IFNγ mediates the resistance of tumor cells to distinct NK cell subsets.

BACKGROUND: Immune checkpoint blockade targeting the adaptive immune system has revolutionized the treatment of cancer. Despite impressive clinical benefits observed, patient subgroups remain non-responsive underscoring the necessity for combinational therapies harnessing additional immune cells. Natural killer (NK) cells are emerging tools for cancer therapy. However, only subpopulations of NK cells that are differentially controlled by inhibitory receptors exert reactivity against particular cancer types. How to leverage the complete anti-tumor potential of all NK cell subsets without favoring the emergence of NK cell-resistant tumor cells remains unresolved. METHODS: We performed a genome-wide CRISPR/Cas9 knockout resistance screen in melanoma cells in co-cultures with human primary NK cells. We comprehensively evaluated factors regulating tumor resistance and susceptibility by focusing on NK cell subsets in an allogenic setting. Moreover, we tested therapeutic blocking antibodies currently used in clinical trials. RESULTS: Melanoma cells deficient in antigen-presenting or the IFNγ-signaling pathways were depleted in remaining NK cell-co-cultured melanoma cells and displayed enhanced sensitivity to NK cells. Treatment with IFNγ induced potent resistance of melanoma cells to resting, IL-2-cultured and ADCC-activated NK cells that depended on B2M required for the expression of both classical and non-classical MHC-I. IFNγ-induced expression of HLA-E mediated the resistance of melanoma cells to the NKG2A+ KIR- and partially to the NKG2A+ KIR+ NK cell subset. The expression of classical MHC-I by itself was sufficient for the inhibition of the NKG2A- KIR+, but not the NKG2A+ KIR+ NK cell subset. Treatment of NK cells with monalizumab, an NKG2A blocking mAb, enhanced the reactivity of a corresponding subset of NK cells. The combination of monalizumab with lirilumab, blocking KIR2 receptors, together with DX9, blocking KIR3DL1, was required to restore cytotoxicity of all NK cell subsets against IFNγ-induced resistant tumor cells in melanoma and tumors of different origins. CONCLUSION: Our data reveal that in the context of NK cells, IFNγ induces the resistance of tumor cells by the upregulation of classical and non-classical MHC-I. Moreover, we reveal insights into NK cell subset reactivity and propose a therapeutic strategy involving combinational monalizumab/lirilumab/DX9 treatment to fully restore the antitumor response across NK cell subsets.

The immunoglobulin superfamily ligand B7H6 subjects T cell responses to NK cell surveillance.

Understanding the mechanisms that regulate T cell immunity is critical for the development of effective therapies for diseases associated with T cell dysfunction, including autoimmune diseases, chronic infections, and cancer. Co-inhibitory "checkpoint molecules," such as programmed cell death protein-1, balance excessive or prolonged immune activation by T cell-intrinsic signaling. Here, by screening for mediators of natural killer (NK) cell recognition on T cells, we identified the immunoglobulin superfamily ligand B7H6 to be highly expressed by activated T cells, including patient-infused CD19-targeting chimeric antigen receptor (CAR) T cells. Unlike other checkpoint molecules, B7H6 mediated NKp30-dependent recognition and subsequent cytolysis of activated T cells by NK cells. B7H6+ T cells were prevalent in the tissue and blood of several diseases, and their abundance in tumor tissue positively correlated with clinical response in a cohort of patients with immune checkpoint inhibitor-treated esophageal cancer. In humanized mouse models, NK cell surveillance via B7H6 limited the persistence and antitumor activity of CAR T cells, and its genetic deletion enhanced T cell proliferation and persistence. Together, we provide evidence of B7H6 protein expression by activated T cells and suggest the B7H6-NKp30 axis as a therapeutically actionable NK cell-dependent immune checkpoint that regulates human T cell function.

Vitamin A-treated natural killer cells reduce interferon-gamma production and support regulatory T-cell differentiation.

