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PDF file - 123K, Supplemental Figure 3 - Increased ERK activation and macrophage infiltration in rapid progressing KS lesions.

D6 is an atypical chemokine receptor acting as a decoy and scavenger for inflammatory CC chemokines expressed in lymphatic endothelial cells. Here, we report that D6 is expressed in Kaposi sarcoma (KS), a tumor ontogenetically related to the lymphatic endothelium. Both in human tumors and in an experimental model, D6 expression levels were inversely correlated with tumor aggressiveness and increased infiltration of proangiogenic macrophages. Inhibition of monocyte recruitment reduced the growth of tumors, while adoptive transfer of wild-type, but not CCR2−/− macrophages, increased the growth rate of D6-competent neoplasms. In the KS model with the B-Raf V600E–activating mutation, inhibition of B-Raf or the downstream ERK pathway induced D6 expression; in progressing human KS tumors, the activation of ERK correlates with reduced levels of D6 expression. These results indicate that activation of the K-Ras–B-Raf–ERK pathway during KS progression downregulates D6 expression, which unleashes chemokine-mediated macrophage recruitment and their acquisition of an M2-like phenotype supporting angiogenesis and tumor growth. Combined targeting of CCR2 and the ERK pathway should be considered as a therapeutic option for patients with KS. Cancer Immunol Res; 2(7); 679–89. ©2014 AACR.

PDF file - 90K, Supplemental Figure 2 - D6- and D6+KS tumors have equal necrotic areas.

PDF file - 123K, Supplemental Figure 1 - D6+KS but not D6- KS cells display chemokine scavenging activity.

Neutrophils play a key role in defence against infection and in the activation and regulation of innate and adaptive immunity. In cancer, tumour-associated neutrophils (TANs) have emerged as an important component of the tumour microenvironment. Here, they can exert dual functions. TANs can be part of tumour-promoting inflammation by driving angiogenesis, extracellular matrix remodelling, metastasis and immunosuppression. Conversely, neutrophils can also mediate antitumour responses by direct killing of tumour cells and by participating in cellular networks that mediate antitumour resistance. Neutrophil diversity and plasticity underlie the dual potential of TANs in the tumour microenvironment. Myeloid checkpoints as well as the tumour and tissue contexture shape neutrophil function in response to conventional therapies and immunotherapy. We surmise that neutrophils can provide tools to tailor current immunotherapy strategies and pave the way to myeloid cell-centred therapeutic strategies, which would be complementary to current approaches.

Background Immunotherapeutic early-phase clinical trials (ieCTs) increasingly adopt large expansion cohorts exploring novel agents across different tumor types. High-grade glioma (HGG) patients are usually excluded from these trials. Methods Data of patients with recurrent HGGs treated within multicohort ieCTs between February 2014 and August 2019 (experimental group, EG) at our Phase I Unit were retrospectively reviewed and compared to a matched control group (CG) of patients treated with standard therapies. We retrospectively evaluated clinical, laboratory, and molecular parameters through univariate and multivariate analysis. A prospective characterization of circulating leukocyte subpopulations was performed in the latest twenty patients enrolled in the EG, with a statistical significance cutoff of P < .1. Results Thirty HGG patients were treated into six ieCTs. Fifteen patients received monotherapies (anti-PD-1, anti-CSF-1R, anti-TGFβ, anti-cereblon), fifteen patients combination regimens (anti-PD-L1 + anti-CD38, anti-PD-1 + anti-CSF-1R). In the EG, median progression-free survival and overall survival (OS) from treatment initiation were 1.8 and 8.6 months; twelve patients survived more than 12 months, and two of them more than 6 years. Univariate analysis identified O6-methylguanine DNA methyltransferase (MGMT) promoter methylation and total protein value at six weeks as significantly correlated with a better outcome. Decreased circulating neutrophils and increased conventional dendritic cells levels lead to significantly better OS. Conclusions A subgroup of EG patients achieved remarkably durable disease control. MGMT promoter methylation identifies patients who benefit more from immunotherapy. Monitoring dynamic changes of innate immune cell populations may help to predict clinical outcomes.

