CXCR6 positions cytotoxic T cells to receive critical survival signals in the tumor microenvironment.

Cytotoxic T lymphocyte (CTL) responses against tumors are maintained by stem-like memory cells that self-renew but also give rise to effector-like cells. The latter gradually lose their anti-tumor activity and acquire an epigenetically fixed, hypofunctional state, leading to tumor tolerance. Here, we show that the conversion of stem-like into effector-like CTLs involves a major chemotactic reprogramming that includes the upregulation of chemokine receptor CXCR6. This receptor positions effector-like CTLs in a discrete perivascular niche of the tumor stroma that is densely occupied by CCR7+ dendritic cells (DCs) expressing the CXCR6 ligand CXCL16. CCR7+ DCs also express and trans-present the survival cytokine interleukin-15 (IL-15). CXCR6 expression and IL-15 trans-presentation are critical for the survival and local expansion of effector-like CTLs in the tumor microenvironment to maximize their anti-tumor activity before progressing to irreversible dysfunction. These observations reveal a cellular and molecular checkpoint that determines the magnitude and outcome of anti-tumor immune responses.

Structural Basis for Allosteric Ligand Recognition in the Human CC Chemokine Receptor 7.

The CC chemokine receptor 7 (CCR7) balances immunity and tolerance by homeostatic trafficking of immune cells. In cancer, CCR7-mediated trafficking leads to lymph node metastasis, suggesting the receptor as a promising therapeutic target. Here, we present the crystal structure of human CCR7 fused to the protein Sialidase NanA by using data up to 2.1 Å resolution. The structure shows the ligand Cmp2105 bound to an intracellular allosteric binding pocket. A sulfonamide group, characteristic for various chemokine receptor ligands, binds to a patch of conserved residues in the Gi protein binding region between transmembrane helix 7 and helix 8. We demonstrate how structural data can be used in combination with a compound repository and automated thermal stability screening to identify and modulate allosteric chemokine receptor antagonists. We detect both novel (CS-1 and CS-2) and clinically relevant (CXCR1-CXCR2 phase-II antagonist Navarixin) CCR7 modulators with implications for multi-target strategies against cancer.

CCR7 Chemokine Receptor-Inducible lnc-Dpf3 Restrains Dendritic Cell Migration by Inhibiting HIF-1α-Mediated Glycolysis.

CCR7 chemokine receptor stimulation induces rapid but transient dendritic cell (DC) migration toward draining lymph nodes, which is critical for the initiation of protective immunity and maintenance of immune homeostasis. The mechanisms for terminating CCR7-mediated DC migration remain incompletely understood. Here we have identified a long non-coding RNA lnc-Dpf3 whose feedback restrained CCR7-mediated DC migration. CCR7 stimulation upregulated lnc-Dpf3 via removing N6-methyladenosine (m6A) modification to prevent RNA degradation. DC-specific lnc-Dpf3 deficiency increased CCR7-mediated DC migration, leading to exaggerated adaptive immune responses and inflammatory injuries. Mechanistically, CCR7 stimulation activated the HIF-1α transcription factor pathway in DCs, leading to metabolic reprogramming toward glycolysis for DC migration. lnc-Dpf3 directly bound to HIF-1α and suppressed HIF-1α-dependent transcription of the glycolytic gene Ldha, thus inhibiting DC glycolytic metabolism and migratory capacity. We demonstrate a critical role for CCR7-inducible lnc-Dpf3 in coupling epigenetic and metabolic pathways to feedback-control DC migration and inflammatory responses.

Rapid anti-myeloma activity by T cells expressing an anti-BCMA CAR with a human heavy-chain-only antigen-binding domain.