Natural killer (NK) cells are innate cytotoxic lymphocytes that contribute to immune responses against stressed, transformed, or infected cells. NK cell effector functions are regulated by microenvironmental factors, including cytokines, metabolites, and nutrients. Vitamin A is an essential micronutrient that plays an indispensable role in embryogenesis and development, but was also reported to regulate immune responses. However, the role of vitamin A in regulating NK cell functions remains poorly understood. Here, we show that the most prevalent vitamin A metabolite, all-trans retinoic acid (atRA), induces transcriptional and functional changes in NK cells leading to altered metabolism and reduced IFN-γ production in response to a wide range of stimuli. atRA-exposed NK cells display a reduced ability to support dendritic cell (DC) maturation and to eliminate immature DCs. Moreover, they support the polarization and proliferation of regulatory T cells. These results imply that in vitamin A-enriched environments, NK cells can acquire functions that might promote tolerogenic immunity and/or immunosuppression.

Hepatitis D infection induces IFN-ß-mediated NK cell activation and TRAIL-dependent cytotoxicity.

Background and aims: The co-infection of hepatitis B (HBV) patients with the hepatitis D virus (HDV) causes the most severe form of viral hepatitis and thus drastically worsens the course of the disease. Therapy options for HBV/HDV patients are still limited. Here, we investigated the potential of natural killer (NK) cells that are crucial drivers of the innate immune response against viruses to target HDV-infected hepatocytes. Methods: We established in vitro co-culture models using HDV-infected hepatoma cell lines and human peripheral blood NK cells. We determined NK cell activation by flow cytometry, transcriptome analysis, bead-based cytokine immunoassays, and NK cell-mediated effects on T cells by flow cytometry. We validated the mechanisms using CRISPR/Cas9-mediated gene deletions. Moreover, we assessed the frequencies and phenotype of NK cells in peripheral blood of HBV and HDV superinfected patients. Results: Upon co-culture with HDV-infected hepatic cell lines, NK cells upregulated activation markers, interferon-stimulated genes (ISGs) including the death receptor ligand tumor necrosis factor-related apoptosis-inducing ligand (TRAIL), produced interferon (IFN)-γ and eliminated HDV-infected cells via the TRAIL-TRAIL-R2 axis. We identified IFN-ß released by HDV-infected cells as an important enhancer of NK cell activity. In line with our in vitro data, we observed activation of peripheral blood NK cells from HBV/HDV co-infected, but not HBV mono-infected patients. Conclusion: Our data demonstrate NK cell activation in HDV infection and their potential to eliminate HDV-infected hepatoma cells via the TRAIL/TRAIL-R2 axis which implies a high relevance of NK cells for the design of novel anti-viral therapies.

Liver sinusoidal endothelial cells orchestrate NK cell recruitment and activation in acute inflammatory liver injury.

Liver sinusoidal endothelial cells (LSECs) rapidly clear lipopolysaccharide (LPS) from the bloodstream and establish intimate contact with immune cells. However, their role in regulating liver inflammation remains poorly understood. We show that LSECs modify their chemokine expression profile driven by LPS or interferon-γ (IFN-γ), resulting in the production of the myeloid- or lymphoid-attracting chemokines CCL2 and CXCL10, respectively, which accumulate in the serum of LPS-challenged animals. Natural killer (NK) cell exposure to LSECs in vitro primes NK cells for higher production of IFN-γ in response to interleukin-12 (IL-12) and IL-18. In livers of LPS-injected mice, NK cells are the major producers of this cytokine. In turn, LSECs require exposure to IFN-γ for CXCL10 expression, and endothelial-specific Cxcl10 gene deletion curtails NK cell accumulation in the inflamed livers. Thus, LSECs respond to both LPS and immune-derived signals and fuel a positive feedback loop of immune cell attraction and activation in the inflamed liver tissue.

Microenvironmental signals shaping NK-cell reactivity in cancer.

