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Massive parallel sequencing uncovers actionable FGFR2-PPHLN1 fusion and ARAF mutations in intrahepatic cholangiocarcinoma

2015-01-29 00:03:16

Nature Communications; 22 January 2015 ; DOI: 10.1038/ncomms7087



Daniela Sia, Bojan Losic, Agrin Moeini, Laia Cabellos, Ke Hao, Kate Revill, Dennis Bonal, Oriana Miltiadous, Zhongyang Zhang, Yujin Hoshida, Helena Cornella, Mireia Castillo-Martin, Roser Pinyol, Yumi Kasai, Sasan Roayaie, Swan N. Thung, Josep Fuster, Myron E. Schwartz, Samuel Waxman, Carlos Cordon-Cardo, Eric Schadt, Vincenzo Mazzaferro, Josep M. Llovet



Abstract



Intrahepatic cholangiocarcinoma (iCCA) is a fatal bile duct cancer with dismal prognosis and limited therapeutic options. By performing RNA- and exome-sequencing analyses, we report a novel fusion event, FGFR2-PPHLN1 (16%), and damaging mutations in the ARAF oncogene (11%). Here we demonstrate that the chromosomal translocation t(10;12)(q26;q12) leading to FGFR2-PPHLN1 fusion possesses transforming and oncogenic activity, which is successfully inhibited by a selective FGFR2 inhibitor in vitro. Among the ARAF mutations, N217I and G322S lead to activation of the pathway and N217I shows oncogenic potential in vitro. Screening of a cohort of 107 iCCA patients reveals that FGFR2 fusions represent the most recurrent targetable alteration (45%, 17/107), while they are rarely present in other primary liver tumours (0/100 of hepatocellular carcinoma (HCC); 1/21 of mixed iCCA-HCC). Taken together, around 70% of iCCA patients harbour at least one actionable molecular alteration (FGFR2 fusions, IDH1/2, ARAF, KRAS, BRAF and FGF19) that is amenable for therapeutic targeting.



Introduction



Intrahepatic cholangiocarcinoma (iCCA) is an aggressive malignancy with a 5-year survival rate less than 10%. Surgery is the only curative option for iCCA, although it is limited to patients with early-stage disease2. The majority of iCCA patients are diagnosed at more advanced stages, where there is no accepted standard of care. Therefore, there is an unmet need to develop a first-line therapy for these patients.



A deeper knowledge of the molecular mechanisms underlying this disease is crucial for the development of new effective targeted therapies. Of late, there has been a significant improvement in our understanding of the molecular basis of iCCA. Large-scale molecular profiling studies have enabled the first molecular classifications to emerge. Furthermore, deep sequencing studies have provided a preliminary description of somatic mutations for iCCA: novel mutations in chromatin remodeling genes (BAP1, ARID1A and PBRM1) have been recently unveiled, whereas frequent mutations in KRAS, IDH1 and IDH2 have been confirmed. Nonetheless, none of the targetable mutations have been explored yet in early clinical trials.



In addition to somatic mutations, somatic gene fusions are able to drive the development of human cancers, though their translational relevance has been mostly limited to haematological
malignancies. The recent discovery of novel fusion events associated with different types of solid tumours, such as prostate, lung and breast cancer has increased the interest in these genetic alterations. In fact, one of such fusions (the EML–ALK fusion in lung cancer) has emerged as a druggable target and its inhibition leads to survival improvements. Interestingly, a variety of FGFR2 gene fusions have been recently identified in iCCA, suggesting that these events may represent novel candidate therapeutic targets and that similar strategies could be used for its clinical management.



Massive parallel sequencing technology allows the characterization of cellular transcriptomes and genomes at single-base resolution, including the detection of somatic gene mutations and
intragenic fusions that may lead to oncogenic molecular pathway activation. To uncover candidate oncogenes that may represent novel targets for iCCA therapy, we profile a cohort of 122 iCCA
cases by performing RNA and DNA sequencing (discovery set: seven and eight paired iCCA samples analysed, respectively; screening cohort: 114 iCCA tissues). We discover a novel recurrent oncogenic fusion gene, FGFR2–PPHLN1 (16%) and damaging mutations in the oncogene ARAF (11%). The screening of a large iCCA cohort reveals that around 70% of the tumours harbour at least one targetable molecular alteration (for example, FGFR2 fusions, KRAS/BRAF/EGFR/IDH mutations) with ~45% of patients analysed positive for at least one FGFR2 rearrangement. Significantly, the transforming and oncogenic activity of the FGFR2–PPHLN1 fusion can be successfully inhibited by a selective FGFR2 inhibitor (BGJ398) in vitro. Together our work unveils a large fraction of iCCA patients with specific targetable molecular alterations and reveals FGFR2 rearrangements as the most recurrent molecular alteration event reported so far in this disease.



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Key Words



Biological sciences | Cancer | Genetics