Multiple myeloma (MM) is a rarely curable malignancy of plasma cells. MM expresses B cell maturation antigen (BCMA). We developed a fully human anti-BCMA chimeric antigen receptor (CAR) with a heavy-chain-only antigen-recognition domain, a 4-1BB domain, and a CD3ζ domain. The CAR was designated FHVH33-CD8BBZ. We conducted the first-in-humans clinical trial of T cells expressing FHVH33-CD8BBZ (FHVH-T). Twenty-five patients with relapsed MM were treated. The stringent complete response rate (sCR) was 52%. Median progression-free survival (PFS) was 78 weeks. Of 24 evaluable patients, 6 (25%) had a maximum cytokine-release syndrome (CRS) grade of 3; no patients had CRS of greater than grade 3. Most anti-MM activity occurred within 2-4 weeks of FHVH-T infusion as shown by decreases in the rapidly changing MM markers serum free light chains, urine light chains, and bone marrow plasma cells. Blood CAR+ cell levels peaked during the time that MM elimination was occurring, between 7 and 15 days after FHVH-T infusion. C-C chemokine receptor type 7 (CCR7) expression on infusion CD4+ FHVH-T correlated with peak blood FHVH-T levels. Single-cell RNA sequencing revealed a shift toward more differentiated FHVH-T after infusion. Anti-CAR antibody responses were detected in 4 of 12 patients assessed. FHVH-T has powerful, rapid, and durable anti-MM activity.

Chemokine receptor 7 contributes to T- and B-cell filtering in ageing bladder, cystitis and bladder cancer.

BACKGROUND: Research has suggested significant correlations among ageing, immune microenvironment, inflammation and tumours. However, the relationships among ageing, immune microenvironment, cystitis and bladder urothelial carcinoma (BLCA) in the bladder have rarely been reported. METHODS: Bladder single-cell and transcriptomic data from young and old mice were used for immune landscape analysis. Transcriptome, single-cell and The Cancer Genome Atlas Program datasets of BLCA and interstitial cystitis/bladder pain syndrome (IC/BPS) were used to analyse immune cell infiltration and molecular expression. Bladder tissues from mice, IC/BPS and BLCA were collected to validate the results. RESULTS: Eight types of immune cells (macrophages, B-cells, dendritic cells, T-cells, monocytes, natural killer cells, γδ T-cells and ILC2) were identified in the bladder of mice. Aged mice bladder tissues had a significantly higher number of T-cells, γδ T-cells, ILC2 and B-cells than those in the young group (P < 0.05). Three types of T-cells (NK T-cells, γδ T-cells and naïve T-cells) and three types of B-cells (follicular B-cells, plasma and memory B-cells) were identified in aged mice bladder. Chemokine receptor 7 (CCR7) is highly expressed in aged bladder, IC/BPS and BLCA (P < 0.05). CCR7 is likely to be involved in T- and B-cell infiltration in aged bladder, IC/BPS and BLCA. Interestingly, the high CCR7 expression on BLCA cell membranes was a prognostic protective factor. CONCLUSIONS: In this study, we characterised the expression profiles of immune cells in bladder tissues of aged and young mice and demonstrated that CCR7-mediated T- and B-cell filtration contributes to the development of bladder ageing, IC/BPS and BLCA.

Astragaloside IV targeting autophagy of T cells improves inflammation of asthma.

Astragaloside IV (AST) has been confirmed to have antiasthmatic effects. However, the underline mechanism is unclear. The study aimed to explore the treatment mechanism of AST based on autophagy of memory T cells. AST treatment significantly decreased the number of T effector cells in asthma mice blood and the nude mice that received AST-treated TCMs had relieved inflammation compared with the untreated group; meanwhile, we found that AST significantly decreased the autophagy level and inhibited OX40/OX40L signal pathway of lymphocytes. The results highlighted that AST regulated autophagy to inhibit differentiation of effector T-cell phenotype.

CCL21-Ser expression in melanoma cells recruits CCR7+ naïve T cells to tumor tissues and promotes tumor growth.

CCL21-Ser, a chemokine encoded by the Ccl21a gene, is constitutively expressed in the thymic epithelial cells and stromal cells of secondary lymphoid organs. It regulates immune cell migration and survival through its receptor CCR7. Herein, using CCL21-Ser-expressing melanoma cells and the Ccl21a-deficient mice, we demonstrated the functional role of cancer cell-derived CCL21-Ser in melanoma growth in vivo. The B16-F10 tumor growth was significantly decreased in Ccl21a-deficient mice compared with that in wild-type mice, indicating that host-derived CCL21-Ser contributes to melanoma proliferation in vivo. In Ccl21a-deficient mice, tumor growth of melanoma cells expressing CCL21-Ser was significantly enhanced, suggesting that CCL21-Ser from melanoma cells promotes tumor growth in the absence of host-derived CCL21-Ser. The increase in tumor growth was associated with an increase in the CCR7+ CD62L+ T cell frequency in the tumor tissue but was inversely correlated with Treg frequency, suggesting that naïve T cells primarily promote tumor growth. Adoptive transfer experiments demonstrated that naïve T cells are preferentially recruited from the blood into tumors with melanoma cell-derived CCL21-Ser expression. These results suggest that CCL21-Ser from melanoma cells promotes the infiltration of CCR7+ naïve T cells into the tumor tissues and creates a tumor microenvironment favorable for melanoma growth.

