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Combined hepatocellular-cholangiocarcinoma derives from liver progenitor cells and depends on senescence and IL-6 trans-signaling

Published:August 17, 2022DOI:https://doi.org/10.1016/j.jhep.2022.07.029

      Highlights

      • The combined HCC-CCA tumor originates from hepatic progenitors.
      • Hepatic progenitors are not the source of HCC.
      • Combined HCC-CCA initiate on the background of liver inflammation.
      • Combined HCC-CCA development is dependent on IL-6 trans-signaling.
      • Senescence contributes to combined HCC-CCA development through IL-6 secretion.

      Background & Aims

      Primary liver cancers include hepatocellular carcinoma (HCC), intrahepatic cholangiocarcinoma (CCA) and combined HCC-CCA tumors (cHCC-CCA). It has been suggested, but not unequivocally proven, that hepatic progenitor cells (HPCs) can contribute to hepatocarcinogenesis. We aimed to determine whether HPCs contribute to HCC, cHCC-CCA or both types of tumors.

      Methods

      To trace progenitor cells during hepatocarcinogenesis, we generated Mdr2-KO mice that harbor a yellow fluorescent protein (YFP) reporter gene driven by the Foxl1 promoter which is expressed specifically in progenitor cells. These mice (Mdr2-KOFoxl1-CRE;RosaYFP) develop chronic inflammation and HCCs by the age of 14-16 months, followed by cHCC-CCA tumors at the age of 18 months.

      Results

      In this Mdr2-KOFoxl1-CRE;RosaYFP mouse model, liver progenitor cells are the source of cHCC-CCA tumors, but not the source of HCC. Ablating the progenitors, caused reduction of cHCC-CCA tumors but did not affect HCCs. RNA-sequencing revealed enrichment of the IL-6 signaling pathway in cHCC-CCA tumors compared to HCC tumors. Single-cell RNA-sequencing (scRNA-seq) analysis revealed that IL-6 is expressed by immune and parenchymal cells during senescence, and that IL-6 is part of the senescence-associated secretory phenotype. Administration of an anti-IL-6 antibody to Mdr2-KOFoxl1-CRE;RosaYFP mice inhibited the development of cHCC-CCA tumors. Blocking IL-6 trans-signaling led to a decrease in the number and size of cHCC-CCA tumors, indicating their dependence on this pathway. Furthermore, the administration of a senolytic agent inhibited IL-6 and the development of cHCC-CCA tumors.

      Conclusion

      Our results demonstrate that cHCC-CCA, but not HCC tumors, originate from HPCs, and that IL-6, which derives in part from cells in senescence, plays an important role in this process via IL-6 trans-signaling. These findings could be applied to develop new therapeutic approaches for cHCC-CCA tumors.

      Lay summary

      Combined hepatocellular carcinoma–cholangiocarcinoma is the third most prevalent type of primary liver cancer (i.e. a cancer that originates in the liver). Herein, we show that this type of cancer originates in stem cells in the liver and that it depends on inflammatory signaling. Specifically, we identify a cytokine called IL-6 that appears to be important in the development of these tumors. Our results could be used for the development of novel treatments for these aggressive tumors.

