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1.
J Immunol ; 208(7): 1802-1812, 2022 04 01.
Artigo em Inglês | MEDLINE | ID: mdl-35288470

RESUMO

NK cell receptors allow NK cells to recognize targets such as tumor cells. Many of them are expressed on a subset of NK cells, independently of each other, which creates a vast diversity of receptor combinations. Whether these combinations influence NK cell antitumor responses is not well understood. We addressed this question in the C57BL/6 mouse model and analyzed the individual effector response of 444 mouse NK cell subsets, defined by combinations of 12 receptors, against tumor cell lines originating from different tissues and mouse strains. We found a wide range of reactivity among NK subsets, but the same hierarchy of responses was observed for the different tumor types, showing that the repertoire of NK cell receptors does not encode for different tumor specificities but for different intrinsic reactivities. The coexpression of CD27, NKG2A, and DNAM-1 identified subsets with relative cytotoxic specialization, whereas reciprocally, CD11b and KLRG1 defined the best IFN-γ producers. The expression of educating receptors Ly49C, Ly49I, and NKG2A was also strongly correlated with IFN-γ production, but this effect was suppressed by unengaged receptors Ly49A, Ly49F, and Ly49G2. Finally, IL-15 coordinated NK cell effector functions, but education and unbound inhibitory receptors retained some influence on their response. Collectively, these data refine our understanding of the mechanisms governing NK cell reactivity, which could help design new NK cell therapy protocols.


Assuntos
Interferon gama , Células Matadoras Naturais , Animais , Linhagem Celular Tumoral , Interferon gama/metabolismo , Células Matadoras Naturais/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Receptores de Células Matadoras Naturais/metabolismo
2.
J Hepatol ; 71(6): 1086-1098, 2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31349000

RESUMO

BACKGROUND & AIMS: Liver macrophages can be involved in both pathogen clearance and/or pathogenesis. To get further insight on their role during chronic hepatitis B virus (HBV) infections, our aim was to phenotypically and functionally characterize in vivo and ex vivo the interplay between HBV, primary human liver macrophages (PLMs) and primary blood monocytes differentiated into pro-inflammatory or anti-inflammatory macrophages (M1-MDMs or M2-MDMs, respectively). METHODS: PLMs or primary blood monocytes, either ex vivo differentiated into M1-MDMs or M2-MDMs, were exposed to HBV and their activation followed by ELISA or quantitative reverse transcription PCR (RT-qPCR). Liver biopsies from HBV-infected patients were analysed by RT-qPCR or immunohistochemistry. Viral parameters in HBV-infected primary human hepatocytes and differentiated HepaRG cells were followed by ELISA, qPCR and RT-qPCR analyses. RESULTS: HBc protein was present within the macrophages of liver biopsies taken from HBV-infected patients. Macrophages from HBV-infected patients also expressed higher levels of anti-inflammatory macrophage markers than those from non-infected patients. Ex vivo exposure of naive PLMs to HBV led to reduced secretion of pro-inflammatory cytokines. Upon exposure to HBV or HBV-producing cells during differentiation and activation, M1-MDMs secreted less IL-6 and IL-1ß, whereas M2-MDMs secreted more IL-10 when exposed to HBV during activation. Finally, cytokines produced by M1-MDMs, but not those produced by HBV-exposed M1-MDMs, decreased HBV infection of hepatocytes. CONCLUSIONS: Altogether, our data strongly suggest that HBV modulates liver macrophage functions to favour the establishment of infection. LAY SUMMARY: Hepatitis B virus modulates liver macrophage function in order to favour the establishment and likely maintenance of infection. It impairs the production of the antiviral cytokine IL-1ß, while promoting that of IL-10 in the microenvironment. This phenotype can be recapitulated in naive liver macrophages or monocyte-derived-macrophages ex vivo by short exposure to the virus or cells replicating the virus, thus suggesting an "easy to implement" mechanism of inhibition.


Assuntos
Diferenciação Celular/imunologia , Vírus da Hepatite B/fisiologia , Hepatite B Crônica , Células de Kupffer , Ativação de Macrófagos/imunologia , Monócitos , Células Cultivadas , DNA Viral/isolamento & purificação , Hepatite B Crônica/imunologia , Hepatite B Crônica/patologia , Humanos , Imuno-Histoquímica , Imunomodulação , Interleucina-10 , Interleucina-1beta , Células de Kupffer/imunologia , Células de Kupffer/patologia , Monócitos/imunologia , Monócitos/patologia , Sistema Fagocitário Mononuclear/imunologia
3.
Artigo em Inglês | MEDLINE | ID: mdl-29439958

