Abstract

Cancer stem cells (CSCs) possess the capacity for self-renewal and the potential to differentiate into non-CSCs. The recent discoveries of dynamic equilibrium between CSCs and non-CSCs revealed the significance of acquiring CSC-like properties in non-CSCs as an important process in progression of cancer. The mechanism underlying acquisition of CSC-like properties has mainly been investigated in the context of epithelial–mesenchymal transition. Here, we demonstrate the dedifferentiation process may be an alternative mechanism in acquisition of CSC-like properties in human colorectal cancer cells. By exploring the single-cell gene expression analysis of organoids developed from CD44+ CSCs, we identified TWIST1 as a key molecule for maintaining the undifferentiated state of cancer cells. Consistent with the finding, we found that TGF-beta signaling pathway, a regulator of TWIST1, was specifically activated in the undifferentiated CD44+ CSCs in human colorectal cancer using microarray-based gene expression analysis and quantitative pathology imaging system. Furthermore, we showed that external stimulation with TGF-beta and the induction of TWIST1 converted CD44 non-CSCs into the undifferentiated CD44+ CSCs, leading to the significant increment of CSCs in xenograft models. This study strongly suggests dedifferentiation driven by TGF-beta signaling enhances stem cell properties in human colorectal cancer.

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References

  1. 1.

    Reya T, Morrison SJ, Clarke MF, Weissman IL. Stem cells, cancer, and cancer stem cells. Nature. 2001;414:105–11.

  2. 2.

    Bonnet D, Dick JE. Human acute myeloid leukemia is organized as a hierarchy that originates from a primitive hematopoietic cell. Nat Med. 1997;3:730–7.

  3. 3.

    Al-Hajj M, Wicha MS, Benito-Hernandez A, Morrison SJ, Clarke MF. Prospective identification of tumorigenic breast cancer cells. Proc Natl Acad Sci USA. 2003;100:3983–8.

  4. 4.

    Dalerba P, Dylla SJ, Park IK, Liu R, Wang X, Cho RW, et al. Phenotypic characterization of human colorectal cancer stem cells. Proc Natl Acad Sci USA. 2007;104:10158–63.

  5. 5.

    Lee CJ, Dosch J, Simeone DM. Pancreatic cancer stem cells. J Clin Oncol. 2008;26:2806–12.

  6. 6.

    Zhang S, Balch C, Chan MW, Lai HC, Matei D, Schilder JM, et al. Identification and characterization of ovarian cancer-initiating cells from primary human tumors. Cancer Res. 2008;68:4311–20.

  7. 7.

    Patrawala L, Calhoun T, Schneider-Broussard R, Li H, Bhatia B, Tang S, et al. Highly purified CD44+prostate cancer cells from xenograft human tumors are enriched in tumorigenic and metastatic progenitor cells. Oncogene. 2006;25:1696–708.

  8. 8.

    Dalerba P, Kalisky T, Sahoo D, Rajendran PS, Rothenberg ME, Leyrat AA, et al. Single-cell dissection of transcriptional heterogeneity in human colon tumors. Nat Biotechnol. 2011;29:1120–7.

  9. 9.

    Vermeulen L, Todaro M, de Sousa Mello F, Sprick MR, Kemper K, Perez Alea M, et al. Single-cell cloning of colon cancer stem cells reveals a multi-lineage differentiation capacity. Proc Natl Acad Sci USA. 2008;105:13427–32.

  10. 10.

    Massague J. TGFbeta signalling in context. Nat Rev Mol Cell Biol. 2012;13:616–30.

  11. 11.

    De Craene B, Berx G. Regulatory networks defining EMT during cancer initiation and progression. Nat Rev Cancer. 2013;13:97–110.

  12. 12.

    Mani SA, Guo W, Liao MJ, Eaton EN, Ayyanan A, Zhou AY, et al. The epithelial-mesenchymal transition generates cells with properties of stem cells. Cell. 2008;133:704–15.

  13. 13.

    Morel AP, Lievre M, Thomas C, Hinkal G, Ansieau S, Puisieux A. Generation of breast cancer stem cells through epithelial-mesenchymal transition. PLoS ONE. 2008;3:e2888.

  14. 14.

