RESUMO
The functional diversity of natural killer (NK) cell repertoires stems from differentiation, homeostatic, receptor-ligand interactions and adaptive-like responses to viral infections. In the present study, we generated a single-cell transcriptional reference map of healthy human blood- and tissue-derived NK cells, with temporal resolution and fate-specific expression of gene-regulatory networks defining NK cell differentiation. Transfer learning facilitated incorporation of tumor-infiltrating NK cell transcriptomes (39 datasets, 7 solid tumors, 427 patients) into the reference map to analyze tumor microenvironment (TME)-induced perturbations. Of the six functionally distinct NK cell states identified, a dysfunctional stressed CD56bright state susceptible to TME-induced immunosuppression and a cytotoxic TME-resistant effector CD56dim state were commonly enriched across tumor types, the ratio of which was predictive of patient outcome in malignant melanoma and osteosarcoma. This resource may inform the design of new NK cell therapies and can be extended through transfer learning to interrogate new datasets from experimental perturbations or disease conditions.
Assuntos
Células Matadoras Naturais , Linfócitos do Interstício Tumoral , Neoplasias , Transcriptoma , Microambiente Tumoral , Humanos , Células Matadoras Naturais/imunologia , Células Matadoras Naturais/metabolismo , Microambiente Tumoral/imunologia , Neoplasias/imunologia , Neoplasias/genética , Linfócitos do Interstício Tumoral/imunologia , Linfócitos do Interstício Tumoral/metabolismo , Perfilação da Expressão Gênica , Análise de Célula Única , Redes Reguladoras de Genes , Antígeno CD56/metabolismo , Regulação Neoplásica da Expressão Gênica , Diferenciação CelularRESUMO
Substantial numbers of B cell leukemia and lymphoma patients relapse due to antigen loss or heterogeneity after anti-CD19 chimeric antigen receptor (CAR) T cell therapy. To overcome antigen escape and address antigen heterogeneity, we engineered induced pluripotent stem cell-derived NK cells to express both an NK cell-optimized anti-CD19 CAR for direct targeting and a high affinity, non-cleavable CD16 to augment antibody-dependent cellular cytotoxicity. In addition, we introduced a membrane-bound IL-15/IL-15R fusion protein to promote in vivo persistence. These engineered cells, termed iDuo NK cells, displayed robust CAR-mediated cytotoxic activity that could be further enhanced with therapeutic antibodies targeting B cell malignancies. In multiple in vitro and xenogeneic adoptive transfer models, iDuo NK cells exhibited robust anti-lymphoma activity. Furthermore, iDuo NK cells effectively eliminated both CD19+ and CD19- lymphoma cells and displayed a unique propensity for targeting malignant cells over healthy cells that expressed CD19, features not achievable with anti-CAR19 T cells. iDuo NK cells combined with therapeutic antibodies represent a promising approach to prevent relapse due to antigen loss and tumor heterogeneity in patients with B cell malignancies.
Assuntos
Leucemia , Neoplasias , Humanos , Deriva e Deslocamento Antigênicos , Leucemia/terapia , Células Matadoras NaturaisRESUMO
Adoptive transfer of allogeneic NK cells holds great promise for cancer immunotherapy. There is a variety of protocols to expand NK cells in vitro, most of which are based on stimulation with cytokines alone or in combination with feeder cells. Although IL-15 is essential for NK cell homeostasis in vivo, it is commonly used at supraphysiological levels to induce NK cell proliferation in vitro. As a result, adoptive transfer of such IL-15-addicted NK cells is associated with cellular stress because of sudden cytokine withdrawal. In this article, we describe a dose-dependent addiction to IL-15 during in vitro expansion of human NK cells, leading to caspase-3 activation and profound cell death upon IL-15 withdrawal. NK cell addiction to IL-15 was tightly linked to the BCL-2/BIM ratio, which rapidly dropped during IL-15 withdrawal. Furthermore, we observed a proliferation-dependent induction of BIM short, a highly proapoptotic splice variant of BIM in IL-15-activated NK cells. These findings shed new light on the molecular mechanisms involved in NK cell apoptosis following cytokine withdrawal and may guide future NK cell priming strategies in a cell therapy setting.
