RESUMEN
NKG2C+ natural killer (NK) cell plays a vital role in CMV infection control after hematopoietic stem cell transplantation (HSCT). However, the modulation on NKG2C+ NK cell reconstitution is still unclear. NK cell education is affected by the interactions of HLA-I/killer immunoglobulin receptor (KIR). Our aim is to figure out which HLA-I/KIR interaction plays a dominant role in NKG2C+ NK education. Based on allogeneic haploidentical HSCT, we investigated the expansion and function of single KIR positive NKG2C+ NK cells via the interaction of KIR with both donor HLA and recipient HLA at days 30, 90, and 180 after HSCT. KIR2DL2/L3 single-positive/NKG2C+ cells were significantly expanded compared with KIR2DL1 or KIR3DL1 single-positive/NKG2C+ cells when donors and recipients were both HLA-C1/C1 or HLA-C1C1BW4 (p < 0.05), with higher NKp30 expression (p < 0.05). Moreover, the proportion of single KIR positive NK cells increased in both NKG2C+/NKG2A- NK cells and conventional NKG2C-/NKG2A- NK cells over time. We also observed that increased proportion of KIR2DL2/L3 single-positive/NKG2C+ NK cells correlated with higher incidence of acute graft-versus-host disease (aGVHD). Our study allows a better understanding of HLA-I/KIR interaction in the NKG2C+ NK cell education after HSCT.
Asunto(s)
Enfermedad Injerto contra Huésped , Trasplante de Células Madre Hematopoyéticas , Enfermedad Injerto contra Huésped/metabolismo , Antígenos HLA-C/inmunología , Trasplante de Células Madre Hematopoyéticas/efectos adversos , Humanos , Incidencia , Células Asesinas Naturales/metabolismo , Receptores KIR/genética , Receptores KIR/metabolismo , Receptores KIR2DL2/genética , Receptores KIR2DL2/metabolismoRESUMEN
The closely related inhibitory killer-cell immunoglobulin-like receptors (KIR), KIR2DL2 and KIR2DL3, regulate the activation of natural killer cells (NK) by interacting with the human leukocyte antigen-C1 (HLA-C1) group of molecules. KIR2DL2, KIR2DL3 and HLA-C1 are highly polymorphic, with this variation being associated with differences in the onset and progression of some human diseases. However, the molecular bases underlying these associations remain unresolved. Here, we determined the crystal structures of KIR2DL2 and KIR2DL3 in complex with HLA-C*07:02 presenting a self-epitope. KIR2DL2 differed from KIR2DL3 in docking modality over HLA-C*07:02 that correlates with variabilty of recognition of HLA-C1 allotypes. Mutagenesis assays indicated differences in the mechanism of HLA-C1 allotype recognition by KIR2DL2 and KIR2DL3. Similarly, HLA-C1 allotypes differed markedly in their capacity to inhibit activation of primary NK cells. These functional differences derive, in part, from KIR2DS2 suggesting KIR2DL2 and KIR2DL3 binding geometries combine with other factors to distinguish HLA-C1 functional recognition.
Asunto(s)
Antígenos HLA-C/metabolismo , Simulación del Acoplamiento Molecular , Receptores KIR2DL2/química , Receptores KIR2DL2/metabolismo , Receptores KIR2DL3/química , Receptores KIR2DL3/metabolismo , Células HEK293 , Humanos , Células Asesinas Naturales/inmunología , Ligandos , Proteínas Mutantes/química , Proteínas Mutantes/metabolismo , Péptidos/química , Unión Proteica , Mapeo de Interacción de ProteínasRESUMEN
Chronic exposure to arsenic causes cancers in various organs including the skin, liver, lung, and bladder in humans, but the mechanisms of the multi-organ carcinogenicity of arsenic remain unknown. Natural killer (NK) cells play important roles in the immune surveillance and elimination of tumor cells. Although accumulating evidence has indicated that arsenic has immunosuppressive properties, little is known about the effects of arsenic on the tumoricidal functions of NK cells. We examined the effects of arsenite on the cytotoxic activities of human and mouse NK cells toward target tumor cells. Exposure of human NK-92 cells and primary mouse NK cells to sublethal doses of arsenite reduced the IL-2-activated cytotoxic activities toward human K562 cells and murine YAC-1 cells, respectively. NK cells recognize target cells via integrated signals from both activating and inhibitory receptors and induce apoptosis of target cells via a granzyme/perforin system. We found that exposure of NK-92 cells to arsenite diminished the IL-2-activated down-regulation of the inhibitory receptors, KIR2DL2 and KIR2DL3, and the up-regulation of granzyme B and lymphotoxin-α. The IL-2-activated increases in secretion of interferon-γ and IL-10 were also slightly reduced by arsenite. Thus, arsenite suppressed the IL-2-activated cytotoxic activity of NK cells by disrupting multiple pathways required for the recognition and killing of target tumor cells. Our findings provide new insights into the roles of NK cell-mediated tumor immunity in cancer development by arsenic.
