CARD9 promotes cholangiocarcinoma by regulating the IL-17A/Hedgehog and the THEM4/AKT/mTOR signaling pathways

Cholangiocarcinoma (CCA) is a highly lethal malignant tumor originating from the bile duct, and its underlying mechanisms are not fully understood. In order to identify key genes in CCA, we downloaded gene expression data from public GSE76297, GSE26566 and TCGA-CHOL datasets. CARD9 was selected as a CCA-related gene from the datasets. Its expression was identified in the CCA tissues via PCR, western blot and immunohistochemistry assays. The loss-and-gain function assay of CARD9 was conducted in CCA cell lines (QBC939 and RBE) and nude mice. This study found a significant upregulation of CARD9 in CCA tissues, which is associated with poor prognosis in patients. Overexpression of CARD9 promotes proliferation and invasion of QBC939 and RBE cells, enhances the growth and development in CCA mouse models, and reduces sensitivity to chemotherapy drugs for CCA. Mechanistically, CARD9 activates the NF-κB and NLRP3 inflammatory signaling pathways, induces excessive release of various inflammatory factors, and triggers a cascade reaction of interleukin-17 (IL-17A). IL-17A promotes the stability of IHH mRNA by regulating HuR, enhances IHH transcription, and activates the Hedgehog signaling pathway to accelerate the progression of CCA. In addition, CARD9 promotes proliferation and invasion of CCA cells by interacting with THEM4, thereby facilitating AKT and mTOR phosphorylation, and accelerating the progression of CCA. Overall, these data suggest that CARD9 is involved in the progression of CCA by regulating the IL-17A/Hedgehog and the THEM4/AKT/mTOR signaling pathways. Therefore, CARD9 may serve as a novel therapeutic target for CCA treatment.