CCL19/CCR7 drives regulatory T cell migration and indicates poor prognosis in gastric cancer.

BACKGROUND: Gastric cancer is associated with significant morbidity and mortality in the world. Blocking programmed cell death protein 1 pathway have been approved for the treatment of a variety of tumors and have achieved remarkable clinical therapeutic effects. However, immune checkpoint inhibitors failed to achieve satisfactory results in gastric cancer. There is a need to identify novel immunotherapy targets in gastric cancer. METHODS: We analysed the correlation between Treg cells and CD8 + T cells in gastric cancer samples. We studied the relationship between chemokines and Treg cells or CD8 + T cells in gastric cancer. We compared CCL19/CCR7 expression in gastric cancer patients in TCGA database. We performed transwell experiments to determine the influence of CCL19 on Treg cells and CD8 + T cells migratory capacity. We conducted survival analysis of CCL19 and CCR7 in gastric cancer database. RESULTS: Treg cells show positive correlation with CD8 + T cells in gastric cancer. Treg cell expression was significantly upregulated in tumor tissues. Patients with high FOXP3 expression had worse overall survival than those with low FOXP3 expression. CCL19 had strong correlation with FOXP3 and weak correlation with CD8A. CCL19 had strong impact on the migratory capacity of Treg cells but weak impact on the migratory capacity of CD8 + T cells. Both CCL19 and CCR7 expression were significantly upregulated in gastric cancer tissues. Survival analysis demonstrated that both CCL19 and CCR7 indicate poor prognosis in gastric cancer. CONCLUSIONS: CCL19/CCR7 may be a potential novel therapeutic target in gastric cancer.

Evaluation of the novel therapeutic anti-CCR7 antibody CAP-100 as an add-on therapy in chronic lymphocytic leukemia patients receiving venetoclax.

The Bruton's tyrosine kinase inhibitor ibrutinib and the B-cell lymphoma 2 anti-apoptotic protein inhibitor venetoclax provide high response rates in chronic lymphocytic leukemia (CLL). However, there is a growing number of patients that relapse after treatment or show refractory disease, thus new targets and agents are still needed. We have previously reported the chemokine receptor CCR7 as a relevant deregulated target in CLL and have developed CAP-100, a novel therapeutic anti-CCR7 antibody that is under evaluation for relapse/refractory CLL (NCT04704323). While CCR7 expression has been shown to be down-modulated in CLL patients treated with ibrutinib, whether venetoclax acts in a similar manner remains unaddressed. Here, we aimed to document the impact of venetoclax on CCR7 expression in CLL cells, as well as on the pre-clinical activity of CAP-100. To this end, we addressed CCR7 expression by flow cytometry and the antibody efficacy by means of in vitro chemotactic and antibody-dependent cell-mediated cytotoxicity (ADCC) assays. Our data indicate that venetoclax treatment did not significantly modify CCR7 expression pattern nor CAP-100 mechanisms of action. Together, these findings support CAP-100 as an adjuvant therapy to venetoclax that may introduce additional modes of action in CLL therapy.

Health prevention intervention for chronic tissue fibrosis: Based on the specific expression of CCL19/21 + mast cell.

