Positively selected enhancer elements endow osteosarcoma cells with metastatic competence

  • Nature Medicine volume 24, pages 176185 (2018)
  • doi:10.1038/nm.4475
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Metastasis results from a complex set of traits acquired by tumor cells, distinct from those necessary for tumorigenesis. Here, we investigate the contribution of enhancer elements to the metastatic phenotype of osteosarcoma. Through epigenomic profiling, we identify substantial differences in enhancer activity between primary and metastatic human tumors and between near isogenic pairs of highly lung metastatic and nonmetastatic osteosarcoma cell lines. We term these regions metastatic variant enhancer loci (Met-VELs). Met-VELs drive coordinated waves of gene expression during metastatic colonization of the lung. Met-VELs cluster nonrandomly in the genome, indicating that activity of these enhancers and expression of their associated gene targets are positively selected. As evidence of this causal association, osteosarcoma lung metastasis is inhibited by global interruptions of Met-VEL-associated gene expression via pharmacologic BET inhibition, by knockdown of AP-1 transcription factors that occupy Met-VELs, and by knockdown or functional inhibition of individual genes activated by Met-VELs, such as that encoding coagulation factor III/tissue factor (F3). We further show that genetic deletion of a single Met-VEL at the F3 locus blocks metastatic cell outgrowth in the lung. These findings indicate that Met-VELs and the genes they regulate play a functional role in metastasis and may be suitable targets for antimetastatic therapies.

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Change history

  • Corrected online 07 February 2018

    In the version of this article initially published, two of the authors are incorrectly identified as John Stamatoyannopolus and Henri Versteeg. The authors' names are John A Stamatoyannopoulos and Henri H Versteeg. Also, the affiliation "Research Laboratory, Istituto Ortopedico Rizzoli, Bologna, Italy" is incorrect. The correct affiliation is "Laboratory of Experimental Oncology, Istituto Ortopedico Rizzoli, Bologna, Italy". The errors have been corrected in the HTML and PDF versions of the article.


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Download references


The authors thank the members of the Tesar, Kaplan, and Helman laboratories for their input throughout the course of the project as well as B. Decker for his input on the manuscript text. Additional support was provided by the Genomics Core Facility of the Case Western Reserve University School of Medicine's Genetics and Genome Sciences Department and the Case Comprehensive Cancer Center (P30CA043703). This work was supported by the Liddy Shriver Sarcoma Initiative (P.C.S., C.K., J.J.M.), the QuadW Foundation (P.C.S.), Sarcoma Foundation of America (P.C.S.), St. Baldrick's Foundation (A.Y.H.), Alex's Lemonade Stand Foundation (A.Y.H.), Hyundai Hope-on-Wheels Program (A.Y.H.), Pediatric Cancer Research Foundation (A.Y.H.), CCCC AYA Oncology Pilot Grant (A.Y.H.), National Institutes of Health (NIH) grants F30 CA186633 (J.J.M.), F30 CA183510 (T.E.M.), T32 GM007250 (J.J.M., T.E.M., S.H.), R01CA193677 (P.C.S.), R01CA204279 (P.C.S.), R01CA160356 (P.C.S.), F31CA192874 (F.A.), R21CA218790 (A.Y.H.), NIH Intramural Visiting Fellow Program 15335 (M.M.L.), and NIH Intramural Research Program (C.K.).

Author information

Author notes

    • Chand Khanna

    Present address: Ethos Discovery, Washington, DC, USA and Ethos Veterinary Health, Woburn, Massachusetts, USA.


  1. Department of Pathology, Case Western Reserve University, Cleveland, Ohio, USA.

    • James J Morrow
    • , Tyler E Miller
    • , Frederick Allen
    •  & Alex Y Huang
  2. Department of Genetics and Genome Sciences, Case Western Reserve University, Cleveland, Ohio, USA.

    • James J Morrow
    • , Ian Bayles
    • , Alina Saiakhova
    • , Cynthia F Bartels
    • , Stevephen Hung
    • , Gursimran Dhillon
    •  & Peter C Scacheri
  3. Altius Institute for Biomedical Sciences, Seattle, Washington, USA.

    • Alister P W Funnell
    •  & John A Stamatoyannopoulos
  4. Pediatric Oncology Branch, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, Maryland, USA.

    • Michael M Lizardo
    • , Arnulfo Mendoza
    • , Lee J Helman
    •  & Chand Khanna
  5. Thrombosis and Hemostasis Division, Department of Internal Medicine, Leiden University Medical Center, Leiden, the Netherlands.

    • Maaike Y Kapteijn
    •  & Henri H Versteeg
  6. Department of Genome Sciences, University of Washington, Seattle, Washington, USA.

    • Daniel R Chee
  7. Department of Pediatrics, Case Western Reserve University, Cleveland, Ohio, USA.

    • Jay T Myers
    •  & Alex Y Huang
  8. Laboratory of Experimental Oncology, Istituto Ortopedico Rizzoli, Bologna, Italy.

    • Marco Gambarotti
    • , Alberto Righi
    • , Piero Picci
    •  & Peter C Scacheri
  9. Case Comprehensive Cancer Center, Case Western Reserve University, Cleveland, Ohio, USA.

    • Analisa DiFeo
  10. Departments of Anatomic Pathology and Molecular Genetics, Cleveland Clinic, Lerner Research Institute and Taussig Cancer Center, Cleveland, Ohio, USA.

    • Brian P Rubin
  11. Genetics Branch, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, Maryland, USA.

    • Paul S Meltzer


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J.J.M., C.K., and P.C.S. conceived the overall experimental design. J.J.M., C.F.B., and G.D. generated ChIP–seq, RNA-seq, and DHS-seq data. J.J.M., A.S., S.H., and P.C.S. completed analyses of ChIP–seq, RNA-seq, and DHS-seq data. J.J.M. and T.E.M. designed and completed the shRNA screening experiment and subsequent analysis. T.E.M. completed functional enrichment analysis of RNA-seq data. J.J.M. and I.B. generated 4C-seq data. J.J.M. and A.S. analyzed 4C-seq data. J.J.M., A.M., and I.B. and M.M.L. completed the in vivo and ex vivo metastasis experiments. J.J.M., J.T.M., and F.A. designed and completed the orthotopic metastasis experiments. J.J.M., D.R.C., and A.P.W.F. designed and completed the TALEN deletion experiments. M.Y.K. completed the in vitro F3 experiments. M.G., A.R., and P.P. provided subject tumor samples and clinical data. B.P.R. assessed F3 staining in subject tissue microarrays. A.D., A.Y.H., P.S.M., L.J.H., H.H.V., J.A.S., C.K., and P.C.S. provided the technical expertise and facilities to complete the experiments. J.J.M. and P.C.S. analyzed all data and wrote the paper. All authors provided intellectual input, edited, and approved the final manuscript.

Competing interests

The authors declare no competing financial interests.

Corresponding author

Correspondence to Peter C Scacheri.

Supplementary information

PDF files

  1. 1.

    Supplementary Figures & Tables

    Supplementary Figures 1–13 and Supplementary Table 1

  2. 2.

    Life Sciences Reporting Summary

Excel files

  1. 1.

    Supplementary Table 2

    Met-VEL gene overlaps across patient tumors and cell lines

  2. 2.

    Supplementary Table 3

    Hairpins used in high-throughput in vivo RNAi screen