RESUMO

We previously reported that Toll-like receptor 9 (TLR9)-CpG oligonucleotides could inhibit the establishment of hepatitis B virus (HBV) infections in hepatocytes. Our aim was to uncover the underlying mechanisms of this inhibition. HepaRG cells, RPMI-B lymphoblastoma cells, and primary plasmacytoid dendritic cells (pDCs) exposed to HBV and TLR9 ligands/agonists in various configurations were used. We observed an inhibition of HBV infection upon TLR9 stimulations only when agonist was applied during inoculation. This inhibition was independent of interleukin-6 (IL-6)/interferon-inducible protein 10 (IP-10) production as well as of TLR9 expression in hepatocytes. We further demonstrated an entry inhibition mechanism by showing a noncovalent binding of TLR9 agonist to HBV particles. Besides inhibiting HBV entry into hepatocytes, this biophysical interaction between HBV virions and TLR9 agonist was responsible for a reduction of alpha interferon (IFN-α) expression by pDCs. Interestingly, subviral particles composed of only HBsAg were able to genuinely inhibit the TLR9 pathway, without titrating TLR9 ligands. To conclude, our data suggest that synthetic TLR9-CpG oligonucleotides can strongly inhibit HBV entry by "coating" HBV virions and thereby preventing their interaction with cellular receptor. This titration effect of TLR9 agonist is also artifactually responsible for the inhibition of TLR9 engagement in pDCs, whereas a genuine inhibition of this innate pathway was confirmed with HBsAg subviral particles.


Assuntos
Células Dendríticas/metabolismo , Vírus da Hepatite B/patogenicidade , Hepatócitos/metabolismo , Hepatócitos/virologia , Interferon-alfa/metabolismo , Oligodesoxirribonucleotídeos/química , Oligodesoxirribonucleotídeos/farmacologia , Receptor Toll-Like 9/metabolismo , Vírion/patogenicidade , Linhagem Celular , Vírus da Hepatite B/efeitos dos fármacos , Humanos , Receptores Toll-Like/metabolismo , Vírion/metabolismo
4.
Acta Neuropathol ; 135(4): 569-579, 2018 04.
Artigo em Inglês | MEDLINE | ID: mdl-29299667

RESUMO

Paraneoplastic cerebellar degenerations with anti-Yo antibodies (Yo-PCD) are rare syndromes caused by an auto-immune response against neuronal antigens (Ags) expressed by tumor cells. However, the mechanisms responsible for such immune tolerance breakdown are unknown. We characterized 26 ovarian carcinomas associated with Yo-PCD for their tumor immune contexture and genetic status of the 2 onconeural Yo-Ags, CDR2 and CDR2L. Yo-PCD tumors differed from the 116 control tumors by more abundant T and B cells infiltration occasionally organized in tertiary lymphoid structures harboring CDR2L protein deposits. Immune cells are mainly in the vicinity of apoptotic tumor cells, revealing tumor immune attack. Moreover, contrary to un-selected ovarian carcinomas, 65% of our Yo-PCD tumors presented at least one somatic mutation in Yo-Ags, with a predominance of missense mutations. Recurrent gains of the CDR2L gene with tumor protein overexpression were also present in 59% of Yo-PCD patients. Overall, each Yo-PCD ovarian carcinomas carried at least one genetic alteration of Yo-Ags. These data demonstrate an association between massive infiltration of Yo-PCD tumors by activated immune effector cells and recurrent gains and/or mutations in autoantigen-encoding genes, suggesting that genetic alterations in tumor cells trigger immune tolerance breakdown and initiation of the auto-immune disease.


Assuntos
Autoantígenos/genética , Proteínas do Tecido Nervoso/genética , Neoplasias Ovarianas/genética , Neoplasias Ovarianas/imunologia , Degeneração Paraneoplásica Cerebelar/genética , Degeneração Paraneoplásica Cerebelar/imunologia , Linfócitos B/imunologia , Linfócitos B/patologia , Carcinoma/genética , Carcinoma/imunologia , Carcinoma/patologia , Estudos de Coortes , Feminino , Expressão Gênica , Humanos , Imunoglobulina G/metabolismo , Pessoa de Meia-Idade , Mutação , Gradação de Tumores , Neoplasias Ovarianas/patologia , Degeneração Paraneoplásica Cerebelar/patologia , Linfócitos T/imunologia , Linfócitos T/patologia
5.
Eur J Immunol ; 46(2): 354-9, 2016 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-26518732

RESUMO

Intestinal DCs orchestrate gut immune homeostasis by dampening proinflammatory T-cell responses and inducing anti-inflammatory IgA responses. Although no specific DC subset has been strictly assigned so far to govern IgA response, some candidate subsets emerge. In particular, plasmacytoid DCs (pDCs), which notoriously promote anti-viral immunity and T-cell tolerance to innocuous antigens (Ags), contribute to IgA induction in response to intestinal viral infection and promote T-cell-independent IgA responses in vitro. Here, using two transgenic mouse models, we show that neither short-term nor long-term pDC depletion alters IgA class switch recombination in Peyer's patches and frequency of IgA plasma cells in intestinal mucosa at steady state, even in the absence of T-cell help. In addition, pDCs are dispensable for induction of intestinal IgA plasma cells in response to oral immunization with T-cell-dependent or T-cell-independent Ags, and are not required for proliferation and IgA switch of Ag-specific B cells in GALT. These results show that pDCs are dispensable for noninfectious IgA responses, and suggest that various DC subsets may play redundant roles in the control of intestinal IgA responses.