      Graphical abstract

      Keywords

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      References

        • He G.
        • Dhar D.
        • Nakagawa H.
        • Font-Burgada J.
        • Ogata H.
        • Jiang Y.
        • et al.
        Identification of liver cancer progenitors whose malignant progression depends on autocrine IL-6 signaling.
        Cell. 2013; 155: 384-396
        • Yamashita T.
        • Wang X.W.
        Cancer stem cells in the development of liver cancer.
        J Clin Invest. 2013; 123: 1911-1918
        • Benhamouche S.
        • Curto M.
        • Saotome I.
        • Gladden A.B.
        • Liu C.H.
        • Giovannini M.
        • et al.
        Nf2/Merlin controls progenitor homeostasis and tumorigenesis in the liver.
        Genes Dev. 2010; 24: 1718-1730
        • Sia D.
        • Villanueva A.
        • Friedman S.L.
        • Llovet J.M.
        Liver cancer cell of origin, molecular class, and effects on patient prognosis.
        Gastroenterology. 2017; 152: 745-761
        • Calderaro J.
        • Ziol M.
        • Paradis V.
        • Zucman-Rossi J.
        Molecular and histological correlations in liver cancer.
        J Hepatol. 2019; 71: 616-630
        • Yuan D.
        • Huang S.
        • Berger E.
        • Liu L.
        • Gross N.
        • Heinzmann F.
        • et al.
        Kupffer cell-derived Tnf triggers cholangiocellular tumorigenesis through JNK due to chronic mitochondrial dysfunction and ROS.
        Cancer Cell. 2017; 31: 771-789 e776
        • Llovet J.M.
        • Zucman-Rossi J.
        • Pikarsky E.
        • Sangro B.
        • Schwartz M.
        • Sherman M.
        • et al.
        Hepatocellular carcinoma.
        Nat Rev Dis Primers. 2016; 2: 16018
        • Galun E.
        Liver inflammation and cancer: the role of tissue microenvironment in generating the tumor-promoting niche (TPN) in the development of hepatocellular carcinoma.
        Hepatology. 2016; 63: 354-356
        • Barashi N.
        • Weiss I.D.
        • Wald O.
        • Wald H.
        • Beider K.
        • Abraham M.
        • et al.
        Inflammation-induced hepatocellular carcinoma is dependent on CCR5 in mice.
        Hepatology. 2013; 58: 1021-1030
        • Zhu Y.
        • Kwong L.N.
        Insights into the origin of intrahepatic cholangiocarcinoma from mouse models.
        Hepatology. 2020; 72: 305-314
        • Lee T.K.
        • Guan X.Y.
        • Ma S.
        Cancer stem cells in hepatocellular carcinoma - from origin to clinical implications.
        Nat Rev Gastroenterol Hepatol. 2022; 19: 26-44
        • Clerbaux L.A.
        • Manco R.
        • Van Hul N.
        • Bouzin C.
        • Sciarra A.
        • Sempoux C.
        • et al.
        Invasive ductular reaction operates hepatobiliary junctions upon hepatocellular injury in rodents and humans.
        Am J Pathol. 2019; 189: 1569-1581
        • Popper H.
        • Kent G.
        • Stein R.
        Ductular cell reaction in the liver in hepatic injury.
        J Mt Sinai Hosp N Y. 1957; 24: 551-556
        • Roskams T.
        • Desmet V.
        Ductular reaction and its diagnostic significance.
        Semin Diagn Pathol. 1998; 15: 259-269
        • Ko S.
        • Russell J.O.
        • Molina L.M.
        • Monga S.P.
        Liver progenitors and adult cell plasticity in hepatic injury and repair: knowns and unknowns.
        Annu Rev Pathol. 2020; 15: 23-50
        • Sato K.
        • Marzioni M.
        • Meng F.
        • Francis H.
        • Glaser S.
        • Alpini G.
        Ductular reaction in liver diseases: pathological mechanisms and translational significances.
        Hepatology. 2019; 69: 420-430
        • Zhao L.
        • Westerhoff M.
        • Pai R.K.
        • Choi W.T.
        • Gao Z.H.
        • Hart J.
        Centrilobular ductular reaction correlates with fibrosis stage and fibrosis progression in non-alcoholic steatohepatitis.
        Mod Pathol. 2018; 31: 150-159
        • Raven A.
        • Lu W.Y.
        • Man T.Y.
        • Ferreira-Gonzalez S.
        • O'Duibhir E.
        • Dwyer B.J.
        • et al.
        Cholangiocytes act as facultative liver stem cells during impaired hepatocyte regeneration.
        Nature. 2017; 547: 350-354
        • Moeini A.
        • Sia D.
        • Zhang Z.
        • Camprecios G.
        • Stueck A.
        • Dong H.
        • et al.
        Mixed hepatocellular cholangiocarcinoma tumors: cholangiolocellular carcinoma is a distinct molecular entity.
        J Hepatol. 2017; 66: 952-961
        • Brunt E.
        • Aishima S.
        • Clavien P.A.
        • Fowler K.
        • Goodman Z.
        • Gores G.
        • et al.
