RESUMO
Chimeric antigen receptors (CARs) are fusion proteins with an extracellular antigen recognition domain and numerous intracellular signaling domains that have been genetically modified. CAR-engineered T lymphocyte-based therapies have shown great success against blood cancers; however, potential fatal toxicity, such as in cytokine release syndrome, and high costs are some shortcomings that limit the clinical application of CAR-engineered T lymphocytes and remain to overcome. Natural killer (NK) cells are the focal point of current immunological research owing to their receptors that prove to be promising immunotherapeutic candidates for treating cancer. However, to date, manipulation of NK cells to treat malignancies has been moderately successful. Recent progress in the biology of NK cell receptors has greatly transformed our understanding of how NK cells recognize and kill tumor and infected cells. CAR-NK cells may serve as an alternative candidate for retargeting cancer because of their unique recognition mechanisms, powerful cytotoxic effects especially on cancer cells in both CAR-dependent and CAR-independent manners and clinical safety. Moreover, NK cells can serve as an 'off-the-shelf product' because NK cells from allogeneic sources can also be used in immunotherapies owing to their reduced risk of alloreactivity. Although ongoing fundamental research is in the beginning stages, this review provides an overview of recent developments implemented to design CAR constructs to stimulate NK activation and manipulate NK receptors for improving the efficiency of immunotherapy against cancer, summarizes the preclinical and clinical advances of CAR-NK cells against both hematological malignancies and solid tumors and confronts current challenges and obstacles of their applications. In addition, this review provides insights into prospective novel approaches that further enhance the efficiency of CAR-NK therapies and highlights potential questions that require to be addressed in the future.
Assuntos
Imunoterapia Adotiva/métodos , Células Matadoras Naturais/imunologia , Receptores de Antígenos Quiméricos/imunologia , Receptores de Células Matadoras Naturais/imunologia , Citotoxicidade Celular Dependente de Anticorpos , Apoptose , Ensaios Clínicos como Assunto , Citocinas/fisiologia , Citotoxicidade Imunológica , Desenho de Fármacos , Proteína Ligante Fas/fisiologia , Previsões , Proteínas Ligadas por GPI/fisiologia , Antígenos HLA/imunologia , Humanos , Células Matadoras Naturais/química , Células Matadoras Naturais/transplante , Lentivirus/genética , Ligantes , Macrófagos/imunologia , Subfamília K de Receptores Semelhantes a Lectina de Células NK/fisiologia , Neoplasias/terapia , Receptores de Antígenos Quiméricos/genética , Receptores de IgG/fisiologia , Receptores de Células Matadoras Naturais/classificação , Tolerância a Antígenos Próprios , Subpopulações de Linfócitos T/imunologia , Transdução Genética , Microambiente Tumoral , Receptor fas/fisiologiaRESUMO
Melanoma is the most aggressive and deadliest form of skin cancer, and its prognosis is very poor. Although the early detection is responsive to many treatments, metastatic melanoma is refractory to most of them. In the United States, skin melanoma is the fifth most common type of cancer in men and the sixth in women. Current treatment modalities, depending on the cancer stage, consist primarily of surgical excision, chemotherapy, adjuvant therapy, targeted therapies, and immunotherapy. Despite the wide range of therapeutic options and the steadily increasing response rates, a large subset of the treated patients relapse and develop resistance to further treatments. One novel approach in preclinical and clinical trials in immunotherapy is the adaptation of natural killer (NK) cells against resistant cancer cells. NK cells can kill a variety of cancer cell types, as well as the cancer stem cells, while leaving normal cells intact. In skin melanoma, as in most cancers, NK cells in the tumor microenvironment (TME) are functionally impaired. Several factors underlie the defective cause of NK cells, one of which is the dysregulation of the activating receptor NKG2D. This is the dominant receptor in regulating the cytotoxic activity, cytokine production, and regulation of other receptors expressed on NK cells and other lymphocytes. The defective NK cells in cancer models were associated with tumor growth and metastasis. In this review, we discuss the role of NK cells and their phenotypic variants in skin melanoma. Using bioinformatics, we have further analyzed the expression of NKG2D, confirming its low transcript levels in patients with skin melanoma. Furthermore, we show that the CD133 subset of cancer stem cells expresses low levels of NKG2D. Based on these findings we discuss the potential therapeutic approaches that can be exploited to upregulate NKG2D in patients' NK cells and restore their anti-melanoma effects, resulting in tumor regression and prolonged survival.