Since the postulation of the "missing-self" concept, much progress has been made in defining requirements for NK-cell activation. Unlike T lymphocytes that process signals from receptors in a hierarchic manner dominated by the T-cell receptors, NK cells integrate receptor signals more "democratically." Signals originate not only the downstream of cell-surface receptors triggered by membrane-bound ligands or cytokines, but are also mediated by specialized microenvironmental sensors that perceive the cellular surrounding by detecting metabolites or the availability of oxygen. Thus, NK-cell effector functions are driven in an organ and disease-dependent manner. Here, we review the latest findings on how NK-cell reactivity in cancer is determined by the reception and integration of complex signals. Finally, we discuss how this knowledge can be exploited to guide novel combinatorial approaches for NK-cell-based anticancer therapies.

Lyve-1 deficiency enhances the hepatic immune microenvironment entailing altered susceptibility to melanoma liver metastasis.

BACKGROUND: Hyaluronan receptor LYVE-1 is expressed by liver sinusoidal endothelial cells (LSEC), lymphatic endothelial cells and specialized macrophages. Besides binding to hyaluronan, LYVE-1 can mediate adhesion of leukocytes and cancer cells to endothelial cells. Here, we assessed the impact of LYVE-1 on physiological liver functions and metastasis. METHODS: Mice with deficiency of Lyve-1 (Lyve-1-KO) were analyzed using histology, immunofluorescence, microarray analysis, plasma proteomics and flow cytometry. Liver metastasis was studied by intrasplenic/intravenous injection of melanoma (B16F10 luc2, WT31) or colorectal carcinoma (MC38). RESULTS: Hepatic architecture, liver size, endothelial differentiation and angiocrine functions were unaltered in Lyve-1-KO. Hyaluronan plasma levels were significantly increased in Lyve-1-KO. Besides, plasma proteomics revealed increased carbonic anhydrase-2 and decreased FXIIIA. Furthermore, gene expression analysis of LSEC indicated regulation of immunological pathways. Therefore, liver metastasis of highly and weakly immunogenic tumors, i.e. melanoma and colorectal carcinoma (CRC), was analyzed. Hepatic metastasis of B16F10 luc2 and WT31 melanoma cells, but not MC38 CRC cells, was significantly reduced in Lyve-1-KO mice. In vivo retention assays with B16F10 luc2 cells were unaltered between Lyve-1-KO and control mice. However, in tumor-free Lyve-1-KO livers numbers of hepatic CD4+, CD8+ and regulatory T cells were increased. In addition, iron deposition was found in F4/80+ liver macrophages known to exert pro-inflammatory effects. CONCLUSION: Lyve-1 deficiency controlled hepatic metastasis in a tumor cell-specific manner leading to reduced growth of hepatic metastases of melanoma, but not CRC. Anti-tumorigenic effects are likely due to enhancement of the premetastatic hepatic immune microenvironment influencing early liver metastasis formation.

Targeting of Scavenger Receptors Stabilin-1 and Stabilin-2 Ameliorates Atherosclerosis by a Plasma Proteome Switch Mediating Monocyte/Macrophage Suppression.