    Scheel C, Eaton EN, Li SH, Chaffer CL, Reinhardt F, Kah KJ, et al. Paracrine and autocrine signals induce and maintain mesenchymal and stem cell states in the breast. Cell. 2011;145:926–40.

  15. 15.

    Chaffer CL, Marjanovic ND, Lee T, Bell G, Kleer CG, Reinhardt F, et al. Poised chromatin at the ZEB1 promoter enables breast cancer cell plasticity and enhances tumorigenicity. Cell. 2013;154:61–74.

  16. 16.

    Beck B, Lapouge G, Rorive S, Drogat B, Desaedelaere K, Delafaille S, et al. Different levels of Twist1 regulate skin tumor initiation, stemness, and progression. Cell Stem Cell. 2015;16:67–79.

  17. 17.

    Beerling E, Seinstra D, de Wit E, Kester L, van der Velden D, Maynard C, et al. Plasticity between epithelial and mesenchymal states unlinks EMT from metastasis-enhancing stem cell capacity. Cell Rep. 2016;14:2281–8.

  18. 18.

    Kalluri R. The biology and function of fibroblasts in cancer. Nat Rev Cancer. 2016;16:582–98.

  19. 19.

    Sato T, Stange DE, Ferrante M, Vries RG, Van Es JH, Van den Brink S, et al. Long-term expansion of epithelial organoids from human colon, adenoma, adenocarcinoma, and Barrett’s epithelium. Gastroenterology. 2011;141:1762–72.

  20. 20.

    Fujii M, Shimokawa M, Date S, Takano A, Matano M, Nanki K, et al. A colorectal tumor organoid library demonstrates progressive loss of niche factor requirements during tumorigenesis. Cell Stem Cell. 2016;18:827–38.

  21. 21.

    Ricci-Vitiani L, Lombardi DG, Pilozzi E, Biffoni M, Todaro M, Peschle C, et al. Identification and expansion of human colon-cancer-initiating cells. Nature. 2007;445:111–5.

  22. 22.

    O’Brien CA, Pollett A, Gallinger S, Dick JE. A human colon cancer cell capable of initiating tumor growth in immunodeficient mice. Nature. 2007;445:106–10.

  23. 23.

    Vermeulen L, De Sousa EMF, van der Heijden M, Cameron K, de Jong JH, Borovski T, et al. Wnt activity defines colon cancer stem cells and is regulated by the microenvironment. Nat Cell Biol. 2010;12:468–76.

  24. 24.

    Medema JP, Vermeulen L. Microenvironmental regulation of stem cells in intestinal homeostasis and cancer. Nature. 2011;474:318–26.

  25. 25.

    Ikushima H, Todo T, Ino Y, Takahashi M, Miyazawa K, Miyazono K. Autocrine TGF-beta signaling maintains tumorigenicity of glioma-initiating cells through Sry-related HMG-box factors. Cell Stem Cell. 2009;5:504–14.

  26. 26.

    Suva ML, Rheinbay E, Gillespie SM, Patel AP, Wakimoto H, Rabkin SD, et al. Reconstructing and reprogramming the tumor-propagating potential of glioblastoma stem-like cells. Cell. 2014;157:580–94.

  27. 27.

    Boumahdi S, Driessens G, Lapouge G, Rorive S, Nassar D, Le Mercier M, et al. SOX2 controls tumour initiation and cancer stem-cell functions in squamous-cell carcinoma. Nature. 2014;511:246–50.

  28. 28.

    Zhu P, Wang Y, He L, Huang G, Du Y, Zhang G, et al. ZIC2-dependent OCT4 activation drives self-renewal of human liver cancer stem cells. J Clin Invest. 2015;125:3795–808.

  29. 29.

    Mullen AC, Orlando DA, Newman JJ, Loven J, Kumar RM, Bilodeau S, et al. Master transcription factors determine cell-type-specific responses to TGF-beta signaling. Cell. 2011;147:565–76.

  30. 30.

    Oshimori N, Fuchs E. The harmonies played by TGF-beta in stem cell biology. Cell Stem Cell. 2012;11:751–64.

  31. 31.

    Jiang J, Ng HH. TGFbeta and SMADs talk to NANOG in human embryonic stem cells. Cell Stem Cell. 2008;3:127–8.

  32. 32.

