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1.
Nat Commun ; 12(1): 832, 2021 02 05.
Artigo em Inglês | MEDLINE | ID: mdl-33547304

RESUMO

The two T cell inhibitory receptors PD-1 and TIM-3 are co-expressed during exhausted T cell differentiation, and recent evidence suggests that their crosstalk regulates T cell exhaustion and immunotherapy efficacy; however, the molecular mechanism is unclear. Here we show that PD-1 contributes to the persistence of PD-1+TIM-3+ T cells by binding to the TIM-3 ligand galectin-9 (Gal-9) and attenuates Gal-9/TIM-3-induced cell death. Anti-Gal-9 therapy selectively expands intratumoral TIM-3+ cytotoxic CD8 T cells and immunosuppressive regulatory T cells (Treg cells). The combination of anti-Gal-9 and an agonistic antibody to the co-stimulatory receptor GITR (glucocorticoid-induced tumor necrosis factor receptor-related protein) that depletes Treg cells induces synergistic antitumor activity. Gal-9 expression and secretion are promoted by interferon ß and γ, and high Gal-9 expression correlates with poor prognosis in multiple human cancers. Our work uncovers a function for PD-1 in exhausted T cell survival and suggests Gal-9 as a promising target for immunotherapy.


Assuntos
Adenocarcinoma/terapia , Neoplasias do Colo/terapia , Galectinas/imunologia , Regulação Neoplásica da Expressão Gênica/imunologia , Proteína Relacionada a TNFR Induzida por Glucocorticoide/imunologia , Receptor Celular 2 do Vírus da Hepatite A/imunologia , Receptor de Morte Celular Programada 1/imunologia , Adenocarcinoma/genética , Adenocarcinoma/imunologia , Adenocarcinoma/mortalidade , Animais , Anticorpos/farmacologia , Antineoplásicos Imunológicos/farmacologia , Neoplasias do Colo/genética , Neoplasias do Colo/imunologia , Neoplasias do Colo/mortalidade , Galectinas/antagonistas & inibidores , Galectinas/genética , Proteína Relacionada a TNFR Induzida por Glucocorticoide/agonistas , Proteína Relacionada a TNFR Induzida por Glucocorticoide/genética , Receptor Celular 2 do Vírus da Hepatite A/genética , Humanos , Imunoterapia/métodos , Células Jurkat , Melanoma Experimental/genética , Melanoma Experimental/imunologia , Melanoma Experimental/mortalidade , Melanoma Experimental/terapia , Camundongos , Camundongos Endogâmicos BALB C , Receptor de Morte Celular Programada 1/genética , Ligação Proteica , Transdução de Sinais , Neoplasias Cutâneas/genética , Neoplasias Cutâneas/imunologia , Neoplasias Cutâneas/mortalidade , Neoplasias Cutâneas/terapia , Análise de Sobrevida , Linfócitos T Citotóxicos/efeitos dos fármacos , Linfócitos T Citotóxicos/imunologia , Linfócitos T Citotóxicos/patologia , Linfócitos T Reguladores/efeitos dos fármacos , Linfócitos T Reguladores/imunologia , Linfócitos T Reguladores/patologia
2.
Chem Commun (Camb) ; 57(4): 504-507, 2021 Jan 14.
Artigo em Inglês | MEDLINE | ID: mdl-33331360

RESUMO

A novel STING agonist, CDGSF, ipsilaterally modified with phosphorothioate and fluorine, was synthesized. The phosphorothioate in CDGSF might be a site for covalent conjugation. Injection of CDGSF generated an immunogenic ("hot") tumor microenvironment to suppress melanoma, more efficiently than dithio CDG. In particular, immunization with SARS-CoV-2 spike protein using CDGSF as an adjuvant elicited an exceptionally high antibody titer and a robust T cell response, overcoming the drawbacks of aluminum hydroxide. These results highlighted the therapeutic potential of CDGSF for cancer immunotherapy and the adjuvant potential of the STING agonist in the SARS-CoV-2 vaccine for the first time.