Assuntos
Proteína 11 Semelhante a Bcl-2/metabolismo , Proliferação de Células/efeitos dos fármacos , Interleucina-15/farmacologia , Células Matadoras Naturais/efeitos dos fármacos , Apoptose , Proteína 11 Semelhante a Bcl-2/genética , Caspase 3/metabolismo , Morte Celular/efeitos dos fármacos , Morte Celular/imunologia , Humanos , Interleucina-15/imunologia , Células K562 , Células Matadoras Naturais/imunologia , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2/metabolismoRESUMO
Newton's third law of motion states that for every action on a physical object there is an equal and opposite reaction. The dynamic change in functional potential of natural killer (NK) cells during education bears many features of such classical mechanics. Cumulative physical interactions between cells, under a constant influence of homeostatic drivers of differentiation, lead to a reactive spectrum that ultimately shapes the functionality of each NK cell. Inhibitory signaling from an array of self-specific receptors appear not only to suppress self-reactivity but also aid in the persistence of effector functions over time, thereby allowing the cell to gradually build up a functional potential. Conversely, the frequent non-cytolytic interactions between normal cells in the absence of such inhibitory signaling result in continuous stimulation of the cells and attenuation of effector function. Although an innate cell, the degree to which the fate of the NK cell is predetermined versus its ability to adapt to its own environment can be revealed through a Newtonian view of NK cell education, one which is both chronological and dynamic. As such, the development of NK cell functional diversity is the product of qualitatively different physical interactions with host cells, rather than simply the sum of their signals or an imprint based on intrinsically different transcriptional programs.
Assuntos
Comunicação Celular/imunologia , Diferenciação Celular/imunologia , Células Matadoras Naturais/imunologia , Receptores KIR/imunologia , Animais , Antígenos de Diferenciação de Linfócitos T/imunologia , Antígenos de Diferenciação de Linfócitos T/metabolismo , Humanos , Interleucina-15/imunologia , Interleucina-15/metabolismo , Células Matadoras Naturais/metabolismo , Modelos Imunológicos , Receptores KIR/metabolismo , Transdução de Sinais/imunologiaRESUMO
Acute and latent human CMV cause profound changes in the NK cell repertoire, with expansion and differentiation of educated NK cells expressing self-specific inhibitory killer cell Ig-like receptors. In this study, we addressed whether such CMV-induced imprints on the donor NK cell repertoire influenced the outcome of allogeneic stem cell transplantation. Hierarchical clustering of high-resolution immunophenotyping data covering key NK cell parameters, including frequencies of CD56(bright), NKG2A(+), NKG2C(+), and CD57(+) NK cell subsets, as well as the size of the educated NK cell subset, was linked to clinical outcomes. Clusters defining naive (NKG2A(+)CD57(-)NKG2C(-)) NK cell repertoires in the donor were associated with decreased risk for relapse in recipients with acute myeloid leukemia and myelodysplastic syndrome (hazard ratio [HR], 0.09; 95% confidence interval [CI]: 0.03-0.27; p < 0.001). Furthermore, recipients with naive repertoires at 9-12 mo after hematopoietic stem cell transplantation had increased disease-free survival (HR, 7.2; 95% CI: 1.6-33; p = 0.01) and increased overall survival (HR, 9.3; 95% CI: 1.1-77, p = 0.04). Conversely, patients with a relative increase in differentiated NK cells at 9-12 mo displayed a higher rate of late relapses (HR, 8.41; 95% CI: 6.7-11; p = 0.02), reduced disease-free survival (HR, 0.12; 95% CI: 0.12-0.74; p = 0.02), and reduced overall survival (HR, 0.07; 95% CI: 0.01-0.69; p = 0.02). Thus, our data suggest that naive donor NK cell repertoires are associated with protection against leukemia relapse after allogeneic HSCT.
