Modulation of biliary cancer chemo-resistance through microRNA-mediated rewiring of the expansion of CD133+ cells

Pietro Carotenuto, Somaieh Hedayat, Matteo Fassan, Vincenzo Cardinale, Andrea Lampis, Vincenza Guzzardo, Caterina Vicentini, Aldo Scarpa, Luciano Cascione, Daniele Costantini, Guido Carpino, Domenico Alvaro, Michele Ghidini, Francesco Trevisani, Robert Te Poele, Massimiliano Salati, Sofia Ventura, Georgios Vlachogiannis, Jens C Hahne, Luke BoulterStuart J Forbes, Rachel Guest, Umberto Cillo, Ian Said-Huntingford, Ruwaida Begum, Elizabeth Smyth, Vasiliki Michalarea, David Cunningham, Lorenza Rimassa, Armando Santoro, Massimo Roncalli, Vladimir Kirnkin, Paul Clarke, Paul Workman, Nicola Valeri, Chiara Braconi

Research output: Contribution to journalArticlepeer-review

Abstract / Description of output

BACKGROUND AND AIMS: Changes in single microRNA (MIR) expression have been associated with chemo-resistance in Biliary Tract Cancer (BTC). However, a global assessment of the dynamic role of the microRNome has never been performed to identify potential therapeutic targets that are functionally relevant in the BTC cell response to chemotherapy. APPROACH AND RESULTS: high-throughput-screening of 997 LNA-MIR-inhibitors was performed in 6 CCA cell lines treated with Cisplatin-Gemcitabine (CG) seeking changes in cell viability. Validation experiments were performed with miRvana probes. MIR and gene expression was assessed by TaqMan-assay, RNA-sequencing and in-situ-hybridization in 4 indepedent cohorts of human BTC. Knock-out of microRNA was achieved by CRISPR-CAS9 in CCLP cells (MIR1249KO) and tested for effects on chemotherapy sensitivity in-vitro and in-vivo. High-throughput-screening revealed that MIR1249-inhibition enhanced chemotherapy sensitivity across all cell lines. MIR1249 expression was increased in 41% of cases in human BTC. In validation experiments, MIR1249-inhibition did not alter cell viability in untreated or DMSO-treated cells; however it did increase CG effect. MIR1249 expression was increased in CD133+ biliary cancer cells freshly isolated from the stem niche of human BTC, as well as in CD133+ chemo-resistant CCLP cells. MIR1249 modulated the chemotherapy-induced enrichment of CD133+ cells by controlling their clonal expansion via the Wnt-regulator FZD8. MIR1249KO cells had impaired expansion of the CD133+ subclone and its enrichment after chemotherapy, reduced expression of Cancer-Stem-Cell markers, and increased chemosensitivity. MIR1249KO xenograft BTC models showed tumour shrinkage after exposure to weekly CG, while WT models showed only stable disease over treatment.

CONCLUSIONS: MIR1249 mediates resistance to CG in BTC and may be tested as a novel target for therapeutics.

Original languageEnglish
Early online date26 Dec 2019
Publication statusE-pub ahead of print - 26 Dec 2019


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