Asunto(s)
Arsenitos/toxicidad , Citotoxicidad Inmunológica/efectos de los fármacos , Interleucina-2/farmacología , Células Asesinas Naturales/efectos de los fármacos , Leucemia Mielógena Crónica BCR-ABL Positiva/inmunología , Compuestos de Sodio/toxicidad , Escape del Tumor/efectos de los fármacos , Animales , Técnicas de Cocultivo , Granzimas/genética , Granzimas/metabolismo , Humanos , Interferón gamma/genética , Interferón gamma/metabolismo , Interleucina-10/metabolismo , Células K562 , Células Asesinas Naturales/inmunología , Células Asesinas Naturales/metabolismo , Leucemia Mielógena Crónica BCR-ABL Positiva/metabolismo , Leucemia Mielógena Crónica BCR-ABL Positiva/patología , Linfotoxina-alfa/metabolismo , Ratones , Ratones Endogámicos C57BL , Receptores KIR2DL2/genética , Receptores KIR2DL2/metabolismo , Receptores KIR2DL3/genética , Receptores KIR2DL3/metabolismoRESUMEN
Specific killer cell immunoglobulin-like receptor (KIR) in women with recurrent pregnancy loss (RPL) and HLA ligands in couples invoke a susceptibility to RPL. However, the relationship between KIR2DL2 and its cognate ligand HLA-C1 has not been explored. In this prospective cohort study, 160 Caucasian women with RPL and 99 partners were included. KIR/HLA-C typing, NK assay, Th1/Th2 intracellular cytokine ratios, 25-(OH)-vitamin D level, and the presence of autoantibodies were analyzed. KIR2DL2 positive women (P = 0.023) and their partners (P = 0.017) had lower allele frequencies of HLA-C1 than those of KIR2DL2 negative women. KIR2DL2 positive women had significantly lower genotype frequency of HLA-C1C1 as compared to the North American Caucasian population controls (P < 0.05). In the partners of KIR2DL2 positive women, there was a substantially higher frequency of HLA-C2C2 than controls (P = 0.016). Besides, KIR2DL2 negative women had a higher prevalence of anti-ssDNA antibody as compared with that of KIR2DL2 positive women (P = 0.043). There were no differences in the distribution of HLA-C genotypes based on KIR2DL2, regardless of pregnancy outcome in women with RPL and their partners while on immunomodulation treatment. In conclusion, decreased ligands for inhibitory KIRs (inhKIR) could lead to insufficient inhibition of maternal uterine NK cells toward the trophoblast, thereby contributing to the pathogenesis of RPL. Specific KIR and HLA-C genotyping may predict the reproductive outcome of women with RPL.
Asunto(s)
Aborto Habitual/genética , Predisposición Genética a la Enfermedad , Antígenos HLA-C/genética , Factores Inmunológicos/administración & dosificación , Receptores KIR2DL2/metabolismo , Aborto Habitual/sangre , Aborto Habitual/inmunología , Aborto Habitual/prevención & control , Adulto , Alelos , Autoanticuerpos/sangre , Autoanticuerpos/inmunología , Estudios de Casos y Controles , ADN de Cadena Simple/inmunología , Femenino , Frecuencia de los Genes/inmunología , Antígenos HLA-C/metabolismo , Humanos , Células Asesinas Naturales/efectos de los fármacos , Células Asesinas Naturales/inmunología , Células Asesinas Naturales/metabolismo , Masculino , Embarazo , Resultado del Embarazo , Estudios Prospectivos , Receptores KIR2DL2/análisis , Transducción de Señal/efectos de los fármacos , Transducción de Señal/genética , Transducción de Señal/inmunología , Resultado del TratamientoRESUMEN
Natural Killer (NK) cell activation requires integration of inhibitory and activating signaling. Inhibitory signals are determined by members of the killer cell immunoglobulin-like receptor (KIR) family, which have major histocompatibility complex (MHC) class I ligands. Loss of this inhibitory signal leads to NK cell activation. Thus, down-regulation of MHC I during viral infection or cancer induces NK cell activation. However, NK cell activation in the presence of MHC-I has been demonstrated for HLA-C*0102 through changes in its peptide content: "peptide antagonism." Here we identify an antagonist peptide for HLA-C*0304 suggesting that peptide antagonism is a generalizable phenomenon and, using a combination of mathematical modeling, confocal imaging, and immune-assays, we quantitatively determine mechanisms that underlie peptide antagonism in inhibitory KIR2DL2/3 signaling. These data provide a mechanism for NK cell activation based on a reduction of inhibitory signaling in the presence of preserved levels of MHC class I.