Assuntos
Aloenxertos/imunologia , Transplante de Células-Tronco Hematopoéticas , Células Matadoras Naturais/imunologia , Leucemia/imunologia , Subpopulações de Linfócitos/imunologia , Adolescente , Adulto , Idoso , Criança , Análise por Conglomerados , Infecções por Citomegalovirus/complicações , Infecções por Citomegalovirus/imunologia , Intervalo Livre de Doença , Feminino , Citometria de Fluxo , Humanos , Imunofenotipagem , Estimativa de Kaplan-Meier , Leucemia/mortalidade , Leucemia/cirurgia , Masculino , Pessoa de Meia-Idade , Recidiva , Doadores de Tecidos , Adulto JovemRESUMO
The ability of NK cells to specifically recognize cells lacking expression of self-MHC class I molecules was discovered over 30 years ago. It provided the foundation for the "missing self" hypothesis. Research in the two past decades has contributed to a detailed understanding of the molecular mechanisms that determine the specificity and strength of NK cell-mediated "missing self" responses to tumor cells. However, in light of the recent remarkable breakthroughs in clinical cancer immunotherapy, the cytolytic potential of NK cells still remains largely untapped in clinical settings. There is abundant evidence demonstrating partial or complete loss of HLA class I expression in a wide spectrum of human tumor types. Such loss may result from immune selection of escape variants by tumor-specific CD8 T cells and has more recently also been linked to acquired resistance to checkpoint inhibition therapy. In the present review, we discuss the early predictions of the "missing self" hypothesis, its molecular basis and outline the potential for NK cell-based adoptive immunotherapy to convert checkpoint inhibitor therapy-resistant patients into clinical responders.
Assuntos
Antígeno CTLA-4/antagonistas & inibidores , Células Matadoras Naturais/imunologia , Neoplasias/tratamento farmacológico , Receptor de Morte Celular Programada 1/antagonistas & inibidores , Antígenos de Histocompatibilidade Classe I/imunologia , Humanos , Neoplasias/imunologia , Receptores KIR/genética , Linfócitos T Citotóxicos/imunologiaRESUMO
The MHC class Ib molecule HLA-G has previously been reported to be the ligand for the NK cell receptor killer Ig-like receptor (KIR)2DL4, but this remains controversial. In this study, we investigated IFN-γ production by freshly isolated NK cells in response to both soluble and solid-phase ligands, including anti-KIR2DL4 mAbs and rHLA-G. Although freshly isolated CD56(bright) NK cells produced IFN-γ in response to soluble HLA-G preparations, the response was found to be absolutely dependent on the presence of small numbers of contaminating CD56(-), CD14(-), CD11c(+) myeloid dendritic cells (mDCs). HLA-G tetramers bound only to the contaminating mDCs in the NK preparations, and Abs to KIR2DL4 and HLA-G did not block NK cell IFN-γ production. NK cells did not respond to plate-bound HLA-G. Freshly isolated NK cells also produced IFN-γ in response to unpurified soluble anti-KIR2DL4 mAb but not to low endotoxin affinity-purified Ab. The data suggest that previous reports of functional interactions between KIR2DL4 and HLA-G may have resulted from the use of purified NK cells that were contaminated with mDCs and HLA-G preparations that were contaminated with material capable of stimulating mDCs to produce cytokines that stimulate NK cells to produce IFN-γ.