Assuntos
Linfócitos B/imunologia , Células Dendríticas/imunologia , Imunoglobulina A/metabolismo , Mucosa Intestinal/imunologia , Plasmócitos/imunologia , Animais , Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos/genética , Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos/metabolismo , Homeostase , Humanos , Tolerância Imunológica , Imunização , Switching de Imunoglobulina , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Transgênicos , Linfócitos T/imunologia , Fator de Transcrição 4
6.
J Autoimmun ; 67: 8-18, 2016 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-26341385

RESUMO

Plasmacytoid dendritic cells (pDCs) exhibit both innate and adaptive functions. In particular they are the main source of type I IFNs and directly impact T cell responses through antigen presentation. We have previously demonstrated that during experimental autoimmune encephalomyelitis (EAE) initiation, myelin-antigen presentation by pDCs is associated with suppressive Treg development and results in attenuated EAE. Here, we show that pDCs transferred during acute disease phase confer recovery from EAE. Clinical improvement is associated with migration of injected pDCs into inflamed CNS and is dependent on the subsequent and selective chemerin-mediated recruitment of endogenous pDCs to the CNS. The protective effect requires pDC pre-loading with myelin antigen, and is associated with the modulation of CNS-infiltrating pDC phenotype and inhibition of CNS encephalitogenic T cells. This study may pave the way for novel pDC-based cell therapies in autoimmune diseases, aiming at specifically modulating pathogenic cells that induce and sustain autoimmune inflammation.


Assuntos
Transferência Adotiva , Quimiotaxia/imunologia , Células Dendríticas/imunologia , Células Dendríticas/metabolismo , Encefalomielite Autoimune Experimental/imunologia , Encefalomielite Autoimune Experimental/metabolismo , Animais , Autoantígenos/imunologia , Terapia Baseada em Transplante de Células e Tecidos , Quimiocinas/metabolismo , Modelos Animais de Doenças , Encefalomielite Autoimune Experimental/patologia , Encefalomielite Autoimune Experimental/terapia , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Camundongos , Camundongos Knockout , Bainha de Mielina/imunologia , Receptores de Quimiocinas , Receptores Acoplados a Proteínas G/metabolismo , Medula Espinal/imunologia , Medula Espinal/metabolismo , Medula Espinal/patologia
7.
J Immunol ; 193(7): 3398-408, 2014 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-25194054

RESUMO

The stimulation of TLRs by pathogen-derived molecules leads to the production of proinflammatory cytokines. Because uncontrolled inflammation can be life threatening, TLR regulation is important; however, few studies have identified the signaling pathways that contribute to the modulation of TLR expression. In this study, we examined the relationship between activation and the transcriptional regulation of TLR9. We demonstrate that infection of primary human epithelial cells, B cells, and plasmacytoid dendritic cells with dsDNA viruses induces a regulatory temporary negative-feedback loop that blocks TLR9 transcription and function. TLR9 transcriptional downregulation was dependent on TLR9 signaling and was not induced by TLR5 or other NF-κB activators, such as TNF-α. Engagement of the TLR9 receptor induced the recruitment of a suppressive complex, consisting of NF-κBp65 and HDAC3, to an NF-κB cis element on the TLR9 promoter. Knockdown of HDAC3 blocked the transient suppression in which TLR9 function was restored. These results provide a framework for understanding the complex pathways involved in transcriptional regulation of TLR9, immune induction, and inflammation against viruses.


Assuntos
Infecções por Vírus de DNA/imunologia , Vírus de DNA/imunologia , Regiões Promotoras Genéticas/imunologia , Receptor Toll-Like 9/imunologia , Transcrição Gênica/imunologia , Animais , Infecções por Vírus de DNA/genética , Infecções por Vírus de DNA/patologia , Células Dendríticas/imunologia , Células Dendríticas/patologia , Células Epiteliais/imunologia , Células Epiteliais/patologia , Feminino , Técnicas de Silenciamento de Genes , Células HEK293 , Histona Desacetilases/genética , Histona Desacetilases/imunologia , Humanos , Masculino , Camundongos , Células NIH 3T3 , Plasmócitos/imunologia , Plasmócitos/patologia , Receptor Toll-Like 9/genética , Fator de Transcrição RelA/genética , Fator de Transcrição RelA/imunologia , Transcrição Gênica/genética
8.
Oncoimmunology ; 13(1): 2372118, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-38939518

RESUMO

The need for reliable biomarkers to predict clinical benefit from anti-PD1 treatment in metastatic melanoma (MM) patients remains unmet. Several parameters have been considered in the tumor environment or the blood, but none has yet achieved sufficient accuracy for routine clinical practice. Whole blood samples from MM patients receiving second-line anti-PD1 treatment (NCT02626065), collected longitudinally, were analyzed by flow cytometry to assess the immune cell subsets absolute numbers, the expression of immune checkpoints or ligands on T cells and the functionality of innate immune cells and T cells. Clinical response was assessed according to Progression-Free Survival (PFS) status at one-year following initiation of anti-PD1 (responders: PFS > 1 year; non-responders: PFS ≤ 1 year). At baseline, several phenotypic and functional alterations in blood immune cells were observed in MM patients compared to healthy donors, but only the proportion of polyfunctional memory CD4+ T cells was associated with response to anti-PD1. Under treatment, a decreased frequency of HVEM on CD4+ and CD8+ T cells after 3 months of treatment identified responding patients, whereas its receptor BTLA was not modulated. Both reduced proportion of CD69-expressing CD4+ and CD8+ T cells and increased number of polyfunctional blood memory T cells after 3 months of treatment were associated with response to anti-PD1. Of upmost importance, the combination of changes of all these markers accurately discriminated between responding and non-responding patients. These results suggest that drugs targeting HVEM/BTLA pathway may be of interest to improve anti-PD1 efficacy.


