RESUMO
Immune checkpoint blockades (ICBs) have revolutionized cancer therapy through unleashing anti-tumor adaptive immunity. Despite that, they are usually effective only in a small subset of patients and relapse can occur in patients who initially respond to the treatment. Recent breakthroughs in this field have identified innate immune checkpoints harnessed by cancer cells to escape immunosurveillance from innate immunity. MHC1 appears to be such a molecule expressed on cancer cells which can transmit a negative signal to innate immune cells through interaction with leukocyte immunoglobulin like receptor B1 (LILRB1). The review aims to summarize the current understanding of MHC1/LILRB1 axis on mediating cancer immune evasion with an emphasis on the therapeutic potential to block this axis for cancer therapy. Nevertheless, one should note that this field is still in its infancy and more studies are warranted to further verify the effectiveness and safety in clinical as well as the potential to combine with existing immune checkpoints.
Assuntos
Imunidade Inata , Receptor B1 de Leucócitos Semelhante a Imunoglobulina , Neoplasias , Humanos , Receptor B1 de Leucócitos Semelhante a Imunoglobulina/metabolismo , Receptor B1 de Leucócitos Semelhante a Imunoglobulina/imunologia , Neoplasias/imunologia , Neoplasias/terapia , Animais , Inibidores de Checkpoint Imunológico/uso terapêutico , Evasão Tumoral , Antígenos de Histocompatibilidade Classe I/imunologia , Antígenos de Histocompatibilidade Classe I/metabolismo , Imunoterapia/métodos , Transdução de Sinais , Antígenos CDRESUMO
Background: Placental malaria (PM) is associated with a higher susceptibility of infants to Plasmodium falciparum (Pf) malaria. A hypothesis of immune tolerance has been suggested but no clear explanation has been provided so far. Our goal was to investigate the involvement of inhibitory receptors LILRB1 and LILRB2, known to drive immune evasion upon ligation with pathogen and/or host ligands, in PM-induced immune tolerance. Method: Infants of women with or without PM were enrolled in Allada, southern Benin, and followed-up for 24 months. Antibodies with specificity for five blood stage parasite antigens were quantified by ELISA, and the frequency of immune cell subsets was quantified by flow cytometry. LILRB1 or LILRB2 expression was assessed on cells collected at 18 and 24 months of age. Findings: Infants born to women with PM had a higher risk of developing symptomatic malaria than those born to women without PM (IRR=1.53, p=0.040), and such infants displayed a lower frequency of non-classical monocytes (OR=0.74, p=0.01) that overexpressed LILRB2 (OR=1.36, p=0.002). Moreover, infants born to women with PM had lower levels of cytophilic IgG and higher levels of IL-10 during active infection. Interpretation: Modulation of IgG and IL-10 levels could impair monocyte functions (opsonisation/phagocytosis) in infants born to women with PM, possibly contributing to their higher susceptibility to malaria. The long-lasting effect of PM on infants' monocytes was notable, raising questions about the capacity of ligands such as Rifins or HLA-I molecules to bind to LILRB1 and LILRB2 and to modulate immune responses, and about the reprogramming of neonatal monocytes/macrophages.
Assuntos
Antimaláricos , Malária Falciparum , Glicoproteínas de Membrana , Placenta , Receptores Imunológicos , Anticorpos Antiprotozoários , Feminino , Humanos , Imunoglobulina G/sangue , Lactente , Recém-Nascido , Interleucina-10 , Receptor B1 de Leucócitos Semelhante a Imunoglobulina/genética , Receptor B1 de Leucócitos Semelhante a Imunoglobulina/imunologia , Glicoproteínas de Membrana/genética , Glicoproteínas de Membrana/imunologia , Monócitos/metabolismo , Placenta/parasitologia , Plasmodium falciparum , Gravidez , Receptores Imunológicos/genética , Receptores Imunológicos/imunologiaRESUMO
Natural killer (NK) cells play a crucial role in the control of human viral infections but their activity is significantly impaired in patients infected with chronic hepatitis B (CHB). The mechanism that contributes to NK cell dysfunction in CHB needs further elucidation. In this study, we analyzed the expression and function of the novel inhibitory receptor immunoglobulin-like transcript-2 (ILT2) on NK cells from 131 CHB patients and 36 healthy controls. We observed that ILT2 expression on circulating CD56dimCD16+NK cells was increased in immune-tolerant, immune-active and HBeAg-negative hepatitis patients compared with inactive carriers and controls. The frequency of ILT2+CD56dimNK cells was positively correlated with serum viral load in immune-tolerant patients. The percentage of ILT2+CD56dimNK cells decreased along with HBV load in CHB patients who received antiviral therapy. Functional analysis showed that ILT2+CD56dimNK cells in CHB patients had significantly reduced degranulation and IFN-γ production. Upregulation of ILT2 was associated with high levels of apoptosis in CD56dimCD16+NK cells from CHB patients. ILT2 blockade was shown to increase the cytotoxicity and IFN-γ production of CD56dimNK cells in some CHB patients. Finally, ILT2 was found to be moderately upregulated by TGF-ß1, which was increased in immune-tolerant, immune-active and HBeAg-negative hepatitis patients. Our results show that chronic HBV infection increases the levels of the inhibitory receptor ILT2 on CD56dimNK cells and inhibits their functions, providing a new mechanism of NK-cell disability in CHB patients.