Chronic tissue fibrosis is a common pathological feature of connective tissue diseases and malignant tumors, and its prevention has been a major focus of relevant research.However, the details of the mechanism of action of tissue-colonizing immune cells in fibroblast migration are unclear. In this study, connective tissue disease tissue specimens and solid tumor specimens were selected to observe the relationship between mast cells and interstitial fibrosis and the expression characteristics of mast cells. Our findings suggest that the number of mast cells in the tissue correlates with the degree of pathological fibrosis and that mast cells specifically express the chemokines CCL19 and CCL21, especially CCL19. CCR7+ fibroblasts are highly expressed in mast cell clusters. The mast cell line HMC-1 regulates CD14+ monocyte-derived fibroblasts via CCL19. In disease tissue fibrosis, mast cell activation may increase the expression of chemokines, especially CCL19, in the tissue, thereby inducing a large number of CCR7-positive fibroblasts to migrate to specific tissues. This study lays a foundation for the mechanism of tissue fibrosis and provides evidence for the mechanism by which mast cells induce fibroblast migration.Through the experimental results of this paper, we can combine the induction factors of chronic tissue fibrosis and put forward targeted health prevention strategies.

Heparin Specifically Interacts with Basic BBXB Motifs of the Chemokine CCL21 to Define CCR7 Signaling.

Chemokines are critically involved in controlling directed leukocyte migration. Spatiotemporal secretion together with local retention processes establish and maintain local chemokine gradients that guide directional cell migration. Extracellular matrix proteins, particularly glycosaminoglycans (GAGs), locally retain chemokines through electrochemical interactions. The two chemokines CCL19 and CCL21 guide CCR7-expressing leukocytes, such as antigen-bearing dendritic cells and T lymphocytes, to draining lymph nodes to initiate adaptive immune responses. CCL21-in contrast to CCL19-is characterized by a unique extended C-terminus composed of highly charged residues to facilitate interactions with GAGs. Notably, both chemokines can trigger common, but also ligand-biased signaling through the same receptor. The underlying molecular mechanism of ligand-biased CCR7 signaling is poorly understood. Using a series of naturally occurring chemokine variants in combination with newly designed site-specific chemokine mutants, we herein assessed CCR7 signaling, as well as GAG interactions. We demonstrate that the charged chemokine C-terminus does not fully confer CCL21-biased CCR7 signaling. Besides the positively charged C-terminus, CCL21 also possesses specific BBXB motifs comprising basic amino acids. We show that CCL21 variants where individual BBXB motifs are mutated retain their capability to trigger G-protein-dependent CCR7 signaling, but lose their ability to interact with heparin. Moreover, we show that heparin specifically interacts with CCL21, but not with CCL19, and thereby competes with ligand-binding to CCR7 and prevents signaling. Hence, we provide evidence that soluble heparin, but not the other GAGs, complexes with CCL21 to define CCR7 signaling in a ligand-dependent manner.

iNKT cells with high PLZF expression are recruited into the lung via CCL21-CCR7 signaling to facilitate the development of asthma tolerance in mice.

Establishment of immune tolerance is crucial to protect humans against asthma. Promyelocytic leukemia zinc finger (PLZF) is an emerging suppressor of inflammatory responses. CCL21-CCR7 signaling mediates tolerance development. However, whether PLZF and CCL21-CCR7 are required for the development of asthma tolerance is unknown. Here, we found that Zbtb16 (coding PLZF) and Ccl21 were upregulated in OVA-induced asthma tolerance (OT) lungs by RNA-seq. PLZF physically interacted with GATA3 and its expression was higher in GATA3+ Th2 cells and ILC2s in OT lungs. Zbtb16-knockdown in lymphocytes promoted the differentiation of CD3e+ CD4+ T cells, particularly those producing IL-4 and IL-5. Moreover, iNKT cells with high expression of PLZF were recruited into the lungs via draining lymph nodes during tolerance. Blockade of CCL21-CCR7 signaling in OT mice decreased the PLZF+  cell population, abolished CCR7-induced PLZF+ iNKT recruitment to the lungs, enhanced Th2responses and exacerbated lung pathology. In OT mice, respiratory syncytial virus (RSV) infection impeded PLZF+  cell and CCR7+ PLZF+ iNKT cellrecruitment to the lungs and increased airway resistance. Collectively, these results indicate that PLZF could interact with GATA3 and restrain differentiation of IL-4- and IL-5-producing T cells, iNKT cells with high PLZF expression are recruited to the lungs via CCL21-CCR7 signaling to facilitate the development of asthma tolerance.

Effect of ibrutinib on CCR7 expression and functionality in chronic lymphocytic leukemia and its implication for the activity of CAP-100, a novel therapeutic anti-CCR7 antibody.