Assuntos
Melanoma , Receptor de Morte Celular Programada 1 , Receptores Imunológicos , Membro 14 de Receptores do Fator de Necrose Tumoral , Adulto , Idoso , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/efeitos dos fármacos , Linfócitos T CD8-Positivos/metabolismo , Inibidores de Checkpoint Imunológico/uso terapêutico , Inibidores de Checkpoint Imunológico/farmacologia , Melanoma/tratamento farmacológico , Melanoma/imunologia , Melanoma/patologia , Receptor de Morte Celular Programada 1/antagonistas & inibidores , Receptor de Morte Celular Programada 1/metabolismo , Receptores Imunológicos/metabolismo , Membro 14 de Receptores do Fator de Necrose Tumoral/metabolismo , Resultado do Tratamento
9.
Sci Transl Med ; 16(731): eadd1834, 2024 Jan 24.
Artigo em Inglês | MEDLINE | ID: mdl-38266104

RESUMO

Tumor-associated macrophages (TAMs) are a critical determinant of resistance to PD-1/PD-L1 blockade. This phase 1 study (MEDIPLEX, NCT02777710) investigated the safety and efficacy of pexidartinib, a CSF-1R-directed tyrosine kinase inhibitor (TKI), and durvalumab (anti-PD-L1) in patients with advanced colorectal and pancreatic carcinoma with the aim to enhance responses to PD-L1 blockade by eliminating CSF-1-dependent suppressive TAM. Forty-seven patients were enrolled. No unexpected toxicities were observed, one (2%) high microsatellite instability CRC patient had a partial response, and seven (15%) patients experienced stable disease as their best response. Increase of CSF-1 concentrations and decrease of CD14lowCD16high monocytes in peripheral blood mononuclear cells (PBMCs) confirmed CSF-1R engagement. Treatment decreased blood dendritic cell (DC) subsets and impaired IFN-λ/IL-29 production by type 1 conventional DCs in ex vivo TLR3-stimulated PBMCs. Pexidartinib also targets c-KIT and FLT3, both key growth factor receptors of DC development and maturation. In patients, FLT3-L concentrations increased with pexidartinib treatment, and AKT phosphorylation induced by FLT3-L ex vivo stimulation was abrogated by pexidartinib in human blood DC subsets. In addition, pexidartinib impaired the FLT3-L- but not GM-CSF-dependent generation of DC subsets from murine bone marrow (BM) progenitors in vitro and decreased DC frequency in BM and tumor-draining lymph node in vivo. Our results demonstrate that pexidartinib, through the inhibition of FLT3 signaling, has a deleterious effect on DC differentiation, which may explain the limited antitumor clinical activity observed in this study. This work suggests that inhibition of FLT3 should be considered when combining TKIs with immune checkpoint inhibitors.


Assuntos
Aminopiridinas , Anticorpos Monoclonais , Antígeno B7-H1 , Neoplasias Pancreáticas , Pirróis , Humanos , Animais , Camundongos , Fator Estimulador de Colônias de Macrófagos , Leucócitos Mononucleares , Receptores Proteína Tirosina Quinases , Tirosina Quinase 3 Semelhante a fms
10.
Int J Cancer ; 133(3): 771-8, 2013 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-23389942

RESUMO

We previously reported that plasmacytoid dendritic cells (pDCs) infiltrating breast tumors are impaired for their interferon-α (IFN-α) production, resulting in local regulatory T cells amplification. We designed our study to decipher molecular mechanisms of such functional defect of tumor-associated pDC (TApDC) in breast cancer. We demonstrate that besides IFN-α, the production by Toll-like receptor (TLR)-activated healthy pDC of IFN-ß and TNF-α but not IP-10/CXCL10 nor MIP1-α/CCL3 is impaired by the breast tumor environment. Importantly, we identified TGF-ß and TNF-α as major soluble factors involved in TApDC functional alteration. Indeed, recombinant TGF-ß1 and TNF-α synergistically blocked IFN-α production of TLR-activated pDC, and neutralization of TGF-ß and TNF-α in tumor-derived supernatants restored pDCs' IFN-α production. The involvment of tumor-derived TGF-ß was further confirmed in situ by the detection of phosphorylated Smad2 in the nuclei of TApDC in breast tumor tissues. Mechanisms of type I IFN inhibition did not involve TLR downregulation but the inhibition of IRF-7 expression and nuclear translocation in pDC after their exposure to tumor-derived supernatants or recombinant TGF-ß1 and TNF-α. Our findings indicate that targeting TApDC to restore their IFN-α production might be an achievable strategy to induce antitumor immunity in breast cancer by combining TLR7/9-based immunotherapy with TGF-ß and TNF-α antagonists.