        cHCC-CCA: consensus terminology for primary liver carcinomas with both hepatocytic and cholangiocytic differentation.
        Hepatology. 2018; 68: 113-126
        • Mauad T.H.
        • van Nieuwkerk C.M.
        • Dingemans K.P.
        • Smit J.J.
        • Schinkel A.H.
        • Notenboom R.G.
        • et al.
        Mice with homozygous disruption of the mdr2 P-glycoprotein gene. A novel animal model for studies of nonsuppurative inflammatory cholangitis and hepatocarcinogenesis.
        Am J Pathol. 1994; 145: 1237-1245
        • Potikha T.
        • Stoyanov E.
        • Pappo O.
        • Frolov A.
        • Mizrahi L.
        • Olam D.
        • et al.
        Interstrain differences in chronic hepatitis and tumor development in a murine model of inflammation-mediated hepatocarcinogenesis.
        Hepatology. 2013; 58: 192-204
        • Ma S.
        • Chan K.W.
        • Hu L.
        • Lee T.K.
        • Wo J.Y.
        • Ng I.O.
        • et al.
        Identification and characterization of tumorigenic liver cancer stem/progenitor cells.
        Gastroenterology. 2007; 132: 2542-2556
        • Shin S.
        • Walton G.
        • Aoki R.
        • Brondell K.
        • Schug J.
        • Fox A.
        • et al.
        Foxl1-Cre-marked adult hepatic progenitors have clonogenic and bilineage differentiation potential.
        Genes Dev. 2011; 25: 1185-1192
        • Govaere O.
        • Komuta M.
        • Berkers J.
        • Spee B.
        • Janssen C.
        • de Luca F.
        • et al.
        Keratin 19: a key role player in the invasion of human hepatocellular carcinomas.
        Gut. 2014; 63: 674-685
        • Ikenaga N.
        • Liu S.B.
        • Sverdlov D.Y.
        • Yoshida S.
        • Nasser I.
        • Ke Q.
        • et al.
        A new Mdr2(-/-) mouse model of sclerosing cholangitis with rapid fibrosis progression, early-onset portal hypertension, and liver cancer.
        Am J Pathol. 2015; 185: 325-334
        • Sackett S.D.
        • Li Z.
        • Hurtt R.
        • Gao Y.
        • Wells R.G.
        • Brondell K.
        • et al.
        Foxl1 is a marker of bipotential hepatic progenitor cells in mice.
        Hepatology. 2009; 49: 920-929
        • Fickert P.
        • Fuchsbichler A.
        • Wagner M.
        • Zollner G.
        • Kaser A.
        • Tilg H.
        • et al.
        Regurgitation of bile acids from leaky bile ducts causes sclerosing cholangitis in Mdr2 (Abcb4) knockout mice.
        Gastroenterology. 2004; 127: 261-274
        • Llovet J.M.
        • Kelley R.K.
        • Villanueva A.
        • Singal A.G.
        • Pikarsky E.
        • Roayaie S.
        • et al.
        Hepatocellular carcinoma.
        Nat Rev Dis Primers. 2021; 7: 6
        • Ng I.O.
        • Na J.
        • Lai E.C.
        • Fan S.T.
        • Ng M.
        Ki-67 antigen expression in hepatocellular carcinoma using monoclonal antibody MIB1. A comparison with proliferating cell nuclear antigen.
        Am J Clin Pathol. 1995; 104: 313-318
        • Katzenellenbogen M.
        • Mizrahi L.
        • Pappo O.
        • Klopstock N.
        • Olam D.
        • Jacob-Hirsch J.
        • et al.
        Molecular mechanisms of liver carcinogenesis in the mdr2-knockout mice.
        Mol Cancer Res. 2007; 5: 1159-1170
        • Smit J.J.
        • Schinkel A.H.
        • Oude Elferink R.P.
        • Groen A.K.
        • Wagenaar E.
        • van Deemter L.
        • et al.
        Homozygous disruption of the murine mdr2 P-glycoprotein gene leads to a complete absence of phospholipid from bile and to liver disease.
        Cell. 1993; 75: 451-462
        • Zhou T.
        • Wu N.
        • Meng F.
        • Venter J.
        • Giang T.K.
        • Francis H.
        • et al.
        Knockout of secretin receptor reduces biliary damage and liver fibrosis in Mdr2(-/-) mice by diminishing senescence of cholangiocytes.
        Lab Invest. 2018; 98: 1449-1464
        • Lee T.K.
        • Castilho A.
        • Cheung V.C.
        • Tang K.H.
        • Ma S.
        • Ng I.O.
        CD24(+) liver tumor-initiating cells drive self-renewal and tumor initiation through STAT3-mediated NANOG regulation.
        Cell Stem Cell. 2011; 9: 50-63
        • Wan S.
        • Zhao E.
        • Kryczek I.
        • Vatan L.
        • Sadovskaya A.
        • Ludema G.
        • et al.
        Tumor-associated macrophages produce interleukin 6 and signal via STAT3 to promote expansion of human hepatocellular carcinoma stem cells.
        Gastroenterology. 2014; 147: 1393-1404
        • Tirnitz-Parker J.E.
        • Tonkin J.N.
        • Knight B.
        • Olynyk J.K.
        • Yeoh G.C.
        Isolation, culture and immortalisation of hepatic oval cells from adult mice fed a choline-deficient, ethionine-supplemented diet.