The atypical chemokine receptor ACKR2, formerly named D6, is a scavenger chemokine receptor with a non-redundant role in the control of inflammation and immunity. The scavenging activity of ACKR2 depends on its trafficking properties, which require actin cytoskeleton rearrangements downstream of a &beta, arrestin1-Rac1-PAK1-LIMK1-cofilin-dependent signaling pathway. We here demonstrate that in basal conditions, ACKR2 trafficking properties require intact actin and microtubules networks. The dynamic turnover of actin filaments is required to sustain ACKR2 constitutive endocytosis, while both actin and microtubule networks are involved in processes regulating ACKR2 constitutive sorting to rapid, Rab4-dependent and slow, Rab11-dependent recycling pathways, respectively. After chemokine engagement, ACKR2 requires myosin Vb activity to promote its trafficking from Rab11-positive recycling endosomes to the plasma membrane, which sustains its scavenging activity. Other than cofilin phosphorylation, induction of the &beta, arrestin1-dependent signaling pathway by ACKR2 agonists also leads to the rearrangement of microtubules, which is required to support the myosin Vb-dependent ACKR2 upregulation and its scavenging properties. Disruption of the actin-based cytoskeleton by the apoptosis-inducing agent staurosporine results in impaired ACKR2 internalization and chemokine degradation that is consistent with the emerging scavenging-independent activity of the receptor in apoptotic neutrophils instrumental for promoting efficient efferocytosis during the resolution of inflammation. In conclusion, we provide evidence that ACKR2 activates a &beta, arrestin1-dependent signaling pathway, triggering both the actin and the microtubule cytoskeletal networks, which control its trafficking and scavenger properties.

Neutrophils play a key role in defence against infection and in the activation and regulation of innate and adaptive immunity. In cancer, tumour-associated neutrophils (TANs) have emerged as an important component of the tumour microenvironment. Here, they can exert dual functions. TANs can be part of tumour-promoting inflammation by driving angiogenesis, extracellular matrix remodelling, metastasis and immunosuppression. Conversely, neutrophils can also mediate antitumour responses by direct killing of tumour cells and by participating in cellular networks that mediate antitumour resistance. Neutrophil diversity and plasticity underlie the dual potential of TANs in the tumour microenvironment. Myeloid checkpoints as well as the tumour and tissue contexture shape neutrophil function in response to conventional therapies and immunotherapy. We surmise that neutrophils can provide tools to tailor current immunotherapy strategies and pave the way to myeloid cell-centred therapeutic strategies, which would be complementary to current approaches.

Neutrophils are an important component of the tumor microenvironment, and their infiltration has been associated with a poor prognosis for most human tumors. However, neutrophils have been shown to be endowed with both protumor and antitumor activities, reflecting their heterogeneity and plasticity in cancer. A growing body of studies has demonstrated that chemokines and chemokine receptors, which are fundamental regulators of neutrophils trafficking, can affect neutrophil maturation and effector functions. Here, we review human and mouse data suggesting that targeting chemokines or chemokine receptors can modulate neutrophil activity and improve their antitumor properties and the efficiency of immunotherapy.

As main drivers of leukocyte recruitment during inflammatory reactions, chemokines act as mediatrs of alarmins in priming host defense responses after tissue exposure to toxic or infectious agents, immunomediated damage, and in inflammation-driven tumors. Chemokines can therefore be considered alarm signals generated by tissues in a broad number of conditions, and mechanisms controlling chemokines biological activities are therefore key to regulate tissue reactions induced by alarmins. By transporting, presenting or scavenging different sets of chemokines, atypical chemokine receptors represent an emerign subfamily of chemokine receptors which operates in tissues as chemokine gatekeepers in order to establish and shape their gradients and coordinate leukocyte recruitment.

Atypical chemokine receptors (ACKRs) are regulators of leukocyte traffic, inflammation, and immunity. ACKR2 is a scavenger for most inflammatory CC chemokines and is a negative regulator of inflammation. Here we report that ACKR2 is expressed in hematopoietic precursors and downregulated during myeloid differentiation. Genetic inactivation of ACKR2 results in increased levels of inflammatory chemokine receptors and release from the bone marrow of neutrophils with increased anti-metastatic activity. In a model of NeuT-driven primary mammary carcinogenesis ACKR2 deficiency is associated with increased primary tumor growth and protection against metastasis. ACKR2 deficiency results in neutrophil-mediated protection against metastasis in mice orthotopically transplanted with 4T1 mammary carcinoma and intravenously injected with B16F10 melanoma cell lines. Thus, ACKR2 is a key regulator (checkpoint) of mouse myeloid differentiation and function and its targeting unleashes the anti-metastatic activity of neutrophils in mice., The atypical chemokine receptor ACKR2 regulates immune responses. Here the authors confirm that ACKR2 depletion promotes primary tumor growth but show it has an anti-metastatic effect in mouse models of breast cancer by affecting myeloid differentiation and unleashing the anti-metastatic activity of neutrophils.