Assuntos
Neoplasias da Mama/metabolismo , Células Dendríticas/metabolismo , Interferon-alfa/biossíntese , Fator de Crescimento Transformador beta/farmacologia , Fator de Necrose Tumoral alfa/farmacologia , Quimiocina CCL3/biossíntese , Quimiocina CXCL10/biossíntese , Feminino , Humanos , Fator Regulador 7 de Interferon/biossíntese , Interferon beta/biossíntese , Fosforilação , Transporte Proteico , Proteínas Recombinantes/farmacologia , Proteína Smad2/metabolismo , Receptor 7 Toll-Like/metabolismo , Receptor Toll-Like 9/metabolismo , Fator de Crescimento Transformador beta/metabolismo , Fator de Necrose Tumoral alfa/biossíntese , Fator de Necrose Tumoral alfa/metabolismo
11.
Blood ; 118(19): 5130-40, 2011 Nov 10.
Artigo em Inglês | MEDLINE | ID: mdl-21937703

RESUMO

Absent in peripheral tissues during homeostasis, human plasmacytoid dendritic cells (pDCs) are described in inflamed skin or mucosa. Here, we report that, unlike blood pDCs, a subset of tonsil pDCs express functional CCR6 and CCR10, and their respective ligands CCL20 and CCL27are detected in inflamed epithelia contacting blood dendritic cell antigen 2(+) pDCs. Moreover, pDCs are recruited to imiquimod-treated skin tumors in WT but not CCR6-deficient mice, and competitive adoptive transfers reveal that CCR6-deficient pDCs are impaired in homing to inflamed skin tumors after intravenous transfer. On IL-3 culture, CCR6 and CCR10 expression is induced on human blood pDCs that become responsive to CCL20 and CCL27/CCL28, respectively. Interestingly, unlike myeloid DC, blood pDCs initially up-regulate CCR7 expression and CCL19 responsiveness on IL-3 ± CpG-B and then acquire functional CCR6 and CCR10. Finally, IL-3-differentiated CCR6(+) CCR10(+) pDCs secrete high levels of IFN-α in response to virus. Overall, we propose an unexpected pDCs migratory model that may best apply for mucosal-associated lymphoid tissues. After CCR7-mediated extravasation into lymphoid tissues draining inflamed epithelia, blood pDCs may be instructed to up-regulate CCR6 and/or CCR10 allowing their homing into inflamed epithelia (in mucosae or skin). At this site, pDCs can then produce IFN-α contributing to pathogen clearance and/or local inflammation.


Assuntos
Células Dendríticas/imunologia , Inflamação/imunologia , Receptores CCR10/metabolismo , Receptores CCR6/metabolismo , Transferência Adotiva , Animais , Diferenciação Celular/imunologia , Movimento Celular/imunologia , Quimiocina CCL19/farmacologia , Quimiocina CCL20/farmacologia , Células Dendríticas/patologia , Epitélio/imunologia , Epitélio/patologia , Feminino , Humanos , Inflamação/patologia , Interferon-alfa/biossíntese , Interleucina-3/farmacologia , Ligantes , Tecido Linfoide/imunologia , Tecido Linfoide/patologia , Melanoma Experimental/imunologia , Melanoma Experimental/patologia , Glicoproteínas de Membrana/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Modelos Imunológicos , Tonsila Palatina/citologia , Tonsila Palatina/imunologia , Receptores CCR6/deficiência , Receptores CCR6/genética , Receptor 7 Toll-Like/metabolismo
12.
Trends Immunol ; 31(10): 391-7, 2010 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-20832362

RESUMO

HIV-1, hepatitis B virus, hepatitis C virus, and human papillomavirus type 16 cause persistent infections that frequently precede cancer development. Virions of these viruses are weak inducers of interferon-α and impair Toll-like receptor (TLR)9 function. Loss of TLR9 responsiveness also occurs in tumors without viral etiology such as breast, ovary, and head and neck carcinomas. Recent reports have suggested that viruses and components of the tumor microenviroment interact with regulatory receptors on plasmacytoid dendritic cells (pDCs) to impair TLR7 and TLR9 signaling, and to downregulate TLR9 gene expression. The limited responsiveness of pDCs might contribute to reduced innate immune responses during chronic viral infections and oncogenesis, and represent a target for new therapeutic approaches based on TLR agonists.