Assuntos
Imunoterapia , Melanoma , Subfamília K de Receptores Semelhantes a Lectina de Células NK/fisiologia , Neoplasias Cutâneas , Humanos , Células Matadoras Naturais , Melanoma/terapia , Neoplasias Cutâneas/terapia , Microambiente TumoralRESUMO
BACKGROUND: Myeloid leukemia represents a heterogeneous group of cancers of blood and bone marrow which arise from clonal expansion of hematopoietic myeloid lineage cells. Acute myeloid leukemia (AML) has traditionally been treated with multi-agent chemotherapy, but conventional therapies have not improved the long-term survival for decades. Chronic myeloid leukemia (CML) is an indolent disease which requires lifelong treatment, is associated with significant side effects, and carries a risk of progression to potentially lethal blast crises. RECENT FINDINGS: Recent advances in molecular biology, virology, and immunology have enabled researchers to grow and modify T lymphocytes ex-vivo. Chimeric antigen receptor (CAR) T-cell therapy has been shown to specifically target cells of lymphoid lineage and induce remission in acute lymphoblastic leukemia (ALL) patients. While the success of CAR T-cells against ALL is considered a defining moment in modern oncology, similar efficacy against myeloid leukemia cells remains elusive. Over the past 10 years, numerous CAR T-cells have been developed that can target novel myeloid antigens, and many clinical trials are finally starting to yield encouraging results. In this review, we present the recent advances in this field and discuss strategies for future development of myeloid targeting CAR T-cell therapy. CONCLUSIONS: The field of CAR T-cell therapy has rapidly evolved over the past few years. It represents a radically new approach towards cancers, and with continued refinement it may become a viable therapeutic option for patients of acute and chronic myeloid leukemia.
Assuntos
Imunoterapia Adotiva/métodos , Leucemia Mielogênica Crônica BCR-ABL Positiva/terapia , Leucemia Mieloide Aguda/terapia , Receptores de Antígenos Quiméricos/imunologia , Humanos , Subunidade alfa de Receptor de Interleucina-3/fisiologia , Subfamília K de Receptores Semelhantes a Lectina de Células NK/fisiologia , Lectina 3 Semelhante a Ig de Ligação ao Ácido Siálico/fisiologiaRESUMO
Hepatocellular carcinoma is a common malignant tumor with high mortality. Its malignant proliferation, invasion, and metastasis are closely related to the cellular immune function of the patients. NKG2D is a key activated and type II membrane protein molecule expressed on the surface of almost all NK cells. The human NKG2D gene is 270 kb long, located at 12p12.3-p13.1, and contains 10 exons and 9 introns. The three-dimensional structure of the NKG2D monomeric protein contains two alpha-helices, two beta-lamellae, and four disulfide bonds, and its' signal of activation is transmitted mainly by the adaptor protein (DAP). NKG2D ligands, including MICA, MICB, and ULBPs, can be widely expressed in hepatoma cells. After a combination of NKG2D and DAP10 in the form of homologous two polymers, the YxxM motif in the cytoplasm is phosphorylated and then signaling pathways are also gradually activated, such as PI3K, PLCγ2, JNK-cJunN, and others. Activated NK cells can enhance the sensitivity to hepatoma cells and specifically dissolve by releasing a variety of cytokines (TNF-α and IFN-γ), perforin, and high expression of FasL, CD16, and TRAIL. NK cells may specifically bind to the over-expressed MICA, MICB, and ULBPs of hepatocellular carcinoma cells through the surface activating receptor NKG2D, which can help to accurately identify hepatoma, play a critical role in anti-hepatoma via the pathway of cytotoxic effects, and obviously delay the poor progress of hepatocellular carcinoma.
Assuntos
Carcinoma Hepatocelular/metabolismo , Subfamília K de Receptores Semelhantes a Lectina de Células NK/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Citocinas/metabolismo , Humanos , Células Matadoras Naturais/metabolismo , Ligantes , Neoplasias Hepáticas/metabolismo , Subfamília K de Receptores Semelhantes a Lectina de Células NK/fisiologia , Transdução de Sinais/imunologiaRESUMO
BACKGROUND: Studies have shown that CD4CD25Foxp3Treg cells suppress NKG2D expression on NK cells via a cell contact-dependent mechanism and increased TGF-ß and IL-10 production in some cancer models. We herein aimed to explore whether CD4CD25Foxp3Tregs suppress NKG2D-mediated NK cell cytotoxicity in peripheral blood and elucidate the exact mechanism underlying this phenomenon. METHODS: To explore the function of NKG2D, NK cell cultures were treated with an NKG2D-blocking antibody to block these receptors. Additionally, TGF-ß- and IL-10-blocking antibodies were added to NK and CD4CD25Foxp3Treg cell cocultures to evaluate whether the latter cells suppress NKG2D expression of NK cells via increasing the production of TGF-ß and IL-10. The expression of NKG2D on NK cells was detected by 3-color flow cytometry, and NK cell activity was assessed by 3 assays: a nonradioactive cytotoxicity assay, an ELISA measuring IFN-γ production and a flow cytometry assay to evaluate CD107a expression. RESULTS: Blocking NKG2D decreased NK cell cytotoxicity, IFN-γ production and CD107a expression. Moreover, blocking TGF-ß and IL-10 substantially increased the NKG2D expression in NK and CD4CD25Foxp3Treg cell cocultures. Similarly, blocking TGF-ß and IL-10 enhanced NK cell cytotoxicity, IFN-γ production and CD107a expression; Transwell insert assays also revealed increased IFN-γ production and CD107a and NKG2D expression. CONCLUSION: CD4CD25Foxp3Tregs suppress NKG2D-mediated NK cell cytotoxicity in peripheral blood via a cell contact-dependent mechanism and increased TGF-ß and IL-10 production.