Asunto(s)
Células Asesinas Naturales/inmunología , Receptores KIR2DL2/metabolismo , Antígenos/inmunología , Células Cultivadas , Análisis por Conglomerados , Antígenos HLA-C/metabolismo , Humanos , Activación de Linfocitos , Microscopía Confocal , Modelos Teóricos , Péptidos/inmunología , Unión Proteica , Receptor Cross-Talk , Transducción de SeñalRESUMEN
BACKGROUND: Lassa virus (LASV) is the etiologic agent of an acute hemorrhagic fever endemic in West Africa. Natural killer (NK) cells control viral infections in part through the interaction between killer cell immunoglobulin-like receptors (KIRs) and their ligands. LASV infection is associated with defective immune responses, including inhibition of NK cell activity in the presence of MHC-class 1+-infected target cells. METHODS: We compared individual KIR and HLA-class 1 genotypes of 68 healthy volunteers to 51 patients infected with LASV in Sierra Leone, including 37 survivors and 14 fatalities. Next, potential HLA-C1, HLA-C2, and HLA-Bw4 binding epitopes were in silico screened among LASV nucleoprotein (NP) and envelope glycoprotein (GP). Selected 10-mer peptides were then tested in peptide-HLA stabilization, KIR binding and polyfunction assays. FINDINGS: LASV-infected patients were similar to healthy controls, except for the inhibitory KIR2DL2 gene. We found a specific increase in the HLA-C1:KIR2DL2 interaction in fatalities (10/11) as compared to survivors (12/19) and controls (19/29). We also identified that strong of NP and GP viral epitopes was only observed with HLA-C molecules, and associated with strong inhibition of degranulation in the presence of KIR2DL+ NK cells. This inhibitory effect significantly increased in the presence of the vGP420 variant, detected in 28.1% of LASV sequences. INTERPRETATION: Our finding suggests that presentation of specific LASV epitopes by HLA-C alleles to the inhibitory KIR2DL2 receptor on NK cells could potentially prevent the killing of infected cells and provides insights into the mechanisms by which LASV can escape NK-cell-mediated immune pressure.
Asunto(s)
Epítopos/inmunología , Antígenos HLA-C/inmunología , Células Asesinas Naturales/inmunología , Células Asesinas Naturales/metabolismo , Fiebre de Lassa/inmunología , Fiebre de Lassa/metabolismo , Virus Lassa/inmunología , Receptores KIR2DL2/metabolismo , Antígenos Virales/inmunología , Línea Celular , Citotoxicidad Inmunológica , Mapeo Epitopo/métodos , Genotipo , Antígenos HLA-C/genética , Humanos , Tolerancia Inmunológica , Inmunomodulación , Inmunofenotipificación , Fiebre de Lassa/genética , Fiebre de Lassa/virología , Unión Proteica , Receptores KIR2DL2/genéticaRESUMEN
The killer cell immunoglobulin-like receptor (KIR) KIR2DS2 induces natural killer (NK) cell activation upon ligation and in genetic studies is associated with protection against certain cancers and viral infections. One of the difficulties in understanding KIR2DS2 has been that ligands have been hard to define. In part, this is because the high sequence homology between KIR2DS2 and KIR2DL3/KIR2DL2 has made it difficult to make antibodies that specifically detect NK cells expressing KIR2DS2. Using transfected NK cell line (NKL) cells and primary human samples, we report the identification of a novel antibody combination which allows identification of NK cells with relatively high expression of KIR2DS2. This separation is sufficient to examine primary human NK cell activation in response to KIR2DS2 specific ligands.
Asunto(s)
Anticuerpos/metabolismo , Inmunofenotipificación/métodos , Células Asesinas Naturales/metabolismo , Neoplasias/inmunología , Receptores KIR/genética , Virosis/inmunología , Separación Celular , Células Cultivadas , Citometría de Flujo , Heterocigoto , Humanos , Vigilancia Inmunológica , Activación de Linfocitos , Receptores KIR/inmunología , Receptores KIR/metabolismo , Receptores KIR2DL2/genética , Receptores KIR2DL2/inmunología , Receptores KIR2DL2/metabolismo , Receptores KIR2DL3/genética , Receptores KIR2DL3/inmunología , Receptores KIR2DL3/metabolismoRESUMEN
Inhibitory KIRs play a central role in regulating NK cell activity. KIR2DL2/3 bind to HLA-C molecules, but the modulation of these interactions by viral infections and presentation of viral epitopes is not well-understood. We investigated whether the frequencies of KIR2DL2/3+ NK cells recognizing HLA-C*03:04/viral peptide complexes were impacted by YFV vaccination or HIV-1 and HCV infection. Ex vivo HLA class I tetramer staining of primary human NK cells derived from YFV-vaccinated individuals, or HIV-1- or HCV-infected individuals revealed that the YFV/HLA-C*03:04-NS2A4-13-tetramer bound to a larger proportion of KIR2DL2/3+ NK cells compared to HIV-1/HLA-C*03:04-Gag296-304- or HCV/HLA-C*03:04-Core136-144-tetramers. The YFV/HLA-C*03:04-NS2A4-13-tetramer also exhibited a stronger avidity to KIR2DL2/3 compared to the other tested tetramers. The proportional frequencies of KIR2DL2/3+ NK cells binding to the three tested HLA-C*03:04 tetramers were identical between YFV-vaccinated individuals or HIV-1- or HCV-infected individuals, and remained stable following YFV vaccination. These data demonstrate consistent hierarchies in the frequency of primary KIR2DL2/3+ NK cells binding HLA-C*03:04/peptide complexes that were determined by the HLA-C-presented peptide and not modulated by the underlying viral infection or vaccination.