        Int J Biochem Cell Biol. 2007; 39: 2226-2239
        • Almale L.
        • Garcia-Alvaro M.
        • Martinez-Palacian A.
        • Garcia-Bravo M.
        • Lazcanoiturburu N.
        • Addante A.
        • et al.
        c-Met signaling is essential for mouse adult liver progenitor cells expansion after transforming growth factor-beta-induced epithelial-mesenchymal transition and regulates cell phenotypic switch.
        Stem Cells. 2019; 37: 1108-1118
        • Fischer M.
        • Goldschmitt J.
        • Peschel C.
        • Brakenhoff J.P.
        • Kallen K.J.
        • Wollmer A.
        • et al.
        A bioactive designer cytokine for human hematopoietic progenitor cell expansion.
        Nat Biotechnol. 1997; 15: 142-145
        • Schumertl T.
        • Lokau J.
        • Rose-John S.
        • Garbers C.
        Function and proteolytic generation of the soluble interleukin-6 receptor in health and disease.
        Biochim Biophys Acta Mol Cell Res. 2022; 1869: 119143
        • Garbers C.
        • Heink S.
        • Korn T.
        • Rose-John S.
        Interleukin-6: designing specific therapeutics for a complex cytokine.
        Nat Rev Drug Discov. 2018; 17: 395-412
        • Faget D.V.
        • Ren Q.
        • Stewart S.A.
        Unmasking senescence: context-dependent effects of SASP in cancer.
        Nat Rev Cancer. 2019; 19: 439-453
        • Yousefzadeh M.J.
        • Flores R.R.
        • Zhu Y.
        • Schmiechen Z.C.
        • Brooks R.W.
        • Trussoni C.E.
        • et al.
        An aged immune system drives senescence and ageing of solid organs.
        Nature. 2021; 594: 100-105
        • Childs B.G.
        • Durik M.
        • Baker D.J.
        • van Deursen J.M.
        Cellular senescence in aging and age-related disease: from mechanisms to therapy.
        Nat Med. 2015; 21: 1424-1435
        • Shriki A.
        • Lanton T.
        • Sonnenblick A.
        • Levkovitch-Siany O.
        • Eidelshtein D.
        • Abramovitch R.
        • et al.
        Multiple roles of IL-6 in hepatic injury, steatosis, and senescence aggregate to suppress tumorigenesis.
        Cancer Res. 2021; 81: 4766-4777
        • Eggert T.
        • Wolter K.
        • Ji J.
        • Ma C.
        • Yevsa T.
        • Klotz S.
        • et al.
        Distinct functions of senescence-associated immune responses in liver tumor surveillance and tumor progression.
        Cancer Cell. 2016; 30: 533-547
        • Kang T.W.
        • Yevsa T.
        • Woller N.
        • Hoenicke L.
        • Wuestefeld T.
        • Dauch D.
        • et al.
        Senescence surveillance of pre-malignant hepatocytes limits liver cancer development.
        Nature. 2011; 479: 547-551
        • Martinez-Zamudio R.I.
        • Roux P.F.
        • de Freitas J.
        • Robinson L.
        • Dore G.
        • Sun B.
        • et al.
        AP-1 imprints a reversible transcriptional programme of senescent cells.
        Nat Cell Biol. 2020; 22: 842-855
        • Salmonowicz H.
        • Passos J.F.
        Detecting senescence: a new method for an old pigment.
        Aging Cell. 2017; 16: 432-434
        • Birch J.
        • Gil J.
        Senescence and the SASP: many therapeutic avenues.
        Genes Dev. 2020; 34: 1565-1576
        • Konopleva M.
        • Contractor R.
        • Tsao T.
        • Samudio I.
        • Ruvolo P.P.
        • Kitada S.
        • et al.
        Mechanisms of apoptosis sensitivity and resistance to the BH3 mimetic ABT-737 in acute myeloid leukemia.
        Cancer Cell. 2006; 10: 375-388
        • Spolverato G.
        • Bagante F.
        • Tsilimigras D.
        • Ejaz A.
        • Cloyd J.
        • Pawlik T.M.
        Management and outcomes among patients with mixed hepatocholangiocellular carcinoma: a population-based analysis.
        J Surg Oncol. 2019; 119: 278-287
        • Tirnitz-Parker J.E.E.
        • Forbes S.J.
        • Olynyk J.K.
        • Ramm G.A.
        Cellular plasticity in liver regeneration: spotlight on cholangiocytes.
        Hepatology. 2019; 69: 2286-2289
        • Schaub J.R.
        • Huppert K.A.
        • Kurial S.N.T.
        • Hsu B.Y.
        • Cast A.E.
        • Donnelly B.
        • et al.
        De novo formation of the biliary system by TGFbeta-mediated hepatocyte transdifferentiation.
        Nature. 2018; 557: 247-251
        • Hickson L.J.
        • Langhi Prata L.G.P.
        • Bobart S.A.
        • Evans T.K.
        • Giorgadze N.
        • Hashmi S.K.
        • et al.
        Senolytics decrease senescent cells in humans: preliminary report from a clinical trial of Dasatinib plus Quercetin in individuals with diabetic kidney disease.
        EBioMedicine. 2019; 47: 446-456