Dysregulated lipid metabolism in hepatocellular carcinoma cancer stem cells

Abstract

According to the stem cell theory for cancer, hepatocellular carcinomas are sustained by a group of cancer stem cells (CSCs) which are responsible for resistance to chemotherapy. In the present study we aimed to examine lipid metabolism in cancer stem cells induced by long-term treatment with sorafenib and its relationship with acquisition of a CSC-like phenotype. Two cell lines (HepG2SF1 and Huh7SF1) were generated by incubation with a step-wise increase of sorafenib concentrations for 10 months. These cell lines displayed stem-like characteristics like increase in the expression of ABCB1A, Nanog and Oct4 as well as an E-cadherin/N-cadherin switch. HepG2SF1 and Huh7SF1 cells showed intracellular accumulation of neutral lipids, assessed by flow cytometry and confocal microscopy. The exam of lipid metabolism revealed that HepG2SF1 and Huh7SF1 cells increased the expression of the enzymes involved in de novo lipid synthesis ATP–citrate lyase (ACLY), acetyl-CoA carboxylase (ACC) and fatty acid synthase (FASN) and that of the fatty acid transporter CD36. In addition, these CSC-like cells had enhanced expression of the lipogenic transcription factor SREBP1c. Analysis of the key metabolic sensor AMP-activated kinase (AMPK) demonstrated that both AMPK phosphorylation and levels were decreased in the CSC-like cells compared to their parental cells. Interestingly, transfection of HepG2SF1 and Huh7SF1 cells with AMPK, restored the levels of the lipogenic enzymes and SREBP1c and decreased the intracellular lipid accumulation. Furthermore, AMPK transfection decreased the stemness markers and inhibited the E-cadherin/N-cadherin switch. Targeting AMPK and lipid metabolism of hepatocellular cancer stem cells is a promising strategy to face stemness and chemotherapy resistance.

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References

  1. 1.

    Schneller D, Angel P (2019) Cellular origin of hepatocellular carcinoma. In: Tirnitz-Parker JEE (ed) Hepatocellular carcinoma. Codon Publications, Brisbane. https://doi.org/10.15586/hepatocellularcarcinoma.2019.ch1

    Google Scholar 

  2. 2.

    Wang N, Wang S, Li MY, Hu BG, Liu LP, Yang SL, Yang S, Gong Z, Lai PBS, Chen GG (2018) Cancer stem cells in hepatocellular carcinoma: an overview and promising therapeutic strategies. Ther Adv Med Oncol. https://doi.org/10.1177/1758835918816287

    Article  PubMed  PubMed Central  Google Scholar 

  3. 3.

    Flores-Tellez TN, Villa-Trevino S, Pina-Vazquez C (2017) Road to stemness in hepatocellular carcinoma. World J Gastroenterol 23(37):6750–6776. https://doi.org/10.3748/wjg.v23.i37.6750

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  4. 4.

    Huo Y, Chen WS, Lee J, Feng GS, Newton IG (2019) Stress conditions induced by locoregional therapies stimulate enrichment and proliferation of liver cancer stem cells. J Vasc Interv Radiol 30(12):2016–2025. https://doi.org/10.1016/j.jvir.2019.02.026

    Article  PubMed  Google Scholar 

  5. 5.

    Dongre A, Weinberg RA (2019) New insights into the mechanisms of epithelial-mesenchymal transition and implications for cancer. Nat Rev Mol Cell Biol 20(2):69–84. https://doi.org/10.1038/s41580-018-0080-4

    CAS  Article  PubMed  Google Scholar 

  6. 6.

    Korshunov DA, Kondakova IV, Shashova EE (2019) Modern perspective on metabolic reprogramming in malignant neoplasms. Biochemistry 84(10):1129–1142. https://doi.org/10.1134/S000629791910002X

    CAS  Article  PubMed  Google Scholar 

  7. 7.

    Koundouros N, Poulogiannis G (2019) Reprogramming of fatty acid metabolism in cancer. Br J Cancer. https://doi.org/10.1038/s41416-019-0650-z

    Article  PubMed  PubMed Central  Google Scholar 

  8. 8.

    Kim WY (2019) Therapeutic targeting of lipid synthesis metabolism for selective elimination of cancer stem cells. Arch Pharm Res 42(1):25–39. https://doi.org/10.1007/s12272-018-1098-z

    CAS  Article  PubMed  Google Scholar 

  9. 9.