    Merlos-Suarez A, Barriga FM, Jung P, Iglesias M, Cespedes MV, Rossell D, et al. The intestinal stem cell signature identifies colorectal cancer stem cells and predicts disease relapse. Cell Stem Cell. 2011;8:511–24.

  33. 33.

    Shvab A, Haase G, Ben-Shmuel A, Gavert N, Brabletz T, Dedhar S, et al. Induction of the intestinal stem cell signature gene SMOC-2 is required for L1-mediated colon cancer progression. Oncogene. 2016;35:549–57.

  34. 34.

    Barriga FM, Montagni E, Mana M, Mendez-Lago M, Hernando-Momblona X, Sevillano M, et al. Mex3a marks a slowly dividing subpopulation of Lgr5+intestinal stem cells. Cell Stem Cell. 2017;20:801–16 e807.

  35. 35.

    de Sousa EMF, Colak S, Buikhuisen J, Koster J, Cameron K, de Jong JH, et al. Methylation of cancer-stem-cell-associated Wnt target genes predicts poor prognosis in colorectal cancer patients. Cell Stem Cell. 2011;9:476–85.

  36. 36.

    Jung P, Sommer C, Barriga FM, Buczacki SJ, Hernando-Momblona X, Sevillano M, et al. Isolation of human colon stem cells using surface expression of PTK7. Stem Cell Rep. 2015;5:979–87.

  37. 37.

    Trapnell C, Cacchiarelli D, Grimsby J, Pokharel P, Li S, Morse M, et al. The dynamics and regulators of cell fate decisions are revealed by pseudotemporal ordering of single cells. Nat Biotechnol. 2014;32:381–6.

  38. 38.

    Jorissen RN, Gibbs P, Christie M, Prakash S, Lipton L, Desai J, et al. Metastasis-associated gene expression changes predict poor outcomes in patients with dukes stage B and C colorectal cancer. Clin Cancer Res. 2009;15:7642–51.

  39. 39.

    Wu Y, Tran T, Dwabe S, Sarkissyan M, Kim J, Nava M, et al. A83-01 inhibits TGF-beta-induced upregulation of Wnt3 and epithelial to mesenchymal transition in HER2-overexpressing breast cancer cells. Breast Cancer Res Treat. 2017;163:449–60.

  40. 40.

    Ovadya Y, Krizhanovsky V. A new twist in kidney fibrosis. Nat Med. 2015;21:975–7.

  41. 41.

    Brabletz T, Kalluri R, Nieto MA, Weinberg RA. EMT in cancer. Nat Rev Cancer. 2018;18:128–34.

  42. 42.

    Foroutan M, Cursons J, Hediyeh-Zadeh S, Thompson EW, Davis MJ. A Transcriptional program for detecting TGFbeta-induced EMT in cancer. Mol Cancer Res. 2017;15:619–31.

  43. 43.

    Yaeger R, Chatila WK, Lipsyc MD, Hechtman JF, Cercek A, Sanchez-Vega F, et al. Clinical sequencing defines the genomic landscape of metastatic colorectal cancer. Cancer Cell. 2018;33:125–36 e123.

  44. 44.

    Pino MS, Kikuchi H, Zeng M, Herraiz MT, Sperduti I, Berger D, et al. Epithelial to mesenchymal transition is impaired in colon cancer cells with microsatellite instability. Gastroenterology. 2010;138:1406–17.

  45. 45.

    Fleming NI, Jorissen RN, Mouradov D, Christie M, Sakthianandeswaren A, Palmieri M, et al. SMAD2, SMAD3 and SMAD4 mutations in colorectal cancer. Cancer Res. 2013;73:725–35.

  46. 46.

    Valcourt U, Kowanetz M, Niimi H, Heldin CH, Moustakas A. TGF-beta and the Smad signaling pathway support transcriptomic reprogramming during epithelial-mesenchymal cell transition. Mol Biol Cell. 2005;16:1987–2002.

  47. 47.

    Derynck R, Zhang YE. Smad-dependent and Smad-independent pathways in TGF-beta family signalling. Nature. 2003;425:577–84.

  48. 48.

    Zhang B, Halder SK, Kashikar ND, Cho YJ, Datta A, Gorden DL, et al. Antimetastatic role of Smad4 signaling in colorectal cancer. Gastroenterology. 2010;138:969–80. e961–3

  49. 49.