Assuntos
Hepatite Crônica/complicações , Neoplasias/imunologia , Neoplasias/virologia , Transdução de Sinais , Receptor 7 Toll-Like/imunologia , Receptor Toll-Like 9/imunologia , Animais , Células Dendríticas/imunologia , Células Dendríticas/metabolismo , Humanos , Neoplasias/tratamento farmacológico , Neoplasias/etiologia , Receptor 7 Toll-Like/agonistas , Receptor 7 Toll-Like/metabolismo , Receptor Toll-Like 9/agonistas , Receptor Toll-Like 9/metabolismo
13.
Methods Mol Biol ; 2618: 17-35, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-36905506

RESUMO

Dendritic cells (DCs) play a key role in the antitumor immunity, as they are at the interface of innate and adaptive immunity. This important task can only be performed thanks to the broad range of mechanisms that DCs can perform to activate other immune cells. As DCs are well known for their outstanding capacity to prime and activate T cells through antigen presentation, DCs were intensively investigated during the past decades. Numerous studies have identified new DC subsets, leading to a large variety of subsets commonly separated into cDC1, cDC2, pDCs, mature DCs, Langerhans cells, monocyte-derived DCs, Axl-DCs, and several other subsets. Here, we review the specific phenotypes, functions, and localization within the tumor microenvironment (TME) of human DC subsets thanks to flow cytometry and immunofluorescence but also with the help of high-output technologies such as single-cell RNA sequencing and imaging mass cytometry (IMC).


Assuntos
Células Dendríticas , Microambiente Tumoral , Humanos , Imunidade Adaptativa , Apresentação de Antígeno , Fenótipo
14.
Methods Mol Biol ; 2618: 187-197, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-36905518

RESUMO

Dendritic cells (DCs) are professional antigen-presenting cells (APCs) that have the ability to orchestrate adaptive and innate immune responses by antigen phagocytosis and T cell activation across different inflammatory settings such as tumor development. As specific DC identity and how these cells interact with their neighbors is still not fully understood, it remains a challenge to unravel DC heterogeneity, particularly in human cancers. In this chapter, we describe a protocol to isolate and characterize tumor-infiltrating DCs.


Assuntos
Células Dendríticas , Neoplasias , Humanos , Camundongos , Animais , Ativação Linfocitária , Neoplasias/patologia , Fagocitose
15.
Nat Cell Biol ; 25(12): 1736-1745, 2023 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-38036749

RESUMO

Myeloid cell infiltration of solid tumours generally associates with poor patient prognosis and disease severity1-13. Therefore, understanding the regulation of myeloid cell differentiation during cancer is crucial to counteract their pro-tumourigenic role. Bone marrow (BM) haematopoiesis is a tightly regulated process for the production of all immune cells in accordance to tissue needs14. Myeloid cells differentiate during haematopoiesis from multipotent haematopoietic stem and progenitor cells (HSPCs)15-17. HSPCs can sense inflammatory signals from the periphery during infections18-21 or inflammatory disorders22-27. In these settings, HSPC expansion is associated with increased myeloid differentiation28,29. During carcinogenesis, the elevation of haematopoietic growth factors supports the expansion and differentiation of committed myeloid progenitors5,30. However, it is unclear whether cancer-related inflammation also triggers demand-adapted haematopoiesis at the level of multipotent HSPCs. In the BM, HSPCs reside within the haematopoietic niche which delivers HSC maintenance and differentiation cues31-35. Mesenchymal stem cells (MSCs) are a major cellular component of the BM niche and contribute to HSC homeostasis36-41. Modifications of MSCs in systemic disorders have been associated with HSC differentiation towards myeloid cells22,42. It is unknown if MSCs are regulated in the context of solid tumours and if their myeloid supportive activity is impacted by cancer-induced systemic changes. Here, using unbiased transcriptomic analysis and in situ imaging of HSCs and the BM niche during breast cancer, we show that both HSCs and MSCs are transcriptionally and spatially modified. We demonstrate that breast tumour can distantly remodel the cellular cross-talks in the BM niche leading to increased myelopoiesis.


Assuntos
Medula Óssea , Neoplasias da Mama , Humanos , Feminino , Neoplasias da Mama/patologia , Células-Tronco Hematopoéticas/metabolismo , Células-Tronco Multipotentes/metabolismo , Diferenciação Celular , Nicho de Células-Tronco , Células da Medula Óssea
16.
Clin Transl Immunology ; 11(5): e1382, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-35517992

RESUMO

Objectives: Plasmacytoid DCs (pDCs) play a critical yet enigmatic role in antitumor immunity through their pleiotropic immunomodulatory functions. Despite proof of pDC diversity in several physiological or pathological contexts, pDCs have been studied as a whole population so far in cancer. The assessment of individual pDC subsets is needed to fully grasp their involvement in cancer immunity, especially in melanoma where pDC subsets are largely unknown and remain to be uncovered. Methods: We explored for the first time the features of diverse circulating and tumor-infiltrating pDC subsets in melanoma patients using multi-parametric flow cytometry, and assessed their clinical relevance. Based on CD80, PDL1, CD2, LAG3 and Axl markers, we provided an integrated overview of the frequency, basal activation status and functional features of pDC subsets in melanoma patients together with their relationship to clinical outcome. Results: Strikingly, we demonstrated that P3-pDCs (CD80+PDL1-) accumulated within the tumor of melanoma patients and negatively correlated with clinical outcomes. The basal activation status, diversification towards P1-/P2-/P3-pDCs and functionality of several pDC subsets upon TLR7/TLR9 triggering were perturbed in melanoma patients, and were differentially linked to clinical outcome. Conclusion: Our study shed light for the first time on the phenotypic and functional heterogeneity of pDCs in the blood and tumor of melanoma patients and their potential involvement in shaping clinical outcomes. Such novelty brightens our understanding of pDC complexity, and prompts the further deciphering of pDCs' features to better apprehend and exploit these potent immune players. It highlights the importance of considering pDC diversity when developing pDC-based therapeutic strategies to ensure optimal clinical success.