    Intlekofer AM, Finley LWS (2019) Metabolic signatures of cancer cells and stem cells. Nat Metab 1(2):177–188. https://doi.org/10.1038/s42255-019-0032-0

    Article  PubMed  PubMed Central  Google Scholar 

  10. 10.

    Warburg O (1956) On respiratory impairment in cancer cells. Science 124(3215):269–270

    CAS  PubMed  Google Scholar 

  11. 11.

    Viale A, Pettazzoni P, Lyssiotis CA, Ying H, Sanchez N, Marchesini M, Carugo A, Green T, Seth S, Giuliani V, Kost-Alimova M, Muller F, Colla S, Nezi L, Genovese G, Deem AK, Kapoor A, Yao W, Brunetto E, Kang Y, Yuan M, Asara JM, Wang YA, Heffernan TP, Kimmelman AC, Wang H, Fleming JB, Cantley LC, DePinho RA, Draetta GF (2014) Oncogene ablation-resistant pancreatic cancer cells depend on mitochondrial function. Nature 514(7524):628–632. https://doi.org/10.1038/nature13611

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  12. 12.

    Alptekin A, Ye B, Ding HF (2017) Transcriptional regulation of stem cell and cancer stem cell metabolism. Curr Stem Cell Rep 3(1):19–27. https://doi.org/10.1007/s40778-017-0071-y

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  13. 13.

    Munir R, Lisec J, Swinnen JV, Zaidi N (2019) Lipid metabolism in cancer cells under metabolic stress. Br J Cancer 120(12):1090–1098. https://doi.org/10.1038/s41416-019-0451-4

    Article  PubMed  Google Scholar 

  14. 14.

    Hardie DG, Schaffer BE, Brunet A (2016) AMPK: an energy-sensing pathway with multiple inputs and outputs. Trends Cell Biol 26(3):190–201. https://doi.org/10.1016/j.tcb.2015.10.013

    CAS  Article  PubMed  Google Scholar 

  15. 15.

    Bort A, Sanchez BG, Mateos-Gomez PA, Vara-Ciruelos D, Rodriguez-Henche N, Diaz-Laviada I (2019) Targeting AMP-activated kinase impacts hepatocellular cancer stem cells induced by long-term treatment with sorafenib. Mol Oncol 13(5):1311–1331. https://doi.org/10.1002/1878-0261.12488

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  16. 16.

    van Schaijik B, Davis PF, Wickremesekera AC, Tan ST, Itinteang T (2018) Subcellular localisation of the stem cell markers OCT4, SOX2, NANOG, KLF4 and c-MYC in cancer: a review. J Clin Pathol 71(1):88–91. https://doi.org/10.1136/jclinpath-2017-204815

    CAS  Article  PubMed  Google Scholar 

  17. 17.

    Yin X, Zhang BH, Zheng SS, Gao DM, Qiu SJ, Wu WZ, Ren ZG (2015) Coexpression of gene Oct4 and Nanog initiates stem cell characteristics in hepatocellular carcinoma and promotes epithelial-mesenchymal transition through activation of Stat3/Snail signaling. J Hematol Oncol 8:23. https://doi.org/10.1186/s13045-015-0119-3

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  18. 18.

    Loh YH, Wu Q, Chew JL, Vega VB, Zhang W, Chen X, Bourque G, George J, Leong B, Liu J, Wong KY, Sung KW, Lee CW, Zhao XD, Chiu KP, Lipovich L, Kuznetsov VA, Robson P, Stanton LW, Wei CL, Ruan Y, Lim B, Ng HH (2006) The Oct4 and Nanog transcription network regulates pluripotency in mouse embryonic stem cells. Nat Genet 38(4):431–440. https://doi.org/10.1038/ng1760

    CAS  Article  PubMed  Google Scholar 

  19. 19.

    Kahraman DC, Kahraman T, Cetin-Atalay R (2019) Targeting PI3K/Akt/mTOR pathway identifies differential expression and functional role of IL8 in liver cancer stem cell enrichment. Mol Cancer Ther 18(11):2146–2157. https://doi.org/10.1158/1535-7163.MCT-19-0004

    CAS  Article  PubMed  Google Scholar 

  20. 20.

    Zhang XL, Jia Q, Lv L, Deng T, Gao J (2015) Tumorspheres derived from HCC cells are enriched with cancer stem cell-like cells and present high chemoresistance dependent on the Akt pathway. Anticancer Agents Med Chem 15(6):755–763. https://doi.org/10.2174/1871520615666150202111721

    CAS  Article  PubMed  Google Scholar 

  21. 21.