CAP-100 is a novel therapeutic antibody directed against the ligand binding site of human CCR7. This chemokine receptor is overexpressed in chronic lymphocytic leukemia (CLL) and orchestrates the homing of CLL cells into the lymph node. Previous studies, on a very limited number of samples, hypothesized that the Bruton's tyrosine kinase inhibitor (BTKi) ibrutinib might induce loss of surface CCR7 levels in CLL cells. CAP-100 will be evaluated in clinical trials as a therapy for relapse/refractory CLL patients, who have received at least two systemic therapies (NCT04704323). As nowadays many relapse/refractory CLL patients will have received ibrutinib as a prior therapy, we aimed to investigate in a large cohort of CLL patients the impact of this BTKi on CCR7 expression and functionality as well as on the therapeutic activity of CAP-100. Our data confirm that ibrutinib moderately down-regulates the very high expression of CCR7 in CLL cells but has no apparent effect on CCR7-induced chemotaxis. Moreover, CLL cells are perfectly targetable by CAP-100 which led to a complete inhibition of CCR7-mediated migration and induced strong target cell killing through antibody-dependent cell-mediated cytotoxicity, irrespective of previous or contemporary ibrutinib administration. Together, these results validate the therapeutic utility of CAP-100 as a next-line single-agent therapy for CLL patients who failed to ibrutinib and confirm that CAP-100 and ibrutinib have complementary non-overlapping mechanisms of action, potentially allowing for combination therapy.

ccr7 affects both morphogenesis and differentiation during early Xenopus embryogenesis.

Chemokines play important roles in early embryogenesis, including morphogenesis and cell differentiation, before the immune system is established. We characterized Xenopus laevis CC-type chemokine receptor 7 S (ccr7.S) to clarify its role during early development. ccr7 transcripts were detected ubiquitously in early embryos. Dorsal overexpression of ccr7.S inhibited gastrulation, and ccr7.S mRNA-injected embryos had short axes and widely opened neural folds. Because the Keller sandwich explants of the injected embryos elongated well, ccr7.S might affect cell migration, but not convergent extension movements. Ventral ccr7.S overexpression induced secondary axes and chrd.1 upregulation in gastrula-stage embryos. Animal cap assays showed increased expression of neural and cement gland marker genes at later stages. Ccr7.S knockdown reduced chrd.1 expression and inhibited gastrulation at the dorsal side. Our findings suggest that ccr7.S plays important roles in morphogenetic movement and cell differentiation.

CCR7 alterations associated with inferior outcome of adult T-cell leukemia/lymphoma under mogamulizumab treatment.

Adult T-cell leukemia/lymphoma (ATL) patients have a very poor prognosis. The humanized anti-CCR4 therapeutic monoclonal antibody, mogamulizumab, is a key agent for ATL treatment. Our previous integrated molecular analysis demonstrated that among all the driver genes in ATL, CCR7 gene alterations were significantly associated with clinical response to mogamulizumab. Accordingly, here we investigated the detailed clinical impact of CCR7 alterations in a larger cohort of ATL patients. These CCR7 alterations, most of which lead to C-terminus truncations, were observed in 27 of 223 patients (12%). For patients receiving mogamulizumab but not allogeneic hematopoietic stem cell transplantation (HSCT), CCR7 alterations were significantly associated with worse survival (median survival from the first dose of mogamulizumab of 0.7 years for 12 patients with CCR7 alterations vs. 1.6 years for 72 patients without, p = 0.020). On the other hand, the presence or absence of CCR7 alterations had no significant impact on survival in the entire cohort (median overall survival of 1.4 and 1.8 years, respectively, p = 0.901), or on the survival of patients receiving allogeneic HSCT (median survival from the day of transplantation of 0.9 years for 6 patients with CCR7 alterations and 1.4 years for 48 without, p = 0.543). Multivariate analysis indicated that patients with CCR4 alterations but lacking CCR7 alterations (n = 20) had significantly better survival after receiving mogamulizumab-containing treatments (hazard ratio for survival, 0.437, 95% confidence interval, 0.192-0.994). This study contributes to the establishment of precision medicine for ATL.

Trichinella spiralis nurse cell formation is regulated via CCR7+ dendritic cells.

