Cutting Edge: PDGF-DD Binding to NKp44 Costimulates TLR9 Signaling and Proinflammatory Cytokine Secretion in Human Plasmacytoid Dendritic Cells.

NKp44 is a human receptor originally found on activated NK cells, group 1 and group 3 innate lymphoid cells that binds dimers of platelet-derived growth factor D (PDGF-DD). NKp44 is also expressed on tissue plasmacytoid dendritic cells (PDCs), but NKp44-PDGF-DD interaction on PDCs remains unstudied. Engagement of NKp44 with PDGF-DD in vitro enhanced PDC secretion of IFN-α, TNF, and IL-6 in response to the TLR9 ligand CpG-ODN, but not TLR7/8 ligands. In tissues, PDCs were found in close contact with PDGF-DD-expressing cells in the high endothelial venules and epithelium of tonsils, melanomas, and skin lesions infected with Molluscum contagiosum. Recombinant PDGF-DD enhanced the serum IFN-α response to systemic HSV-1 infection in a humanized mouse model. We conclude that NKp44 integrates with TLR9 signaling to enhance PDC cytokine production. These findings may have bearings for immune responses to TLR9-based adjuvants, therapy for tumors expressing PDGF-DD, and infections with DNA viruses that induce PDGF-DD expression to enhance viral spread.

Tailoring Natural Killer cell immunotherapy to the tumour microenvironment.

Natural killer (NK) cells are cytotoxic and cytokine-secreting cells that can mediate potent anti-tumour activity. Accumulating evidence indicates that NK cell functions are severely compromised within the confines of the tumour microenvironment thus impairing the efficacy and development of NK cell-based therapies. Here we review the various cellular and molecular pathways that tumours have supplanted to evade NK cell surveillance. We highlight novel strategies designed to alleviate or circumvent the immunosuppressive conditions of the tumour microenvironment in order to emancipate NK cell function and stifle the inexorable growth and metastasis of malignant cells.

TARM1 Is a Novel Leukocyte Receptor Complex-Encoded ITAM Receptor That Costimulates Proinflammatory Cytokine Secretion by Macrophages and Neutrophils.

We identified a novel, evolutionarily conserved receptor encoded within the human leukocyte receptor complex and syntenic region of mouse chromosome 7, named T cell-interacting, activating receptor on myeloid cells-1 (TARM1). The transmembrane region of TARM1 contained a conserved arginine residue, consistent with association with a signaling adaptor. TARM1 associated with the ITAM adaptor FcRγ but not with DAP10 or DAP12. In healthy mice, TARM1 is constitutively expressed on the cell surface of mature and immature CD11b(+)Gr-1(+) neutrophils within the bone marrow. Following i.p. LPS treatment or systemic bacterial challenge, TARM1 expression was upregulated by neutrophils and inflammatory monocytes and TARM1(+) cells were rapidly recruited to sites of inflammation. TARM1 expression was also upregulated by bone marrow-derived macrophages and dendritic cells following stimulation with TLR agonists in vitro. Ligation of TARM1 receptor in the presence of TLR ligands, such as LPS, enhanced the secretion of proinflammatory cytokines by macrophages and primary mouse neutrophils, whereas TARM1 stimulation alone had no effect. Finally, an immobilized TARM1-Fc fusion protein suppressed CD4(+) T cell activation and proliferation in vitro. These results suggest that a putative T cell ligand can interact with TARM1 receptor, resulting in bidirectional signaling and raising the T cell activation threshold while costimulating the release of proinflammatory cytokines by macrophages and neutrophils.

IL17RB and IL17REL Expression Are Associated with Improved Prognosis in HPV-Infected Head and Neck Squamous Cell Carcinomas.

