Abstract 6722: OFD1 as a novel biomarker in cholangiocarcinoma: insights into ciliary dysfunction and tumor growth

Abstract Cholangiocarcinoma (CCA) is a rare and aggressive solid tumor originating from cholangiocytes lining the biliary ducts. It is the second most common primary hepatic malignancy, with an increasing global incidence and limited therapeutic options. Recent studies suggest that the primary cilium acts as a tumor suppressor in the biliary tract by regulating key cancer-related signaling pathways. Moreover, evidence links the reduction or loss of primary cilia to increased aggressiveness in various human cancers. Here, we aimed to identify and functionally characterize primary cilia-associated genes with potential translational relevance for CCA prognosis and treatment. Our study included 120 CCA patients, comprising 51% (36) with intrahepatic CCA and 49% (34) with extrahepatic CCA. Whole-transcriptome and whole-exome profiling were conducted on laser micro-dissected formalin-fixed paraffin-embedded (FFPE) tissue specimens. Bioinformatic analysis identified OFD1 as the top-ranked cilium-associated gene (fold-change ≥ 2.5, p < 0.01) among differentially expressed genes in tumor, stromal, and normal tissues. We investigated the role of OFD1 in cilia formation and cell proliferation through in vitro experiments in healthy human cholangiocytes and CCA cell lines. Immunofluorescence staining with ciliary markers revealed that OFD1 silencing promoted ciliogenesis and inhibited tumor growth in CCA cell lines. Western blot analysis confirmed a significant upregulation of ciliary proteins, including PCM1, ARL13B, and acetylated tubulin, following OFD1 silencing. Furthermore, CRISPR/Cas9-mediated OFD1 depletion in CCA cells led to a marked reduction in cell proliferation and restored ciliogenesis. Conversely, OFD1 overexpression in normal cholangiocytes resulted in increased cell proliferation and migration, along with suppression of ciliogenesis. Clinically, higher OFD1 expression was significantly associated with poorer overall survival in CCA patients (10 vs. 39 months; HR 2.33, 95% CI 1.01-5.31, p = 0.03). In conclusion, we demonstrated that OFD1 functions as an oncogene in CCA, influencing primary cilia formation and tumor cell growth. Higher OFD1 expression is a significant predictor of worse survival outcomes, identifying OFD1 as a promising biomarker and potential therapeutic target in CCA. Citation Format: Fabiola Piscopo, Anna Barbato, Massimiliano Salati, Carla Pollastro, Giorgia Gitano, Alberto Marino, Lorenzo Evangelista, Luigi Ferrante, Antonella Iuliano, Luca Reggiani-Bonetti, Maria Salatiello, Pasquale Pisapia, Umberto Malapelle, Giancarlo Troncone, Massimo Dominici, Brunella Franco, Pietro Carotenuto. OFD1 as a novel biomarker in cholangiocarcinoma: insights into ciliary dysfunction and tumor growth [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 6722.