Assuntos
Antígenos CD/imunologia , Hepatite B Crônica , Receptor B1 de Leucócitos Semelhante a Imunoglobulina/imunologia , Antígeno CD56/imunologia , Proteínas Ligadas por GPI/imunologia , Antígenos E da Hepatite B , Vírus da Hepatite B , Humanos , Interferon gama/metabolismo , Células Matadoras Naturais , Receptores de IgG/imunologiaRESUMO
Recombination of antibody genes in B cells can involve distant genomic loci and contribute a foreign antigen-binding element to form hybrid antibodies with broad reactivity for Plasmodium falciparum. So far, antibodies containing the extracellular domain of the LAIR1 and LILRB1 receptors represent unique examples of cross-chromosomal antibody diversification. Here, we devise a technique to profile non-VDJ elements from distant genes in antibody transcripts. Independent of the preexposure of donors to malaria parasites, non-VDJ inserts were detected in 80% of individuals at frequencies of 1 in 104 to 105 B cells. We detected insertions in heavy, but not in light chain or T cell receptor transcripts. We classify the insertions into four types depending on the insert origin and destination: 1) mitochondrial and 2) nuclear DNA inserts integrated at VDJ junctions; 3) inserts originating from telomere proximal genes; and 4) fragile sites incorporated between J-to-constant junctions. The latter class of inserts was exclusively found in memory and in in vitro activated B cells, while all other classes were already detected in naïve B cells. More than 10% of inserts preserved the reading frame, including transcripts with signs of antigen-driven affinity maturation. Collectively, our study unravels a mechanism of antibody diversification that is layered on the classical V(D)J and switch recombination.
Assuntos
Diversidade de Anticorpos , Linfócitos B , Genes de Imunoglobulinas , Anticorpos Antiprotozoários/genética , Antígenos CD/imunologia , Linfócitos B/imunologia , Genômica , Humanos , Cadeias Leves de Imunoglobulina/genética , Receptor B1 de Leucócitos Semelhante a Imunoglobulina/imunologia , Mutagênese Insercional , Plasmodium falciparum , Receptores de Antígenos de Linfócitos T/genética , Receptores Imunológicos/imunologiaRESUMO
HLA-G: ILT2 has recently been positioned as a major immune checkpoint in urologic cancers. In clear cell renal cell carcinoma (ccRCC), tumor-infiltrating CD8+ T cells expressing ILT2 are a highly cytotoxic cell population, distinct from PD1+ T cells, and whose function is inhibited by HLA-G+ targets. Here we report that ILT2 receptor can also be expressed by CD4+ T cells in urologic cancer patients. In the course of deciphering the role of these ILT2+CD4+ T cells, we found a statistical association between the tumor context and these T cells, and a positive correlation between the levels of peripheral and intra-tumoral CD4+ILT2+ T cells. Phenotypic analyses revealed that CD4+ILT2+ T cells express memory T cell (CD27-CD28-CD57+) and cytotoxicity (Tbet+Perforin+KLRG1+NKp80+GPR56+) markers, consistent with a CD4+CTL phenotype. Functional assays showed that ccRCC-infiltrating CD4+ILT2+ T cells indeed have high cytolytic properties and therefore function as proper CD4+CTLs, but are selectively inhibited by HLA-G+ targets. Clinical relevance was provided by immunohistochemical analyses on ccRCC tumor lesions with HLA-G+ HLA class II+ tumor cells next to CD4+ T cell infiltrates. Our findings provide evidence supporting that ILT2+ T cells constitute a reservoir of intratumor cytotoxic T cells that is not targeted by the current checkpoint inhibitors, but could be by anti-HLA-G/anti-ILT2 antibodies as novel immunotherapy in HLA-G+ tumors.
Assuntos
Antígenos CD/imunologia , Linfócitos T CD4-Positivos/imunologia , Carcinoma de Células Renais/imunologia , Antígenos HLA-G/imunologia , Neoplasias Renais/imunologia , Receptor B1 de Leucócitos Semelhante a Imunoglobulina/imunologia , Linfócitos T Citotóxicos/imunologia , Idoso , Antineoplásicos/farmacologia , Linfócitos T CD8-Positivos/imunologia , Carcinoma de Células Renais/tratamento farmacológico , Feminino , Humanos , Neoplasias Renais/tratamento farmacológico , Masculino , Células T de Memória/imunologia , Pessoa de Meia-IdadeRESUMO
Some Plasmodium falciparum repetitive interspersed families of polypeptides (RIFINs)-variant surface antigens that are expressed on infected erythrocytes1-bind to the inhibitory receptor LAIR1, and insertion of DNA that encodes LAIR1 into immunoglobulin genes generates RIFIN-specific antibodies2,3. Here we address the general relevance of this finding by searching for antibodies that incorporate LILRB1, another inhibitory receptor that binds to ß2 microglobulin and RIFINs through their apical domains4,5. By screening plasma from a cohort of donors from Mali, we identified individuals with LILRB1-containing antibodies. B cell clones isolated from three donors showed large DNA insertions in the switch region that encodes non-apical LILRB1 extracellular domain 3 and 4 (D3D4) or D3 alone in the variable-constant (VH-CH1) elbow. Through mass spectrometry and binding assays, we identified a large set of RIFINs that bind to LILRB1 D3. Crystal and cryo-electron microscopy structures of a RIFIN in complex with either LILRB1 D3D4 or a D3D4-containing antibody Fab revealed a mode of RIFIN-LILRB1 D3 interaction that is similar to that of RIFIN-LAIR1. The Fab showed an unconventional triangular architecture with the inserted LILRB1 domains opening up the VH-CH1 elbow without affecting VH-VL or CH1-CL pairing. Collectively, these findings show that RIFINs bind to LILRB1 through D3 and illustrate, with a naturally selected example, the general principle of creating novel antibodies by inserting receptor domains into the VH-CH1 elbow.