    Wang Y, Viscarra J, Kim SJ, Sul HS (2015) Transcriptional regulation of hepatic lipogenesis. Nat Rev Mol Cell Biol 16(11):678–689. https://doi.org/10.1038/nrm4074

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  22. 22.

    Raghow R, Dong Q, Elam MB (2019) Phosphorylation dependent proteostasis of sterol regulatory element binding proteins. Biochim Biophys Acta Mol Cell Biol Lipids 1864(8):1145–1156. https://doi.org/10.1016/j.bbalip.2019.04.015

    CAS  Article  PubMed  Google Scholar 

  23. 23.

    Li Y, Xu S, Mihaylova MM, Zheng B, Hou X, Jiang B, Park O, Luo Z, Lefai E, Shyy JY, Gao B, Wierzbicki M, Verbeuren TJ, Shaw RJ, Cohen RA, Zang M (2011) AMPK phosphorylates and inhibits SREBP activity to attenuate hepatic steatosis and atherosclerosis in diet-induced insulin-resistant mice. Cell Metab 13(4):376–388. https://doi.org/10.1016/j.cmet.2011.03.009

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  24. 24.

    Glatz JF, Luiken JJ (2017) From fat to FAT (CD36/SR-B2): understanding the regulation of cellular fatty acid uptake. Biochimie 136:21–26. https://doi.org/10.1016/j.biochi.2016.12.007

    CAS  Article  PubMed  Google Scholar 

  25. 25.

    Lee YK, Park JE, Lee M, Hardwick JP (2018) Hepatic lipid homeostasis by peroxisome proliferator-activated receptor gamma 2. Liver Res 2(4):209–215. https://doi.org/10.1016/j.livres.2018.12.001

    Article  PubMed  PubMed Central  Google Scholar 

  26. 26.

    Lee SH, Lee JH, Lee HY, Min KJ (2019) Sirtuin signaling in cellular senescence and aging. BMB Rep 52(1):24–34

    CAS  Article  Google Scholar 

  27. 27.

    Ling S, Tian Y, Zhang H, Jia K, Feng T, Sun D, Gao Z, Xu F, Hou Z, Li Y, Wang L (2014) Metformin reverses multidrug resistance in human hepatocellular carcinoma Bel7402/5fluorouracil cells. Mol Med Rep 10(6):2891–2897. https://doi.org/10.3892/mmr.2014.2614

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  28. 28.

    Ponugoti B, Kim DH, Xiao Z, Smith Z, Miao J, Zang M, Wu SY, Chiang CM, Veenstra TD, Kemper JK (2010) SIRT1 deacetylates and inhibits SREBP-1C activity in regulation of hepatic lipid metabolism. J Biol Chem 285(44):33959–33970. https://doi.org/10.1074/jbc.M110.122978

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  29. 29.

    Ruderman NB, Xu XJ, Nelson L, Cacicedo JM, Saha AK, Lan F, Ido Y (2010) AMPK and SIRT1: a long-standing partnership? Am J Physiol Endocrinol Metab 298(4):E751–E760. https://doi.org/10.1152/ajpendo.00745.2009

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  30. 30.

    Yu A, Dang W (2017) Regulation of stem cell aging by SIRT1—linking metabolic signaling to epigenetic modifications. Mol Cell Endocrinol 455:75–82. https://doi.org/10.1016/j.mce.2017.03.031

    CAS  Article  PubMed  Google Scholar 

  31. 31.

    Hayes CN, Zhang P, Chayama K (2019) The role of lipids in hepatocellular carcinoma. In: Tirnitz-Parker JEE (ed) Hepatocellular carcinoma. Codon Publications, Brisbane. https://doi.org/10.15586/hepatocellularcarcinoma.2019.ch5

    Google Scholar 

  32. 32.

    Visweswaran M, Arfuso F, Warrier S, Dharmarajan A (2019) Concise review: aberrant lipid metabolism as an emerging therapeutic strategy to target cancer stem cells. Stem Cells.https://doi.org/10.1002/stem.3101

    Article  PubMed  Google Scholar 

  33. 33.

    Giampietri C, Petrungaro S, Cordella M, Tabolacci C, Tomaipitinca L, Facchiano A, Eramo A, Filippini A, Facchiano F, Ziparo E (2017) Lipid storage and autophagy in melanoma cancer cells. Int J Mol Sci.https://doi.org/10.3390/ijms18061271

    Article  PubMed  PubMed Central  Google Scholar 

  34. 34.