Assuntos
Antígenos HLA-G/imunologia , Interferon gama/imunologia , Células Matadoras Naturais/imunologia , Receptores KIR2DL4/imunologia , Receptores KIR2DL4/metabolismo , Anticorpos Monoclonais/imunologia , Antígenos CD/imunologia , Antígenos CD/metabolismo , Células Cultivadas , Células Dendríticas/imunologia , Células Dendríticas/metabolismo , Humanos , Interferon gama/metabolismo , Interleucina-12/imunologia , Interleucina-12/metabolismo , Células Matadoras Naturais/metabolismo , Glicoproteínas de Membrana/imunologia , Glicoproteínas de Membrana/metabolismo , Células Mieloides/imunologia , Células Mieloides/metabolismo , Receptores Imunológicos/imunologia , Receptores Imunológicos/metabolismoRESUMO
UNLABELLED: The RV144 HIV-1 vaccine trial demonstrated partial efficacy of 31% against HIV-1 infection. Studies into possible correlates of protection found that antibodies specific to the V1 and V2 (V1/V2) region of envelope correlated inversely with infection risk and that viruses isolated from trial participants contained genetic signatures of vaccine-induced pressure in the V1/V2 region. We explored the hypothesis that the genetic signatures in V1 and V2 could be partly attributed to selection by vaccine-primed T cells. We performed a T-cell-based sieve analysis of breakthrough viruses in the RV144 trial and found evidence of predicted HLA binding escape that was greater in vaccine versus placebo recipients. The predicted escape depended on class I HLA A*02- and A*11-restricted epitopes in the MN strain rgp120 vaccine immunogen. Though we hypothesized that this was indicative of postacquisition selection pressure, we also found that vaccine efficacy (VE) was greater in A*02-positive (A*02(+)) participants than in A*02(-) participants (VE = 54% versus 3%, P = 0.05). Vaccine efficacy against viruses with a lysine residue at site 169, important to antibody binding and implicated in vaccine-induced immune pressure, was also greater in A*02(+) participants (VE = 74% versus 15%, P = 0.02). Additionally, a reanalysis of vaccine-induced immune responses that focused on those that were shown to correlate with infection risk suggested that the humoral responses may have differed in A*02(+) participants. These exploratory and hypothesis-generating analyses indicate there may be an association between a class I HLA allele and vaccine efficacy, highlighting the importance of considering HLA alleles and host immune genetics in HIV vaccine trials. IMPORTANCE: The RV144 trial was the first to show efficacy against HIV-1 infection. Subsequently, much effort has been directed toward understanding the mechanisms of protection. Here, we conducted a T-cell-based sieve analysis, which compared the genetic sequences of viruses isolated from infected vaccine and placebo recipients. Though we hypothesized that the observed sieve effect indicated postacquisition T-cell selection, we also found that vaccine efficacy was greater for participants who expressed HLA A*02, an allele implicated in the sieve analysis. Though HLA alleles have been associated with disease progression and viral load in HIV-1 infection, these data are the first to suggest the association of a class I HLA allele and vaccine efficacy. While these statistical analyses do not provide mechanistic evidence of protection in RV144, they generate testable hypotheses for the HIV vaccine community and they highlight the importance of assessing the impact of host immune genetics in vaccine-induced immunity and protection. (This study has been registered at ClinicalTrials.gov under registration no. NCT00223080.).
Assuntos
Vacinas contra a AIDS/imunologia , Infecções por HIV/prevenção & controle , HIV-1/imunologia , Antígeno HLA-A2/genética , Antígeno HLA-A2/imunologia , Vacinas contra a AIDS/administração & dosagem , Estudos de Coortes , Estudos de Associação Genética , Proteína gp120 do Envelope de HIV/genética , Proteína gp120 do Envelope de HIV/imunologia , Infecções por HIV/imunologia , Humanos , Linfócitos T/imunologiaRESUMO
Killer Ig-like receptors (KIRs) are innate immune receptors expressed by NK and T cells classically associated with the detection of missing self through loss of their respective MHC ligand. Some KIR specificities for allelic classical class I MHC (MHC-I) have been described, whereas other KIR receptor-ligand relationships, including those associated with nonclassical MHC-I, have yet to be clearly defined. We report in this article that KIR3DL2 and KIR2DS4 and the nonclassical Ag HLA-F, expressed as a free form devoid of peptide, physically and functionally interact. These interactions extend to include classical MHC-I open conformers as ligands, defining new relationships between KIR receptors and MHC-I. The data collectively suggest a broader, previously unrecognized interaction between MHC-I open conformers--including prototypical HLA-F--and KIR receptors, acting in an immunoregulatory capacity centered on the inflammatory response.