Assuntos
Anticorpos/química , Anticorpos/imunologia , Antígenos de Protozoários/química , Antígenos de Protozoários/imunologia , Microscopia Crioeletrônica , Receptor B1 de Leucócitos Semelhante a Imunoglobulina/química , Plasmodium falciparum/química , Plasmodium falciparum/imunologia , Adolescente , Adulto , Sequência de Aminoácidos , Anticorpos/ultraestrutura , Especificidade de Anticorpos , Antígenos de Protozoários/ultraestrutura , Sítios de Ligação de Anticorpos , Criança , Pré-Escolar , Estudos de Coortes , Humanos , Lactente , Receptor B1 de Leucócitos Semelhante a Imunoglobulina/imunologia , Mali , Modelos Moleculares , Plasmodium falciparum/genética , Plasmodium falciparum/ultraestrutura , Domínios Proteicos , Adulto JovemRESUMO
Human leukocyte antigen G (HLA-G) mediates maternal-fetal immune tolerance. It is also considered an immune checkpoint in cancer since it may mediate immune evasion and thus promote tumor growth. HLA-G is, therefore, a potential target for immunotherapy. However, existing monoclonal antibodies directed against HLA-G lack sufficient specificity and are not suitable for immune checkpoint inhibition in a clinical setting. For this reason, it is essential that alternative approaches are explored to block the interaction between HLA-G and its receptors. In this review, we discuss the structure and peptide presentation of HLA-G, and its interaction with the receptors Ig-like transcript (ILT) 2, ILT4, and Killer cell immunoglobulin-like receptor 2DL4 (KIR2DL4). Based on our findings, we propose three alternative strategies to block the interaction between HLA-G and its receptors in cancer immunotherapy: (1) prevention of HLA-G dimerization, (2) targeting the peptide-binding groove of HLA-G, and (3) targeting the HLA-G receptors. These strategies should be an important focus of future studies that aim to develop immune checkpoint inhibitors to block the interaction between HLA-G and its receptors for the treatment of cancer.
Assuntos
Antígenos CD/imunologia , Antígenos HLA-G/imunologia , Imunoterapia , Receptor B1 de Leucócitos Semelhante a Imunoglobulina/imunologia , Glicoproteínas de Membrana/metabolismo , Proteínas de Neoplasias/imunologia , Neoplasias , Receptores Imunológicos/metabolismo , Receptores KIR2DL4/imunologia , Humanos , Neoplasias/imunologia , Neoplasias/patologia , Neoplasias/terapiaRESUMO
The Plasmodium species that cause malaria are obligate intracellular parasites, and disease symptoms occur when these parasites replicate in human blood. Despite the risk of immune detection, the parasite delivers proteins that bind to host receptors on the cell surfaces of infected erythrocytes. In the causative parasite of the most deadly form of malaria in humans, Plasmodium falciparum, RIFINs form the largest family of surface proteins displayed by erythrocytes1. Some RIFINs can bind to inhibitory immune receptors, and these RIFINs act as targets for unusual antibodies that contain a LAIR1 ectodomain2-4 or as ligands for LILRB15. RIFINs stimulate the activation of and signalling by LILRB15, which could potentially lead to the dampening of human immune responses. Here, to understand how RIFINs activate LILRB1-mediated signalling, we determine the structure of a RIFIN bound to LILRB1. We show that this RIFIN mimics the natural activating ligand of LILRB1, MHC class I, in its LILRB1-binding mode. A single mutation in the RIFIN disrupts the complex, blocks LILRB1 binding of all tested RIFINs and abolishes signalling in a reporter assay. In a supported lipid bilayer system, which mimics the activation of natural killer (NK) cells by antibody-dependent cell-mediated cytotoxicity, both RIFIN and MHC are recruited to the immunological synapse of NK cells and reduce the activation of NK cells, as measured by the mobilization of perforin. Therefore, LILRB1-binding RIFINs mimic the binding mode of the natural ligand of LILRB1 and suppress the function of NK cells.