Asunto(s)
Antígenos HLA-C/inmunología , Células Asesinas Naturales/inmunología , Células Asesinas Naturales/metabolismo , Péptidos/metabolismo , Receptores KIR2DL2/metabolismo , Receptores KIR2DL3/metabolismo , Citometría de Flujo , Antígenos HLA-C/química , Antígenos HLA-C/metabolismo , Humanos , Leucocitos Mononucleares/inmunología , Leucocitos Mononucleares/metabolismo , Complejos Multiproteicos , Péptidos/química , Unión Proteica , Vacunación , Vacunas/inmunologíaRESUMEN
Gamma delta (γδ) T cells, which possess potent cytotoxicity against a wide range of cancer cells, have become a potential avenue for adoptive immunotherapy. Decitabine (DAC) has been reported to enhance the immunogenicity of tumor cells, thereby reinstating endogenous immune recognition and tumor lysis. However, DAC has also been demonstrated to have direct effects on immune cells. In this study, we report that DAC inhibits γδ T cell proliferation. In addition, DAC increases the number of KIR2DL2/3-positive γδ T cells, which are less cytotoxic than the KIR2DL2/3-negative γδ T cells. We found that DAC upregulated KIR2DL2/3 expression in KIR2DL2/3-negative γδ T cells by inhibiting KIR2DL2/3 promoter methylation, which enhances the binding of KIR2DL2/3 promoter to Sp-1 and activates KIR2DL2/3 gene expression. Our data demonstrated that DAC can inhibit the function of human γδ T cells at both cellular and molecular levels, which confirms and extrapolates the results of previous studies showing that DAC can negatively regulate the function of NK cells and αß T cells of the immune system.
Asunto(s)
Antineoplásicos/farmacología , Decitabina/farmacología , Inmunoterapia Adoptiva/métodos , Leucemia Mieloide Aguda/terapia , Síndromes Mielodisplásicos/terapia , Receptores KIR2DL2/metabolismo , Linfocitos T/inmunología , Adulto , Anciano , Anciano de 80 o más Años , Antineoplásicos/uso terapéutico , Proliferación Celular/efectos de los fármacos , Células Cultivadas , Citotoxicidad Inmunológica , Metilación de ADN , Decitabina/uso terapéutico , Regulación de la Expresión Génica , Humanos , Leucemia Mieloide Aguda/inmunología , Masculino , Persona de Mediana Edad , Síndromes Mielodisplásicos/inmunología , Regiones Promotoras Genéticas/genética , Unión Proteica , Proteínas Quinasas/metabolismo , Receptores de Antígenos de Linfocitos T gamma-delta/metabolismo , Receptores KIR2DL2/genética , Linfocitos T/efectos de los fármacos , Linfocitos T/trasplanteRESUMEN
To exploit autologous NK cells for cancer immunotherapy, it is highly relevant to circumvent killer cell immunoglobulin-like receptor (KIR)-mediated self-inhibition of human NK cells by HLA-I-expressing tumor cells. Here, we show that stimulation of NK cells with IL-12/15/18 for two days led to downregulation of surface expression of the inhibitory KIR2DL2/L3, KIR2DL1 and KIR3DL1 receptors on peripheral blood NK cells. Downregulation of KIR expression was attributed to decreased KIR mRNA levels which could be re-induced already 3 days after re-culture in IL-2. Reduced KIR2DL2/L3 expression on IL-12/15/18-activated NK cells resulted in less inhibition upon antibody-mediated KIR engagement and increased CD16-dependent cytotoxicity in redirected lysis assays. Most importantly, downregulated KIR2DL2/L3 expression enabled enhanced cytotoxicity of IL-12/15/18-stimulated NK cells against tumor cells expressing cognate HLA-I molecules. NK cells pre-activated with IL-12/15/18 were previously shown to exert potent anti-tumor activity and memory-like long-lived functionality, mediating remission in a subset of acute myeloid leukemia (AML) patients in a clinical trial. Our study reveals a novel mechanism of IL-12/15/18 in improving the cytotoxicity of NK cells by reducing their sensitivity to inhibition by self-HLA-I due to decreased KIR expression, highlighting the potency of IL-12/15/18-activated NK cells for anti-tumor immunotherapy protocols.
