T cell memory relies on the generation of antigen-specific progenitors with stem-like properties. However, the identity of these progenitors has remained unclear, precluding a full understanding of the differentiation trajectories that underpin the heterogeneity of antigen-experienced T cells. We used a systematic approach guided by single-cell RNA-sequencing data to map the organizational structure of the human CD8+ memory T cell pool under physiological conditions. We identified two previously unrecognized subsets of clonally, epigenetically, functionally, phenotypically and transcriptionally distinct stem-like CD8+ memory T cells. Progenitors lacking the inhibitory receptors programmed death-1 (PD-1) and T cell immunoreceptor with Ig and ITIM domains (TIGIT) were committed to a functional lineage, whereas progenitors expressing PD-1 and TIGIT were committed to a dysfunctional, exhausted-like lineage. Collectively, these data reveal the existence of parallel differentiation programs in the human CD8+ memory T cell pool, with potentially broad implications for the development of immunotherapies and vaccines.

Center for Cancer Research National Cancer Institute Bethesda MD USA

Center of Life Sciences Skolkovo Institute of Science and Technology Moscow Russia

Central European Institute of Technology Brno Czech Republic

Department of Biomedical Sciences Humanitas University Pieve Emanuele Milan Italy

Department of Medical Biotechnologies and Translational Medicine University of Milan Milan Italy

Division of Cancer and Genetics Cardiff University School of Medicine Cardiff UK

Division of Infection and Immunity Cardiff University School of Medicine Cardiff UK

Genomic Unit Humanitas Clinical and Research Center IRCCS Rozzano Milan Italy

Humanitas Flow Cytometry Core Humanitas Clinical and Research Center IRCCS Rozzano Milan Italy

Innovative Immunotherapies Unit Division of Immunology Transplantation and Infectious Diseases IRCCS San Raffaele Scientific Institute Milan Italy

Institute of Genetic and Biomedical Research UoS Milan National Research Council Rozzano Milan Italy

Laboratory of Translational Immunology Humanitas Clinical and Research Center IRCCS Rozzano Milan Italy

Pirogov Russian National Research Medical University Moscow Russia

Regensburg Center for Interventional Immunology Regensburg Germany

Shemyakin and Ovchinnikov Institute of Bioorganic Chemistry Moscow Russia

St Jude Children's Research Hospital Memphis TN USA

Systems Immunity Research Institute Cardiff University School of Medicine Cardiff UK

Unit of Clinical and Experimental Immunology Humanitas Clinical and Research Center IRCCS Rozzano Milan Italy

University of Regensburg Regensburg Germany

Vaccine and Infectious Disease Division Fred Hutchinson Cancer Research Center Seattle WA USA

Komentář v

Nat Immunol. 2020 Dec;21(12):1484-1485. doi: 10.1038/s41590-020-00815-y. PubMed

Zobrazit více v PubMed

Kaech SM, Wherry EJ, Ahmed R. Effector and memory T-cell differentiation: implications for vaccine development. Nat Rev Immunol. 2002;2:251–262. PubMed

Gattinoni L, Speiser DE, Lichterfeld M, Bonini C. T memory stem cells in health and disease. Nat Med. 2017;23:18–27. PubMed PMC

Lugli E, Galletti G, Boi SK, Youngblood BA. Stem, effector, and hybrid states of memory CD8+ T cells. Trends Immunol. 2020;41:17–28. PubMed PMC

Gattinoni L, et al. A human memory T cell subset with stem cell-like properties. Nat Med. 2011;17:1290–1297. PubMed PMC

Biasco L, et al. In vivo tracking of T cells in humans unveils decade-long survival and activity of genetically modified T memory stem cells. Sci Transl Med. 2015;7:273ra13 PubMed

Mahnke YD, Brodie TM, Sallusto F, Roederer M, Lugli E. The who’s who of T-cell differentiation: human memory T-cell subsets. Eur J Immunol. 2013;43:2797–2809. PubMed

Graef P, et al. Serial transfer of single-cell-derived immunocompetence reveals stemness of CD8+ central memory T cells. Immunity. 2014;41:116–126. PubMed

Wherry EJ. T cell exhaustion. Nat Immunol. 2011;12:492–499. PubMed

Angelosanto JM, Blackburn SD, Crawford A, Wherry EJ. Progressive loss of memory T cell potential and commitment to exhaustion during chronic viral infection. J Virol. 2012;86:8161–8170. PubMed PMC

Schietinger A, et al. Tumor-specific T cell dysfunction is a dynamic antigen-driven differentiation program initiated early during tumorigenesis. Immunity. 2016;45:389–401. PubMed PMC

Philip M, et al. Chromatin states define tumour-specific T cell dysfunction and reprogramming. Nature. 2017;545:452–456. PubMed PMC

Sen DR, et al. The epigenetic landscape of T cell exhaustion. Science. 2016;354:1165–1169. PubMed PMC

Im SJ, et al. Defining CD8+ T cells that provide the proliferative burst after PD-1 therapy. Nature. 2016;537:417–421. PubMed PMC