    Guinney J, Dienstmann R, Wang X, de Reynies A, Schlicker A, Soneson C, et al. The consensus molecular subtypes of colorectal cancer. Nat Med. 2015;21:1350–6.

  50. 50.

    Wang D, Rai B, Qi F, Liu T, Wang J, Wang X, et al. Influence of the Twist gene on the invasion and metastasis of colon cancer. Oncol Rep. 2018;39:31–44.

  51. 51.

    Ben-Porath I, Thomson MW, Carey VJ, Ge R, Bell GW, Regev A, et al. An embryonic stem cell-like gene expression signature in poorly differentiated aggressive human tumors. Nat Genet. 2008;40:499–507.

  52. 52.

    Kim J, Orkin SH. Embryonic stem cell-specific signatures in cancer: insights into genomic regulatory networks and implications for medicine. Genome Med. 2011;3:75.

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Acknowledgements

We especially thank N Torada, T Ueki, and T Manabe, S Nagai from the Department of Surgery and Oncology, Kyushu University, for providing clinical colorectal cancer tissues. They sincerely thank T. Isobe for development of methods and wonderful lecture. They also thank A. Yurino for providing mice, Yasufumi Uehara for taking confocal microscopy images, T Sugio, T Jiroumaru for performing microarray analysis. Co-workers at our labs, F Hanamura, Yumiko Uehara, K Yamaguchi, K Sagara, K Tsuchihashi, S Arita, supported this work. This study was supported in part by a Grant-in-Aid for Scientific Research (S) (to KA, #16H06391), a Grant-in-Aid for Scientific Research (A) (to KA, #16H02662), a Grant-in-Aid for Scientific Research (C) (to EB, #15K08970), a Grant-in-Aid for Young Scientists (A) (to Y Kikushige #16H06250), a Grant-in-Aid for Young Scientists (B) (to SA, #25860542), a Grant-in-Aid for Scientific Research (B) (to Y Kunisaki, #15H04859), from the JSPS, and a Grant-in-Aid for Scientific Research on Innovative Areas “Stem Cell Aging and Disease” (to TM and Y Kikushige, #25115002) from MEXT, Japan, and the “Project for Cancer Research and Therapeutic Evolution” (to KA, 18cm0106507h0003), and the “Practical Research for Innovative Cancer Control” (to Y Kikushige, 18ck0106196h0003) from Japan Agency for Medical Research and development, AMED, and the Shinnihon Foundation of Advanced Medical Treatment Research (to HA, Y Kikushige, and Y Kunisaki), and a collaborative research grant from AstraZeneca KK (to MN, AZKK Science Promotion Grant).

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Affiliations

  1. Department of Medicine and Biosystemic Sciences, Kyushu University Graduate School of Medical Sciences, Fukuoka, Japan

    • Michitaka Nakano
    • , Yoshikane Kikushige
    • , Kohta Miyawaki
    • , Yuya Kunisaki
    • , Shinichi Mizuno
    • , Katsuto Takenaka
    • , Shingo Tamura
    • , Yuta Okumura
    • , Mamoru Ito
    • , Hiroshi Ariyama
    • , Hitoshi Kusaba
    • , Takahiro Maeda
    • , Eishi Baba
    •  & Koichi Akashi
  2. Department of Comprehensive Clinical Oncology, Faculty of Medical Sciences, Kyushu University, Fukuoka, Japan

    • Eishi Baba
  3. Department of Surgery and Oncology, Kyushu University Graduate School of Medical Sciences, Fukuoka, Japan

    • Masafumi Nakamura

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Conflict of interest

The authors declare that they have no conflict of interest.

Ethics statement

Colorectal cancer tissues of 79 patients were obtained upon resection. All human studies were performed in accordance with Declaration of Helsinki principles and the guidelines of the Research Ethics Committee on Human Experimentation of Kyushu University (#28–95) and written informed consent was received from participants prior to inclusion in the study. Supplementary Tables S1 and S2 provide a summary of the clinical information of the colorectal cancer patients analyzed in this study.

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Correspondence to Eishi Baba.

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https://doi.org/10.1038/s41388-018-0480-0

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