17.
Front Immunol ; 13: 1040600, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-36353633

RESUMO

Subversion of immunity by tumors is a crucial step for their development. Dendritic cells (DCs) are strategic immune cells that orchestrate anti-tumor immune responses but display altered functions in cancer. The bases for such DCs' hijacking are not fully understood. Tumor cells harbor unusual glycosylation patterns of surface glycoproteins and glycolipids. DCs express glycan-binding receptors, named C-type lectin receptors (CLR), allowing them to sense changes in glycan signature of their environment, and subsequently trigger a response. Recognition of tumor glycans by CLRs is crucial for DCs to shape antitumor immunity, and decisive in the orientation of the response. Yet the status of the CLR machinery on DCs in cancer, especially melanoma, remained largely unknown. We explored CLR expression patterns on circulating and tumor-infiltrating cDC1s, cDC2s, and pDCs of melanoma patients, assessed their clinical relevance, and further depicted the correlations between CLR expression profiles and DCs' features. For the first time, we highlighted that the CLR repertoire of circulating and tumor-infiltrating cDC1s, cDC2s, and pDCs was strongly perturbed in melanoma patients, with modulation of DCIR, CLEC-12α and NKp44 on circulating DCs, and perturbation of Dectin-1, CD206, DEC205, DC-SIGN and CLEC-9α on tumor-infiltrating DCs. Furthermore, melanoma tumor cells directly altered CLR expression profiles of healthy DC subsets, and this was associated with specific glycan patterns (Man, Fuc, GlcNAc) that may interact with DCs through CLR molecules. Notably, specific CLR expression profiles on DC subsets correlated with unique DCs' activation status and functionality and were associated with clinical outcome of melanoma patients. Higher proportions of DCIR-, DEC205-, CLEC-12α-expressing cDCs were linked with a better survival, whereas elevated proportions of CD206-, Dectin1-expressing cDCs and NKp44-expressing pDCs were associated with a poor outcome. Thus, melanoma tumor may shape DCs' features by exploiting the plasticity of the CLR machinery. Our study revealed that melanoma manipulates CLR pathways to hijack DC subsets and escape from immune control. It further paved the way to exploit glycan-lectin interactions for the design of innovative therapeutic strategies, which exploit DCs' potentialities while avoiding hijacking by tumor, to properly reshape anti-tumor immunity by manipulating the CLR machinery.


Assuntos
Células Dendríticas , Melanoma , Masculino , Humanos , Lectinas Tipo C/metabolismo , Glicoproteínas de Membrana/metabolismo , Polissacarídeos , Melanoma/metabolismo
18.
J Immunother Cancer ; 10(3)2022 03.
Artigo em Inglês | MEDLINE | ID: mdl-35288462

RESUMO

BACKGROUND: The efficacy of immunotherapies in metastatic melanoma depends on a robust T cell infiltration. Oncogenic alterations of tumor cells have been associated to T cell exclusion. Identifying novel cancer cell-intrinsic non-genetic mechanisms of immune escape, the targeting of which would reinstate T cell recruitment, would allow to restore the response to anti-programmed cell death protein 1 (PD-1) antibody therapy. The epithelial-to-mesenchymal transition (EMT)-inducing transcription factor ZEB1 is a major regulator of melanoma cell plasticity, driving resistance to mitogen-activated protein kinase (MAPK) targeted therapies. We thus wondered whether ZEB1 signaling in melanoma cells may promote immune evasion and resistance to immunotherapy. METHODS: We evaluated the putative correlation between ZEB1 expression in melanoma cells and the composition of the immune infiltrate in a cohort of 60 human melanoma samples by combining transcriptomic (RNA-sequencing) and seven-color spatial multi-immunofluorescence analyses. Algorithm-based spatial reconstitution of tumors allowed the quantification of CD8+, CD4+ T cells number and their activation state (PD-1, Ki67). ZEB1 gain-of-function or loss-of-function approaches were then implemented in syngeneic melanoma mouse models, followed by monitoring of tumor growth, quantification of immune cell populations frequency and function by flow cytometry, cytokines secretion by multiplex analyses. Chromatin-immunoprecipitation was used to demonstrate the direct binding of this transcription factor on the promoters of cytokine-encoding genes. Finally, the sensitivity to anti-PD-1 antibody therapy upon ZEB1 gain-of-function or loss-of-function was evaluated. RESULTS: Combined spatial and transcriptomic analyses of the immune infiltrates in human melanoma samples demonstrated that ZEB1 expression in melanoma cells is associated with decreased CD8+ T cell infiltration, independently of ß-catenin pathway activation. ZEB1 ectopic expression in melanoma cells impairs CD8+ T cell recruitment in syngeneic mouse models, resulting in tumor immune evasion and resistance to immune checkpoint blockade. Mechanistically, we demonstrate that ZEB1 directly represses the secretion of T cell-attracting chemokines, including CXCL10. Finally, Zeb1 knock-out, by promoting CD8+ T cell infiltration, synergizes with anti-PD-1 antibody therapy in promoting tumor regression. CONCLUSIONS: We identify the ZEB1 transcription factor as a key determinant of melanoma immune escape, highlighting a previously unknown therapeutic target to increase efficacy of immunotherapy in melanoma. TRIAL REGISTRATION NUMBER: NCT02828202.


