Somatic gene mutations can alter the vulnerability of cancer cells to T-cell-based immunotherapies. Here we perturbed genes in human melanoma cells to mimic loss-of-function mutations involved in resistance to these therapies, by using a genome-scale CRISPR–Cas9 library that consisted of around 123,000 single-guide RNAs, and profiled genes whose loss in tumour cells impaired the effector function of CD8+ T cells. The genes that were most enriched in the screen have key roles in antigen presentation and interferon-γ signalling, and correlate with cytolytic activity in patient tumours from The Cancer Genome Atlas. Among the genes validated using different cancer cell lines and antigens, we identified multiple loss-of-function mutations in APLNR, encoding the apelin receptor, in patient tumours that were refractory to immunotherapy. We show that APLNR interacts with JAK1, modulating interferon-γ responses in tumours, and that its functional loss reduces the efficacy of adoptive cell transfer and checkpoint blockade immunotherapies in mouse models. Our results link the loss of essential genes for the effector function of CD8+ T cells with the resistance or non-responsiveness of cancer to immunotherapies.

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The research was supported by the Intramural Research Program of the NCI, and by the Cancer Moonshot program for the Center for Cell-based Therapy at the NCI, NIH. The work was also supported by the Milstein Family Foundation. We thank S. A. Rosenberg, K. Hanada, A. Wellstein, C. Hurley and L. M. Weiner for their valuable discussions and intellectual input, M. Kruhlak, Z. Yu, C. Subramaniam, C. Kariya, A. J. Leonardi, N. Ha, H. Xu, M. A. Black and H. Chinnasamy for technical assistance in this project. This work used the computational resources of the NIH HPC Biowulf cluster (http://hpc.nih.gov). The results here are in part based upon data generated by the TCGA Research Network: http://cancergenome.nih.gov/. This study was done in partial fulfilment of a PhD in Tumor Biology to S.J.P. N.E.S. is supported by the NIH through NHGRI (R00-HG008171) and a Sidney Kimmel Scholar Award.

Author information

Author notes

    • Shashank J. Patel
    •  & Neville E. Sanjana

    These authors contributed equally to this work.


  1. National Cancer Institute, National Institutes of Health (NIH), Bethesda, Maryland 20892, USA

    • Shashank J. Patel
    • , Rigel J. Kishton
    • , Arash Eidizadeh
    • , Suman K. Vodnala
    • , Maggie Cam
    • , Jared J. Gartner
    • , Li Jia
    • , Seth M. Steinberg
    • , Tori N. Yamamoto
    • , Anand S. Merchant
    • , Gautam U. Mehta
    • , Anna Chichura
    • , Eric Tran
    • , Robert Eil
    • , Madhusudhanan Sukumar
    • , Eva Perez Guijarro
    • , Chi-Ping Day
    • , Paul Robbins
    • , Steve Feldman
    • , Glenn Merlino
    •  & Nicholas P. Restifo
  2. NIH-Georgetown University Graduate Partnership Program, Georgetown University Medical School, Washington DC 20057, USA

    • Shashank J. Patel
  3. New York Genome Center, New York, New York 10013, USA

    • Neville E. Sanjana
  4. Department of Biology, New York University, New York, New York 10012, USA

    • Neville E. Sanjana
  5. Immunology Graduate Group, University of Pennsylvania, Philadelphia, Pennsylvania 19104, USA

    • Tori N. Yamamoto
  6. Children’s Hospital of Philadelphia and Department of Genetics, University of Pennsylvania, Pennsylvania 19104, USA

    • Ophir Shalem
  7. Broad Institute of MIT and Harvard, Cambridge, Massachusetts 02142, USA

    • Feng Zhang
  8. McGovern Institute for Brain Research, Massachusetts Institute of Technology, Cambridge, Massachusetts 02139, USA

    • Feng Zhang
  9. Center for Cell-based Therapy, Center for Cancer Research, National Institutes of Health (NIH), Bethesda, Maryland 20892, USA

    • Nicholas P. Restifo


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S.J.P., N.E.S., and N.P.R. designed the study and wrote the manuscript. S.J.P. carried out CRISPR screens and validation experiments. N.E.S., O.S. and S.J.P. analysed CRISPR screen data. S.J.P. and N.E.S. analysed human mutation datasets from immunotherapy cohorts. T.N.Y., G.U.M., A.C., M.S. and S.F. assisted in generation of TCR-engineered T cells and CRISPR-edited cells. R.E., A.E., T.N.Y., S.K.V., G.U.M., A.C. and M.S. edited the manuscript. S.J.P., A.E. and S.K.V. carried out mouse experiments. G.M., E.P.G. and C.-P.D. developed B2905 mouse model for anti-CTLA4 experiments. S.K.V. and L.J. analysed RNA-seq data. M.C. and A.S.M. analysed TCGA datasets. J.J.G. performed indel analyses. S.M.S. analysed clinical data. R.J.K. performed western blots and immunoprecipitation experiments. F.Z., E.T. and P.R. contributed reagents. N.P.R. supervised the study.

Competing interests

The authors declare no competing financial interests.

Corresponding authors

Correspondence to Shashank J. Patel or Neville E. Sanjana or Nicholas P. Restifo.

Reviewer Information Nature thanks R. Levine and the other anonymous reviewer(s) for their contribution to the peer review of this work.

Publisher's note: Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

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