Macrophages are a key component of the tumor microenvironment (TME). Blocking Clever-1, a molecule expressed in M2-like macrophages, unleashed macrophage and T-cell–mediated antitumor immunity. Myeloid checkpoints including Clever-1 hold promise as alternative or complementary immunotherapy strategies. See related article by Viitala et al., p. 3289

Glioma is the most common primary brain cancer, and half of patients present a diagnosis of glioblastoma (GBM), its most aggressive and lethal form. Conventional therapies, including surgery, radiotherapy, and chemotherapy, have not resulted in major ameliorations in GBM survival outcome, which remains extremely poor. Recent immunotherapy improvements for other tumors, coupled with growing knowledge of the complex interactions between malignant glioma cells and the immune system, led to an exponential increase in glioma immunotherapy research. However, immunotherapeutic strategies in GBM have not yet reached their full potential, mainly due to the limited understanding of the strong immunosuppressive microenvironment (TME) characterizing this tumor. Glioma-associated macrophages and microglia (GAMs) are key drivers of the local immunosuppression promoting tumor progression and its resistance to immunomodulating therapeutic strategies. Together with other myeloid cells, such as dendritic cells and neutrophils, GAMs actively shape glioma TME, modulate anti-tumoral immune response and support angiogenesis, tumor cell invasion and proliferation. In this review, we discuss the role of myeloid cells in the complex TME of glioma and the available clinical data on therapeutic strategies focusing on approaches that affect myeloid cells activity in GBM.

The chemokine system is a fundamental component of cancer-related inflammation involved in all stages of cancer development. It controls not only leukocyte infiltration in primary tumors but also angiogenesis, cancer cell proliferation, and migration to metastatic sites. Atypical chemokine receptors are a new, emerging class of regulators of the chemokine system. They control chemokine bioavailability by scavenging, transporting, or storing chemokines. They can also regulate the activity of canonical chemokine receptors with which they share the ligands by forming heterodimers or by modulating their expression levels or signaling activity. Here, we summarize recent results about the role of these receptors (atypical chemokine receptor 1/Duffy antigen receptor for chemokine, atypical chemokine receptor 2/D6, atypical chemokine receptor 3/CXC-chemokine receptor 7, and atypical chemokine receptor 4/CC-chemokine receptor-like 1) on the tumorigenesis process, indicating that their effects are strictly dependent on the cell type on which they are expressed and on their coexpression with other chemokine receptors. Indeed, atypical chemokine receptors inhibit tumor growth and progression through their activity as negative regulators of chemokine bioavailability, whereas, on the contrary, they can promote tumorigenesis when they regulate the signaling of other chemokine receptors, such as CXC-chemokine receptor 4. Thus, atypical chemokine receptors are key components of the regulatory network of inflammation and immunity in cancer and may have a major effect on anti-inflammatory and immunotherapeutic strategies.