The chemokine receptor CCR7 is a well-established homing receptor for dendritic cells (DCs) and T-cells. Interaction with the CCL19 and CCL21 ligands promotes priming of immune responses in lymphoid tissues; however, the mechanism underlying CCR7-induced immune responses against helminth parasite infection remains unknown. Thus, we examined the role of CCR7 in generating protective immune responses against intracellular Trichinella spiralis infection. The results showed significantly increased CCR7, CCL19 and CCL21 expression in the muscle tissue compared to that in the intestinal tissue in T. spiralis-infected mice. The CCR7-expressing DC population increased in the mesenteric and peripheral lymph nodes (PLNs) during T. spiralis infection. Notably, the number of CCR7-expressing cells in PLNs increased by more than 30% at 28 days post-infection; however, this increase was significantly inhibited in CCR7-blocked mice treated with CCR7-specific antibodies. T helper 2 (Th2)-and regulatory T (Treg )-related cytokine levels were also reduced by CCR7-specific antibody treatment. CCR7-blocked mice lost their resistance to T. spiralis infection in the muscle phase but not in the intestinal phase. Furthermore, fewer eosinophils around the nurse cells and reduced total and T. spiralis-specific IgE in the serum were observed in CCR7-blocked mice compared to those infected with only T. spiralis. CCR7 blockade led to the T. spiralis infection-induced suppression of Th2- and Treg -related cytokine production in vitro. These results suggest that CCR7 in DCs might play an essential role in host defence mechanisms against T. spiralis infection, particularly in the muscle stage of the infection, by accelerating Th2 and Treg cell responses.

WGCNA and molecular docking reveal key hub genes and potential natural inhibitor in interstitial cystitis/bladder pain syndrome.

INTRODUCTION AND HYPOTHESIS: The etiology and treatment of interstitial cystitis/bladder pain syndrome are still controversial. The purpose of this study is to determine the key genes and specific regulatory pathways related to it and to find potential drug-active components through integrated bioinformatics. METHODS: The data set GSE11783 was downloaded from GEO database. The modules significantly related to interstitial cystitis/bladder pain syndrome were identified by weighted correlation network analysis. The genes in the key modules were analyzed by functional enrichment and protein interaction by Cytoscape software, and finally the core hub genes were screened. Furthermore, the molecular docking verification of active components and key proteins was carried out by using AutoDock Vin software. RESULTS: Among the 14 modules derived from WGCNA, turquoise module had the highest correlation with IC/BPS (r = 0.85, P < 0.001). The genes in the module were mainly enriched in the biological processes such as the interaction between cytokines and cytokine receptors and chemokine signaling pathway. The genes in the related modules of differentially expressed genes and WGCNA traits were intersected to obtain the core hub genes. Protein-protein interaction network analysis showed that the key genes were upregulated genes CCR7 and CCL19. In terms of molecular docking, triptolide, the active component in the traditional anti-inflammatory drug Tripterygium wilfordii, can form effective molecular binding with both core hub genes. CONCLUSIONS: Our study identified the core hub genes CCR7 and CCL19, which acted as essential components in interstitial cystitis/bladder pain syndrome. Furthermore, CCR7 and CCL19 can form effective binding with triptolide, which will provide new insights into the development of new therapies for interstitial cystitis/bladder pain syndrome.

The C-terminal peptide of CCL21 drastically augments CCL21 activity through the dendritic cell lymph node homing receptor CCR7 by interaction with the receptor N-terminus.

The endogenous chemokines CCL19 and CCL21 signal via their common receptor CCR7. CCL21 is the main lymph node homing chemokine, but a weak chemo-attractant compared to CCL19. Here we show that the 41-amino acid positively charged peptide, released through C-terminal cleavage of CCL21, C21TP, boosts the immune cell recruiting activity of CCL21 by up to 25-fold and the signaling activity via CCR7 by ~ 100-fold. Such boosting is unprecedented. Despite the presence of multiple basic glycosaminoglycan (GAG) binding motifs, C21TP boosting of CCL21 signaling does not involve interference with GAG mediated cell-surface retention. Instead, boosting is directly dependent on O-glycosylations in the CCR7 N-terminus. As dictated by the two-step binding model, the initial chemokine binding involves interaction of the chemokine fold with the receptor N-terminus, followed by insertion of the chemokine N-terminus deep into the receptor binding pocket. Our data suggest that apart from a role in initial chemokine binding, the receptor N-terminus also partakes in a gating mechanism, which could give rise to a reduced ligand activity, presumably through affecting the ligand positioning. Based on experiments that support a direct interaction of C21TP with the glycosylated CCR7 N-terminus, we propose that electrostatic interactions between the positively charged peptide and sialylated O-glycans in CCR7 N-terminus may create a more accessible version of the receptor and thus guide chemokine docking to generate a more favorable chemokine-receptor interaction, giving rise to the peptide boosting effect.