Assuntos
Fatores de Transcrição Forkhead/imunologia , Subunidade alfa de Receptor de Interleucina-2/imunologia , Células Matadoras Naturais/imunologia , Subfamília K de Receptores Semelhantes a Lectina de Células NK/fisiologia , Linfócitos T Reguladores/imunologia , Animais , Citotoxicidade Imunológica , Humanos , Ativação Linfocitária , CamundongosRESUMO
Graft-versus-host disease (GVHD) is a major barrier to the widespread use of allogeneic hematopoietic stem cell transplantation (allo-HSCT) for treating hematologic malignancies. Myeloid-derived suppressor cells (MDSCs) have been recognized as crucial immunosuppressive cells in various pathologic settings. Here, we investigated whether the unique functional properties of MDSCs could be harnessed to control allo-HSCT-associated GVHD. Using multiple murine GVHD/GVL models including both MHC-mismatched and miHA-mismatched, we demonstrated that treatment with CD115+ MDSCs efficiently suppressed GVHD but did not significantly impair graft-versus-leukemia (GVL) activity, leading to 80 and 67% protection in treated mice in GVHD and GVL models, respectively. The mechanism for this dissociation of GVHD from GVL, specifically the emergence of donor-derived NKG2D+ CD8 T cells with a memory phenotype in MDSC-treated recipient mice, was identified. NKG2D expression on donor T cells was required for eradication of allogeneic lymphoma cells. Furthermore, long-term surviving MDSC recipients that exhibited cytolytic activities against allogeneic leukemia cells had a significantly increased percentage of T regulatory cells and, more importantly, NKG2D+ CD8 T cells. These findings indicate that MDSCs can be used as a novel cell-based therapy to suppress GVHD while maintaining GVL activities through selective induction of NKG2D+ CD8 memory T cells.
Assuntos
Doença Enxerto-Hospedeiro/prevenção & controle , Efeito Enxerto vs Leucemia , Células Supressoras Mieloides/fisiologia , Animais , Complexo CD3/fisiologia , Citotoxicidade Imunológica , Feminino , Transplante de Células-Tronco Hematopoéticas , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Subfamília K de Receptores Semelhantes a Lectina de Células NK/fisiologia , Linfócitos T/imunologia , Transplante HomólogoRESUMO
It has already been shown that neutralization of the activating NK cell receptor NKG2D in combination with co-stimulation blockade prolongs graft survival of vascularized transplants. In order to clarify the underlying cellular mechanisms, we transplanted complete MHC-disparate BALB/c-derived cardiac grafts into C57BL/6 wildtypes or mice deficient for NKG2D (Klrk1-/- ). Although median survival was 8 days for both recipient groups, we detected already at day 5 posttransplantation significantly greater intragraft frequencies of NKp46+ NK cells in Klrk1-/- recipients than in wildtypes. This was followed by a significantly greater infiltration of CD4+ , but a lesser infiltration of CD8+ T cell frequencies. Contrary to published observations, co-stimulation blockade with CTLA4-Ig resulted in a significant acceleration of cardiac rejection by Klrk1-/- recipients, and this result was confirmed by applying a neutralizing antibody against NKG2D to wildtypes. In both experimental setups, grafts derived from Klrk1-/- recipients were characterized by significantly higher levels of interferon-γ mRNA, and both CD4+ and CD8+ T cells displayed a greater capacity for degranulation and interferon-γ production. In summary, our results clearly illustrate that NKG2D expression in the recipient is important for cardiac allograft survival, thus supporting the hypothesis that impairment of NK cells prevents the establishment of graft acceptance.
Assuntos
Linfócitos T CD4-Positivos/imunologia , Rejeição de Enxerto/etiologia , Sobrevivência de Enxerto/imunologia , Transplante de Coração/efeitos adversos , Subfamília K de Receptores Semelhantes a Lectina de Células NK/fisiologia , Animais , Rejeição de Enxerto/metabolismo , Rejeição de Enxerto/patologia , Interferon gama/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Knockout , Complicações Pós-Operatórias , Taxa de Sobrevida , Transplante HomólogoRESUMO
Endoplasmic reticulum (ER) stress is commonly observed in intestinal epithelial cells (IECs) and can, if excessive, cause spontaneous intestinal inflammation as shown by mice with IEC-specific deletion of X-box-binding protein 1 (Xbp1), an unfolded protein response-related transcription factor. In this study, Xbp1 deletion in the epithelium (Xbp1ΔIEC ) is shown to cause increased expression of natural killer group 2 member D (NKG2D) ligand (NKG2DL) mouse UL16-binding protein (ULBP)-like transcript 1 and its human orthologue cytomegalovirus ULBP via ER stress-related transcription factor C/EBP homology protein. Increased NKG2DL expression on mouse IECs is associated with increased numbers of intraepithelial NKG2D-expressing group 1 innate lymphoid cells (ILCs; NK cells or ILC1). Blockade of NKG2D suppresses cytolysis against ER-stressed epithelial cells in vitro and spontaneous enteritis in vivo. Pharmacological depletion of NK1.1+ cells also significantly improved enteritis, whereas enteritis was not ameliorated in Recombinase activating gene 1-/-;Xbp1ΔIEC mice. These experiments reveal innate immune sensing of ER stress in IECs as an important mechanism of intestinal inflammation.