BACKGROUND: Different immunotherapeutic strategies are currently under clinical evaluation in the context of neuroncology. However, a critical barrier to an effective antitumor immunity is still represented by the wide array of immunosuppressive forces exhibited by malignant gliomas (MGs). The present study aims to perform a comprehensive analysis of infiltrating and circulating leukocyte subsets (lymphocytes, macrophages, dendritic cells, neutrophils) in patients with MGs, identifying the immunological profile of the different histo-molecular subtypes, exploring the functional changes during and after anticancer therapies. Here we report first preliminary data on infiltrating and circulating neutrophils. MATERIALS AND METHODS: Patients with a clinical-radiological diagnosis of malignant diffuse glioma referring to the NeuroOncological Surgery Unit of the Humanitas Research Hospital between January 2017 and March 2018 were prospectively enrolled in the study. Fresh tumor tissue specimens were collected the day of surgery; peripheral blood samples (PBS) were collected at baseline and then prospectively every two months during active anticancer treatments and then every six months during follow-up. The analysis of the frequency as well as phenotypic and functional characterization of tumor associated neutrophils (TANs) and of circulating neutrophils were performed by flow cytometry. Data were correlated with histology and the molecular markers isocitrate dehydrogenase (IDH) mutation and O(6)-methylguanine-DNA methyltransferase (MGMT) promoter methylation. RESULTS: As per May 2018, 26 out of 84 patients (MG specimens and PBS) plus PBS of 4 healthy donors were analyzed. 12 had IDH wt GBM (n=7 MGMT methylated; n=5 MGMT unmethylated), 5 IDH mut GBM (n=5 MGMT unmethylated), 4 IDH mut anaplastic astrocytoma (n=3 MGMT methylated; n=1 MGMT undetermined), 4 IDH mut anaplastic oligodendrogliomas (n=4 MGMT methylated). TANs represented 5% (SD= 2,67%) of all CD45+ tumor disaggregated single cell suspensions. No statistically significative differences were found comparing the percentage of TANs (CD45(+)CD11b(+)CD66b(+)CD16(+)) among different histo-molecular subgroups. However, an increased percentage of activated TANs (CD62L(neg)CD16(high)) were found in samples from IDH wt GBM patients with methylated MGMT promoter compared to unmethylated ones (p=0,005). Increased percentages of activated neutrophils were found in PBS from IDH wt GBM patients compared to healthy donors (p=0,005). CONCLUSIONS: An increased number of activated TANs are present in MGMT methylated IDH wt GBM patients compared to unmethylated. IDH wt GBM patients seem to have higher percentages of circulating activated neutrophils compared to healthy donors. Correlation between different subpopulations of TANs, treatment response and clinical outcome will be presented at the meeting.

Chemokines are key mediators of inflammation. In pathological tissues, the main roles of chemokines are to regulate leucocyte accumulation through the activation of oriented cell migration and the activation of limited programs of gene transcription. Through these activities, chemokines exert many crucial functions, including the regulation of angiogenesis. The ‘chemokine system’ is tightly regulated at several levels, such as the post-transcriptional processing of ligands, the regulation of the expression and function of the receptors and through the expression of molecules known as ‘atypical chemokine receptors’, proteins that function as chemokine scavenging and presenting molecules. Several experimental evidence obtained in vitro , in animal models and in human studies, has defined a crucial role of chemokines in cardiovascular diseases. An intense area of research is currently exploring the possibility to develop new effective therapeutic strategies through the identification of chemokine receptor antagonists.

The early events linking diet-induced adipose tissue inflammation and insulin resistance remain poorly understood. In this issue of Cell Metabolism, Kawano et al. (2016) show that infiltration of colonic pro-inflammatory macrophages orchestrated by the intestinal CCL2/CCR2 axis kick-starts this process during high-fat-diet feeding.

Although chemokines are best known for their role in directing cell migration, accumulating evidence indicate their involvement in many other processes. This review focus on the role of chemokines in hematopoiesis with an emphasis on myelopoiesis. Indeed, many chemokine family members are an important component of the cytokine network present in the bone marrow that controls proliferation, retention, and mobilization of hematopoietic progenitors.

Chemokine receptors play an important role in leukocyte migration, both in physiological and pathological conditions, and the interest in new methodologies for their detection is increasing. In this review, we focused on chemokine receptors detection through flow cytometric approaches, including the use of specific antibodies and fluorescent chemokines, and on approaches aimed at the analysis of their functions, from intracellular trafficking to signaling activities.

Studies in preclinical models have demonstrated the superior anti-tumor effect of CpG oligodeoxynucleotides (CpG-ODN) when administered at the tumor site rather than systemically. We evaluated the effect of aerosolized CpG-ODN on lung metastases in mice injected with immunogenic N202.1A mammary carcinoma cells or weakly immunogenic B16 melanoma cells. Upon reaching the bronchoalveolar space, aerosolized CpG-ODN activated a local immune response, as indicated by production of IL-12p40, IFN-γ and IL-1β and by recruitment and maturation of DC cells in bronchoalveolar lavage fluid of mice. Treatment with aerosolized CpG-ODN induced an expansion of CD4+ cells in lung and was more efficacious than systemic i.p. administration against experimental lung metastases of immunogenic N202.1A mammary carcinoma cells, whereas only i.p. delivery of CpG-ODN provided anti-tumor activity, which correlated with NK cell expansion in the lung, against lung metastases of the poorly immunogenic B16 melanoma. The inefficacy of aerosol therapy to induce NK expansion was related to the presence of immunosuppressive macrophages in B16 tumor-bearing lungs, as mice depleted of these cells by clodronate treatment responded to aerosol CpG-ODN through expansion of the NK cell population and significantly reduced numbers of lung metastases. Our results indicate that tumor immunogenicity and the tumor-induced immunosuppressive environment are critical factors to the success of CpG therapy in the lung, and point to the value of routine sampling of the lung immune environment in defining an optimal immunotherapeutic strategy.