Assuntos
Receptor B1 de Leucócitos Semelhante a Imunoglobulina/química , Receptor B1 de Leucócitos Semelhante a Imunoglobulina/imunologia , Malária Falciparum/imunologia , Proteínas de Membrana/química , Proteínas de Membrana/imunologia , Proteínas de Protozoários/química , Proteínas de Protozoários/imunologia , Sequência de Aminoácidos , Animais , Citotoxicidade Celular Dependente de Anticorpos/imunologia , Sítios de Ligação/imunologia , Antígenos de Histocompatibilidade Classe I/imunologia , Humanos , Células Matadoras Naturais/citologia , Células Matadoras Naturais/imunologia , Receptor B1 de Leucócitos Semelhante a Imunoglobulina/metabolismo , Ligantes , Bicamadas Lipídicas , Ativação Linfocitária , Malária Falciparum/parasitologia , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Modelos Moleculares , Mimetismo Molecular/imunologia , Mutação , Perforina/metabolismo , Proteínas de Protozoários/genética , Proteínas de Protozoários/metabolismo , Transdução de SinaisRESUMO
Clear cell renal cell carcinoma (ccRCC) constitutes the most common renal cell carcinoma subtype and has long been recognized as an immunogenic cancer. As such, significant attention has been directed toward optimizing immune-checkpoints (IC)-based therapies. Despite proven benefits, a substantial number of patients remain unresponsive to treatment, suggesting that yet unreported, immunosuppressive mechanisms coexist within tumors and their microenvironment. Here, we comprehensively analyzed and ranked forty-four immune-checkpoints expressed in ccRCC on the basis of in-depth analysis of RNAseq data collected fromâ¯the TCGA database and advanced statistical methods designed to obtain the group of checkpoints that best discriminates tumor from healthy tissues. Immunohistochemistry and flow cytometry confirmed and enlarged the bioinformatics results. In particular, by using the recursive feature elimination method, we show that HLA-G, B7H3, PDL-1 and ILT2 are the most relevant genes that characterize ccRCC. Notably, ILT2 expression was detected for the first time on tumor cells. The levels of other ligand-receptor pairs such as CD70:CD27; 4-1BB:4-1BBL; CD40:CD40L; CD86:CTLA4; MHC-II:Lag3; CD200:CD200R; CD244:CD48 were also found highly expressed in tumors compared to adjacent non-tumor tissues. Collectively, our approach provides a comprehensible classification of forty-four IC expressed in ccRCC, some of which were never reported before to be co-expressed in ccRCC. In addition, the algorithms used allowed identifying the most relevant group that best discriminates tumor from healthy tissues. The data can potentially assist on the choice of valuable immune-therapy targets which hold potential for the development of more effective anti-tumor treatments.
Assuntos
Antígenos CD/imunologia , Biomarcadores Tumorais/imunologia , Carcinoma de Células Renais/imunologia , Antígenos HLA-G/imunologia , Neoplasias Renais/imunologia , Receptor B1 de Leucócitos Semelhante a Imunoglobulina/imunologia , Glicoproteínas de Membrana/imunologia , Receptores Imunológicos/imunologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Carcinoma de Células Renais/patologia , Estudos de Casos e Controles , Feminino , Seguimentos , Perfilação da Expressão Gênica , Humanos , Neoplasias Renais/patologia , Masculino , Pessoa de Meia-Idade , Prognóstico , Estudos RetrospectivosRESUMO
Invariant Natural Killer T (iNKT) cells are a small and distinct population of T cells crucial in immunomodulation. After activation by alpha-GalactosylCeramide (αGC), an exogenic glycolipid antigen, iNKT cells can rapidly release cytokines to enhance specific anti-tumor activity. Several human clinical trials on iNKT cell-based anti-cancer are ongoing, however results are not as striking as in murine models. Given that iNKT-based immunotherapies are dependent mainly on antigen-presenting cells (APC), a human tolerogenic molecule with no murine homolog, such as Human Leucocyte Antigen G (HLA-G), could contribute to this discrepancy. HLA-G is a well-known immune checkpoint molecule involved in fetal-maternal tolerance and in tumor immune escape. HLA-G exerts its immunomodulatory functions through the interaction with immune inhibitory receptors such as ILT2, differentially expressed on immune cell subsets. We hypothesized that HLA-G might inhibit iNKT function directly or by inducing tolerogenic APC leading to iNKT cell anergy, which could impact the results of current clinical trials. Using an ILT2-transduced murine iNKT cell line and human iNKT cells, we demonstrate that iNKT cells are sensitive to HLA-G, which inhibits their cytokine secretion. Furthermore, human HLA-G+ dendritic cells, called DC-10, failed at inducing iNKT cell activation compared to their autologous HLA-Gâ DCs counterparts. Our data show for the first time that the HLA-G/ILT2 ICP is involved in iNKT cell function modulation.