Assuntos
Proteínas de Transporte/fisiologia , Retículo Endoplasmático/fisiologia , Antígenos de Histocompatibilidade Classe I/fisiologia , Inflamação/fisiopatologia , Mucosa Intestinal/fisiopatologia , Subfamília K de Receptores Semelhantes a Lectina de Células NK/fisiologia , Animais , Proteínas de Transporte/metabolismo , Retículo Endoplasmático/metabolismo , Enterite/etiologia , Enterite/metabolismo , Enterite/fisiopatologia , Deleção de Genes , Antígenos de Histocompatibilidade Classe I/metabolismo , Inflamação/metabolismo , Mucosa Intestinal/metabolismo , Proteínas de Membrana , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Transgênicos , Estresse Fisiológico/fisiologia , Regulação para Cima , Proteína 1 de Ligação a X-Box/fisiologiaRESUMO
Mice housed in an enriched environment display a tumor-resistant phenotype due to eustress stimulation. However, the mechanisms underlying enriched environment-induced protection against cancers remain largely unexplained. In this study, we observed a significant antitumor effect induced by enriched environment in murine pancreatic cancer and lung cancer models. This effect remained intact in T/B lymphocyte-deficient Rag1-/- mice, but was nearly eliminated in natural killer (NK) cell-deficient Beige mice or in antibody-mediated NK-cell-depleted mice, suggesting a predominant role of NK cells in enriched environment-induced tumor inhibition. Exposure to enriched environment enhanced NK-cell activity against tumors and promoted tumoral infiltration of NK cells. Enriched environment increased the expression levels of CCR5 and NKG2D (KLRK1) in NK cells; blocking their function effectively blunted the enriched environment-induced enhancement of tumoral infiltration and cytotoxic activity of NK cells. Moreover, blockade of ß-adrenergic signaling or chemical sympathectomy abolished the effects of enriched environment on NK cells and attenuated the antitumor effect of enriched environment. Taken together, our results provide new insight into the mechanism by which eustress exerts a beneficial effect against cancer. Cancer Res; 77(7); 1611-22. ©2017 AACR.
Assuntos
Citotoxicidade Imunológica , Abrigo para Animais , Células Matadoras Naturais/imunologia , Subfamília K de Receptores Semelhantes a Lectina de Células NK/fisiologia , Receptores CCR5/fisiologia , Sistema Nervoso Simpático/fisiologia , Animais , Linhagem Celular Tumoral , Meio Ambiente , Leptina/fisiologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Neoplasias/terapiaRESUMO
NK cells, which are highly enriched in the liver, are potent regulators of antiviral T cells and immunopathology in persistent viral infection. We investigated the role of the NKG2D axis in T cell/NK cell interactions in hepatitis B. Activated and hepatitis B virus (HBV)-specific T cells, particularly the CD4 fraction, expressed NKG2D ligands (NKG2DL), which were not found on T cells from healthy controls (p < 0.001). NKG2DL-expressing T cells were strikingly enriched within HBV-infected livers compared with the periphery or to healthy livers (p < 0.001). NKG2D+NK cells were also increased and preferentially activated in the HBV-infected liver (p < 0.001), in direct proportion to the percentage of MICA/B-expressing CD4 T cells colocated within freshly isolated liver tissue (p < 0.001). This suggests that NKG2DL induced on T cells within a diseased organ can calibrate NKG2D-dependent activation of local NK cells; furthermore, NKG2D blockade could rescue HBV-specific and MICA/B-expressing T cells from HBV-infected livers. To our knowledge, this is the first ex vivo demonstration that non-virally infected human T cells can express NKG2DL, with implications for stress surveillance by the large number of NKG2D-expressing NK cells sequestered in the liver.