ACRs (atypical chemokine receptors) were initially referred to as ‘silent’ receptors on the basis of a lack of signalling and functional activities that are typically observed with conventional chemokine receptors. Although ACRs do not directly induce cell migration, they indirectly control leucocyte recruitment by shaping chemokine gradients in tissues through degradation, transcytosis or local concentration of their cognate ligands. Recent evidence also suggests that these biological activities are supported by G-protein-independent, β-arrestin-dependent signalling events. In the present article, we review current knowledge on structural and signalling properties of ACRs that are changing our view on this entire class of receptors from silent to endogenous β-arrestin-biased signalling receptors.

The atypical chemokine receptor D6 was initially called “silent” on the basis of lack of conventional signaling events that lead to directional cell migration. It has emerged that D6 is able to bind and drive to degradative compartments most inflammatory CC chemokines and that is able to convey G-protein independent signaling events to optimize its scavenging activity. We here summarize the knowledge available today on D6 structural and signaling properties and its essential role for the control of inflammatory cells traffic and proper development of the adaptive immune response.

D6 is an atypical chemokine receptor involved in chemokine degradation and resolution of acute inflammatory responses in mice. Emerging evidence suggests that D6 might behave differently in human chronic inflammatory conditions. We, therefore, investigated the involvement of D6 in the immune responses in COPD, a chronic inflammatory condition of the lung.D6 expression was quantified by immunohistochemistry in surgical resected lung specimens from 16 patients with COPD (FEV(1), 57% ± 6% predicted) and 18 control subjects with normal lung function (nine smokers and nine nonsmokers). BAL was also obtained and analyzed by flow cytometry, immunofluorescence, and molecular analysis for further assessment of D6 involvement.D6 expression in the lung was mainly detected in alveolar macrophages (AMs). The percentage of D6(+) AMs was markedly increased in patients with COPD as compared with both smoker and nonsmoker control subjects (P.0005 for both). D6 expression was detected at both transcript and protein level in AMs but not in monocyte-derived macrophages. Finally, D6 expression was positively correlated with markers of immune activation (CD8(+) T lymphocytes, IL-32, tumor necrosis factor-α, B-cell activating factor of the tumor necrosis factor family, phospho-p38 mitogen-activated protein kinase) and negatively with lung function (FEV(1), FEV(1)/FVC).D6 is expressed in AMs from patients with COPD, and its expression correlates with the degree of functional impairment and markers of immune activation. Upregulation of D6 in AMs could indicate that, besides its known scavenger activity in acute inflammation, D6 may have additional roles in chronic inflammatory conditions possibly promoting immune activation.

The lymphatic system plays an important role in the induction of the immune response by transporting antigens, inflammatory mediators, and leukocytes from peripheral tissues to draining lymph nodes (LNs). It is emerging that lymphatic endothelial cells (LECs) are playing an active role in this context via the expression of chemokines, inflammatory mediators promoting cell migration, and chemokine receptors. Particularly, LECs express atypical chemokine receptors (ACKRs), which are unable to promote conventional signaling and cell migration whilst they are involved in the regulation of chemokine availability. Here we provide a summary of the data on the role of ACKR2 expressed by lymphatics, indicating an essential role for this ACKRs in the regulation of the inflammation and the immune response in different pathological conditions, including infection, allergy, and cancer.

Chemokines and their receptors are key mediators of the inflammatory process regulating leukocyte extravasation and directional migration into inflamed and infected tissues. The control of chemokine availability within inflamed tissues is necessary to attain a resolving environment and when this fails chronic inflammation ensues. Accordingly, vertebrates have adopted a number of mechanisms for removing chemokines from inflamed sites to help precipitate resolution. Over the past 15 years it has become apparent that essential players in this process are the members of the atypical chemokine receptor family. Broadly speaking this family is expressed on stromal cell types and scavenges chemokines to either limit their spatial availability or to remove them from in vivo sites. Here we provide a brief review of these atypical chemokine receptors and discuss their involvement in the resolution of inflammatory responses and the therapeutic implications of our current knowledge.