Assuntos
Antígenos de Histocompatibilidade Classe I/imunologia , Células Matadoras Naturais/imunologia , Células Matadoras Naturais/metabolismo , Receptores Imunológicos/metabolismo , Receptores de Células Matadoras Naturais/metabolismo , Linhagem Celular , Citotoxicidade Imunológica , Expressão Gênica , Antígenos de Histocompatibilidade Classe I/química , Antígenos de Histocompatibilidade Classe I/metabolismo , Humanos , Ligantes , Ligação Proteica/imunologia , Multimerização Proteica , Receptores KIR3DL2/metabolismo , Receptores de Células Matadoras Naturais/genética , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Ressonância de Plasmônio de SuperfícieRESUMO
Peptides that are presented by MHC class I (MHC-I) are processed from two potential sources, as follows: newly synthesized endogenous proteins for direct presentation on the surface of most nucleated cells and exogenous proteins for cross-presentation typically by professional APCs. In this study, we present data that implicate the nonclassical HLA-F and open conformers of MHC-I expressed on activated cells in a pathway for the presentation of exogenous proteins by MHC-I. This pathway is distinguished from the conventional endogenous pathway by its independence from TAP and tapasin and its sensitivity to inhibitors of lysosomal enzymes, and further distinguished by its dependence on MHC-I allotype-specific epitope recognition for Ag uptake. Thus, our data from in vitro experiments collectively support a previously unrecognized model of Ag cross-presentation mediated by HLA-F and MHC-I open conformers on activated lymphocytes and monocytes, which may significantly contribute to the regulation of immune system functions and the immune defense.
Assuntos
Apresentação Cruzada , Antígenos de Histocompatibilidade Classe I/imunologia , Modelos Imunológicos , Sequência de Aminoácidos , Antígenos de Superfície/imunologia , Antígenos Virais/química , Antígenos Virais/imunologia , Linfócitos B/imunologia , Linhagem Celular Tumoral , Endossomos/efeitos dos fármacos , Endossomos/enzimologia , Endossomos/imunologia , Inibidores Enzimáticos/farmacologia , Epitopos/imunologia , Epitopos de Linfócito T , Antígenos de Histocompatibilidade Classe I/química , Humanos , Células Matadoras Naturais/imunologia , Ativação Linfocitária , Linfócitos/imunologia , Linfoma Difuso de Grandes Células B/patologia , Dados de Sequência Molecular , Monócitos/imunologia , Fragmentos de Peptídeos/imunologia , Conformação Proteica , Transporte Proteico , Linfócitos T Citotóxicos/imunologiaRESUMO
The functionality of natural killer (NK) cells is tuned during education and is associated with remodeling of the lysosomal compartment. We hypothesized that genetic variation in killer cell immunoglobulin-like receptor (KIR) and HLA, which is known to influence the functional strength of NK cells, fine-tunes the payload of effector molecules stored in secretory lysosomes. To address this possibility, we performed a high-resolution analysis of KIR and HLA class I genes in 365 blood donors and linked genotypes to granzyme B loading and functional phenotypes. We found that granzyme B levels varied across individuals but were stable over time in each individual and genetically determined by allelic variation in HLA class I genes. A broad mapping of surface receptors and lysosomal effector molecules revealed that DNAM-1 and granzyme B levels served as robust metric of the functional state in NK cells. Variation in granzyme B levels at rest was tightly linked to the lytic hit and downstream killing of major histocompatibility complex-deficient target cells. Together, these data provide insights into how variation in genetically hardwired receptor pairs tunes the releasable granzyme B pool in NK cells, resulting in predictable hierarchies in global NK cell function.