Asunto(s)
Vacunas contra el Cáncer/inmunología , Inmunoterapia Adoptiva/métodos , Células Asesinas Naturales/inmunología , Leucemia Mieloide Aguda/terapia , Receptores KIR2DL2/metabolismo , Receptores KIR2DL3/metabolismo , Receptores KIR3DL1/metabolismo , Animales , Citotoxicidad Celular Dependiente de Anticuerpos , Línea Celular Tumoral , Regulación hacia Abajo , Antígenos HLA/metabolismo , Humanos , Interleucina-12/metabolismo , Interleucina-15/metabolismo , Interleucina-18/metabolismo , Células Asesinas Naturales/trasplante , Leucemia Mieloide Aguda/inmunología , Activación de Linfocitos , RatonesRESUMEN
The functions of activating members of the killer cell Ig-like receptor (KIR) family are not fully understood, as the ligands for these receptors are largely unidentified. In this study, we report that KIR2DS2 reporter cells recognize a ligand expressed by cancer cell lines. All cancer targets recognized by KIR2DS2 were also recognized by KIR2DL2 and KIR2DL3 reporters. Trogocytosis of membrane proteins from the cancer targets was observed with responding reporter cells, indicating the formation of KIR2DS2 ligand-specific immunological synapses. HLA-C typing of target cells showed that KIR2DS2 recognition was independent of the HLA C1 or C2 group, whereas targets cells that were only recognized by KIR2DL3 expressed C1 group alleles. Anti-HLA class I Abs blocked KIR2DL3 responses toward C1-expressing targets, but they did not block KIR2DS2 recognition of cancer cells. Small interfering RNA knockdown of ß2-microglobulin reduced the expression of class I H chain on the cancer targets by >97%, but it did not reduce the KIR2DS2 reporter responses, indicating a ß2-microglobulin-independent ligand for KIR2DS2. Importantly, KIR2DL3 responses toward some KIR2DS2 ligand-expressing cells were also undiminished after ß2-microglobulin knockdown, and they were not blocked by anti-HLA class I Abs, suggesting that KIR2DL3, in addition to the traditional HLA-C ligands, can bind to the same ß2-microglobulin-independent ligand as KIR2DS2. These observations indicate the existence of a novel, presently uncharacterized ligand for the activating NK cell receptor KIR2DS2. Molecular identification of this ligand may lead to improved KIR-HLA mismatching in hematopoietic stem cell transplantation therapy for leukemia and new, more specific NK cell-based cancer therapies.
Asunto(s)
Neoplasias/metabolismo , Receptores KIR2DL2/metabolismo , Línea Celular Tumoral , Separación Celular , Citometría de Flujo , Técnicas de Silenciamiento del Gen , Humanos , LigandosRESUMEN
Natural killer (NK) cells control viral infection in part through the interaction between killer cell immunoglobulin-like receptors (KIRs) and their human leukocyte antigen (HLA) ligands. We investigated 504 anti-retroviral (ART)-free Japanese patients chronically infected with HIV-1 and identified two KIR/HLA combinations, KIR2DL2/HLA-C∗12:02 and KIR2DL2/HLA-C∗14:03, that impact suppression of HIV-1 replication. KIR2DL2+ NK cells suppressed viral replication in HLA-C∗14:03+ or HLA-C∗12:02+ cells to a significantly greater extent than did KIR2DL2- NK cells in vitro. Functional analysis showed that the binding between HIV-1-derived peptide and HLA-C∗14:03 or HLA-C∗12:02 influenced KIR2DL2+ NK cell activity through reduced expression of the peptide-HLA (pHLA) complex on the cell surface (i.e., reduced KIR2DL2 ligand expression), rather than through reduced binding affinity of KIR2DL2 to the respective pHLA complexes. Thus, KIR2DL2/HLA-C∗12:02 and KIR2DL2/HLA-C∗14:03 compound genotypes have protective effects on control of HIV-1 through a mechanism involving KIR2DL2-mediated NK cell recognition of virus-infected cells, providing additional understanding of NK cells in HIV-1 infection.
