Innate lymphoid cells (ILCs) are emerging as a family of effectors and regulators of innate immunity and tissue remodeling. Interleukin 22 (IL-22)- and IL-17-producing ILCs, which depend on the transcription factor RORγt, express CD127 (IL-7 receptor α-chain) and the natural killer cell marker CD161. Here we describe another lineage-negative CD127(+)CD161(+) ILC population found in humans that expressed the chemoattractant receptor CRTH2. These cells responded in vitro to IL-2 plus IL-25 and IL-33 by producing IL-13. CRTH2(+) ILCs were present in fetal and adult lung and gut. In fetal gut, these cells expressed IL-13 but not IL-17 or IL-22. There was enrichment for CRTH2(+) ILCs in nasal polyps of chronic rhinosinusitis, a typical type 2 inflammatory disease. Our data identify a unique type of human ILC that provides an innate source of T helper type 2 (T(H)2) cytokines.

Tissue-resident memory T cells (T cells) in the airways mediate protection against respiratory infection. We characterized T cells expressing integrin α (CD103) that reside within the epithelial barrier of human lungs. These cells had specialized profiles of chemokine receptors and adhesion molecules, consistent with their unique localization. Lung T cells were poised for rapid responsiveness by constitutive expression of deployment-ready mRNA encoding effector molecules, but they also expressed many inhibitory regulators, suggestive of programmed restraint. A distinct set of transcription factors was active in CD103 T cells, including Notch. Genetic and pharmacological experiments with mice revealed that Notch activity was required for the maintenance of CD103 T cells. We have thus identified specialized programs underlying the residence, persistence, vigilance and tight control of human lung T cells.

Resident memory T cells (T) reside in the lung epithelium and mediate protective immunity against respiratory pathogens. Although lung CD8 T have been extensively characterized, the properties of CD4 T remain unclear. Here we determined the transcriptional signature of CD4 T, identified by the expression of CD103, retrieved from human lung resection material. Various tissue homing molecules were specifically upregulated on CD4 T, whereas expression of tissue egress and lymph node homing molecules were low. CD103 T expressed low levels of T-bet, only a small portion expressed Eomesodermin (Eomes), and although the mRNA levels for Hobit were increased, protein expression was absent. On the other hand, the CD103 T showed a Notch signature. CD4CD103 T constitutively expressed high transcript levels of numerous cytotoxic mediators that was functionally reflected by a fast recall response, magnitude of cytokine production, and a high degree of polyfunctionality. Interestingly, the superior cytokine production appears to be because of an accessible interferon-γ (IFNγ) locus and was partially because of rapid translation of preformed mRNA. Our studies provide a molecular understanding of the maintenance and potential function of CD4 T in the human lung. Understanding the specific properties of CD4 T is required to rationally improve vaccine design.

The human lung T cell compartment contains many CD8 + T cells specific for respiratory viruses, suggesting that the lung is protected from recurring respiratory infections by a resident T cell pool. The entry site for respiratory viruses is the epithelium, in which a subset of lung CD8 + T cells expressing CD103 (αE integrin) resides. Here, we determined the specificity and function of CD103 + CD8 + T cells in protecting human lung against viral infection. Mononuclear cells were isolated from human blood and lung resection samples. Variable numbers of CD103 + CD8 + T cells were retrieved from the lung tissue. Interestingly, expression of CD103 was seen only in lung CD8 + T cells specific for influenza but not in those specific for EBV or CMV. CD103 + and influenza-reactive cells preferentially expressed NKG2A, an inhibitor of CD8 + T cell cytotoxic function. In contrast to CD103 -CD8 + T cells, most CD103 + CD8 + cells did not contain perforin or granzyme B. However, they could quickly upregulate these cytotoxic mediators when exposed to a type I IFN milieu or via contact with their specific antigen. This mechanism may provide a rapid and efficient response to influenza infection, without inducing cytotoxic damage to the delicate epithelial barrier.