Changes in the cellular secretome are implicated in virus infection, malignancy, and anti-tumor immunity. We analyzed the association between transcriptional signatures (TS) from 24 different immune and stromal cell types on the prognosis of HPV-infected and HPV-free head and neck squamous carcinoma (HNSCC) patients from The Cancer Genome Atlas (TCGA) cohort. We found that HPV-positive HNSCC patients have tumors with elevated immune cell TS and improved prognosis, which was specifically associated with an increased tumor abundance of memory B and activated natural killer (NK) cell TS, compared to HPV-free HNSCC patients. HPV-infected patients upregulated many transcripts encoding secreted factors, such as growth factors, hormones, chemokines and cytokines, and their cognate receptors. Analysis of secretome transcripts and cognate receptors revealed that tumor expression of IL17RB and IL17REL are associated with a higher viral load and memory B and activated NK cell TS, as well as improved prognosis in HPV-infected HNSCC patients. The transcriptional parameters that we describe may be optimized to improve prognosis and risk stratification in the clinic and provide insights into gene and cellular targets that may potentially enhance anti-tumor immunity mediated by NK cells and memory B cells in HPV-infected HNSCC patients.

The extended human leukocyte receptor complex: diverse ways of modulating immune responses.

The leukocyte receptor complex (LRC) and its extended region comprise a large set of genes encoding immunoglobulin superfamily (IgSF) receptors, interspersed with other loci. Although the external Ig-like domains of these molecules are related, they have evolved to bind a wide array of different ligands. Comparison of the organization and functions of the different receptors encoded in the LRC provides insight into their roles in immune recognition, their evolution, and their relevance to disease. In addition, these molecules provide classic examples of inhibitory receptors paired, side by side, with activating receptors that couple with adapter proteins, such as DAP12. Some of these activating receptors can be considered as bifunctional sensors that can perceive changes in the state of their ligands that favors an inhibitory rather than activating response, whereas other receptors have evolved different means, acting as transporters or even molecular chaperones to achieve immune repression. We briefly summarize the complement of receptors encoded in this region of chromosome 19 and discuss the many diverse and versatile mechanisms they have evolved to restrain immune responses.

A Transcriptional Signature of PDGF-DD Activated Natural Killer Cells Predicts More Favorable Prognosis in Low-Grade Glioma.

The binding of platelet-derived growth factor D (PDGF-DD) to the NKp44 receptor activates a distinct transcriptional program in primary IL-2 expanded human natural killer (NK) cells. We were interested in knowing if the PDGF-DD-NKp44 pathway of NK cell activation might play a clinically relevant role in anti-tumor immunity. In order to address this question, we determined transcriptional signatures unique to resting, IL-2 expanded, and PDGF-DD activated, NK cells, in addition to different T cell subsets, and established the abundance of these immune cell phenotypes in The Cancer Genome Atlas (TCGA) low-grade glioma (LGG) dataset using CIBERSORT. Our results show that LGG patient tumors enriched for either the PDGF-DD activated NK cell or memory CD8+ T cell phenotypes are associated with a more favorable prognosis. Combined cell phenotype analyses revealed that patients with LGG tumors enriched for the PDGF-DD activated NK cell phenotype and the CD4+ T helper cell phenotype had a more favorable prognosis. High expression of transcripts encoding members of the killer cell lectin-like receptor (KLR) family, such as KLRK1 and KLRC2, KLRC3 and KLRC4 in LGG tumors were associated with more favorable prognosis, suggesting that these NK cell family receptors may play a prominent role in LGG anti-tumor immunity. Finally, many of the TCGA findings were reciprocated in LGG patients from the Chinese Glioma Genome Atlas (CGGA) dataset. Our results provide transcriptomic evidence that PDGF-DD activated NK cells and KLR family receptors may play an important clinical role in immune surveillance of LGG.

A Transcriptional Signature of IL-2 Expanded Natural Killer Cells Predicts More Favorable Prognosis in Bladder Cancer.