Assuntos
Melanoma , Homeobox 1 de Ligação a E-box em Dedo de Zinco , Animais , Transição Epitelial-Mesenquimal/fisiologia , Humanos , Imunoterapia , Melanoma/tratamento farmacológico , Melanoma/genética , Melanoma/patologia , Camundongos , Oncogenes , Homeobox 1 de Ligação a E-box em Dedo de Zinco/genética
19.
Cancers (Basel) ; 14(5)2022 Feb 25.
Artigo em Inglês | MEDLINE | ID: mdl-35267497

RESUMO

BACKGROUND: Following disappointing results with PD-1/PD-L1 inhibitors in ovarian cancer, it is essential to explore other immune targets. The aim of this study is to describe the tumor immune microenvironment (TME) according to genomic instability in high grade serous ovarian carcinoma (HGSOC) patients receiving primary debulking surgery followed by carboplatin-paclitaxel chemotherapy +/- nintedanib. METHODS: 103 HGSOC patients' tumor samples from phase III AGO-OVAR-12 were analyzed. A comprehensive analysis of the TME was performed by immunohistochemistry on tissue microarray. Comparative genomic hybridization was carried out to evaluate genomic instability signatures through homologous recombination deficiency (HRD) score, genomic index, and somatic copy number alterations. The relationship between genomic instability and TME was explored. RESULTS: Patients with high intratumoral CD3+ T lymphocytes had longer progression-free survival (32 vs. 19.6 months, p = 0.009) and overall survival (OS) (median not reached). High HLA-E expression on tumor cells was associated with a longer OS (median OS not reached vs. 52.9 months, p = 0.002). HRD profile was associated with high HLA-E expression on tumor cells and an improved OS. In the multivariate analysis, residual tumor, intratumoral CD3, and HLA-E on tumor cells were more predictive than other parameters. CONCLUSIONS: Our results suggest HLA-E/CD94-NKG2A/2C is a potential immune target particularly in the HRD positive ovarian carcinoma subgroup.

20.
BMC Cancer ; 11: 213, 2011 May 30.
Artigo em Inglês | MEDLINE | ID: mdl-21624121

RESUMO

BACKGROUND: Chemokines and chemokine receptors are major actors of leukocytes trafficking and some have been shown to play an important role in cancer metastasis. Chemokines CCL19, CCL20 and CCL21 and their receptors CCR6 and CCR7, were assessed as potential biomarkers of metastatic dissemination in primary breast cancer. METHODS: Biomarker expression levels were evaluated using immunohistochemistry on paraffin-embedded tissue sections of breast cancer (n = 207). RESULTS: CCR6 was expressed by tumor cells in 35% of cases. CCR7 was expressed by spindle shaped stromal cells in 43% of cases but not by tumor cells in this series. CCL19 was the only chemokine found expressed in a significant number of breast cancers and was expressed by both tumor cells and dendritic cells (DC). CCR6, CCL19 and CCR7 expression correlated with histologic features of aggressive disease. CCR6 expression was associated with shorter relapse-free survival (RFS) in univariate and but not in multivariate analysis (p = 0.0316 and 0.055 respectively), and was not associated with shorter overall survival (OS). Expression of CCR7 was not significantly associated with shorter RFS or OS. The presence of CCL19-expressing DC was associated with shorter RFS in univariate and multivariate analysis (p = 0.042 and 0.020 respectively) but not with shorter OS. CONCLUSION: These results suggest a contribution of CCR6 expression on tumor cells and CCL19-expressing DC in breast cancer dissemination. In our series, unlike what was previously published, CCR7 was exclusively expressed on stromal cells and was not associated with survival.


Assuntos
Neoplasias da Mama/diagnóstico , Neoplasias da Mama/fisiopatologia , Regulação Neoplásica da Expressão Gênica , Ligantes , Receptores CCR6/metabolismo , Receptores CCR7/metabolismo , Adulto , Idoso , Idoso de 80 Anos ou mais , Neoplasias da Mama/mortalidade , Quimiocinas C/metabolismo , Feminino , Humanos , Linfonodos/metabolismo , Linfonodos/patologia , Metástase Linfática/patologia , Pessoa de Meia-Idade , Prognóstico , Células Estromais/metabolismo , Células Estromais/patologia , Análise de Sobrevida
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