BACKGROUND: Scavenger receptors Stabilin-1 (Stab1) and Stabilin-2 (Stab2) are preferentially expressed by liver sinusoidal endothelial cells. They mediate the clearance of circulating plasma molecules controlling distant organ homeostasis. Studies suggest that Stab1 and Stab2 may affect atherosclerosis. Although subsets of tissue macrophages also express Stab1, hematopoietic Stab1 deficiency does not modulate atherogenesis. Here, we comprehensively studied how targeting Stab1 and Stab2 affects atherosclerosis. METHODS: ApoE-KO mice were interbred with Stab1-KO and Stab2-KO mice and fed a Western diet. For antibody targeting, Ldlr-KO mice were also used. Unbiased plasma proteomics were performed and independently confirmed. Ligand binding studies comprised glutathione-S-transferase-pulldown and endocytosis assays. Plasma proteome effects on monocytes were studied by single-cell RNA sequencing in vivo, and by gene expression analyses of Stabilin ligand-stimulated and plasma-stimulated bone marrow-derived monocytes/macrophages in vitro. RESULTS: Spontaneous and Western diet-associated atherogenesis was significantly reduced in ApoE-Stab1-KO and ApoE-Stab2-KO mice. Similarly, inhibition of Stab1 or Stab2 by monoclonal antibodies significantly reduced Western diet-associated atherosclerosis in ApoE-KO and Ldlr-KO mice. Although neither plasma lipid levels nor circulating immune cell numbers were decisively altered, plasma proteomics revealed a switch in the plasma proteome, consisting of 231 dysregulated proteins comparing wildtype with Stab1/2-single and Stab1/2-double KO, and of 41 proteins comparing ApoE-, ApoE-Stab1-, and ApoE-Stab2-KO. Among this broad spectrum of common, but also disparate scavenger receptor ligand candidates, periostin, reelin, and TGFBi (transforming growth factor, ß-induced), known to modulate atherosclerosis, were independently confirmed as novel circulating ligands of Stab1/2. Single-cell RNA sequencing of circulating myeloid cells of ApoE-, ApoE-Stab1-, and ApoE-Stab2-KO mice showed transcriptomic alterations in patrolling (Ccr2-/Cx3cr1++/Ly6Clo) and inflammatory (Ccr2+/Cx3cr1+/Ly6Chi) monocytes, including downregulation of proatherogenic transcription factor Egr1. In wildtype bone marrow-derived monocytes/macrophages, ligand exposure alone did not alter Egr1 expression in vitro. However, exposure to plasma from ApoE-Stab1-KO and ApoE-Stab2-KO mice showed a reverted proatherogenic macrophage activation compared with ApoE-KO plasma, including downregulation of Egr1 in vitro. CONCLUSIONS: Inhibition of Stab1/Stab2 mediates an anti-inflammatory switch in the plasma proteome, including direct Stabilin ligands. The altered plasma proteome suppresses both patrolling and inflammatory monocytes and, thus, systemically protects against atherogenesis. Altogether, anti-Stab1- and anti-Stab2-targeted therapies provide a novel approach for the future treatment of atherosclerosis.

Endothelial RBPJ Is Essential for the Education of Tumor-Associated Macrophages.

Epithelial ovarian cancer (EOC) is one of the most lethal gynecologic cancers worldwide. EOC cells educate tumor-associated macrophages (TAM) through CD44-mediated cholesterol depletion to generate an immunosuppressive tumor microenvironment (TME). In addition, tumor cells frequently activate Notch1 receptors on endothelial cells (EC) to facilitate metastasis. However, further work is required to establish whether the endothelium also influences the education of recruited monocytes. Here, we report that canonical Notch signaling through RBPJ in ECs is an important player in the education of TAMs and EOC progression. Deletion of Rbpj in the endothelium of adult mice reduced infiltration of monocyte-derived macrophages into the TME of EOC and prevented the acquisition of a typical TAM gene signature; this was associated with stronger cytotoxic activity of T cells and decreased tumor burden. Mechanistically, CXCL2 was identified as a novel Notch/RBPJ target gene that regulated the expression of CD44 on monocytes and subsequent cholesterol depletion of TAMs. Bioinformatic analysis of ovarian cancer patient data showed that increased CXCL2 expression is accompanied by higher expression of CD44 and TAM education. Together, these findings indicate that EOC cells induce the tumor endothelium to secrete CXCL2 to establish an immunosuppressive microenvironment. SIGNIFICANCE: Endothelial Notch signaling favors immunosuppression by increasing CXCL2 secretion to stimulate CD44 expression in macrophages, facilitating their education by tumor cells.

NK Cells Under Hypoxia: The Two Faces of Vascularization in Tumor and Pregnancy.