Assuntos
Células Matadoras Naturais , Receptores KIR , Granzimas/genética , Granzimas/metabolismo , Receptores KIR/genética , Receptores KIR/metabolismo , Antígenos de Histocompatibilidade Classe I/metabolismo , GenótipoRESUMO
HLA-F has low levels of polymorphism in humans and is highly conserved among primates, suggesting a conserved function in the immune response. In this study, we probed the structure of HLA-F on the surface of B lymphoblastoid cell lines and activated lymphocytes by direct measurement of peptide binding to native HLA-F. Our findings suggested that HLA-F is expressed independently of bound peptide, at least in regard to peptide complexity profiles similar to those of either HLA-E or classical MHC class I (MHC-I). As a further probe of native HLA-F structure, we used a number of complementary approaches to explore the interactions of HLA-F with other molecules, at the cell surface, intracellularly, and in direct physical biochemical measurements. This analysis demonstrated that HLA-F surface expression was coincident with MHC-I H chain (HC) expression and was downregulated upon perturbation of MHC-I HC structure. It was further possible to directly demonstrate that MHC-I would interact with HLA-F only when in the form of an open conformer free of peptide and not as a trimeric complex. This interaction was directly observed by coimmunoprecipitation and by surface plasmon resonance and indirectly on the surface of cells through coincident tetramer and MHC-I HC colocalization. These data suggest that HLA-F is expressed independently of peptide and that a physical interaction specific to MHC-I HC plays a role in the function of MHC-I HC expression in activated lymphocytes.
Assuntos
Linfócitos B/metabolismo , Antígenos HLA/química , Antígenos HLA/metabolismo , Antígenos de Histocompatibilidade Classe I/metabolismo , Linfócitos B/imunologia , Western Blotting , Linhagem Celular , Separação Celular , Citometria de Fluxo , Imunofluorescência , Antígenos de Histocompatibilidade Classe I/química , Humanos , Imunoprecipitação , Espectrometria de Massas , Ligação Proteica , Conformação ProteicaRESUMO
Allogeneic natural killer (NK) cell adoptive transfer is a promising treatment for several cancers but is less effective for the treatment of multiple myeloma. In this study, we report on quadruple gene-engineered induced pluripotent stem cell (iPSC)-derived NK cells designed for mass production from a renewable source and for dual targeting against multiple myeloma through the introduction of an NK cell-optimized chimeric antigen receptor (CAR) specific for B cell maturation antigen (BCMA) and a high affinity, non-cleavable CD16 to augment antibody-dependent cellular cytotoxicity when combined with therapeutic anti-CD38 antibodies. Additionally, these cells express a membrane-bound interleukin-15 fusion molecule to enhance function and persistence along with knock out of CD38 to prevent antibody-mediated fratricide and enhance NK cell metabolic fitness. In various preclinical models, including xenogeneic adoptive transfer models, quadruple gene-engineered NK cells consistently demonstrate durable antitumor activity independent of exogenous cytokine support. Results presented here support clinical translation of this off-the-shelf strategy for effective treatment of multiple myeloma.
Assuntos
Mieloma Múltiplo , Humanos , Mieloma Múltiplo/genética , Mieloma Múltiplo/terapia , Células Matadoras Naturais , Antígeno de Maturação de Linfócitos B , Receptores de Células Matadoras Naturais , Subfamília D de Receptores Semelhantes a Lectina de Células NKRESUMO
Cytotoxic lymphocytes, including natural killer (NK) cells and T cells are distinguished by their ability to eliminate target cells through release of secretory lysosomes. Conventional lysosomes and secretory lysosomes are part of the pleomorphic endolysosomal system and characterized by its highly dynamic nature. Several calcium-permeable TRP calcium channels play an essential role in endolysosomal calcium signaling to ensure proper function of these organelles. In NK cells, the expression of self MHC-specific inhibitory receptors dynamically tunes their secretory potential in a non-transcriptional, calcium-dependent manner. New insights suggest that TRPML1-mediated lysosomal calcium fluxes are tightly interconnected to NK cell functionality through modulation of granzyme B and perforin content of the secretory lysosome. Lysosomal TRP channels show a subset-specific expression pattern during NK differentiation, which is paralleled with gradually increased loading of effector molecules in secretory lysosomes. Methodological advances, including organellar patch-clamping, specific pharmacological modulators, and genetically-encoded calcium indicators open up new possibilities to investigate how TRP channels influence communication between intracellular organelles in immune cells. This review discusses our current understanding of lysosome biogenesis in NK cells with an emphasis on the TRP mucolipin family and the implications for NK cell functionality and cancer immunotherapy.