Assuntos
Antígenos CD/imunologia , Células Dendríticas/imunologia , Antígenos HLA-G/imunologia , Receptor B1 de Leucócitos Semelhante a Imunoglobulina/imunologia , Células T Matadoras Naturais/imunologia , Animais , Células Apresentadoras de Antígenos/imunologia , Células Cultivadas , Citocinas/imunologia , Humanos , Tolerância Imunológica/imunologia , Ativação Linfocitária/imunologia , Camundongos , Camundongos Endogâmicos C57BLRESUMO
Only some cancer patients respond to the immune-checkpoint inhibitors being used in the clinic, and other therapeutic targets are sought. Here, we investigated the HLA-G/ILT2 checkpoint in clear-cell renal-cell carcinoma (ccRCC) patients and focused on tumor-infiltrating CD8+ T lymphocytes (TIL) expressing the HLA-G receptor ILT2. Using transcriptomics and flow cytometry, we characterized both peripheral blood and tumor-infiltrating CD8+ILT2+ T cells from cancer patients as late-differentiated CD27-CD28-CD57+ cytotoxic effectors. We observed a clear dichotomy between CD8+ILT2+ and CD8+PD-1+ TIL subsets. These subsets, which were sometimes present at comparable frequencies in TIL populations, barely overlapped phenotypically and were distinguished by expression of exclusive sets of surface molecules that included checkpoint molecules and activating and inhibitory receptors. CD8+ILT2+ TILs displayed a more mature phenotype and higher expression of cytotoxic molecules. In ex vivo functional experiments with both peripheral blood T cells and TILs, CD8+ILT2+ T cells displayed significantly higher cytotoxicity and IFNγ production than their ILT2- (peripheral blood mononuclear cells, PBMC) and PD-1+ (TILs) counterparts. HLA-G expression by target cells specifically inhibited CD8+ILT2+ T-cell cytotoxicity, but not that of their CD8+ILT2- (PBMC) or CD8+PD-1+ (TIL) counterparts, an effect counteracted by blocking the HLA-G/ILT2 interaction. CD8+ILT2+ TILs may therefore constitute an untapped reservoir of fully differentiated cytotoxic T cells within the tumor microenvironment, independent of the PD1+ TILs targeted by immune therapies, and specifically inhibited by HLA-G. These results emphasize the potential of therapeutically targeting the HLA-G/ILT2 checkpoint in HLA-G+ tumors, either concomitantly with anti-PD-1/PD-L1 or in cases of nonresponsiveness to anti-PD-1/PD-L1.
Assuntos
Antígenos CD/imunologia , Linfócitos T CD8-Positivos/imunologia , Antígenos HLA-G/metabolismo , Neoplasias Renais/imunologia , Receptor B1 de Leucócitos Semelhante a Imunoglobulina/imunologia , Receptor de Morte Celular Programada 1/imunologia , Microambiente Tumoral , Neoplasias da Bexiga Urinária/imunologia , Antineoplásicos Imunológicos/uso terapêutico , Perfilação da Expressão Gênica , Humanos , Neoplasias Renais/tratamento farmacológico , Neoplasias Renais/metabolismo , Receptor B1 de Leucócitos Semelhante a Imunoglobulina/antagonistas & inibidores , Linfócitos do Interstício Tumoral/imunologia , Receptor de Morte Celular Programada 1/antagonistas & inibidores , Linfócitos T Citotóxicos/imunologia , Neoplasias da Bexiga Urinária/tratamento farmacológico , Neoplasias da Bexiga Urinária/metabolismoRESUMO
Immune checkpoint inhibitors are among the newest, cutting-edge methods for the treatment of cancer. Currently, they primarily influence T cell adaptive immunotherapy targeting the PD-1/PD-L1 and CTLA-4/B7 signalling pathways. These inhibitors fight cancer by reactivating the patient's own adaptive immune system, with good results in many cancers. With the discovery of the "Don't Eat Me" molecule, CD47, antibody-based drugs that target the macrophage-related innate immunosuppressive signalling pathway, CD47-SIRPα, have been developed and have achieved stunning results in the laboratory and the clinic, but there remain unexplained instances of tumour immune escape. While investigating the immunological tolerance of cancer to anti-CD47 antibodies, a second "Don't Eat Me" molecule on tumour cells, beta 2 microglobulin (ß2m), a component of MHC class I, was described. Some tumour cells reduce their surface expression of MHC class I to escape T cell recognition. However, other tumour cells highly express ß2m complexed with the MHC class I heavy chain to send a "Don't Eat Me" signal by binding to leucocyte immunoglobulin-like receptor family B, member 1 (LILRB1) on macrophages, leading to a loss of immune surveillance. Investigating the mechanisms underlying this immunosuppressive MHC class I-LILRB1 signalling axis in tumour-associated macrophages will be useful in developing therapies to restore macrophage function and control MHC class I signalling in patient tumours. The goal is to promote adaptive immunity while suppressing the innate immune response to tumours. This work will identify new therapeutic targets for the development of pharmaceutical-based tumour immunotherapy.
Assuntos
Antígenos CD/imunologia , Tolerância Imunológica/imunologia , Receptor B1 de Leucócitos Semelhante a Imunoglobulina/imunologia , Neoplasias/terapia , Evasão Tumoral/imunologia , Microglobulina beta-2/imunologia , Imunidade Adaptativa/imunologia , Antígenos de Histocompatibilidade Classe I/imunologia , Humanos , Imunidade Inata/imunologia , Macrófagos/imunologia , Transdução de Sinais/imunologia , Linfócitos T/imunologiaRESUMO
Elicitation of tumor cell killing by CD8+ T cells is an effective therapeutic approach for cancer. In addition to using immune checkpoint blockade to reinvigorate existing but unresponsive tumor-specific T cells, alternative therapeutic approaches have been developed, including stimulation of polyclonal T cell cytolytic activity against tumors using bispecific T cell engager (BiTE) molecules that simultaneously engage the TCR complex and a tumor-associated Ag. BiTE molecules are efficacious against hematologic tumors and are currently being explored as an immunotherapy for solid tumors. To understand mechanisms regulating BiTE molecule--mediated CD8+ T cell activity against solid tumors, we sought to define human CD8+ T cell populations that efficiently respond to BiTE molecule stimulation and identify factors regulating their cytolytic activity. We find that human CD45RA+CCR7- CD8+ T cells are highly responsive to BiTE molecule stimulation, are enriched in genes associated with cytolytic effector function, and express multiple unique inhibitory receptors, including leukocyte Ig-like receptor B1 (LILRB1). LILRB1 and programmed cell death protein 1 (PD1) were found to be expressed by distinct CD8+ T cell populations, suggesting different roles in regulating the antitumor response. Engaging LILRB1 with its ligand HLA-G on tumor cells significantly inhibited BiTE molecule-induced CD8+ T cell activation. Blockades of LILRB1 and PD1 induced greater CD8+ T cell activation than either treatment alone. Together, our data suggest that LILRB1 functions as a negative regulator of human CD8+ effector T cells and that blocking LILRB1 represents a unique strategy to enhance BiTE molecule therapeutic activity against solid tumors.