Chemokines are a large family of secreted cytokines whose main function is to mediate leukocyte directional migration. Most cancers contain chemokines and express chemokine receptors as a consequence of the activity of deregulated transcription factors or tumor-suppressor genes. Indeed chemokines expression at the tumor site dictates leukocyte infiltration and angiogenesis, while chemokine receptors expression by tumor cells promotes their growth and matastatization. Chemokines also have several indirect effects on tumor growth and are a relevant element in the cancer-related inflammation. In this chapter we will describe technical approaches available to study the role of chemokines in leukocyte infiltration and tumor metastatization in murine tumor models.

The role of neutrophils in cancer and metastasis is still debated and controversial since they have been shown to be endowed with both pro- and antitumor functions. These contradictory results seem to be now explained by recent discoveries of tumor-associated neutrophils plasticity and multiple neutrophil subsets. Chemokines and chemokine receptors are known to tightly regulate the release of neutrophils from the bone marrow, their passage into circulation and transmigration into the tissues as well as tumor infiltration. It is emerging that chemokine receptors are differentially expressed by neutrophil subsets and they affect not only their recruitment but also their effector functions. Here we are resuming human and murine data suggesting that therapeutic modulation of neutrophil activity through the targeting of specific chemokines or chemokine receptors can improve their anti-tumoral properties.

Chemokine receptors are differentially expressed on leukocyte subpopulations dictating their ability to migrate both in physiological and pathological conditions. Their expression is modulated during leukocyte differentiation and maturation and they can be used as markers to identify and characterize the frequency and the activation state of leukocytes present in a tissue. Here, we will describe flow cytometry approaches to detect chemokine receptors identifying subpopulations of circulating monocytes and neutrophils.

Chemokines are a large family of secreted cytokines whose main function is to mediate leukocyte directional migration. Most cancers contain chemokines and express chemokine receptors as a consequence of the activity of deregulated transcription factors or tumor-suppressor genes. Indeed chemokines expression at the tumor site dictates leukocyte infiltration and angiogenesis, while chemokine receptors expression by tumor cells promotes their growth and matastatization. Chemokines also have several indirect effects on tumor growth and are a relevant element in the cancer-related inflammation. In this chapter we will describe technical approaches available to study the role of chemokines in leukocyte infiltration and tumor metastatization in murine tumor models.

Chemokine receptors are differentially expressed on leukocyte subpopulations dictating their ability to migrate both in physiological and pathological conditions. Their expression is modulated during leukocyte differentiation and maturation and they can be used as markers to identify and characterize the frequency and the activation state of leukocytes present in a tissue. Here, we will describe flow cytometry approaches to detect chemokine receptors identifying subpopulations of circulating monocytes and neutrophils.

The chemokine system includes a set of G protein-uncoupled receptors characterized by arrestin-biased signaling, recently named atypical chemokine receptors (ACKRs). ACKRs are emerging as a distinct subfamily of chemokine receptors with a significant impact on the leukocyte recruitment in vivo in face of their inability to support cell migration. ACKRs control chemokine distribution in tissues by different mechanisms, including degradation, transcytosis, and presentation to typical chemokine receptors. The unbalanced activation of the β-arrestin signaling module provides a new conceptual element in the understanding of the molecular mechanisms underlying their loss of chemotactic activity and gain of chemokine gradient-shaping functions.

IL-1R2 was the first decoy receptor to be described. Subsequently receptors which act as pure decoys or scavengers or trigger dampening of cytokine signaling have been described for cytokines and chemokines. Here we review the current understanding of the mode of action and significance in pathology of the chemokine atypical receptor ACKR2, the IL-1 decoy receptor IL-1R2 and the atypical IL-1 receptor family IL-1R8. Decoy and scavenger receptors with no or atypical signaling have emerged as a general strategy conserved in evolution to tune the action of cytokines, chemokines and growth factors.