Environmental conditions greatly shape the phenotype and function of immune cells. Specifically, hypoxic conditions that exist within tissues and organs have been reported to affect both the adaptive and the innate immune system. Natural killer (NK) cells belong to the innate immune system. They are among the first immune cells responding to infections and are involved in tumor surveillance. NK cells produce cytokines that shape other innate and adaptive immune cells, and they produce cytolytic molecules leading to target cell killing. Therefore, they are not only involved in steady state tissue homeostasis, but also in pathogen and tumor clearance. Hence, understanding the role of NK cells in pathological and physiological immune biology is an emerging field. To date, it remains incompletely understood how the tissue microenvironment shapes NK cell phenotype and function. In particular, the impact of low oxygen concentrations in tissues on NK cell reactivity has not been systematically dissected. Here, we present a comprehensive review focusing on two highly compelling hypoxic tissue environments, the tumor microenvironment (pathological) and the decidua (physiological) and compare their impact on NK cell reactivity.

Caspase-8 in endothelial cells maintains gut homeostasis and prevents small bowel inflammation in mice.

The gut has a specific vascular barrier that controls trafficking of antigens and microbiota into the bloodstream. However, the molecular mechanisms regulating the maintenance of this vascular barrier remain elusive. Here, we identified Caspase-8 as a pro-survival factor in mature intestinal endothelial cells that is required to actively maintain vascular homeostasis in the small intestine in an organ-specific manner. In particular, we find that deletion of Caspase-8 in endothelial cells results in small intestinal hemorrhages and bowel inflammation, while all other organs remained unaffected. We also show that Caspase-8 seems to be particularly needed in lymphatic endothelial cells to maintain gut homeostasis. Our work demonstrates that endothelial cell dysfunction, leading to the breakdown of the gut-vascular barrier, is an active driver of chronic small intestinal inflammation, highlighting the role of the intestinal vasculature as a safeguard of organ function.

Radiotherapy orchestrates natural killer cell dependent antitumor immune responses through CXCL8.

Radiotherapy is a mainstay cancer therapy whose antitumor effects partially depend on T cell responses. However, the role of Natural Killer (NK) cells in radiotherapy remains unclear. Here, using a reverse translational approach, we show a central role of NK cells in the radiation-induced immune response involving a CXCL8/IL-8-dependent mechanism. In a randomized controlled pancreatic cancer trial, CXCL8 increased under radiotherapy, and NK cell positively correlated with prolonged overall survival. Accordingly, NK cells preferentially infiltrated irradiated pancreatic tumors and exhibited CD56dim-like cytotoxic transcriptomic states. In experimental models, NF-κB and mTOR orchestrated radiation-induced CXCL8 secretion from tumor cells with senescence features causing directional migration of CD56dim NK cells, thus linking senescence-associated CXCL8 release to innate immune surveillance of human tumors. Moreover, combined high-dose radiotherapy and adoptive NK cell transfer improved tumor control over monotherapies in xenografted mice, suggesting NK cells combined with radiotherapy as a rational cancer treatment strategy.

Human ILC3 Exert TRAIL-Mediated Cytotoxicity Towards Cancer Cells.

Group 3 helper Innate Lymphoid Cells (ILC3s) are cytokine-producing lymphocytes that respond to stress signals released during disturbed tissue homeostasis and infection. Upon activation, ILC3s secrete IL-22 and IL-17, and orchestrate immune responses against extracellular pathogens. Their role in cancer remains poorly explored. To determine their anti-cancer effector potential, we co-cultured cytokine-activated human ILC3s with cancer cells of different origins. ILC3s were able to directly respond to tumor cells, resulting in enhanced IFN-γ production. Upon tumor cell encounter, ILC3s maintained expression of the transcription factor RORγt, indicating that ILC3s preserved their identity. ILC3s were able to directly kill both hepatocellular carcinoma and melanoma tumor cells expressing cell-death receptor TRAILR2, through the activation of Caspase-8 in target cells. Moreover, liver-derived cytokine-activated ILC3s also expressed TRAIL and were able to eliminate hepatoblastoma cells. Together, our data reveal that ILC3s can participate in anti-tumor immune response through direct recognition of tumor cells resulting in IFN-γ release and TRAIL-dependent cytotoxicity. Thus, ILC3s might be ancillary players of anti-tumor immunity in tissues, acting as primary responders against transformed or metastasizing cells, which might be further exploited for therapies against cancer.