Assuntos
Anticorpos Biespecíficos/farmacologia , Antígenos CD/imunologia , Imunoterapia/métodos , Receptor B1 de Leucócitos Semelhante a Imunoglobulina/imunologia , Neoplasias/imunologia , Linfócitos T Citotóxicos/imunologia , Anticorpos Biespecíficos/imunologia , Humanos , Receptor B1 de Leucócitos Semelhante a Imunoglobulina/antagonistas & inibidores , Ativação Linfocitária/efeitos dos fármacos , Ativação Linfocitária/imunologia , Subpopulações de Linfócitos T/imunologia , Células Tumorais CultivadasRESUMO
X-ray crystallographic studies of class I peptide-MHC molecules (pMHC) continue to provide important insights into immune recognition, however their success depends on generation of diffraction-quality crystals, which remains a significant challenge. While protein engineering techniques such as surface-entropy reduction and lysine methylation have proven utility in facilitating and/or improving protein crystallisation, they risk affecting the conformation and biochemistry of the class I MHC antigen binding groove. An attractive alternative is the use of noncovalent crystallisation chaperones, however these have not been developed for pMHC. Here we describe a method for promoting class I pMHC crystallisation, by exploiting its natural ligand interaction with the immunoregulatory receptor LILRB1 as a novel crystallisation chaperone. First, focussing on a model HIV-1-derived HLA-A2-restricted peptide, we determined a 2.4â¯Å HLA-A2/LILRB1 structure, which validated that co-crystallisation with LILRB1 does not alter conformation of the antigenic peptide. We then demonstrated that addition of LILRB1 enhanced the crystallisation of multiple peptide-HLA-A2 complexes, and identified a generic condition for initial co-crystallisation. LILRB1 chaperone-based crystallisation enabled structure determination for HLA-A2 complexes previously intransigent to crystallisation, including both conventional and post-translationally-modified peptides, of diverse lengths. Since both the LILRB1 recognition interface on the HLA-A2 α3 domain molecule and HLA-A2-mediated crystal contacts are predominantly conserved across class I MHC molecules, the approach we outline could prove applicable to a diverse range of class I pMHC. LILRB1 chaperone-mediated crystallisation should expedite molecular insights into the immunobiology of diverse immune-related diseases and immunotherapeutic strategies, particularly involving class I pMHC complexes that are challenging to crystallise.
Assuntos
Antígenos CD/química , Cristalografia por Raios X/métodos , Antígeno HLA-A2/química , Receptor B1 de Leucócitos Semelhante a Imunoglobulina/química , Chaperonas Moleculares/química , Antígenos CD/imunologia , Sítios de Ligação , Cristalização , HIV-1/imunologia , Antígeno HLA-A2/imunologia , Humanos , Epitopos Imunodominantes , Receptor B1 de Leucócitos Semelhante a Imunoglobulina/imunologia , Ligantes , Modelos Moleculares , Chaperonas Moleculares/imunologia , Fosforilação , Ligação Proteica , Conformação Proteica , Relação Estrutura-AtividadeRESUMO
BACKGROUND: Active infections with cytomegalovirus (CMV) increase NK cell expression of the inhibitory receptor LIR-1 and the activating receptor NKG2C in transplant recipients. However, the effects of CMV on NK cells are different in HIV patients stable on antiretroviral therapy (ART) and have not been analyzed in young HIV patients beginning ART. METHODOLOGY: We followed a cohort of 78 Indonesian HIV patients beginning ART. CMV antibodies were measured in plasma before ART (baseline), and after 1, 3, 6, and 12 months. CMV DNA was sought in blood granulocytes at baseline by quantitative PCR assay and a deletion in the NKG2C gene was identified by PCR. NK cell profiles were monitored by flow cytometry in 19 patients stratified by the presence of CMV DNA. Healthy controls (n = 17) were assessed once. RESULTS: All 78 patients were CMV seropositive and 41 had detectable CMV DNA. CMV DNA+ patients had higher proportions of total NK cells and CD16+ NK cells at baseline, but similar expression of LIR-1 and NKp30 on NK cells on ART. However, levels of CMV antibody were inversely related to median LIR-1 expression on NK cells. A dramatic elevation in cells expressing NKG2C was restricted to CMV DNA+ patients heterozygous for the NKG2C deletion. Patients with High NKG2C expression had lower levels of CMV antibodies. CONCLUSION: A subpopulation of NK cells expressing NKG2C was induced by CMV replication in HIV patients heterozygous for a deletion in this gene. Individuals with an abundant NKG2C+ and LIR-1+ NK cells displayed lower levels of CMV reactive antibody.
