analyzing the target protein levels. Wound-healing assay has been used to evaluate the ability of cells to migrate. Colony formation assay has been conducted to assess the ability of cancer cells to form colonies. Flow cytometry analysis has been executed to examine cell death induction. The findings were validated in a minimum of three separate experiments and the quantitative data has been presented as the mean±SEM. Student's t-tests were used to evaluate variances between two groups in paired samples. A p-value lower than 0.05 was considered as statistically significant (*p<0.05;**p<0.01;***p<0.001). Results: Our results show that treating iCCA cells with either free GlaB or HA-GlaB results in a significant decrease in cell proliferation, viability, migration, colony-forming ability, and Gli1 protein levels in a dose- and time-dependent manner (0.05<0.001). Moreover, the drug is able to induce iCCA cell death after the administration, compared to the controls. Conclusion: A new natural compound, called Glabrescione B, has the ability to hinder the growth, survival, invasiveness of iCCA cells and trigger their death. Collectively, this information offers understanding of a novel and possibly advantageous natural chemical compound for treating iCCA in vivo.