Assuntos
Linfócitos T CD4-Positivos/imunologia , Hepatite B Crônica/imunologia , Células Matadoras Naturais/imunologia , Fígado/imunologia , Subfamília K de Receptores Semelhantes a Lectina de Células NK/fisiologia , Adulto , Comunicação Celular , Células Cultivadas , Feminino , Humanos , Ligantes , Fígado/virologia , Ativação Linfocitária , Masculino , Subfamília K de Receptores Semelhantes a Lectina de Células NK/antagonistas & inibidoresRESUMO
BACKGROUND: Neuroblastoma (NB) is the most common solid extracranial tumor in childhood. Despite advances in therapy, the prognosis is poor and optimized therapies are urgently needed. Therefore, we investigated the antitumor potential of interleukin-15 (IL-15)-activated cytokine-induced killer (CIK) cells against different NB cell lines. PROCEDURE: CIK cells were generated from peripheral blood mononuclear cells by the stimulation with interferon-γ (IFN-γ), IL-2, OKT-3 and IL-15 over a period of 10-12 days. The cytotoxic activity against NB cells was analyzed by nonradioactive Europium release assay before and after blocking of different receptor-ligand interactions relevant in CIK cell-mediated cytotoxicity. RESULTS: The final CIK cell products consisted in median of 83% (range: 75.9-91.9%) CD3+ CD56- T cells, 14% (range: 5.2-20.7%) CD3+ CD56+ NK-like T cells and 2% (range: 0.9-4.8%) CD3- CD56+ NK cells. CIK cells expanded significantly upon ex vivo stimulation with median rates of 22.3-fold for T cells, 58.3-fold for NK-like T cells and 2.5-fold for NK cells. Interestingly, CD25 surface expression increased from less than equal to 1% up to median 79.7%. Cytotoxic activity of CIK cells against NB cells was in median 34.7, 25.9 and 34.8% against the cell lines UKF-NB-3, UKF-NB-4 and SK-N-SH, respectively. In comparison with IL-2-stimulated NK cells, CIK cells showed a significantly higher cytotoxicity. Antibody-mediated blocking of the receptors NKG2D, TRAIL, FasL, DNAM-1, NKp30 and lymphocyte function-associated antigen-1 (LFA-1) significantly reduced lytic activity, indicating that diverse cytotoxic mechanisms might be involved in CIK cell-mediated NB killing. CONCLUSIONS: Unlike the mechanism reported in other malignancies, NKG2D-mediated cytotoxicity does not constitute the major killing mechanism of CIK cells against NB.
Assuntos
Células Matadoras Induzidas por Citocinas/imunologia , Interleucina-15/farmacologia , Neuroblastoma/terapia , Linhagem Celular Tumoral , Citotoxicidade Imunológica , Humanos , Antígeno-1 Associado à Função Linfocitária/fisiologia , Subfamília K de Receptores Semelhantes a Lectina de Células NK/fisiologia , Neuroblastoma/patologiaRESUMO
Natural killer (NK) cells are the primary effectors of the innate immune response against virus-infected cells or cells that have undergone malignant transformation. NK cells recognize their targets through a complex array of activating and inhibitory receptors, which regulate the intensity of the effector response against individual target cells. However, many studies have shown that tumor cells can escape immune cell recognition through a variety of mechanisms, developing resistance to NK cell killing. Using a lentiviral shRNA library, we previously demonstrated that several common signaling pathways modulate susceptibility of tumor cells to NK cell activity. In this study, we focused on one of the genes (PI3KCB), identified in this genetic screen. The PI3KCB gene encodes an isoform of the catalytic subunit of PI3K called P110ß. The PI3K pathway has been linked to diverse cellular functions, but has never been associated with susceptibility to NK cell activity. Gene silencing of PI3KCB resulted in increased susceptibility of several tumor cell lines to NK cell lytic activity and induced increased IFN-γ secretion by NK cells. Treatment of primary tumor cells with two different PI3K inhibitors also increased target cell susceptibility to NK cell activity. These effects are due, at least in part, to modulation of several activating and inhibitory ligands and appear to be correlated with PI3K signaling pathway inhibition. These findings identify a new and important role of PI3KCB in modulating tumor cell susceptibility to NK cells and open the way to future combined target immunotherapies.
Assuntos
Citotoxicidade Imunológica , Células Matadoras Naturais/imunologia , Neoplasias/terapia , Fosfatidilinositol 3-Quinases/fisiologia , Transdução de Sinais/fisiologia , Antígenos de Diferenciação de Linfócitos T/fisiologia , Linhagem Celular Tumoral , Humanos , Imunoterapia , Subfamília K de Receptores Semelhantes a Lectina de Células NK/fisiologia , Neoplasias/imunologia , Fosfatidilinositol 3-Quinases/genética , Inibidores de Fosfoinositídeo-3 QuinaseRESUMO
Hepatocellular carcinoma (HCC) is one of the most frequent causes of cancer-related death globally. Above well-known risk factors for HCC development ranging from various toxins to diseases such as diabetes mellitus, chronic infection with hepatitis B virus and hepatitis C virus (HCV) poses the most serious threat, constituting the cause in more than 80 % of cases. In addition to the viral genes intensively investigated, the pathophysiological importance of host genetic factors has also been greatly and increasingly appreciated. Genome-wide association studies (GWAS) comprehensively search the host genome at the single-nucleotide level, and have successfully identified the genomic region associated with a whole variety of diseases. With respect to HCC, there have been reports from several groups on single nucleotide polymorphisms (SNPs) associated with hepatocarcinogenesis, among which was our GWAS discovering MHC class I polypeptide-related sequence A (MICA) as a susceptibility gene for HCV-induced HCC. MICA is a natural killer (NK) group 2D (NKG2D) ligand, whose interaction with NKG2D triggers NK cell-mediated cytotoxicity toward the target cells, and is a key molecule in tumor immune surveillance as its expression is induced on stressed cells such as transformed tumor cells for the detection by NK cells. In this review, the latest understanding of the MICA-NKG2D system in viral HCC, particularly focused on its antitumor properties and the involvement of MICA SNPs, is summarized, followed by a discussion of targets for state-of-the-art cancer immunotherapy with personalized medicine in view.