Assuntos
Infecções por Citomegalovirus/imunologia , Infecções por HIV/imunologia , Células Matadoras Naturais/imunologia , Adulto , Fármacos Anti-HIV/uso terapêutico , Anticorpos Antivirais/sangue , Antígenos CD/imunologia , Citomegalovirus/genética , Citomegalovirus/imunologia , Infecções por Citomegalovirus/tratamento farmacológico , Infecções por Citomegalovirus/virologia , DNA Viral , Feminino , Genótipo , Infecções por HIV/tratamento farmacológico , Infecções por HIV/virologia , Humanos , Imunoglobulina G/sangue , Receptor B1 de Leucócitos Semelhante a Imunoglobulina/imunologia , Masculino , Pessoa de Meia-Idade , Subfamília C de Receptores Semelhantes a Lectina de Células NK/genética , Subfamília C de Receptores Semelhantes a Lectina de Células NK/imunologia , Carga Viral , Adulto JovemRESUMO
UL18 is a human CMV (HCMV) MHC class I (MHCI) homolog that efficiently inhibits leukocyte immunoglobulin-like receptor subfamily B member 1 (LILRB1)+ NK cells. We found an association of LILRB1 polymorphisms in the regulatory regions and ligand-binding domains with control of HCMV in transplant patients. Naturally occurring LILRB1 variants expressed in model NK cells showed functional differences with UL18 and classical MHCI, but not with HLA-G. The altered functional recognition was recapitulated in binding assays with the binding domains of LILRB1. Each of 4 nonsynonymous substitutions in the first 2 LILRB1 immunoglobulin domains contributed to binding with UL18, classical MHCI, and HLA-G. One of the polymorphisms controlled addition of an N-linked glycan, and that mutation of the glycosylation site altered binding to all ligands tested, including enhancing binding to UL18. Together, these findings indicate that specific LILRB1 alleles that allow for superior immune evasion by HCMV are restricted by mutations that limit LILRB1 expression selectively on NK cells. The polymorphisms also maintained an appropriate interaction with HLA-G, fitting with a principal role of LILRB1 in fetal tolerance.
Assuntos
Antígenos CD , Proteínas do Capsídeo , Infecções por Citomegalovirus , Citomegalovirus , Predisposição Genética para Doença , Antígenos HLA-G , Receptor B1 de Leucócitos Semelhante a Imunoglobulina , Transplante de Órgãos , Polimorfismo Genético , Antígenos CD/genética , Antígenos CD/imunologia , Proteínas do Capsídeo/genética , Proteínas do Capsídeo/imunologia , Linhagem Celular , Citomegalovirus/genética , Citomegalovirus/imunologia , Infecções por Citomegalovirus/genética , Infecções por Citomegalovirus/imunologia , Infecções por Citomegalovirus/patologia , Feminino , Antígenos HLA-G/genética , Antígenos HLA-G/imunologia , Humanos , Células Matadoras Naturais/imunologia , Receptor B1 de Leucócitos Semelhante a Imunoglobulina/genética , Receptor B1 de Leucócitos Semelhante a Imunoglobulina/imunologia , MasculinoRESUMO
Mechanisms of immune regulation may control proliferation of aberrant plasma cells (PCs) in patients with monoclonal gammopathy of undetermined significance (MGUS) preventing progression to active multiple myeloma (MM). We hypothesized that CD85j (LILRB1), an inhibitory immune checkpoint for B cell function, may play a role in MM pathogenesis. In this study, we report that patients with active MM had significantly lower levels of CD85j and its ligand S100A9. Decreased CD85j expression could also be detected in the premalignant condition MGUS, suggesting that loss of CD85j may be an early event promoting tumor immune escape. To gain insight into the molecular mechanisms underlying CD85j functions, we next enforced expression of CD85j in human myeloma cell lines by lentiviral transduction. Interestingly, gene expression profiling of CD85j-overexpressing cells revealed a set of downregulated genes with crucial functions in MM pathogenesis. Furthermore, in vitro functional assays demonstrated that CD85j overexpression increased susceptibility to T cell- and NK-mediated killing. Consistently, ligation of CD85j decreased the number of PCs from individuals with MGUS but not from patients with MM. In conclusion, downregulation of inhibitory immune checkpoints on malignant PCs may provide a novel mechanism of immune escape associated with myeloma pathogenesis.
Assuntos
Antígenos CD/imunologia , Receptor B1 de Leucócitos Semelhante a Imunoglobulina/imunologia , Mieloma Múltiplo/imunologia , Plasmócitos/imunologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Linfócitos B , Linhagem Celular Tumoral , Regulação para Baixo/imunologia , Feminino , Humanos , Células Matadoras Naturais/imunologia , Masculino , Pessoa de Meia-Idade , Linfócitos T/imunologia , Transcriptoma/imunologiaRESUMO
Human Leukocyte Antigen-G (HLA-G), a non-classical, class Ib molecule, has been shown to mediate immunoregulatory functions by inducing apoptosis, inhibits cytotoxicity and differentiation by modulating cytokine secretion. Due to its immune-suppressive function, it facilitates tolerance in feto-maternal interface and transplantation. In contrary, it favours immune evasion of microbes and tumors by inhibiting immune and inflammatory responses. In Tuberculosis (TB), we previously reported differential expression of HLA-G and its receptor Ig-like transcript -2 (ILT-2) in disseminated vs. localized Tuberculosis. The present study explores the impact of HLA-G inhibition on the function of T cells and monocytes, in TB Pleural Effusion (PE), a localized form of TB. Blocking of HLA-G resulted in significant increase in IFN-γ and TNF-α production by CD3+ T cells. Additionally, we observed that HLA-G influences the apoptosis and cytotoxic effect of T cells from TB- PE patients. Next, we checked the impact of interaction between HLA-G and ILT-4 receptor in monocytes derived from TB-PE patients upon blocking and observed significant increase in IFN-γ production. The present study reveals for the first time HLA-G mediated suppression of Th1 cytokines, especially, IFN-γ and TNF-α in TB-PE patients.