ILC1-like NK cells as matchmakers for DC-T cell interactions.

Both innate and adaptive immunity are orchestrated by multiple cell types, specialized cell lineages, and their spatiotemporal encounters. It is thought that adaptive-like NK cell responses to viral infection mainly involve circulating bona fide NK cells. In this issue of Immunity, Flommersfeld et al. (2021) identify a splenic-resident ILC1-like NK cell subset that facilitates CD8+ T cell-DC interactions during anti-viral defense.

NBAS Variants Are Associated with Quantitative and Qualitative NK and B Cell Deficiency.

PURPOSE: Biallelic pathogenic NBAS variants manifest as a multisystem disorder with heterogeneous clinical phenotypes such as recurrent acute liver failure, growth retardation, and susceptibility to infections. This study explores how NBAS-associated disease affects cells of the innate and adaptive immune system. METHODS: Clinical and laboratory parameters were combined with functional multi-parametric immunophenotyping methods in fifteen NBAS-deficient patients to discover possible alterations in their immune system. RESULTS: Our study revealed reduced absolute numbers of mature CD56dim natural killer (NK) cells. Notably, the residual NK cell population in NBAS-deficient patients exerted a lower potential for activation and degranulation in response to K562 target cells, suggesting an NK cell-intrinsic role for NBAS in the release of cytotoxic granules. NBAS-deficient NK cell activation and degranulation was normalized upon pre-activation by IL-2 in vitro, suggesting that functional impairment was reversible. In addition, we observed a reduced number of naïve B cells in the peripheral blood associated with hypogammaglobulinemia. CONCLUSION: In summary, we demonstrate that pathogenic biallelic variants in NBAS are associated with dysfunctional NK cells as well as impaired adaptive humoral immunity.

An intimate encounter: DC3s empower anti-tumor CTLs.

Discrete tissue niches are emerging as essential prerequisites enabling cell communication and function in both homeostasis and disease. In a recent Cell paper, Di Pilato et al. identify a unique dendritic cell-cytotoxic T cell crosstalk within the perivascular space that facilitates T cell survival and proliferation and drives anti-tumor activity.

PPARγ induces PD-L1 expression in MSS+ colorectal cancer cells.

Only a small subset of colorectal cancer (CRC) patients benefits from immunotherapies, comprising blocking antibodies (Abs) against checkpoint receptor "programmed-cell-death-1" (PD1) and its ligand (PD-L1), because most cases lack the required mutational burden and neo-antigen load caused by microsatellite instability (MSI) and/or an inflamed, immune cell-infiltrated PD-L1+ tumor microenvironment. Peroxisome proliferator-activated-receptor-gamma (PPARγ), a metabolic transcription factor stimulated by anti-diabetic drugs, has been previously implicated in pre/clinical responses to immunotherapy. We therefore raised the hypothesis that PPARγ induces PD-L1 on microsatellite stable (MSS) tumor cells to enhance Ab-target engagement and responsiveness to PD-L1 blockage. We found that PPARγ-agonists upregulate PD-L1 mRNA/protein expression in human gastrointestinal cancer cell lines and MSS+ patient-derived tumor organoids (PDOs). Mechanistically, PPARγ bound to and activated DNA-motifs similar to cognate PPARγ-responsive-elements (PPREs) in the proximal -2 kb promoter of the human PD-L1 gene. PPARγ-agonist reduced proliferation and viability of tumor cells in co-cultures with PD-L1 blocking Ab and lymphokine-activated killer cells (LAK) derived from the peripheral blood of CRC patients or healthy donors. Thus, metabolic modifiers improved the antitumoral response of immune checkpoint Ab, proposing novel therapeutic strategies for CRC.

Innate-like NKp30+CD8+ T cells armed with TCR/CAR target tumor heterogeneity.