Objective— Leukocyte infiltration in ischemic areas is a hallmark of myocardial infarction, and overwhelming infiltration of innate immune cells has been shown to promote adverse remodeling and cardiac rupture. Recruitment of inflammatory cells in the ischemic heart depends highly on the family of CC-chemokines and their receptors. Here, we hypothesized that the chemokine decoy receptor D6, which specifically binds and scavenges inflammatory CC-chemokines, might limit inflammation and adverse cardiac remodeling after infarction. Methods and Results— D6 was expressed in human and murine infarcted myocardium. In a murine model of myocardial infarction, D6 deficiency led to increased chemokine (C-C motif) ligand 2 and chemokine (C-C motif) ligand 3 levels in the ischemic heart. D6-deficient (D6 −/−) infarcts displayed increased infiltration of pathogenic neutrophils and Ly6Chi monocytes, associated with strong matrix metalloproteinase-9 and matrix metalloproteinase-2 activities in the ischemic heart. D6 −/− mice were cardiac rupture prone after myocardial infarction, and functional analysis revealed that D6 −/− hearts had features of adverse remodeling with left ventricle dilation and reduced ejection fraction. Bone marrow chimera experiments showed that leukocyte-borne D6 had no role in this setting, and that leukocyte-specific chemokine (C-C motif) receptor 2 deficiency rescued the adverse phenotype observed in D6 −/− mice. Conclusion— We show for the first time that the chemokine decoy receptor D6 limits CC-chemokine–dependent pathogenic inflammation and is required for adequate cardiac remodeling after myocardial infarction.

The atypical chemokine receptor D6 is a decoy and scavenger receptor for most inflammatory CC chemokines and prevents the development of exacerbated inflammatory reactions. Here we report that mice lacking D6 expression in the nonhematopoietic compartment have a selective increase in the number of Ly6Chigh monocytes in the circulation and in secondary lymphoid tissues. Under inflammatory conditions, Ly6Chigh monocytes accumulate in increased number in secondary lymphoid organs of D6−/− mice in a CCR2-dependent manner. Ly6Chigh monocytes derived from D6−/− mice have enhanced immunosuppressive activity, inhibit the development of adaptive immune responses, and partially protect mice from the development of GVHD. Thus, control of CCR2 ligands by D6 regulates the traffic of Ly6Chigh monocytes and controls their immunosuppressive potential.

Chemokines are important components of the hematopoietic niche. The atypical chemokine receptor 1 (ACKR1), expressed on erythrocyte precursors, regulates myeloid differentiation.

Chemokines coordinate leukocyte recruitment during inflammatory and immune responses through the interaction with a distinct subfamily of G protein-coupled receptors. The magnitude of the cellular response elicited by chemokines is dictated by the level of receptor expression at the plasma membrane, which is the balance of finely tuned endocytic and recycling pathways. Recent data have revealed that receptor trafficking properties can drive chemokine receptors to lysosomal degradation or recycling pathways, producing opposite effects on the strength of the intracellular signaling cascade. This review will cover recent advances on the molecular mechanisms underlying chemokine receptor internalization, recycling and degradation pathways, with particular attention to structural motifs present in receptor intracellular domains and their interacting adaptor proteins that modulate receptor trafficking and dictate proper biological response.

Chemokines are a key component of cancer-related inflammation. Chemokines and chemokine receptors are downstream of genetic events that cause neoplastic transformation and are components of chronic inflammatory conditions, which predispose to cancer. Components of the chemokine system affect in a cell autonomous or non-autonomous way multiple pathways of tumor progression, including: leukocyte recruitment and function; cellular senescence; tumor cell proliferation and survival; invasion and metastasis. Available information in preclinical and clinical settings suggests that the chemokine system represents a valuable target for the development of innovative therapeutic strategies.

The decoy receptor D6 plays a nonredundant role in the control of inflammatory processes through scavenging of inflammatory chemokines. However it remains unclear how it is regulated. Here we show that D6 scavenging activity relies on unique trafficking properties. Under resting conditions, D6 constitutively recycled through both a rapid wortmannin (WM)–sensitive and a slower brefeldin A (BFA)–sensitive pathway, maintaining low levels of surface expression that required both Rab4 and Rab11 activities. In contrast to “conventional” chemokine receptors that are down-regulated by cognate ligands, chemokine engagement induced a dose-dependent BFA-sensitive Rab11-dependent D6 re-distribution to the cell membrane and a corresponding increase in chemokine degradation rate. Thus, the energy-expensive constitutive D6 cycling through Rab11 vesicles allows a rapid, ligand concentration–dependent increase of chemokine scavenging activity by receptor redistribution to the plasma membrane. D6 is not regulated at a transcriptional level in a variety of cellular contexts, thus ligand-dependent optimization of its scavenger performance represents a rapid and unique mechanism allowing D6 to control inflammation.

Chemokines play a key role in immune processes by controlling leukocyte recruitment in physiological and pathological condition. This review discusses the regulation of the chemokine system, its role in human diseases, and its potential relevance as a new pharmacological target.