Assuntos
Adjuvantes Imunológicos/administração & dosagem , /prevenção & controle , Melanoma Experimental/tratamento farmacológico , Proteínas de Membrana/agonistas , Nucleotídeos Cíclicos/administração & dosagem , Neoplasias Cutâneas/tratamento farmacológico , Adjuvantes Imunológicos/síntese química , Hidróxido de Alumínio/administração & dosagem , Hidróxido de Alumínio/química , Animais , Anticorpos Antivirais/biossíntese , Linfócitos B/efeitos dos fármacos , Linfócitos B/imunologia , Linfócitos B/virologia , /virologia , ELISPOT , Humanos , Imunoterapia/métodos , Interferon gama/biossíntese , Melanoma Experimental/imunologia , Melanoma Experimental/mortalidade , Melanoma Experimental/patologia , Proteínas de Membrana/genética , Proteínas de Membrana/imunologia , Camundongos , Nucleotídeos Cíclicos/síntese química , /imunologia , Neoplasias Cutâneas/imunologia , Neoplasias Cutâneas/mortalidade , Neoplasias Cutâneas/patologia , Glicoproteína da Espícula de Coronavírus/administração & dosagem , Glicoproteína da Espícula de Coronavírus/química , Glicoproteína da Espícula de Coronavírus/imunologia , Análise de Sobrevida , Linfócitos T/efeitos dos fármacos , Linfócitos T/imunologia , Linfócitos T/virologia , Carga Tumoral/efeitos dos fármacos , Microambiente Tumoral/efeitos dos fármacos , Microambiente Tumoral/imunologia , Vacinação/métodos
3.
PLoS One ; 15(8): e0237646, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32845913

RESUMO

Tumor antigen-primed CD8 T cells differentiate into effector T cells that kill tumor cells rapidly, whereas durable responses of CD8 T cells are required to cope with long-lasting tumor growth. However, it is not well known how persisting CD8 T cells are generated. In this study, we analyzed CD8 T cells primed by antigens in tumor-draining lymph nodes and found that CD8 T cells first differentiated into a CD62L-intermediate (CD62Lint) stage upon antigen stimulation. These cells gave rise to tumor-infiltrating CD62L-CD44high Bcl6- effector T cells and CD62L+CD44highBcl6+ memory-like T cells. Memory-like T cells within the tumor expressed CD127, CXCR3 and had the potential to proliferate significantly when they were transferred into tumor-bearing mice. Bcl6 expression in these T cells was critical because Bcl6-/-CD62L+CD44highCD8T cells within the tumor were defective in expansion after secondary transfer. Taken together, our findings show that CD62L+CD44highBcl6+ cells are generated from highly proliferating CD62Lint T cells and retain high proliferative potential, which contributes to replenishment of effector T cells within the tumor.


Assuntos
Linfócitos T CD8-Positivos/imunologia , Carcinoma Pulmonar de Lewis/patologia , Memória Imunológica/imunologia , Selectina L/metabolismo , Melanoma Experimental/patologia , Receptor de Morte Celular Programada 1/metabolismo , Proteínas Proto-Oncogênicas c-bcl-6/metabolismo , Animais , Carcinoma Pulmonar de Lewis/imunologia , Carcinoma Pulmonar de Lewis/metabolismo , Diferenciação Celular , Feminino , Selectina L/genética , Masculino , Melanoma Experimental/imunologia , Melanoma Experimental/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Receptor de Morte Celular Programada 1/genética , Proteínas Proto-Oncogênicas c-bcl-6/genética , Células Tumorais Cultivadas
4.
Cancer Immunol Immunother ; 69(10): 2157-2162, 2020 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-32638080

RESUMO

Epidemiological evidence suggests that females have an advantage over males in cases of melanoma incidence, progression, and survival. However, the biological mechanisms underlying these sex differences remain unclear. With the knowledge that females generally have a more robust immune system than males, we investigated sex differences in melanoma progression in a B16-F10/BL6 syngeneic mouse model. We observed significantly less tumor volume and growth rate over 14 days in female mice compared to male mice. Furthermore, higher populations of CD4+ and CD8+ T cells, which indicate adaptive immune responses, were found in the circulating blood and tumors of females and corresponded with less tumor growth, and vice versa in males. Our results highlight a mouse model that represents melanoma progression in the human population and displays a higher immune response to melanoma in females compared to males. These findings suggest that the immune system may be one of the mechanisms responsible for sex differences in melanoma.