Activation of natural killer (NK) cell function is regulated by cytokines, such as IL-2, and secreted factors upregulated in the tumor microenvironment, such as platelet-derived growth factor D (PDGF-DD). In order to elucidate a clinical role for these important regulators of NK cell function in antitumor immunity, we generated transcriptional signatures representing resting, IL-2-expanded, and PDGF-DD-activated, NK cell phenotypes and established their abundance in The Cancer Genome Atlas bladder cancer (BLCA) dataset using CIBERSORT. The IL-2-expanded NK cell phenotype was the most abundant in low and high grades of BLCA tumors and was associated with improved prognosis. In contrast, PDGFD expression was associated with numerous cancer hallmark pathways in BLCA tumors compared with normal bladder tissue, and a high tumor abundance of PDGFD transcripts and the PDGF-DD-activated NK cell phenotype were associated with a poor BLCA prognosis. Finally, high tumor expression of transcripts encoding the activating NK cell receptors, KLRK1 and the CD160-TNFRSF14 receptor-ligand pair, was strongly correlated with the IL-2-expanded NK cell phenotype and improved BLCA prognosis. The transcriptional parameters we describe may be optimized to improve BLCA patient prognosis and risk stratification in the clinic and potentially provide gene targets of therapeutic significance for enhancing NK cell antitumor immunity in BLCA.

The Role of NK Cells and Innate Lymphoid Cells in Brain Cancer.

The brain is considered an immune privileged site due to the high selectivity of the blood-brain barrier which restricts the passage of molecules and cells into the brain parenchyma. Recent studies have highlighted active immunosurveillance mechanisms in the brain. Here we review emerging evidence for the contribution of innate lymphoid cells (ILCs) including natural killer (NK) cells to the immunosurveillance of brain cancers focusing on glioblastoma, one of the most aggressive and most common malignant primary brain tumors diagnosed in adults. Moreover, we discuss how the local tissue microenvironment and unique cellular interactions influence ILC functions in the brain and how these interactions might be successfully harnessed for cancer immunotherapy using insights gained from the studies of autoimmunity, aging, and CNS injury.

Comparative genomics indicates the mammalian CD33rSiglec locus evolved by an ancient large-scale inverse duplication and suggests all Siglecs share a common ancestral region.

The CD33-related sialic acid binding Ig-like lectins (CD33rSiglecs) are predominantly inhibitory receptors expressed on leukocytes. They are distinguishable from conserved Siglecs, such as Sialoadhesin and MAG, by their rapid evolution. A comparison of the CD33rSiglec gene cluster in different mammalian species showed that it can be divided into subclusters, A and B. The two subclusters, inverted in relation to each other, each encode a set of CD33rSiglec genes arranged head-to-tail. Two regions of strong correspondence provided evidence for a large-scale inverse duplication, encompassing the framework CEACAM-18 (CE18) and ATPBD3 (ATB3) genes that seeded the mammalian CD33rSiglec cluster. Phylogenetic analysis was consistent with the predicted inversion. Rodents appear to have undergone wholesale loss of CD33rSiglec genes after the inverse duplication. In contrast, CD33rSiglecs expanded in primates and many are now pseudogenes with features consistent with activating receptors. In contrast to mammals, the fish CD33rSiglecs clusters show no evidence of an inverse duplication. They display greater variation in cluster size and structure than mammals. The close arrangement of other Siglecs and CD33rSiglecs in fish is consistent with a common ancestral region for Siglecs. Expansion of mammalian CD33rSiglecs appears to have followed a large inverse duplication of a smaller primordial cluster over 180 million years ago, prior to eutherian/marsupial divergence. Inverse duplications in general could potentially have a stabilizing effect in maintaining the size and structure of large gene clusters, facilitating the rapid evolution of immune gene families.

Innate lymphoid cell sensing of tissue vitality.

Innate lymphoid cells (ILCs) constitute a heterogeneous population of cytokine-secreting cells that colonize different tissues and are heavily reliant on cytokines and other secreted factors for their development, maintenance and effector functions. Most ILCs are tissue resident and differentiate in non-lymphoid peripheral tissues. As tissue-resident sentinels, ILCs must rapidly identify pathogens or malignancy in an effort to return the tissue to homeostasis. Here we review the mechanisms that ILCs employ to sense cytokines and other potent immunoregulatory factors that promote their development in different tissues as well as the ability to distinguish pathogenic versus healthy tissue microenvironments and highlight the importance of these pathways for human disease.

Exploiting NK Cell Surveillance Pathways for Cancer Therapy.