Intratumoral heterogeneity is frequently associated with tumor immune escape, with MHC-class I and antigen expression loss rendering tumor cells invisible to T cell killing, representing a major challenge for the design of successful adoptive transfer protocols for cancer immunotherapy. While CD8+ T cell recognition of tumor cells is based on the detection of MHC-peptide complexes via specific T cell receptors (TCRs), Natural Killer (NK) cells detect tumor-associated NK ligands by an array of NK receptors. We have recently identified a population of innate-like CD8+ T cells marked by the expression of NKp30, a potent natural cytotoxicity activating NK receptor, whose tumor ligand, B7H6, is frequently upregulated on several cancer types. Here, we harnessed the dual-recognition potential of NKp30+CD8+ T cells, by arming these cells with TCRs or chimeric antigen receptors (CARs) targeting Epidermal Growth Factor Receptor 2 (ErbB2, or HER2), a tumor-associated target overexpressed in several malignancies. HER2-specific NKp30+CD8+ T cells killed not only HER2-expressing target cell lines, but also eliminated tumor cells in the absence of MHC-class I or antigen expression, making them especially effective in eliminating heterogeneous tumor cell populations. Our results show that NKp30+CD8+ T cells equipped with a specific TCR or CAR display a dual capacity to recognize and kill target cells, combining the anti-tumor activity of both CD8+ T and NK cells. This dual-recognition capacity allows these effector cells to target tumor heterogeneity, thus improving therapeutic strategies against tumor escape.

Human innate immune cell crosstalk induces melanoma cell senescence.

Mononuclear phagocytes and NK cells constitute the first line of innate immune defense. How these cells interact and join forces against cancer is incompletely understood. Here, we observed an early accumulation of slan+ (6-sulfo LacNAc) non-classical monocytes (slanMo) in stage I melanoma, which was followed by an increase in NK cell numbers in stage III. Accordingly, culture supernatants of slanMo induced migration of primary human NK cells in vitro via the chemotactic cytokine IL-8 (CXCL8), suggesting a role for slanMo in NK cell recruitment into cancer tissues. High levels of TNF-α and IFN-γ were produced in co-cultures of TLR-ligand stimulated slanMo and NK cells, whereas much lower levels were contained in cultures of slanMo and NK cells alone. Moreover, TNF-α and IFN-γ concentrations in slanMo/NK cell co-cultures exceeded those in CD14+ monocyte/NK cell and slanMo/T cell co-cultures. Importantly, TNF-α and IFN-γ that was produced in TLR-ligand stimulated slanMo/NK cell co-cultures induced senescence in different melanoma cell lines, as indicated by reduced melanoma cell proliferation, increased senescence-associated ß-galactosidase expression, p21 upregulation, and induction of a senescence-associated secretory phenotype (SASP). Taken together, we identified a role for slanMo and NK cells in a collaborative innate immune defense against melanoma by generating a tumor senescence-inducing microenvironment. We conclude that enhancing the synergistic innate immune crosstalk of slanMo and NK cells could improve current immunotherapeutic approaches in melanoma.

Type 1 Treg cells promote the generation of CD8+ tissue-resident memory T cells.

Tissue-resident memory T (TRM) cells, functionally distinct from circulating memory T cells, have a critical role in protective immunity in tissues, are more efficacious when elicited after vaccination and yield more effective antitumor immunity, yet the signals that direct development of TRM cells are incompletely understood. Here we show that type 1 regulatory T (Treg) cells, which express the transcription factor T-bet, promote the generation of CD8+ TRM cells. The absence of T-bet-expressing type 1 Treg cells reduces the presence of TRM cells in multiple tissues and increases pathogen burden upon infectious challenge. Using infection models, we show that type 1 Treg cells are specifically recruited to local inflammatory sites via the chemokine receptor CXCR3. Close proximity with effector CD8+ T cells and Treg cell expression of integrin-ß8 endows the bioavailability of transforming growth factor-ß in the microenvironment, thereby promoting the generation of CD8+ TRM cells.