Abstract
The quantity of tumor-infiltrating lymphocytes (TILs) in breast cancer (BC) is a robust prognostic factor for improved patient survival, particularly in triple-negative and HER2-overexpressing BC subtypes1. Although T cells are the predominant TIL population2, the relationship between quantitative and qualitative differences in T cell subpopulations and patient prognosis remains unknown. We performed single-cell RNA sequencing (scRNA-seq) of 6,311 T cells isolated from human BCs and show that significant heterogeneity exists in the infiltrating T cell population. We demonstrate that BCs with a high number of TILs contained CD8+ T cells with features of tissue-resident memory T (TRM) cell differentiation and that these CD8+ TRM cells expressed high levels of immune checkpoint molecules and effector proteins. A CD8+ TRM gene signature developed from the scRNA-seq data was significantly associated with improved patient survival in early-stage triple-negative breast cancer (TNBC) and provided better prognostication than CD8 expression alone. Our data suggest that CD8+ TRM cells contribute to BC immunosurveillance and are the key targets of modulation by immune checkpoint inhibition. Further understanding of the development, maintenance and regulation of TRM cells will be crucial for successful immunotherapeutic development in BC.
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Change history
22 August 2018
In the version of this article originally published, the institution in affiliation 10 was missing. Affiliation 10 was originally listed as Department of Surgery, Royal Melbourne Hospital and Royal Womens’ Hospital, Melbourne, Victoria, Australia. It should have been Department of Surgery, Royal Melbourne Hospital and Royal Womens’ Hospital, University of Melbourne, Melbourne, Victoria, Australia. The error has been corrected in the HTML and PDF versions of this article.
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Acknowledgements
We wish to thank H. Thorne, E. Niedermayr, all the kConFab research nurses and staff, the heads and staff of the Family Cancer Clinics and the Clinical Follow Up Study (which has received funding from the National Health and Medical Research Council of Australia (NHMRC), the National Breast Cancer Foundation (NBCF), Cancer Australia and the National Institute of Health (United States) for their contributions to this research, and the many families who contribute to kConFab. We wish to thank the FACS core facility staff R. Rossi, V. Milovac and S. Curcio, and T. Tan and P. Petrone for additional FACS assistance. We also thank S. Ellis for assistance with confocal imaging, G. M. Arnau for facilitating RNA-seq, and the Anatomical Pathology staff at the Peter MacCallum Cancer Centre. Special thanks also to J. Jabbari and the Australian Genome Research Facility for making the single-cell sequencing possible.
This study was funded by the Breast Cancer Research Foundation (BCRF), NY. S.L. is supported by the Cancer Council Victoria John Colebatch Fellowship and the National Breast Cancer Foundation. P.S. is supported by the NHMRC and the NBCF (Post Graduate Scholarship 1094388), the Cancer Therapeutics CRC and the Peter Mac Foundation. Z.L.T. is supported by the NHMRC (Early Career Fellowship 1106967). D.G. is supported by the Peter Mac Foundation. P.A.B. is supported by the NHMRC (Early Career Fellowship 17-005). S.J.L is supported by the University of Melbourne. S.B.F. is supported by the NHMRC (Practitioner Fellowship 1079329). kConFab is supported by a grant from NBCF, and previously NHMRC, the Queensland Cancer Fund, the Cancer Councils of New South Wales, Victoria, Tasmania and South Australia, and the Cancer Foundation of Western Australia. P.K.D is supported by the NHMRC (Senior Research Fellowship 1136680 and Program Grant 1132373). T.S. is supported by the NHMRC (Program Grant 1054618). P.J.N. is supported by the NHMRC (Program Grant 1132373).
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P.S. conceived and designed the study, provided and collected study materials and samples and patient data, performed experiments, analyzed data and wrote the manuscript. B.V. designed the study, provided and collected study materials and samples and patient data, performed experiments, analyzed data and wrote the manuscript. C.Y. developed analysis methods and software, analyzed data and wrote the manuscript. A.S. developed analysis methods and software, analyzed single-cell sequencing data and wrote the manuscript. C.P.M performed experiments, analyzed data and wrote the manuscript. F.C. analyzed data and wrote the manuscript. R.S. analyzed data and wrote the manuscript. D.J.B performed experiments and wrote the manuscript. Z.L.T. provided and collected study materials and samples, analyzed data and wrote the manuscript. S.D. performed experiments, analyzed data and wrote the manuscript. A.B. performed experiments, analyzed data and wrote the manuscript. L.W. provided and collected study materials and samples and patient data and wrote the manuscript. S.J.L. provided and collected study materials and samples and patient data and wrote the manuscript. C.P. provided and collected study materials and samples and patient data. S.S.N. provided and collected study materials and samples and patient data. A.S.S. provided and collected study materials and samples and patient data. D.E.G. provided and collected study materials and samples and patient data. C.M.T. provided and collected study materials and samples and patient data. P.A.B. analyzed data and wrote the manuscript. S.B.F provided and collected study materials and samples and patient data, analyzed data and wrote the manuscript. kConFab provided and collected study materials and samples. P.K.D. designed the study, analyzed data and wrote the manuscript. T.P.S. developed analysis methods and software, designed the study and wrote the manuscript. L.K.M. designed the study and wrote the manuscript. P.J.N. designed the study and wrote the manuscript. S.L. conceived and designed the study, provided and collected study materials and samples and patient data and wrote the manuscript.
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Supplementary Figures 1–15 and Supplementary Table 1
Supplementary Table 2
CD8 TRM signatures derived from single-cell sequencing.
Supplementary Table 3
Differential expression between all clusters in single-cell sequencing.
Supplementary Table 4
Differential expression between flow sorted CD8+CD69+CD103+ and CD8+CD69+CD103 T cells from 2 primary breast (TNBC and HER2) cancers and 1 liver metastasis (TNBC) from 3 different patients.
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Savas, P., Virassamy, B., Ye, C. et al. Single-cell profiling of breast cancer T cells reveals a tissue-resident memory subset associated with improved prognosis. Nat Med 24, 986–993 (2018). https://doi.org/10.1038/s41591-018-0078-7
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DOI: https://doi.org/10.1038/s41591-018-0078-7
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