Assuntos
Canais de Cálcio/metabolismo , Sinalização do Cálcio/imunologia , Cálcio/metabolismo , Endossomos/metabolismo , Células Matadoras Naturais/imunologia , Lisossomos/metabolismo , Canais de Potencial de Receptor Transitório/metabolismo , Animais , Granzimas/metabolismo , Humanos , Camundongos , Perforina/metabolismoRESUMO
Natural killer (NK) cell repertoires are made up of phenotypically distinct subsets with different functional properties. The molecular programs involved in maintaining NK cell repertoire diversity under homeostatic conditions remain elusive. Here, we show that subset-specific NK cell proliferation kinetics correlate with mTOR activation, and global repertoire diversity is maintained through a high degree of intra-lineage subset plasticity during interleukin (IL)-15-driven homeostatic proliferation in vitro. Slowly cycling sorted KIR+CD56dim NK cells with an induced CD57 phenotype display increased functional potential associated with increased transcription of genes involved in adhesion and immune synapse formation. Rapidly cycling cells upregulate NKG2A, display a general loss of functionality, and a transcriptional signature associated with increased apoptosis/cellular stress, actin-remodeling, and nuclear factor κB (NF-κB) activation. These results shed light on the role of intra-lineage plasticity during NK cell homeostasis and suggest that the functional fate of the cell is tightly linked to the acquired phenotype and transcriptional reprogramming.
Assuntos
Células Matadoras Naturais/metabolismo , Apoptose/genética , Apoptose/fisiologia , Antígeno CD56/metabolismo , Antígenos CD57/metabolismo , Humanos , Interleucina-15/metabolismo , Cinética , NF-kappa B/metabolismo , Subfamília C de Receptores Semelhantes a Lectina de Células NK/metabolismo , Fenótipo , Análise de Sequência de RNA , Sinapses/metabolismoRESUMO
Inhibitory signaling during natural killer (NK) cell education translates into increased responsiveness to activation; however, the intracellular mechanism for functional tuning by inhibitory receptors remains unclear. Secretory lysosomes are part of the acidic lysosomal compartment that mediates intracellular signalling in several cell types. Here we show that educated NK cells expressing self-MHC specific inhibitory killer cell immunoglobulin-like receptors (KIR) accumulate granzyme B in dense-core secretory lysosomes that converge close to the centrosome. This discrete morphological phenotype is independent of transcriptional programs that regulate effector function, metabolism and lysosomal biogenesis. Meanwhile, interference of signaling from acidic Ca2+ stores in primary NK cells reduces target-specific Ca2+-flux, degranulation and cytokine production. Furthermore, inhibition of PI(3,5)P2 synthesis, or genetic silencing of the PI(3,5)P2-regulated lysosomal Ca2+-channel TRPML1, leads to increased granzyme B and enhanced functional potential, thereby mimicking the educated state. These results indicate an intrinsic role for lysosomal remodeling in NK cell education.
