Although human tumours are shaped by the genetic evolution of cancer cells, evidence also suggests that they display hierarchies related to developmental pathways and epigenetic programs in which cancer stem cells (CSCs) can drive tumour growth and give rise to differentiated progeny1. Yet, unbiased evidence for CSCs in solid human malignancies remains elusive. Here we profile 4,347 single cells from six IDH1 or IDH2 mutant human oligodendrogliomas by RNA sequencing (RNA-seq) and reconstruct their developmental programs from genome-wide expression signatures. We infer that most cancer cells are differentiated along two specialized glial programs, whereas a rare subpopulation of cells is undifferentiated and associated with a neural stem cell expression program. Cells with expression signatures for proliferation are highly enriched in this rare subpopulation, consistent with a model in which CSCs are primarily responsible for fuelling the growth of oligodendroglioma in humans. Analysis of copy number variation (CNV) shows that distinct CNV sub-clones within tumours display similar cellular hierarchies, suggesting that the architecture of oligodendroglioma is primarily dictated by developmental programs. Subclonal point mutation analysis supports a similar model, although a full phylogenetic tree would be required to definitively determine the effect of genetic evolution on the inferred hierarchies. Our single-cell analyses provide insight into the cellular architecture of oligodendrogliomas at single-cell resolution and support the cancer stem cell model, with substantial implications for disease management.

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We thank L. Gaffney for graphic support. This work was supported by grants from the National Brain Tumor Society (to M.L.S. and D.N.L.), the Smith Family Foundation (to M.L.S.), NIH-NCI SPORE on brain cancer Career Enhancement Project and Developmental Research Project (to M.L.S.), the Broad Institute Broadnext10 program (to M.L.S. and O.R.R.), the American Cancer Society (to M.L.S.) and start-up funds from the MGH department of Pathology. A.S.V. was supported by the NIH R25 fellowship (NS065743) and research grants from the American Brain Tumor Association and Neurosurgery Research and Education Foundation. I.T. was supported by a Human Frontier Science Program fellowship and a Rothschild fellowship. A.R. was supported by funds from the Howard Hughes Medicine Institute, the Klarman Cell Observatory, STARR cancer consortium, NCI grant 1U24CA180922, by the Koch Institute Support (core) grant P30-CA14051 from the National Cancer Institute, the Ludwig Center and the Broad Institute. A.R. is a scientific advisory board member for ThermoFisher Scientific and Syros Pharmaceuticals and a consultant for Driver Group. Flow cytometry and sorting services were supported by shared instrumentation grant 1S10RR023440-01A1. M.M. was supported by the California Institute of Regenerative Medicine (CIRM) grants RB4-06093 and RN3-06510 and the Virginia and D.K. Ludwig Fund for Cancer Research.

Author information

Author notes

    • Itay Tirosh
    •  & Andrew S. Venteicher

    These authors contributed equally to this work.

    • Aviv Regev
    •  & Mario L. Suvà

    These authors jointly supervised this work.


  1. Broad Institute of Harvard and MIT, Cambridge, Massachusetts 02142, USA

    • Itay Tirosh
    • , Andrew S. Venteicher
    • , Christine Hebert
    • , Leah E. Escalante
    • , Keren Yizhak
    • , Jonathan M. Fisher
    • , Christopher Rodman
    • , Mariella G. Filbin
    • , Cyril Neftel
    • , Jackson Nyman
    • , Benjamin Izar
    • , Joshua M. Francis
    • , Kenneth J. Livak
    • , Dave Gennert
    • , Rahul Satija
    • , Todd R. Golub
    • , Miguel N. Rivera
    • , Gad Getz
    • , Orit Rozenblatt-Rosen
    • , Bradley E. Bernstein
    • , Aviv Regev
    •  & Mario L. Suvà
  2. Department of Pathology and Center for Cancer Research, Massachusetts General Hospital and Harvard Medical School, Boston, Massachusetts 02114, USA

    • Andrew S. Venteicher
    • , Christine Hebert
    • , Leah E. Escalante
    • , Keren Yizhak
    • , Mariella G. Filbin
    • , Cyril Neftel
    • , Niyati Desai
    • , Christina C. Luo
    • , Aanand A. Patel
    • , Maristela L. Onozato
    • , Nicolo Riggi
    • , Ravindra Mylvaganam
    • , A. John Iafrate
    • , Matthew P. Frosch
    • , Miguel N. Rivera
    • , Gad Getz
    • , Bradley E. Bernstein
    • , David N. Louis
    •  & Mario L. Suvà
  3. Department of Neurosurgery, Massachusetts General Hospital and Harvard Medical School, Boston, Massachusetts 02114, USA

    • Andrew S. Venteicher
    • , Anoop P. Patel
    • , Brian V. Nahed
    • , William T. Curry
    • , Robert L. Martuza
    •  & Daniel P. Cahill
  4. Departments of Neurology, Neurosurgery, Pediatrics and Pathology, Stanford University School of Medicine, Stanford, California 94305, USA

    • Christopher Mount
    •  & Michelle Monje
  5. Department of Pediatric Oncology, Dana-Farber Cancer Institute and Children’s Hospital Cancer Center, Boston, Massachusetts 02215, USA

    • Mariella G. Filbin
    •  & Todd R. Golub
  6. Department of Medical Oncology, Dana-Farber Cancer Institute, Boston, Massachusetts 02215, USA

    • Joshua M. Francis
  7. Howard Hughes Medical Institute, Koch Institute, Department of Biology, MIT, Cambridge, Massachusetts 02139, USA

    • Todd R. Golub
    •  & Aviv Regev


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I.T., A.S.V., A.R. and M.L.S. conceived the project, designed the study, and interpreted results. A.S.V., C.H., L.E.E. and C.N. collected single cells and generated single-cell sequencing data. I.T. performed computational analyses. J.M.Fra, K.Y. and G.G. provided support for genomic and genetic analyses. J.M.Fis, C.R. and K.J.L. designed and performed qPCR experiments. C.C.L. and R.M. provided flow cytometry expertise. C.M. and M.M. developed normal human cell cultures used in the study. N.D., N.R., M.N.R., M.L.O. and A.J.I. performed in situ hybridization and FISH experiments. A.P.P., A.A.P., D.G., B.I., J.N., R.S., M.G.F., B.V.N., D.P.C., W.T.C., R.L.M., M.P.F., O.R.R., T. R.G., B.E.B. and D.N.L. provided experimental and analytical support. I.T., A.R. and M.L.S. wrote the manuscript with feedback from all authors.

Competing interests

The authors declare no competing financial interests.

Corresponding authors

Correspondence to Aviv Regev or Mario L. Suvà.

Reviewer Information Nature thanks P. Dirks, J. Rich and the other anonymous reviewer(s) for their contribution to the peer review of this work.

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