Asunto(s)
Infecciones por VIH/inmunología , Infecciones por VIH/virología , VIH-1/fisiología , Antígenos HLA-C/metabolismo , Células Asesinas Naturales/inmunología , Receptores KIR2DL2/metabolismo , Alelos , Pueblo Asiatico , Enfermedad Crónica , Infecciones por VIH/sangre , VIH-1/inmunología , Humanos , Carga Viral , Replicación ViralRESUMEN
We have previously demonstrated that multiple sclerosis (MS) patients with KIR2DL2 expression on Natural killer (NK) cells are more susceptible to herpes simplex virus 1 (HSV-1) infection. We explored cytokine expression by NK cells during HSV-1 infection in association with KIR2DL2 expression. MS KIR2DL2(+) NK cells failed to control HSV-1 infection and secreted high levels of Th17 cytokines, while MS KIR2DL2(-) NK cells released Th1 cytokines, mainly IFN-gamma. Our data showed, for the first time, a peculiar Th17 cytokine secretion by MS KIR2DL2(+) NK cells in the presence of HSV-1 infection, that could be implicated in MS pathogenesis.
Asunto(s)
Citocinas/metabolismo , Herpes Simple , Células Asesinas Naturales/metabolismo , Esclerosis Múltiple Recurrente-Remitente/patología , Receptores KIR2DL2/metabolismo , Adulto , Células Cultivadas , Femenino , Humanos , Células Asesinas Naturales/virología , Activación de Linfocitos , Masculino , Persona de Mediana Edad , Esclerosis Múltiple Recurrente-Remitente/virología , ARN Mensajero/metabolismo , Receptores KIR2DL2/genética , Células Th17/metabolismo , Carga Viral/métodosRESUMEN
BACKGROUND: KIR3DL2, an inhibitory receptor expressed by natural killer cells and a subset of normal CD8(+) T cells, is aberrantly expressed in neoplastic cells in transformed mycosis fungoides and Sézary syndrome. Anti-KIR3DL2 targeted antibody therapy has shown potent activity in preclinical models for these diseases. OBJECTIVES: To examine the expression of KIR3DL2 and its potential use as a therapeutic target in patients with primary cutaneous anaplastic large-cell lymphoma (pcALCL), the most aggressive cutaneous CD30(+) lymphoproliferative disease. METHODS: Samples from 11 patients with pcALCL and three CD30(+) lymphoproliferative disease cell lines - Mac1, Mac2a and Mac2b - were used in KIR3DL2 expression studies using immunohistochemistry, flow cytometry and reverse-transcriptase quantitative polymerase chain reaction. The effect of IPH4102, a monoclonal humanized IgG1 targeting KIR3DL2, was assessed by in vitro cytotoxicity assays against Mac1, Mac2a and Mac2b using allogeneic peripheral blood mononuclear cells as effectors. RESULTS: KIR3DL2 mRNA and protein were found in all human samples of pcALCL, and in the Mac2a and Mac2b cell lines. KIR3DL2 protein expression was present on 85·8 ± 14·0% of CD30(+) skin-infiltrating tumour cells. In vitro functional studies showed that KIR3DL2(+) Mac2a and Mac2b pcALCL lines are sensitive to antibody-derived cytotoxicity mediated by IPH4102, through activation of natural killer cells, in a concentration-dependent manner. CONCLUSIONS: pcALCL tumour cells express KIR3DL2, and we provide preclinical proof of concept for the use of IPH4102, a humanized anti-KIR3DL2 antibody, to treat patients with primary cutaneous CD30(+) ALCL.
Asunto(s)
Linfoma Anaplásico de Células Grandes/tratamiento farmacológico , Receptores KIR2DL2/antagonistas & inhibidores , Neoplasias Cutáneas/tratamiento farmacológico , Adolescente , Adulto , Anciano , Anticuerpos Monoclonales/farmacología , Antineoplásicos/farmacología , Línea Celular Tumoral , Femenino , Humanos , Antígeno Ki-1/metabolismo , Células Asesinas Naturales/fisiología , Leucocitos Mononucleares/metabolismo , Masculino , Persona de Mediana Edad , Receptores KIR2DL2/inmunología , Receptores KIR2DL2/metabolismo , Piel/metabolismo , Células Tumorales Cultivadas , Adulto JovenRESUMEN
AIM: Killer immunoglobulin-like receptors (KIRs) and their interaction with HLA class I ligands have been shown to be associated with Type 1 diabetes mellitus. The aim of our study was to investigate the influence of KIR genes and their HLA-C ligands for susceptibility to Type 1 diabetes in patients from Eastern India. METHODS: A total of 135 patients with Type 1 diabetes and 98 healthy subjects from Eastern India were typed for KIR genes and HLA-C ligands using PCR-based genotyping. The frequencies of these genes were compared between patients and controls. RESULTS: Comparison of KIR genes between Type 1 diabetes patients and healthy subjects revealed significantly different frequencies of KIRs 2DL2 and 2DS4. The presence of HLA-C1 was negatively associated with disease. The presence of both HLA-C1 and -C2 showed a negative association with Type 1 diabetes, whereas the absence of C1 and presence of C2 was positively associated with disease. Stratification analysis of HLA-C ligands and KIRs showed significant associations between Type 1 diabetes and 2DL2+/C1-, 2DL2-/C1+, 2DL3+/C1+, 2DL3+/C1- and 2DS2+/C1-. CONCLUSIONS: Our results suggest that the interaction of KIRs with HLA-C ligands are significant and certain combinations contribute to susceptibility to and protection against Type 1 diabetes.