Assuntos
Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/imunologia , Modelos Animais de Doenças , Linfócitos do Interstício Tumoral/imunologia , Melanoma Experimental/imunologia , Neoplasias Cutâneas/imunologia , Linfócitos T Citotóxicos/imunologia , Animais , Linfócitos T CD4-Positivos/patologia , Linfócitos T CD8-Positivos/patologia , Feminino , Masculino , Melanoma Experimental/patologia , Camundongos , Camundongos Endogâmicos C57BL , Neoplasias Cutâneas/secundário , Linfócitos T Citotóxicos/patologia , Carga Tumoral , Células Tumorais Cultivadas
5.
Cancer Cell ; 37(6): 786-799.e5, 2020 06 08.
Artigo em Inglês | MEDLINE | ID: mdl-32516589

RESUMO

Generation of tumor-infiltrating lymphocytes begins when tumor antigens reach the lymph node (LN) to stimulate T cells, yet we know little of how tumor material is disseminated among the large variety of antigen-presenting dendritic cell (DC) subsets in the LN. Here, we demonstrate that tumor proteins are carried to the LN within discrete vesicles inside DCs and are then transferred among DC subsets. A synapse is formed between interacting DCs and vesicle transfer takes place in the absence of free exosomes. DCs -containing vesicles can uniquely activate T cells, whereas DCs lacking them do not. Understanding this restricted sharing of tumor identity provides substantial room for engineering better anti-tumor immunity.


Assuntos
Apresentação do Antígeno/imunologia , Antígenos de Neoplasias/imunologia , Células Dendríticas/imunologia , Melanoma Experimental/imunologia , Células Mieloides/imunologia , Sinapses/imunologia , Linfócitos T/imunologia , Animais , Células Dendríticas/citologia , Células Dendríticas/metabolismo , Masculino , Melanoma Experimental/metabolismo , Melanoma Experimental/patologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Células Mieloides/citologia , Células Mieloides/metabolismo , Receptores CCR2/fisiologia , Receptores CCR7/fisiologia , Sinapses/metabolismo , Sinapses/patologia , Linfócitos T/citologia , Linfócitos T/metabolismo
6.
Cancer Cell ; 37(6): 818-833.e9, 2020 06 08.
Artigo em Inglês | MEDLINE | ID: mdl-32516591

RESUMO

T cells are central to all currently effective cancer immunotherapies, but the characteristics defining therapeutically effective anti-tumor T cells have not been comprehensively elucidated. Here, we delineate four phenotypic qualities of effective anti-tumor T cells: cell expansion, differentiation, oxidative stress, and genomic stress. Using a CRISPR-Cas9-based genetic screen of primary T cells we measured the multi-phenotypic impact of disrupting 25 T cell receptor-driven kinases. We identified p38 kinase as a central regulator of all four phenotypes and uncovered transcriptional and antioxidant pathways regulated by p38 in T cells. Pharmacological inhibition of p38 improved the efficacy of mouse anti-tumor T cells and enhanced the functionalities of human tumor-reactive and gene-engineered T cells, paving the way for clinically relevant interventions.


Assuntos
Neoplasias da Mama/terapia , Sistemas CRISPR-Cas , Imunoterapia Adotiva/métodos , Melanoma Experimental/terapia , Fenótipo , Linfócitos T/transplante , Proteínas Quinases p38 Ativadas por Mitógeno/antagonistas & inibidores , Animais , Neoplasias da Mama/imunologia , Neoplasias da Mama/patologia , Diferenciação Celular , Feminino , Engenharia Genética , Masculino , Melanoma Experimental/imunologia , Melanoma Experimental/patologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Receptores de Antígenos de Linfócitos T/fisiologia , Linfócitos T/imunologia , Proteínas Quinases p38 Ativadas por Mitógeno/genética
7.
Nat Commun ; 11(1): 3187, 2020 06 24.
Artigo em Inglês | MEDLINE | ID: mdl-32581235

RESUMO

The application of adoptive T cell therapies, including those using chimeric antigen receptor (CAR)-modified T cells, to solid tumors requires combinatorial strategies to overcome immune suppression associated with the tumor microenvironment. Here we test whether the inflammatory nature of oncolytic viruses and their ability to remodel the tumor microenvironment may help to recruit and potentiate the functionality of CAR T cells. Contrary to our hypothesis, VSVmIFNß infection is associated with attrition of murine EGFRvIII CAR T cells in a B16EGFRvIII model, despite inducing a robust proinflammatory shift in the chemokine profile. Mechanistically, type I interferon (IFN) expressed following infection promotes apoptosis, activation, and inhibitory receptor expression, and interferon-insensitive CAR T cells enable combinatorial therapy with VSVmIFNß. Our study uncovers an unexpected mechanism of therapeutic interference, and prompts further investigation into the interaction between CAR T cells and oncolytic viruses to optimize combination therapy.