Leong YA, et al. CXCR5+ follicular cytotoxic T cells control viral infection in B cell follicles. Nat Immunol. 2016;17:1187–1196. PubMed

Utzschneider DT, et al. T cell factor 1-expressing memory-like CD8+ T cells sustain the immune response to chronic viral infections. Immunity. 2016;45:415–427. PubMed

Brummelman J, et al. High-dimensional single cell analysis identifies stem-like cytotoxic CD8+ T cells infiltrating human tumors. J Exp Med. 2018;215:2520–2535. PubMed PMC

He R, et al. Follicular CXCR5-expressing CD8+ T cells curtail chronic viral infection. Nature. 2016;537:412–428. PubMed

Sade-Feldman M, et al. Defining T cell states associated with response to checkpoint immunotherapy in melanoma. Cell. 2018;175:998–1013.:e20. PubMed PMC

Siddiqui I, et al. Intratumoral Tcf1+PD-1+CD8+ T cells with stem-like properties promote tumor control in response to vaccination and checkpoint blockade immunotherapy. Immunity. 2019;50:195–211.:e10. PubMed

Miller BC, et al. Subsets of exhausted CD8+ T cells differentially mediate tumor control and respond to checkpoint blockade. Nat Immunol. 2019;20:326–336. PubMed PMC

Becht E, et al. Dimensionality reduction for visualizing single-cell data using UMAP. Nat Biotechnol. 2018;37:38–44. PubMed

Dusseaux M, et al. Human MAIT cells are xenobiotic-resistant, tissue-targeted, CD161hi IL-17-secreting T cells. Blood. 2011;117:1250–1259. PubMed

Lugli E, et al. Superior T memory stem cell persistence supports long-lived T cell memory. J Clin Invest. 2013;123:594–599. PubMed PMC

Oberdoerffer S, et al. Regulation of CD45 alternative splicing by heterogeneous ribonucleoprotein, hnRNPLL. Science. 2008;321:686–691. PubMed PMC

Lugli E, et al. Identification, isolation and in vitro expansion of human and nonhuman primate T stem cell memory cells. Nat Protoc. 2013;8:33–42. PubMed PMC

Stephen TL, et al. SATB1 expression governs epigenetic repression of PD-1 in tumor-reactive T cells. Immunity. 2017;46:51–64. PubMed PMC

Yao C, et al. Single-cell RNA-seq reveals TOX as a key regulator of CD8+ T cell persistence in chronic infection. Nat Immunol. 2019;20:890–901. PubMed PMC

Alfei F, et al. TOX reinforces the phenotype and longevity of exhausted T cells in chronic viral infection. Nature. 2019;571:265–269. PubMed

Scott AC, et al. TOX is a critical regulator of tumour-specific T cell differentiation. Nature. 2019;571:270–274. PubMed PMC

Khan O, et al. TOX transcriptionally and epigenetically programs CD8+ T cell exhaustion. Nature. 2019;571:211–218. PubMed PMC

Wang X, et al. TOX promotes the exhaustion of antitumor CD8+ T cells by preventing PD1 degradation in hepatocellular carcinoma. J Hepatol. 2019;71:731–741. PubMed

Seo H, et al. TOX and TOX2 transcription factors cooperate with NR4A transcription factors to impose CD8+ T cell exhaustion. Proc Natl Acad Sci. 2019;116:12410–12415. PubMed PMC

Li J, He Y, Hao J, Ni L, Dong C. High levels of Eomes promote exhaustion of anti-tumor CD8+ T cells. Front Immunol. 2018;9:2981. PubMed PMC

Giordano M, et al. Molecular profiling of CD8 T cells in autochthonous melanoma identifies Maf as driver of exhaustion. EMBO J. 2015;34:2042–2058. PubMed PMC

Gattinoni L, et al. Wnt signaling arrests effector T cell differentiation and generates CD8+ memory stem cells. Nat Med. 2009;15:808–813. PubMed PMC

Kondo T, et al. Notch-mediated conversion of activated T cells into stem cell memory-like T cells for adoptive immunotherapy. Nat Commun. 2017;8:15338. PubMed PMC

Widjaja CE, et al. Proteasome activity regulates CD8+ T lymphocyte metabolism and fate specification. J Clin Invest. 2017;127:3609–3623. PubMed PMC

Yang ZZ, et al. TGF-β upregulates CD70 expression and induces exhaustion of effector memory T cells in B-cell non-Hodgkin’s lymphoma. Leukemia. 2014;28:1872–1884. PubMed PMC

Vodnala SK, et al. T cell stemness and dysfunction in tumors are triggered by a common mechanism. Science. 2019;363:eaau0135. PubMed PMC

Henning AN, Roychoudhuri R, Restifo NP. Epigenetic control of CD8+ T cell differentiation. Nat Rev Immunol. 2018;18:340–356. PubMed PMC

Doering TA, et al. Network analysis reveals centrally connected genes and pathways involved in CD8+ T cell exhaustion versus memory. Immunity. 2012;37:1130–1144. PubMed PMC