    The cacner genome atlas programm. https://www.cancer.gov/tcga. Accessed 2 Feb 2020

  35. 35.

    The cancer proteome atlas. https://tcpaportal.org/tcpa/. Accessed 17 Feb 2020

  36. 36.

    Giudetti AM, De Domenico S, Ragusa A, Lunetti P, Gaballo A, Franck J, Simeone P, Nicolardi G, De Nuccio F, Santino A, Capobianco L, Lanuti P, Fournier I, Salzet M, Maffia M, Vergara D (2019) A specific lipid metabolic profile is associated with the epithelial mesenchymal transition program. Biochim Biophys Acta Mol Cell Biol Lipids 1864(3):344–357. https://doi.org/10.1016/j.bbalip.2018.12.011

    CAS  Article  PubMed  Google Scholar 

  37. 37.

    Deng M, Cai X, Long L, Xie L, Ma H, Zhou Y, Liu S, Zeng C (2019) CD36 promotes the epithelial-mesenchymal transition and metastasis in cervical cancer by interacting with TGF-beta. J Transl Med 17(1):352. https://doi.org/10.1186/s12967-019-2098-6

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  38. 38.

    Nath A, Li I, Roberts LR, Chan C (2015) Elevated free fatty acid uptake via CD36 promotes epithelial-mesenchymal transition in hepatocellular carcinoma. Sci Rep 5:14752. https://doi.org/10.1038/srep14752

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  39. 39.

    Hale JS, Otvos B, Sinyuk M, Alvarado AG, Hitomi M, Stoltz K, Wu Q, Flavahan W, Levison B, Johansen ML, Schmitt D, Neltner JM, Huang P, Ren B, Sloan AE, Silverstein RL, Gladson CL, DiDonato JA, Brown JM, McIntyre T, Hazen SL, Horbinski C, Rich JN, Lathia JD (2014) Cancer stem cell-specific scavenger receptor CD36 drives glioblastoma progression. Stem Cells 32(7):1746–1758. https://doi.org/10.1002/stem.1716

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  40. 40.

    Liu L, Liu C, Zhang Q, Shen J, Zhang H, Shan J, Duan G, Guo D, Chen X, Cheng J, Xu Y, Yang Z, Yao C, Lai M, Qian C (2016) SIRT1-mediated transcriptional regulation of SOX2 is important for self-renewal of liver cancer stem cells. Hepatology 64(3):814–827. https://doi.org/10.1002/hep.28690

    CAS  Article  PubMed  Google Scholar 

  41. 41.

    Maehara O, Ohnishi S, Asano A, Suda G, Natsuizaka M, Nakagawa K, Kobayashi M, Sakamoto N, Takeda H (2019) Metformin regulates the expression of CD133 through the AMPK-CEBPbeta pathway in hepatocellular carcinoma cell lines. Neoplasia 21(6):545–556. https://doi.org/10.1016/j.neo.2019.03.007

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  42. 42.

    Saito T, Chiba T, Yuki K, Zen Y, Oshima M, Koide S, Motoyama T, Ogasawara S, Suzuki E, Ooka Y, Tawada A, Tada M, Kanai F, Takiguchi Y, Iwama A, Yokosuka O (2013) Metformin, a diabetes drug, eliminates tumor-initiating hepatocellular carcinoma cells. PLoS ONE 8(7):e70010. https://doi.org/10.1371/journal.pone.0070010

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  43. 43.

    Zhao B, Luo J, Wang Y, Zhou L, Che J, Wang F, Peng S, Zhang G, Shang P (2019) Metformin Suppresses self-renewal ability and tumorigenicity of osteosarcoma stem cells via reactive oxygen species-mediated apoptosis and autophagy. Oxid Med Cell Longev.https://doi.org/10.1155/2019/9290728

    Article  PubMed  PubMed Central  Google Scholar 

  44. 44.

    Vara-Ciruelos D, Dandapani M, Russell FM, Grzes KM, Atrih A, Foretz M, Viollet B, Lamont DJ, Cantrell DA, Hardie DG (2019) Phenformin, but not metformin, delays development of T Cell acute lymphoblastic leukemia/lymphoma via cell-autonomous AMPK activation. Cell Rep 27(3):690–698.e4. https://doi.org/10.1016/j.celrep.2019.03.067

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  45. 45.