Asunto(s)
Diabetes Mellitus Tipo 1/genética , Predisposición Genética a la Enfermedad , Antígenos HLA-C/sangre , Células T Asesinas Naturales/metabolismo , Polimorfismo Genético , Receptores KIR2DL2/genética , Receptores KIR2DL3/genética , Receptores KIR/genética , Alelos , Diabetes Mellitus Tipo 1/sangre , Diabetes Mellitus Tipo 1/inmunología , Diabetes Mellitus Tipo 1/metabolismo , Susceptibilidad a Enfermedades , Regulación de la Expresión Génica , Frecuencia de los Genes , Estudios de Asociación Genética , Antígenos HLA-C/genética , Antígenos HLA-C/metabolismo , Humanos , India , Ligandos , Células T Asesinas Naturales/inmunología , Receptores KIR/agonistas , Receptores KIR/sangre , Receptores KIR/metabolismo , Receptores KIR2DL2/agonistas , Receptores KIR2DL2/sangre , Receptores KIR2DL2/metabolismo , Receptores KIR2DL3/agonistas , Receptores KIR2DL3/sangre , Receptores KIR2DL3/metabolismoAsunto(s)
Antígenos CD/metabolismo , Linfocitos T CD4-Positivos/metabolismo , Granulomatosis con Poliangitis/metabolismo , Subfamília C de Receptores Similares a Lectina de Células NK/metabolismo , Subfamilia K de Receptores Similares a Lectina de Células NK/metabolismo , Receptores Inmunológicos/metabolismo , Receptores KIR2DL2/metabolismo , Receptores KIR2DL3/metabolismo , Adulto , Anciano , Estudios de Casos y Controles , Femenino , Humanos , Receptor Leucocitario Tipo Inmunoglobulina B1 , Masculino , Persona de Mediana Edad , Receptores de Células Asesinas Naturales/metabolismoRESUMEN
The diverse aspects of cutaneous T-cell lymphomas may impede the diagnosis of Sézary syndrome (SS) and mycosis fungoides (MF), in particular, at early stages of the disease. We defined the CD158k/KIR3DL2 molecule as a first positive cell surface marker for Sézary cells (SCs). Here, we designed an optimized flow cytometry gating strategy, allowing the definition of lymphocytes of different sizes and defects of cell surface markers. Quantification by cytomorphology, flow cytometry, or clonal evaluation, gave similar results at initial time points and during the evolution in a prospective study involving 64 consecutive cutaneous T-cell lymphoma or erythrodermic patients. We found that CD158k+ T cells and circulating CD4+ T cells from MF patients exhibited unexpected patterns of cell surface expression with a marked heterogeneity of circulating lymphocytes even at initial diagnosis. Taken together, our results show that a multistep gating of CD158k+ cells is reliable to assess tumor burden in case of SS and suggest that both circulating MF CD4+ T cells and CD158k+ T cells are not homogeneous distinct memory populations. Further phenotypic and functional characterizations of such subsets are needed to better understand the underlying molecular mechanisms leading to the development of these diseases.
Asunto(s)
Biomarcadores de Tumor/metabolismo , Linfoma Cutáneo de Células T/metabolismo , Micosis Fungoide/metabolismo , Células Neoplásicas Circulantes/metabolismo , Receptores KIR2DL2/metabolismo , Síndrome de Sézary/diagnóstico , Anciano , Anciano de 80 o más Años , Anticuerpos Monoclonales/inmunología , Anticuerpos Monoclonales/farmacología , Biomarcadores de Tumor/inmunología , Progresión de la Enfermedad , Estudios de Seguimiento , Heterogeneidad Genética , Humanos , Memoria Inmunológica , Estudios Longitudinales , Subgrupos Linfocitarios/metabolismo , Persona de Mediana Edad , Estudios Prospectivos , Receptores KIR2DL2/inmunología , Síndrome de Sézary/metabolismoRESUMEN
An important checkpoint in the progression of melanoma is the metastasis to lymph nodes. Here, to investigate the role of lymph node NK cells in disease progression, we analyze frequency, phenotype and functions of NK cells from tumour-infiltrated (TILN) and tumour-free ipsilateral lymph nodes (TFLN) of the same patients. We show an expansion of CD56(dim)CD57(dim)CD69 + CCR7 +KIR+ NK cells in TILN. TILN NK cells display robust cytotoxic activity against autologous melanoma cells. In the blood of metastatic melanoma patients, the frequency of NK cells expressing the receptors for CXCL8 receptor is increased compared with healthy subjects, and blood NK cells also express the receptors for CCL2 and IL-6. These factors are produced in high amount in TILN and in vitro switch the phenotype of blood NK cells from healthy donors to the phenotype associated with TILN. Our data suggest that the microenvironment of TILN generates and/or recruits a particularly effective NK cell subset.