Assuntos
Carcinoma Hepatocelular/genética , Antígenos de Histocompatibilidade Classe I/genética , Neoplasias Hepáticas/genética , Subfamília K de Receptores Semelhantes a Lectina de Células NK/fisiologia , Polimorfismo de Nucleotídeo Único , Carcinoma Hepatocelular/imunologia , Carcinoma Hepatocelular/terapia , Carcinoma Hepatocelular/virologia , Cocarcinogênese , Predisposição Genética para Doença , Hepatite B Crônica/complicações , Hepatite C Crônica/complicações , Humanos , Imunoterapia/métodos , Células Matadoras Naturais/imunologia , Neoplasias Hepáticas/imunologia , Neoplasias Hepáticas/terapia , Neoplasias Hepáticas/virologiaAssuntos
Receptores ErbB/imunologia , Imunidade Inata/genética , Linfócitos/imunologia , Animais , Apresentação de Antígeno/genética , Humanos , Camundongos , Subfamília K de Receptores Semelhantes a Lectina de Células NK/fisiologia , Neoplasias/genética , Neoplasias/imunologia , Estabilidade de RNA/genética , Estresse Fisiológico/genética , Estresse Fisiológico/imunologia , Evasão Tumoral/genéticaRESUMO
Natural-killer group 2, member D (NKG2D) is an activating receptor found on activated natural killer cells and on activated T-cells, here termed cytokine-activated killer (CAK) cells. NKG2D ligands are expressed on various human cancer types. Gemcitabine is an anticancer drug which is a less immune-destructive agent than others. Herein, we investigated the clinical efficacy and the underlying mechanisms of a combination of CAK cell infusion therapy and gemcitabine. Twenty-three patients with disseminated carcinomas were treated with chemo-immunotherapy consisting of CAK cell infusion therapy following gemcitabine treatment. To investigate the underlying mechanisms by which CAK cells synergize with gemcitabine, we used enzyme-linked immunosorbent assay, Real-time reverse transcription polymerase chain reaction assay, calcein-release assay, and adherent target detachment assay. Using these assays we determined the NKG2D ligands such as major histocompatibility complex-class I-related chain (MIC)A/B expression in carcinoma cells and the level of cellular cytotoxicity generated by treatment with gemcitabine with/without CAK cells. The tumor responses differed among the patients (n=23). In vitro experiments revealed that MICA/B protein and mRNA expression were up-regulated in several carcinoma cell lines after gemcitabine treatment. Pre-treatment with gemcitabine and subsequent exposure to CAK cells induced greater cytotoxicity than either treatment alone. Inclusion of soluble MICB in CAK cell-mediated cytotoxicity assay significantly reduced cytotoxicity. Our clinical results of gemcitabine-CAK combinatorial therapy demonstrated long-term stable disease despite chemoresistance. In conclusion, the combination of gemcitabine and CAK cells may have clinical therapeutic significance for pancreatic, hepato-biliary tract, and urothelial tract cancer. Our study shows that combining CAK therapy with gemcitabine can lead to successful treatment of metastatic cancer.