Assuntos
Células Matadoras Naturais/metabolismo , Lisossomos/metabolismo , Aminopiridinas/farmacologia , Animais , Granzimas/metabolismo , Compostos Heterocíclicos com 3 Anéis/farmacologia , Humanos , Células K562 , Células Matadoras Naturais/efeitos dos fármacos , Proteína 1 de Membrana Associada ao Lisossomo/genética , Proteína 1 de Membrana Associada ao Lisossomo/metabolismo , Lisossomos/efeitos dos fármacos , Camundongos , Receptores KIR/metabolismo , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/fisiologiaRESUMO
Infection by human cytomegalovirus (HCMV) leads to NKG2C-driven expansion of adaptive natural killer (NK) cells, contributing to host defense. However, approximately 4% of all humans carry a homozygous deletion of the gene that encodes NKG2C (NKG2C(-/-)). Assessment of NK cell repertoires in 60 NKG2C(-/-) donors revealed a broad range of NK cell populations displaying characteristic footprints of adaptive NK cells, including a terminally differentiated phenotype, functional reprogramming, and epigenetic remodeling of the interferon (IFN)-γ promoter. We found that both NKG2C(-) and NKG2C(+) adaptive NK cells expressed high levels of CD2, which synergistically enhanced ERK and S6RP phosphorylation following CD16 ligation. Notably, CD2 co-stimulation was critical for the ability of adaptive NK cells to respond to antibody-coated target cells. These results reveal an unexpected redundancy in the human NK cell response to HCMV and suggest that CD2 provides "signal 2" in antibody-driven adaptive NK cell responses.
Assuntos
Imunidade Adaptativa/imunologia , Antígenos CD2/imunologia , Citomegalovirus/imunologia , Células Matadoras Naturais/imunologia , Subfamília C de Receptores Semelhantes a Lectina de Células NK/genética , Linfócitos T CD8-Positivos/imunologia , Células Cultivadas , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Proteínas Ligadas por GPI/genética , Proteínas Ligadas por GPI/metabolismo , Humanos , Interferon gama/genética , Interferon gama/imunologia , Ativação Linfocitária/imunologia , Subfamília C de Receptores Semelhantes a Lectina de Células NK/deficiência , Receptores de IgG/genética , Receptores de IgG/metabolismo , Proteína S6 Ribossômica/metabolismoRESUMO
Natural killer (NK) cells are innate lymphocytes with a refined ability to recognize transformed cells through a broad array of activating receptors in combination with stochastically expressed inhibitory receptors that recognize MHC-class I. Recent advances in NK cell biology have revealed a high degree of functional plasticity that can be attributed to dynamic cell-to-cell interactions in concert with transcriptional and epigenetic reprogramming. Here, we discuss how new insights into the adaptive behavior of NK cells pave the way for next generation cell therapy based on guided differentiation and selective expansion of particularly cytotoxic NK cell subsets.
Assuntos
Imunoterapia , Células Matadoras Naturais/imunologia , Neoplasias/terapia , HumanosRESUMO
Adaptive natural killer (NK) cell responses to human cytomegalovirus infection are characterized by the expansion of NKG2C(+) NK cells expressing self-specific inhibitory killer-cell immunoglobulin-like receptors (KIRs). Here, we set out to study the HLA class I dependency of such NKG2C(+) NK cell expansions. We demonstrate the expansion of NKG2C(+) NK cells in patients with transporter associated with antigen presentation (TAP) deficiency, who express less than 10% of normal HLA class I levels. In contrast to normal individuals, expanded NKG2C(+) NK cell populations in TAP-deficient patients display a polyclonal KIR profile and remain hyporesponsive to HLA class I-negative target cells. Nonetheless, agonistic stimulation of NKG2C on NK cells from TAP-deficient patients yielded significant responses in terms of degranulation and cytokine production. Thus, while interactions with self-HLA class I molecules likely shape the KIR repertoire of expanding NKG2C(+) NK cells during adaptive NK cell responses in normal individuals, they are not a prerequisite for NKG2C(+) NK cell expansions to occur. The emergence of NKG2C-responsive adaptive NK cells in TAP-deficient patients may contribute to antiviral immunity and potentially explain these patients' low incidence of severe viral infections.