Thommen DS, et al. A transcriptionally and functionally distinct PD-1+ CD8+ T cell pool with predictive potential in non-small-cell lung cancer treated with PD-1 blockade. Nat Med. 2018;24:994–1004. PubMed PMC

Utzschneider DT, et al. High antigen levels induce an exhausted phenotype in a chronic infection without impairing T cell expansion and survival. J Exp Med. 2016;213:1819–1834. PubMed PMC

Fuertes Marraco SA, et al. Long-lasting stem cell-like memory CD8+ T cells with a naive-like profile upon yellow fever vaccination. Sci Transl Med. 2015;7:282ra48 PubMed

Akondy RS, et al. Origin and differentiation of human memory CD8 T cells after vaccination. Nature. 2017;552:362–367. PubMed PMC

Price DA, et al. Avidity for antigen shapes clonal dominance in CD8+ T cell populations specific for persistent DNA viruses. J Exp Med. 2005;202:1349–1361. PubMed PMC

Blank CU, et al. Defining ‘T cell exhaustion’. Nat Rev Immunol. 2019;19:665–674. PubMed PMC

Pauken KE, et al. Epigenetic stability of exhausted T cells limits durability of reinvigoration by PD-1 blockade. Science. 2016;354:1160–1165. PubMed PMC

Ghoneim HE, et al. De novo epigenetic programs inhibit PD-1 blockade-mediated T cell rejuvenation. Cell. 2017;170:142–157.:e19. PubMed PMC

West EE, et al. Tight regulation of memory CD8+ T cells limits their effectiveness during sustained high viral load. Immunity. 2011;35:285–298. PubMed PMC

Lugli E, et al. IL-15 delays suppression and fails to promote immune reconstitution in virally suppressed chronically SIV-infected macaques. Blood. 2011;118:2520–2529. PubMed PMC

Roberto A, et al. Role of naive-derived T memory stem cells in T-cell reconstitution following allogeneic transplantation. Blood. 2015;125:2855–2864. PubMed PMC

Falcone L, Casucci M. Exploiting secreted luciferases to monitor tumor progression in vivo. Methods Mol Biol. 2016;1393:105–111. PubMed

Brummelman J, et al. Development, application and computational analysis of high-dimensional fluorescent antibody panels for single-cell flow cytometry. Nat Protoc. 2019;14:1946–1969. PubMed

Newell EW, et al. Combinatorial tetramer staining and mass cytometry analysis facilitate T-cell epitope mapping and characterization. Nat Biotechnol. 2013;31:623–629. PubMed PMC

Bakker AH, et al. Conditional MHC class I ligands and peptide exchange technology for the human MHC gene products HLA-A1, -A3, -A11, and -B7. Proc Natl Acad Sci. 2008;105:3825–3830. PubMed PMC

Simoni Y, et al. Bystander CD8+ T cells are abundant and phenotypically distinct in human tumour infiltrates. Nature. 2018;557:575–579. PubMed

Satija R, Farrell JA, Gennert D, Schier AF, Regev A. Spatial reconstruction of single-cell gene expression data. Nat Biotechnol. 2015;33:495–502. PubMed PMC

Dobin A, et al. STAR: ultrafast universal RNA-seq aligner. Bioinformatics. 2013;29:15–21. PubMed PMC

Ritchie ME, et al. limma powers differential expression analyses for RNA-sequencing and microarray studies. Nucleic Acids Res. 2015;43:e47. PubMed PMC

Leek JT, Johnson WE, Parker HS, Jaffe AE, Storey JD. The sva package for removing batch effects and other unwanted variation in high-throughput experiments. Bioinformatics. 2012;28:882–883. PubMed PMC

Buenrostro JD, Giresi PG, Zaba LC, Chang HY, Greenleaf WJ. Transposition of native chromatin for fast and sensitive epigenomic profiling of open chromatin, DNA-binding proteins and nucleosome position. Nat Methods. 2013;10:1213–1218. PubMed PMC

Martin M. Cutadapt removes adapter sequences from high-throughput sequencing reads. EMBnetjournal. 2011;17:10–12.

Li H, et al. The Sequence Alignment/Map format and SAMtools. Bioinformatics. 2009;25:2078–2079. PubMed PMC

Zhang Y, et al. Model-based analysis of ChIP-Seq (MACS) Genome Biol. 2008;9:R137. PubMed PMC

Liao Y, Smyth GK, Shi W. featureCounts: an efficient general purpose program for assigning sequence reads to genomic features. Bioinformatics. 2014;30:923–930. PubMed

Capper R, et al. The nature of telomere fusion and a definition of the critical telomere length in human cells. Genes Dev. 2007;21:2495–2508. PubMed PMC

Shugay M, et al. Towards error-free profiling of immune repertoires. Nat Methods. 2014;11:653–655. PubMed

Bolotin DA, et al. MiXCR: software for comprehensive adaptive immunity profiling. Nat Methods. 2015;12:380–381. PubMed

Shugay M, et al. VDJtools: unifying post-analysis of T cell receptor repertoires. PLoS Comput Biol. 2015;11:e1004503. PubMed PMC