Assuntos
Antimetabólitos Antineoplásicos/farmacologia , Células Matadoras Induzidas por Citocinas/imunologia , Desoxicitidina/análogos & derivados , Resistencia a Medicamentos Antineoplásicos , Subfamília K de Receptores Semelhantes a Lectina de Células NK/fisiologia , Neoplasias/terapia , Adulto , Idoso , Idoso de 80 Anos ou mais , Linhagem Celular Tumoral , Terapia Combinada , Citotoxicidade Imunológica , Desoxicitidina/farmacologia , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Metástase Neoplásica , Neoplasias/patologia , Estudos Retrospectivos , GencitabinaRESUMO
Bone marrow mesenchymal stem cells (BMSCs) inhibit immune cell responsiveness, and especially of T lymphocytes. We showed that BMSCs markedly inhibited the proliferation and cytokine production by CD8(+) T cells by a cell-to-cell contact phenomenon and secretion of soluble factors. BMSCs down-regulate the expression of natural killer group 2, member D protein (NKG2D) receptors on CD8(+) T cells when co-cultured with them. Moreover, CD8(+) T cells that express low levels of NKG2D had impaired proliferation after triggering by a mitogen. The major histocompatibility complex (MHC) class I chain-related (MIC) A/B molecule, which is a typical ligand for NKG2D, was expressed on BMSCs, and caused dampening of cell proliferation. Monoclonal antibody blocking experiments targeted to MIC A/B impaired CD8(+) T cell function, as evaluated by proliferation and cytokine production. In addition, the production of prostaglandin E2 (PGE2 ), indoleamine 2, 3-dioxygenase (IDO) and transforming growth factor (TGF)-ß1 were increased when BMSCs were co-cultured with CD8(+) T cells. The addition of specific inhibitors against PGE2 , IDO and TGF-ß partially restored the proliferation of CD8(+) T cells. Our results suggest that BMSCs suppress CD8(+) T cell-mediated activation by suppressing NKG2D expression and secretion of PGE2, IDO and TGF-ß. Our observations further confirm the feasibility of BMSCs as a potential adoptive cellular therapy in immune-mediated diseases such as graft-versus-host disease (GVHD).
Assuntos
Linfócitos T CD8-Positivos/imunologia , Dinoprostona/biossíntese , Indolamina-Pirrol 2,3,-Dioxigenase/biossíntese , Ativação Linfocitária , Células-Tronco Mesenquimais/fisiologia , Subfamília K de Receptores Semelhantes a Lectina de Células NK/fisiologia , Fator de Crescimento Transformador beta/biossíntese , Comunicação Celular , Células Cultivadas , Granzimas/biossíntese , Antígenos de Histocompatibilidade Classe I/fisiologia , Humanos , Interferon gama/biossíntese , Interleucina-2/biossínteseRESUMO
NKG2D plays a major role in controlling immune responses through the regulation of natural killer (NK) cells, αß and γδ T-cell function. This activating receptor recognizes eight distinct ligands (the MHC Class I polypeptide-related sequences (MIC) A andB, and UL16-binding proteins (ULBP)1-6) induced by cellular stress to promote recognition cells perturbed by malignant transformation or microbial infection. Studies into human cytomegalovirus (HCMV) have aided both the identification and characterization of NKG2D ligands (NKG2DLs). HCMV immediate early (IE) gene up regulates NKGDLs, and we now describe the differential activation of ULBP2 and MICA/B by IE1 and IE2 respectively. Despite activation by IE functions, HCMV effectively suppressed cell surface expression of NKGDLs through both the early and late phases of infection. The immune evasion functions UL16, UL142, and microRNA(miR)-UL112 are known to target NKG2DLs. While infection with a UL16 deletion mutant caused the expected increase in MICB and ULBP2 cell surface expression, deletion of UL142 did not have a similar impact on its target, MICA. We therefore performed a systematic screen of the viral genome to search of addition functions that targeted MICA. US18 and US20 were identified as novel NK cell evasion functions capable of acting independently to promote MICA degradation by lysosomal degradation. The most dramatic effect on MICA expression was achieved when US18 and US20 acted in concert. US18 and US20 are the first members of the US12 gene family to have been assigned a function. The US12 family has 10 members encoded sequentially through US12-US21; a genetic arrangement, which is suggestive of an 'accordion' expansion of an ancestral gene in response to a selective pressure. This expansion must have be an ancient event as the whole family is conserved across simian cytomegaloviruses from old world monkeys. The evolutionary benefit bestowed by the combinatorial effect of US18 and US20 on MICA may have contributed to sustaining the US12 gene family.
Assuntos
Citomegalovirus , Antígenos de Histocompatibilidade Classe I/metabolismo , Evasão da Resposta Imune , Células Matadoras Naturais/imunologia , Lisossomos/metabolismo , Proteólise , Proteínas Virais/fisiologia , Adulto , Proteínas de Bactérias/metabolismo , Células Cultivadas , Citomegalovirus/imunologia , Citomegalovirus/patogenicidade , Inibidores Enzimáticos/farmacologia , Humanos , Evasão da Resposta Imune/efeitos dos fármacos , Células Matadoras Naturais/efeitos dos fármacos , Leupeptinas/farmacologia , Proteínas Luminescentes/metabolismo , Lisossomos/efeitos dos fármacos , Macrolídeos/farmacologia , Subfamília K de Receptores Semelhantes a Lectina de Células NK/fisiologia , Proteólise/efeitos dos fármacos , Proteínas Recombinantes/metabolismoRESUMO
Membrane rafts are microdomains of the plasma membrane that have multiple biological functions. The involvement of these structures in the biology of T cells, namely in signal transduction by the TCR, has been widely studied. However, the role of membrane rafts in immunoreceptor signaling in NK cells is less well known. We studied the distribution of the activating NKG2D receptor in lipid rafts by isolating DRMs in a sucrose density gradient or by raft fractionation by ß-OG-selective solubility in the NKL cell line. We found that the NKG2D-DAP10 complex and pVav are recruited into rafts upon receptor stimulation. Qualitative proteomic analysis of these fractions showed that the actin cytoskeleton is involved in this process. In particular, we found that the actin-bundling protein L-plastin plays an important role in the clustering of NKG2D into lipid rafts. Moreover, coengagement of the inhibitory receptor NKG2A partially disrupted NKG2D recruitment into rafts. Furthermore, we demonstrated that L-plastin participates in NKG2D-mediated inhibition of NK cell chemotaxis.