Assuntos
Imunoterapia Adotiva , Interferon beta/metabolismo , Vírus Oncolíticos/metabolismo , Receptores de Antígenos Quiméricos/metabolismo , Linfócitos T/metabolismo , Animais , Apoptose , Linhagem Celular Tumoral , Quimiocinas/metabolismo , Terapia Combinada , Feminino , Interferon beta/genética , Ativação Linfocitária , Melanoma Experimental/imunologia , Melanoma Experimental/terapia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Mutantes , Terapia Viral Oncolítica , Vírus Oncolíticos/genética , Receptor de Interferon alfa e beta/genética , Receptor de Interferon alfa e beta/metabolismo , Receptores de Antígenos de Linfócitos T/metabolismo , Baço/imunologia
8.
J Leukoc Biol ; 107(6): 883-892, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32386455

RESUMO

Nonclassical monocytes maintain vascular homeostasis by patrolling the vascular endothelium, responding to inflammatory signals, and scavenging cellular debris. Nonclassical monocytes also prevent metastatic tumor cells from seeding new tissues, but whether the patrolling function of nonclassical monocytes is required for this process is unknown. To answer this question, we utilized an inducible-knockout mouse that exhibits loss of the integrin-adaptor protein Kindlin-3 specifically in nonclassical monocytes. We show that Kindlin-3-deficient nonclassical monocytes are unable to patrol the vascular endothelium in either the lungs or periphery. We also find that Kindlin-3-deficient nonclassical monocytes cannot firmly adhere to, and instead "slip" along, the vascular endothelium. Loss of patrolling activity by nonclassical monocytes was phenocopied by ablation of LFA-1, an integrin-binding partner of Kindlin-3. When B16F10 murine melanoma tumor cells were introduced into Kindlin-3-deficient mice, nonclassical monocytes showed defective patrolling towards tumor cells and failure to ingest tumor particles in vivo. Consequently, we observed a significant, 4-fold increase in lung tumor metastases in mice possessing Kindlin-3-deficient nonclassical monocytes. Thus, we conclude that the patrolling function of nonclassical monocytes is mediated by Kindlin-3 and essential for these cells to maintain vascular endothelial homeostasis and prevent tumor metastasis to the lung.


Assuntos
Proteínas do Citoesqueleto/genética , Regulação Neoplásica da Expressão Gênica , Antígeno-1 Associado à Função Linfocitária/genética , Melanoma Experimental/genética , Monócitos/imunologia , Fagocitose , Neoplasias Cutâneas/genética , Animais , Medula Óssea/imunologia , Transplante de Medula Óssea , Adesão Celular , Comunicação Celular/imunologia , Proteínas do Citoesqueleto/deficiência , Proteínas do Citoesqueleto/imunologia , Endotélio Vascular/imunologia , Endotélio Vascular/patologia , Feminino , Humanos , Injeções Intravenosas , Pulmão/irrigação sanguínea , Pulmão/imunologia , Pulmão/patologia , Antígeno-1 Associado à Função Linfocitária/imunologia , Melanoma Experimental/imunologia , Melanoma Experimental/secundário , Camundongos , Camundongos Knockout , Monócitos/patologia , Células Neoplásicas Circulantes/imunologia , Células Neoplásicas Circulantes/patologia , Cultura Primária de Células , Transdução de Sinais , Neoplasias Cutâneas/imunologia , Neoplasias Cutâneas/patologia , Irradiação Corporal Total
9.
Cancer Immunol Immunother ; 69(10): 1959-1972, 2020 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-32388678