Resident memory T cells (TRM) inhabit peripheral tissues and are critical for protection against localized infections. Recently, it has become evident that CD103+ TRM are not only important in combating secondary infections, but also for the elimination of tumor cells. In several solid cancers, intratumoral CD103+CD8+ tumor infiltrating lymphocytes (TILs), with TRM properties, are a positive prognostic marker. To better understand the role of TRM in tumors, we performed a detailed characterization of CD8+ and CD4+ TIL phenotype and functional properties in non-small cell lung cancer (NSCLC). Frequencies of CD8+ and CD4+ T cell infiltrates in tumors were comparable, but we observed a sharp contrast in TRM ratios compared to surrounding lung tissue. The majority of both CD4+ and CD8+ TILs expressed CD69 and a subset also expressed CD103, both hallmarks of TRM. While CD103+CD8+ T cells were enriched in tumors, CD103+CD4+ T cell frequencies were decreased compared to surrounding lung tissue. Furthermore, CD103+CD4+ and CD103+CD8+ TILs showed multiple characteristics of TRM, such as elevated expression of CXCR6 and CD49a, and decreased expression of T-bet and Eomes. In line with the immunomodulatory role of the tumor microenvironment, CD8+ and CD4+ TILs expressed high levels of inhibitory receptors 2B4, CTLA-4, and PD-1, with the highest levels found on CD103+ TILs. Strikingly, CD103+CD4+ TILs were the most potent producers of TNF-α and IFN-γ, while other TIL subsets lacked such cytokine production. Whereas, CD103+CD4+PD-1low TILs produced the most effector cytokines, CD103+CD4+PD-1++ and CD69+CD4+PD-1++ TILs produced CXCL13. Furthermore, a large proportion of TILs expressed co-stimulatory receptors CD27 and CD28, unlike lung TRM, suggesting a less differentiated phenotype. Agonistic triggering of these receptors improved cytokine production of CD103+CD4+ and CD69+CD8+ TILs. Our findings thus provide a rationale to target CD103+CD4+ TILs and add co-stimulation to current therapies to improve the efficacy of immunotherapies and cancer vaccines.

CD161(++)IL-18Rα(+)CD8(+) human T cells have recently been identified as a new subset of memory cells but their exact role remains unclear. CD161(++)IL-18Rα(+)CD8(+), mucosal-associated invariant T cells express a semi-invariant TCR Vα7.2-Jα33, which recognizes the MHC-related protein 1. On the basis of properties including the expression of the ABC-B1 transporter, cKit expression and survival after chemotherapy, CD161(++)IL-18Rα(+)CD8(+) T cells have been designated as 'stem' cells. Here we analyse location and functional properties of CD161(++)IL-18Rα(+) CD8(+) T cells and question whether they have other traits that would mark them as genuine 'stem' cells. CD161(++)IL-18Rα(+)CD8(+) T cells were found in peripheral blood, spleen and bone marrow but interestingly hardly at all in lymph nodes (LNs), which may possibly be explained by the finding that these cells express a specific set of chemokine receptors that allows migration to inflamed tissue rather than to LNs. In addition to TCR ligation and co-stimulation, CD161(++)IL-18Rα(+) CD8(+) T cells require cytokines for proliferation. The CD161(++)IL-18Rα(+) CD8(+) pool contains cells reactive towards peptides, derived from both persisting and cleared viruses. Although CD161(++)IL-18Rα(+) CD8(+) T cells express the ABC-B1 transporter, they have shorter telomeres and less telomerase activity and do not express aldehyde dehydrogenase. Finally, CD161(++)IL-18Rα(+) CD8(+) T cells show similarities to terminally differentiated T cells, expressing IFNγ, KLRG1 and the transcription factor Blimp-1. In conclusion, CD161(++)IL-18Rα(+) CD8(+) T cells lack many features of typical 'stem' cells, but appear rather to be a subset of effector-type cells.

Immunotherapy promotes the attack of cancer cells by the immune system; however, it is difficult to detect early responses before changes in tumor size occur. Here, we report the rational design of a fluorogenic peptide able to detect picomolar concentrations of active granzyme B as a biomarker of immune-mediated anticancer action. Through a series of chemical iterations and molecular dynamics simulations, we synthesize a library of FRET peptides and identify probe H5 with an optimal fit into granzyme B. We demonstrate that probe H5 enables the real-time detection of T cell-mediated anticancer activity in mouse tumors and in tumors from lung cancer patients. Furthermore, we show image-based phenotypic screens, which reveal that the AKT kinase inhibitor AZD5363 shows immune-mediated anticancer activity. The reactivity of probe H5 may enable the monitoring of early responses to anticancer treatments using tissue biopsies.

Immune checkpoint inhibitors (ICIs) have provided tremendous clinical benefit in several cancer types. However, systemic activation of the immune system also leads to several immune-related adverse events. Of these, ICI-mediated colitis (IMC) occurs frequently and is the one with the highest absolute fatality. To improve current treatment strategies, it is important to understand the cellular mechanisms that induce this form of colitis. In this review, we discuss important pathways that are altered in IMC in mouse models and in human colon biopsy samples. This reveals a complex interplay between several types of immune cells and the gut microbiome. In addition to a mechanistic understanding, patients at risk should be identifiable before ICI therapy. Here we propose to focus on T-cell subsets that interact with bacteria after inducing epithelial damage. Especially, intestinal resident immune cells are of interest. This may lead to a better understanding of IMC and provides opportunities for prevention and management.