Selective Boosting of CCR7-Acting Chemokines; Short Peptides Boost Chemokines with Short Basic Tails, Longer Peptides Boost Chemokines with Long Basic Tails.

The chemokine receptor CCR7 and its ligands CCL19 and CCL21 regulate the lymph node homing of dendritic cells and naïve T-cells and the following induction of a motile DC-T cell priming state. Although CCL19 and CCL21 bind CCR7 with similar affinities, CCL21 is a weak agonist compared to CCL19. Using a chimeric chemokine, CCL19CCL21N-term|C-term, harboring the N-terminus and the C-terminus of CCL21 attached to the core domain of CCL19, we show that these parts of CCL21 act in a synergistic manner to lower ligand potency and determine the way CCL21 engages with CCR7. We have published that a naturally occurring basic C-terminal fragment of CCL21 (C21TP) boosts the signaling of both CCL19 and CCL21. Boosting occurs as a direct consequence of C21TP binding to the CCR7 N-terminus, which seems to free chemokines with basic C-termini from an unfavorable interaction with negatively charged posttranslational modifications in CCR7. Here, we confirm this using a CCL19-variant lacking the basic C-terminus. This variant displays a 22-fold higher potency at CCR7 compared to WT CCL19 and is highly unaffected by the presence of C21TP. WT CCL19 has a short basic C-terminus, CCL21 a longer one. Here, we propose a way to differentially boost CCL19 and CCL21 activity as short and long versions of C21TP boost CCL19 activity, whereas only a long C21TP version can boost chemokines with a full-length CCL21 C-terminus.

Distinct Chemokine Receptor Expression Profiles in De Novo DLBCL, Transformed Follicular Lymphoma, Richter's Trans-Formed DLBCL and Germinal Center B-Cells.

Chemokine receptors and their ligands have been identified as playing an important role in the development of diffuse large B-cell lymphoma (DLBCL), follicular lymphoma, and Richter syndrome (RS). Our aim was to investigate the different expression profiles in de novo DLBCL, transformed follicular lymphoma (tFL), and RS. Here, we profiled the mRNA expression levels of 18 chemokine receptors (CCR1-CCR9, CXCR1-CXCR7, CX3CR1 and XCR1) using RQ-PCR, as well as immunohistochemistry of seven chemokine receptors (CCR1, CCR4-CCR8 and CXCR2) in RS, de novo DLBCL, and tFL biopsy-derived tissues. Tonsil-derived germinal center B-cells (GC-B) served as non-neoplastic controls. The chemokine receptor expression profiles of de novo DLBCL and tFL substantially differed from those of GC-B, with at least 5-fold higher expression of 15 out of the 18 investigated chemokine receptors (CCR1-CCR9, CXCR1, CXCR2, CXCR6, CXCR7, CX3CR1 and XCR1) in these lymphoma subtypes. Interestingly, the de novo DLBCL and tFL exhibited at least 22-fold higher expression of CCR1, CCR5, CCR8, and CXCR6 compared with RS, whereas no significant difference in chemokine receptor expression profile was detected when comparing de novo DLBCL with tFL. Furthermore, in de novo DLBCL and tFLs, a high expression of CCR7 was associated with a poor overall survival in our study cohort, as well as in an independent patient cohort. Our data indicate that the chemokine receptor expression profile of RS differs substantially from that of de novo DLBCL and tFL. Thus, these multiple dysregulated chemokine receptors could represent novel clinical markers as diagnostic and prognostic tools. Moreover, this study highlights the relevance of chemokine signaling crosstalk in the tumor microenvironment of aggressive lymphomas.