Assuntos
Quimiotaxia de Leucócito/fisiologia , Células Matadoras Naturais/metabolismo , Microdomínios da Membrana/fisiologia , Proteínas dos Microfilamentos/fisiologia , Subfamília K de Receptores Semelhantes a Lectina de Células NK/fisiologia , Citoesqueleto de Actina/fisiologia , Membrana Celular/efeitos dos fármacos , Membrana Celular/metabolismo , Células Cultivadas , Centrifugação com Gradiente de Concentração , Detergentes/farmacologia , Humanos , Células Matadoras Naturais/citologia , Microdomínios da Membrana/efeitos dos fármacos , Proteínas dos Microfilamentos/antagonistas & inibidores , Proteínas dos Microfilamentos/genética , Complexos Multiproteicos , Subfamília C de Receptores Semelhantes a Lectina de Células NK/metabolismo , Proteoma , Interferência de RNA , RNA Interferente Pequeno/farmacologia , Receptores Imunológicos/metabolismo , Transdução de Sinais/imunologiaRESUMO
NK cells are the main cells of the innate immune system that produce IFN-γ, and they express this cytokine at early stages of maturation in response to cytokine stimulation. Conversely, acquisition of IFN-γ-competence in CD4(+) T helper cells requires a differentiation process from naïve toward type 1 (Th1) cells, which is associated with epigenetic remodeling at the IFNG locus. In the present study, we show that the ability of NK cells to produce IFN-γ in response to activating receptor (actR) engagement is gradually acquired during terminal differentiation and is accompanied by progressively higher NF-κB activation in response to actR triggering. Moreover, during the differentiation process NK cells gradually display increasing expression of IFNG and TBX21 (encoding T-bet) transcripts and demethylation at the IFNG promoter. This study provides new insights in the molecular mechanisms underlying NK-cell ability to express IFN-γ upon actR engagement. Thus, we propose that in order to efficiently produce IFN-γ in response to infected or transformed cells, NK cells gain Th1-like features, such as higher IFN-γ competence and epigenetic remodeling of the IFNG promoter, during their terminal differentiation.
Assuntos
Diferenciação Celular , Interferon gama/biossíntese , Células Matadoras Naturais/imunologia , Ilhas de CpG , Metilação de DNA , Humanos , Interferon gama/genética , Células Matadoras Naturais/citologia , NF-kappa B/fisiologia , Subfamília K de Receptores Semelhantes a Lectina de Células NK/fisiologia , Receptor 1 Desencadeador da Citotoxicidade Natural/fisiologia , Receptor 3 Desencadeador da Citotoxicidade Natural/fisiologia , Regiões Promotoras Genéticas , Proteínas com Domínio T/fisiologiaRESUMO
BACKGROUND: The diverse roles of innate immune cells in the pathogenesis of asthma remain to be fully defined. Natural killer (NK) cells are innate lymphocytes that can regulate adaptive immune responses. NK cells are activated in asthma; however, their role in allergic airway inflammation is not fully understood. OBJECTIVE: We investigated the importance of NK cells in house dust mite (HDM)-triggered allergic pulmonary inflammation. Specifically, we aimed to determine the role of the major NK-cell activating receptor NKG2D and NK-cell effector functions mediated by granzyme B. METHODS: Allergic airway inflammation was induced in the airways of mice by repeated intranasal HDM extract administration and responses in wild-type and NKG2D-deficient mice were compared. Adoptive transfer studies were used to identify the cells and mechanisms involved. RESULTS: Mice that lacked NKG2D were resistant to the induction of allergic inflammation and showed little pulmonary eosinophilia, few airway TH2 cells, and no rise in serum IgE after multiple HDM-allergen exposures. However, NKG2D was not required for pulmonary inflammation after a single inoculation of allergen. NKG2D-deficient mice showed no alteration in responses to respiratory virus infection. Transfer of wild-type NK cells (but not CD3(+) cells) into NKG2D-deficient mice restored allergic inflammatory responses only if the NK cells expressed granzyme B. CONCLUSIONS: These studies established a pivotal role for NK-cell NKG2D and granzyme B in the pathogenesis of HDM-induced allergic lung disease, and identified novel therapeutic targets for the prevention and treatment of asthma.