Natural killer (NK) cells can evoke potent anti-tumour activity. This function is largely mediated through a battery of specialised cell-surface receptors which probe the tissue microenvironment for changes in surface and secretory phenotypes that may alert to the presence of infection or malignancy. These receptors have the potential to arouse the robust cytotoxic and cytokine-secreting functions of NK cells and so must be tightly regulated to prevent autoimmunity. However, such functions also hold great promise for clinical intervention. In this review, we highlight some of the latest breakthroughs in fundamental NK cell receptor biology that have illuminated our understanding of the molecular strategies NK cells employ to perceive malignant cells from normal healthy cells. Moreover, we highlight how these sophisticated tumour recognition strategies are being harnessed for cancer immunotherapies in the clinic.

The Natural Cytotoxicity Receptors in Health and Disease.

The Natural Cytotoxicity Receptors (NCRs), NKp46, NKp44, and NKp30, were some of the first human activating Natural Killer (NK) cell receptors involved in the non-MHC-restricted recognition of tumor cells to be cloned over 20 years ago. Since this time many host- and pathogen-encoded ligands have been proposed to bind the NCRs and regulate the cytotoxic and cytokine-secreting functions of tissue NK cells. This diverse set of NCR ligands can manifest on the surface of tumor or virus-infected cells or can be secreted extracellularly, suggesting a remarkable NCR polyfunctionality that regulates the activity of NK cells in different tissue compartments during steady state or inflammation. Moreover, the NCRs can also be expressed by other innate and adaptive immune cell subsets under certain tissue conditions potentially conferring NK recognition programs to these cells. Here we review NCR biology in health and disease with particular reference to how this important class of receptors regulates the functions of tissue NK cells as well as confer NK cell recognition patterns to other innate and adaptive lymphocyte subsets. Finally, we highlight how NCR biology is being harnessed for novel therapeutic interventions particularly for enhanced tumor surveillance.

OSCAR is a collagen receptor that costimulates osteoclastogenesis in DAP12-deficient humans and mice.

Osteoclasts are terminally differentiated leukocytes that erode the mineralized bone matrix. Osteoclastogenesis requires costimulatory receptor signaling through adaptors containing immunoreceptor tyrosine-based activation motifs (ITAMs), such as Fc receptor common γ (FcRγ) and DNAX-activating protein of 12 kDa. Identification of these ITAM-containing receptors and their ligands remains a high research priority, since the stimuli for osteoclastogenesis are only partly defined. Osteoclast-associated receptor (OSCAR) was proposed to be a potent FcRγ-associated costimulatory receptor expressed by preosteoclasts in vitro, but OSCAR lacks a cognate ligand and its role in vivo has been unclear. Using samples from mice and patients deficient in various ITAM signaling pathways, we show here that OSCAR costimulates one of the major FcRγ-associated pathways required for osteoclastogenesis in vivo. Furthermore, we found that OSCAR binds to specific motifs within fibrillar collagens in the ECM that become revealed on nonquiescent bone surfaces in which osteoclasts undergo maturation and terminal differentiation in vivo. OSCAR promoted osteoclastogenesis in vivo, and OSCAR binding to its collagen motif led to signaling that increased numbers of osteoclasts in culture. Thus, our results suggest that ITAM-containing receptors can respond to exposed ligands in collagen, leading to the functional differentiation of leukocytes, which provides what we believe to be a new concept for ITAM regulation of cytokine receptors in different tissue microenvironments.

You say ITAM and I say ITIM, let's call the whole thing off: the ambiguity of immunoreceptor signalling.

Paradigms of protein tyrosine kinase (PTK)-mediated immunoreceptor signalling have developed largely from studies of molecules containing immunoreceptor tyrosine-based activation motifs (ITAM) and immunoreceptor tyrosine-based inhibition motifs (ITIM). In some circumstances, ITAM can mediate inhibition and ITIM can propagate activation signals. In addition to classical immune cells, some ITAM- and ITIM-encoding proteins are more widely expressed and are crucial to the development of haemopoietic cells, like osteoclasts and natural killer cells, as well as tissues such as bone and brain. Closer analysis of activating proteins reveals that some potentially encode ITIM within an ITAM. These 'closet' ITIM could, under conditions of partial ITAM phosphorylation, mediate inhibitory signalling.