Beyond FGFR2-rearrangement: a study on the molecular landscape of intrahepatic cholangiocarcinoma

Background. Intrahepatic cholangiocarcinoma (iCCA) is a rare malignancy arising in the epithelium of the biliary tract and characterized by a dismal prognosis. Molecular profiling has revealed two major somatic alterations that characterize iCCA: point mutations in the IDH1 gene, that occur in 10-20% of cases, while FGFR2 rearrangements characterize the 10-15% of iCCAs. Both are currently druggable with inhibitors of IDH1 or pan-FGFR inhibitors, respectively. Despite this, a considerable proportion of iCCAs currently lack of druggable molecular targets. Thus, the identification of novel predictive biomarkers is fundamental to improve the number of patients that could benefit from precision therapy. Aim. The objective of the study was to employ a comprehensive genomic profiling analysis to identify novel prognostic and predictive biomarkers, and possibly, to correlate the identified genomic profiles with the iCCA’s histo-morphological features. Materials and methods. A total of 120 formalin-fixed paraffin-embedded (FFPE) samples were collected from patients diagnosed with iCCA and referred to the Istituto Oncologico Veneto (IOV). All cases were characterized for their histological features, including histotype, according the 2019-WHO classification, differentiation and growth pattern. The genomic profile was analyzed using the AVENIO Tumor Tissue CGP Kit V2, a DNA-based NGS panel capable of studying genomic alterations in 335 cancer-driving genes. The panel investigates single-nucleotide variants (SNVs), insertions and deletions (indels), rearrangements, and copy number aberrations (CNAs), in addition to microsatellite instability (MSI), tumor mutational burden (TMB), and homologous recombination deficiency (HRD). When possible, the identified alterations were validated with orthogonal techniques. Results. In addition to FGFR2 rearrangements and IDH1/IDH2 point mutations, which have previously been identified in the context of iCCA, other potentially druggable alterations were identified. The loss of one or more genes located on chromosome 9p21 was reported in approximately one-third of iCCA cases, with the CDKN2A, CDKN2B, and MTAP genes, located in close proximity, being lost in 33.7%, 28.7%, and 20% of cases, respectively. In all cases, the presence of MTAP deletion was concomitant with the loss of CDKN2A. Other frequent mutations involved BAP1 (17.5%), ARID1A (13.7%), TP53 (13.7%), PBRM1 (6.2%), and PIK3CA (6.2%). Two cases were found to be MSI (confirmed in IHC, where MLH1 loss was detected) and also characterized by a TMB-high. Additionally, three cases presented alterations in the homologous recombination system (HRD). Conclusions. The molecular characterization of iCCA with a CGP approach reveals that other potential predictive and prognostic targets can be identified in iCCA. For example, in other settings, the loss of the 9p21 locus and in particular the concomitant CDKN2A/MTAP deletion has been associated with poor survival and resistance to immunotherapy, due to the establishment of an immune-cold tumor microenvironment. At the same time, preliminary data reported that MTAP-deleted tumors are more sensitive to MAT2A inhibitor drugs. Also, PIK3CA mutations, despite less frequent, represent potentially druggable targets, together with HRD iCCAs