Asunto(s)
Células Asesinas Naturales/inmunología , Ganglios Linfáticos/inmunología , Melanoma/inmunología , Neoplasias Cutáneas/inmunología , Adulto , Anciano , Antígenos CD/metabolismo , Antígenos de Diferenciación de Linfocitos T/metabolismo , Antígeno CD56/metabolismo , Antígenos CD57/metabolismo , Femenino , Humanos , Células Asesinas Naturales/metabolismo , Lectinas Tipo C/metabolismo , Ganglios Linfáticos/patología , Metástasis Linfática , Masculino , Melanoma/patología , Persona de Mediana Edad , Receptores CCR7/metabolismo , Receptores de Interleucina-6/metabolismo , Receptores KIR2DL2/metabolismo , Receptores KIR2DL3/metabolismo , Neoplasias Cutáneas/patología , Microambiente Tumoral/inmunologíaRESUMEN
OBJECTIVE: The aim of this study was to assess the consequence of sequence variations in HLA-C03:04-presented HIV-1 p24 Gag epitopes on binding of the inhibitory natural killer (NK) cell receptor KIR2DL2 to HLA-C03:04. DESIGN: HIV-1 may possibly evade recognition by KIR+ NK cells through selection of sequence variants that interfere with the interactions of inhibitory killer cell immunoglobulin-like receptors (KIRs) and their target ligands on HIV-1 infected cells. KIR2DL2 is an inhibitory NK cell receptor that binds to a family of HLA-C ligands. Here, we investigated whether HIV-1 encodes for HLA-C03:04-restricted epitopes that alter KIR2DL2 binding. METHODS: Tapasin-deficient 721.220 cells expressing HLA-C03:04 were pulsed with overlapping peptides (10mers overlapped by nine amino acids, spanning the entire HIV-1 p24 Gag sequence) to identify peptides that stabilized HLA-C expression. The impact that sequence variation in HLA-C03:04-binding HIV-1 epitopes has on KIR2DL2 binding and KIR2DL2+ NK cell function was determined using KIR2DL2-Fc constructs and NK cell degranulation assays. RESULTS: Several novel HLA-C03:04 binding epitopes were identified within the HIV-1 p24 Gag consensus sequence. Three of these consensus sequence peptides (Gag144-152, Gag163-171 and Gag295-304) enabled binding of KIR2DL2 to HLA-C03:04 and resulted in inhibition of KIR2DL2+ primary NK cells. Furthermore, naturally occurring minor variants of epitope Gag295-304 enhanced KIR2DL2 binding to HLA-C03:04. CONCLUSION: Our data show that naturally occurring sequence variations within HLA-C03:04-restricted HIV-1 p24 Gag epitopes can have a significant impact on the binding of inhibitory KIR receptors and primary NK cell function.
Asunto(s)
Epítopos/genética , Variación Genética , Proteína p24 del Núcleo del VIH/genética , Antígenos HLA-C/metabolismo , Células Asesinas Naturales/inmunología , Receptores KIR2DL2/metabolismo , Degranulación de la Célula , Línea Celular , Humanos , Unión ProteicaRESUMEN
BACKGROUND: Killer cell immunoglobulin-like receptors (KIR) contribute to the pathogenesis of multiple auto-immune diseases via the modulation of NK-, NKT- and T-cells. Thus, we want to know whether the expression pattern of KIR is associated with systemic lupus erythematosus (SLE) susceptibility. METHODS: Here, real-time quantitative PCR and fluorescence-activated cell sorting (FACS) were used to measure the stimulatory KIR (sKIR) and inhibitory KIR (iKIR) mRAN and protein levels on NK-, NKT- and T-cells in both SLE patients and healthy controls. RESULTS: In SLE patients, CD158a/h (KIR2DL1/S1) was highly expressed while CD158b/i/j (KIR2DL2/L3/S2, iKIR/iKIR/sKIR) was lowly expressed in NK- and NKT-cells in patients. The expression levels of KIR2DL1 and KIR2DL2 (iKIRs) were decreased while the expression levels of KIR2DS1 (sKIR) were increased in NK- and NKT-cells in the patients. CONCLUSIONS: We found that SLE patients represent aberrant expression of stimulatory and inhibitory KIRs in NK- and NKT-cells. Consequently, these different expression levels of KIRs may contribute to the abnormal function of these cells, which lead to the risk of SLE.