    Wang X, Jin J, Wan F, Zhao L, Chu H, Chen C, Liao G, Liu J, Yu Y, Teng H, Fang L, Jiang C, Pan W, Xie X, Li J, Lu X, Jiang X, Ge X, Ye D, Wang P (2019) AMPK promotes SPOP-mediated NANOG Degradation to regulate prostate cancer cell stemness. Dev Cell 48(3):345-360 e347. https://doi.org/10.1016/j.devcel.2018.11.033

    CAS  Article  PubMed  Google Scholar 

  46. 46.

    Penfold L, Woods A, Muckett P, Nikitin AY, Kent TR, Zhang S, Graham R, Pollard A, Carling D (2018) CAMKK2 promotes prostate cancer independently of AMPK via increased lipogenesis. Cancer Res 78(24):6747–6761. https://doi.org/10.1158/0008-5472.CAN-18-0585

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  47. 47.

    Yousefnia S, Momenzadeh S, Seyed Forootan F, Ghaedi K, Nasr Esfahani MH (2018) The influence of peroxisome proliferator-activated receptor gamma (PPARgamma) ligands on cancer cell tumorigenicity. Gene 649:14–22. https://doi.org/10.1016/j.gene.2018.01.018

    CAS  Article  PubMed  Google Scholar 

  48. 48.

    Kaur S, Nag A, Gangenahalli G, Sharma K (2019) Peroxisome proliferator activated receptor gamma sensitizes non-small cell lung carcinoma to gamma irradiation induced apoptosis. Front Genet 10:554. https://doi.org/10.3389/fgene.2019.00554

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  49. 49.

    Pestereva E, Kanakasabai S, Bright JJ (2012) PPARgamma agonists regulate the expression of stemness and differentiation genes in brain tumour stem cells. Br J Cancer 106(10):1702–1712. https://doi.org/10.1038/bjc.2012.161

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  50. 50.

    DeLaForest A, Di Furio F, Jing R, Ludwig-Kubinski A, Twaroski K, Urick A, Pulakanti K, Rao S, Duncan SA (2018) HNF4A regulates the formation of hepatic progenitor cells from human iPSC-derived endoderm by facilitating efficient recruitment of RNA Pol II. Genes (Basel).https://doi.org/10.3390/genes10010021

    Article  Google Scholar 

  51. 51.

    Kurakazu I, Akasaki Y, Hayashida M, Tsushima H, Goto N, Sueishi T, Toya M, Kuwahara M, Okazaki K, Duffy T, Lotz MK, Nakashima Y (2019) FOXO1 transcription factor regulates chondrogenic differentiation through transforming growth factor beta1 signaling. J Biol Chem 294(46):17555–17569. https://doi.org/10.1074/jbc.RA119.009409

    Article  PubMed  Google Scholar 

  52. 52.

    Niculite CM, Enciu AM, Hinescu ME (2019) CD 36: Focus on epigenetic and post-transcriptional regulation. Front Genet 10:680. https://doi.org/10.3389/fgene.2019.00680

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  53. 53.

    Hanai JI, Doro N, Seth P, Sukhatme VP (2013) ATP citrate lyase knockdown impacts cancer stem cells in vitro. Cell Death Dis 4:e696. https://doi.org/10.1038/cddis.2013.215

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  54. 54.

    Begicevic RR, Arfuso F, Falasca M (2019) Bioactive lipids in cancer stem cells. World J Stem Cells 11(9):693–704. https://doi.org/10.4252/wjsc.v11.i9.693

    Article  PubMed  PubMed Central  Google Scholar 

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Acknowledgements

The authors would like to thank Fundación Tatiana Pérez de Guzmán el Bueno (Grant Nº Patrocinio 2019-001) for financial support into their research. Authors acknowledge Dr. G. Hardie for providing AMPK plasmids.

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IDL conceived and supervised the study; AB and IDL designed experiments; BS, IdeM and AB performed experiments; BS and PMG analyzed data; IDL wrote the manuscript.

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Correspondence to Inés Diaz-Laviada.

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Bort, A., Sánchez, B.G., de Miguel, I. et al. Dysregulated lipid metabolism in hepatocellular carcinoma cancer stem cells. Mol Biol Rep 47, 2635–2647 (2020). https://doi.org/10.1007/s11033-020-05352-3

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Keywords

  • HepG2
  • Huh7
  • Sorafenib
  • cancer stem cells
  • Hepatocellular carcinoma
  • Lipid metabolism