Oncogenic dependency on β-catenin in liver cancer cell lines correlates with pathway activation

// Zhihu Ding 1 , Chaomei Shi 1 , Lan Jiang 1 , Tatiana Tolstykh 1 , Hui Cao 1 , Dinesh S. Bangari 2 , Susan Ryan 2 , Mikhail Levit 3 , Taiguang Jin 4 , Karl Mamaat 1, 8 , Qunyan Yu 1 , Hui Qu 1 , Joern Hopke 5 , May Cindhuchao 5 , Dietmar Hoffmann 5 , Fangxian Sun 1 , Mike W. Helms 6 , Kerstin Jahn-Hofmann 6 , Sabine Scheidler 6 , Liang Schweizer 4, 9 , Douglas D. Fang 7, 10 , Jack Pollard 1 , Christopher Winter 1 and Dmitri Wiederschain 1 1 Sanofi Oncology Therapeutic Area, Cambridge, MA, USA 2 Sanofi Translational In Vivo Models, Framingham, MA, USA 3 Sanofi Translational Sciences, Cambridge, MA, USA 4 Sanofi Asia Pacific R&D Hub, Shanghai, People’s Republic of China 5 Sanofi Biologics Research/Molecular Screening Technology, Cambridge, MA, USA 6 Sanofi Biologics Research/Nucleic Acid Therapeutics, Frankfurt am Main, Germany 7 Discovery Services, WuXi AppTec Co., Shanghai, China 8 Current address: Leica Biosystems, Boston, MA 9 Current address: Harbour BioMed, Shanghai, People’s Republic of China 10 Current address: Ascentage Pharma Group, Ltd., Suzhou, People’s Republic of China Correspondence to: Dmitri Wiederschain, email: Dmitri.Wiederschain@sanofi.com Keywords: hepatocellular carcinoma, β-catenin, cell proliferation, siRNA, phenotypic rescue Received: August 16, 2017      Accepted: September 13, 2017      Published: September 28, 2017 ABSTRACT Hepatocellular carcinoma (HCC) represents a serious public health challenge with few therapeutic options available to cancer patients.Wnt/β-catenin pathway is thought to play a significant role in HCC pathogenesis. In this study, we confirmed high frequency of CTNNB1 (β-catenin) mutations in two independent cohorts of HCC patients and demonstrated significant upregulation of β-catenin protein in the overwhelming majority of HCC patient samples, patient-derived xenografts (PDX) and established cell lines. Using genetic tools validated for target specificity through phenotypic rescue experiments, we went on to investigate oncogenic dependency on β-catenin in an extensive collection of human HCC cells lines. Our results demonstrate that dependency on β-catenin generally tracks with its activation status. HCC cell lines that harbored activating mutations in CTNNB1 or displayed elevated levels of non-phosphorylated (active) β-catenin were significantly more sensitive to β-catenin siRNA treatment than cell lines with wild-type CTNNB1 and lower active β-catenin. Finally, significant therapeutic benefit of β-catenin knock-down was demonstrated in established HCC tumor xenografts using doxycycline-inducible shRNA system. β-catenin downregulation and tumor growth inhibition was associated with reduction in AXIN2, direct transcriptional target of β-catenin, and decreased cancer cell proliferation as measured by Ki67 staining. Taken together, our data highlight fundamental importance of aberrant β-catenin signaling in the maintenance of oncogenic phenotype in HCC.