Assuntos
Anticorpos Bloqueadores/farmacologia , Antígenos HLA-G/imunologia , Interferon gama/imunologia , Mycobacterium tuberculosis/imunologia , Derrame Pleural/imunologia , Células Th1/efeitos dos fármacos , Tuberculose Pleural/imunologia , Fator de Necrose Tumoral alfa/imunologia , Antígenos CD/imunologia , Antígenos CD/metabolismo , Apoptose/efeitos dos fármacos , Células Cultivadas , Antígenos HLA-G/metabolismo , Interações Hospedeiro-Patógeno , Humanos , Interferon gama/metabolismo , Receptor B1 de Leucócitos Semelhante a Imunoglobulina/imunologia , Receptor B1 de Leucócitos Semelhante a Imunoglobulina/metabolismo , Glicoproteínas de Membrana/imunologia , Glicoproteínas de Membrana/metabolismo , Monócitos/efeitos dos fármacos , Monócitos/imunologia , Monócitos/metabolismo , Monócitos/microbiologia , Perforina/imunologia , Perforina/metabolismo , Derrame Pleural/metabolismo , Derrame Pleural/microbiologia , Derrame Pleural/patologia , Receptores Imunológicos/imunologia , Receptores Imunológicos/metabolismo , Células Th1/imunologia , Células Th1/metabolismo , Células Th1/microbiologia , Tuberculose Pleural/metabolismo , Tuberculose Pleural/microbiologia , Tuberculose Pleural/patologia , Fator de Necrose Tumoral alfa/metabolismoRESUMO
One of the cardinal features of chronic lymphocytic leukemia (CLL) is its association with a profound immunosuppression. NK cell function is markedly impaired in CLL patients, who show a significant dysregulation of the expression of activating and inhibitory receptors. Here, we analyzed the role of the novel inhibitory receptor Ig-like transcript 2 (ILT2, also termed LIR-1, LILRB1) in the regulation of NK cells in CLL. Our results show that ILT2 expression was significantly decreased on leukemic cells and increased on NK cells of CLL patients, particularly in those with advanced disease and with bad prognostic features, such as those carrying chromosome del(11q). The immunomodulatory drug lenalidomide may regulate the expression of ILT2 and its ligands in CLL since it significantly increased the expression of ILT2 and partially reestablished the expression of its ligands on leukemic cells. Furthermore, lenalidomide significantly increased the activation and proliferation of NK cells, which was strongly enhanced by ILT2 blockade. Combining ILT2 blockade and lenalidomide activated NK cell cytotoxicity resulting in increased elimination of leukemic cells from CLL patients. Overall, we describe herein the role of an inhibitory receptor involved in the suppression of NK cell activity in CLL, which is restored by ILT2 blockade in combination with lenalidomide, suggesting that it may be an interesting therapeutic strategy to be explored in this disease.
Assuntos
Anticorpos Bloqueadores/uso terapêutico , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Células Matadoras Naturais/efeitos dos fármacos , Lenalidomida/uso terapêutico , Leucemia Linfocítica Crônica de Células B/tratamento farmacológico , Receptor B1 de Leucócitos Semelhante a Imunoglobulina/antagonistas & inibidores , Idoso , Anticorpos Bloqueadores/administração & dosagem , Anticorpos Bloqueadores/imunologia , Antígenos CD/imunologia , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Deleção Cromossômica , Sinergismo Farmacológico , Feminino , Humanos , Fatores Imunológicos/administração & dosagem , Fatores Imunológicos/uso terapêutico , Células Matadoras Naturais/imunologia , Lenalidomida/administração & dosagem , Leucemia Linfocítica Crônica de Células B/genética , Leucemia Linfocítica Crônica de Células B/imunologia , Receptor B1 de Leucócitos Semelhante a Imunoglobulina/imunologia , Masculino , Pessoa de Meia-IdadeRESUMO
Exciting progress in the field of cancer immunotherapy has renewed the urgency of the need for basic studies of immunoregulation in both adaptive cell lineages and innate cell lineages. Here we found a central role for major histocompatibility complex (MHC) class I in controlling the phagocytic function of macrophages. Our results demonstrated that expression of the common MHC class I component ß2-microglobulin (ß2M) by cancer cells directly protected them from phagocytosis. We further showed that this protection was mediated by the inhibitory receptor LILRB1, whose expression was upregulated on the surface of macrophages, including tumor-associated macrophages. Disruption of either MHC class I or LILRB1 potentiated phagocytosis of tumor cells both in vitro and in vivo, which defines the MHC class I-LILRB1 signaling axis as an important regulator of the effector function of innate immune cells, a potential biomarker for therapeutic response to agents directed against the signal-regulatory protein CD47 and a potential target of anti-cancer immunotherapy.