RESUMO

Cancer vaccine development has proven challenging with the exception of some virally induced cancers for which prophylactic vaccines exist. Currently, there is only one FDA approved vaccine for the treatment of prostate cancer and as such prostate cancer continues to present a significant unmet medical need. In this study, we examine the effectiveness of a therapeutic cancer vaccine that combines the ISCOMATRIX™ adjuvant (ISCOMATRIX) with the Toll-like receptor 3 agonist, polyinosinic-polycytidylic acid (Poly I:C), and Flt3L, FMS-like tyrosine kinase 3 ligand. We employed the TRAMP-C1 (transgenic adenocarcinoma of the mouse prostate) model of prostate cancer and the self-protein mPAP (prostatic acid phosphatase) as the tumor antigen. ISCOMATRIX™-mPAP-Poly I:C-Flt3L was delivered in a therapeutic prime-boost regime that was consistently able to achieve complete tumor regression in 60% of animals treated and these tumor-free animals were protected upon rechallenge. Investigations into the underlying immunological mechanisms contributing to the effectiveness of this vaccine identified that both innate and adaptive responses are elicited and required. NK cells, CD4+ T cells and interferon-γ were all found to be critical for tumor control while tumor infiltrating CD8+ T cells became disabled by an immunosuppressive microenvironment. There is potential for broader application of this cancer vaccine, as we have been able to demonstrate effectiveness in two additional cancer models; melanoma (B16-OVA) and a model of B cell lymphoma (Eµ-myc-GFP-OVA).


Assuntos
Adjuvantes Imunológicos/administração & dosagem , Antígenos de Neoplasias/imunologia , Linfócitos T CD8-Positivos/imunologia , Vacinas Anticâncer/administração & dosagem , Colesterol/administração & dosagem , Melanoma Experimental/imunologia , Fosfolipídeos/administração & dosagem , Neoplasias da Próstata/imunologia , Saponinas/administração & dosagem , Animais , Apoptose , Linfócitos T CD8-Positivos/efeitos dos fármacos , Proliferação de Células , Modelos Animais de Doenças , Combinação de Medicamentos , Humanos , Interferon gama/metabolismo , Masculino , Melanoma Experimental/tratamento farmacológico , Melanoma Experimental/patologia , Proteínas de Membrana/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Ovalbumina/imunologia , Poli I-C/administração & dosagem , Neoplasias da Próstata/tratamento farmacológico , Neoplasias da Próstata/patologia , Células Tumorais Cultivadas , Microambiente Tumoral/efeitos dos fármacos , Microambiente Tumoral/imunologia , Ensaios Antitumorais Modelo de Xenoenxerto
10.
Nat Med ; 26(7): 1125-1134, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32451499

RESUMO

Understanding of the factors governing immune responses in cancer remains incomplete, limiting patient benefit. In this study, we used mass cytometry to define the systemic immune landscape in response to tumor development across five tissues in eight mouse tumor models. Systemic immunity was dramatically altered across models and time, with consistent findings in the peripheral blood of patients with breast cancer. Changes in peripheral tissues differed from those in the tumor microenvironment. Mice with tumor-experienced immune systems mounted dampened responses to orthogonal challenges, including reduced T cell activation during viral or bacterial infection. Antigen-presenting cells (APCs) mounted weaker responses in this context, whereas promoting APC activation rescued T cell activity. Systemic immune changes were reversed with surgical tumor resection, and many were prevented by interleukin-1 or granulocyte colony-stimulating factor blockade, revealing remarkable plasticity in the systemic immune state. These results demonstrate that tumor development dynamically reshapes the composition and function of the immune macroenvironment.


Assuntos
Infecções Bacterianas/imunologia , Neoplasias da Mama/imunologia , Melanoma Experimental/imunologia , Microambiente Tumoral/imunologia , Animais , Células Apresentadoras de Antígenos/imunologia , Infecções Bacterianas/microbiologia , Infecções Bacterianas/patologia , Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Modelos Animais de Doenças , Feminino , Fator Estimulador de Colônias de Granulócitos e Macrófagos/genética , Fator Estimulador de Colônias de Granulócitos e Macrófagos/imunologia , Humanos , Ativação Linfocitária/imunologia , Melanoma Experimental/genética , Melanoma Experimental/patologia , Camundongos , Linfócitos T/imunologia , Microambiente Tumoral/genética
11.
Cancer Immunol Immunother ; 69(10): 2075-2088, 2020 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-32447411

RESUMO

In line with the ongoing phase I trial (NCT03784625) dedicated to melanoma targeted radionuclide therapy (TRT), we explore the interplay between immune system and the melanin ligand [131I]ICF01012 alone or combined with immunotherapy (immune checkpoint inhibitors, ICI) in preclinical models. Here we demonstrate that [131I]ICF01012 induces immunogenic cell death, characterized by a significant increase in cell surface-exposed annexin A1 and calreticulin. Additionally, [131I]ICF01012 increases survival in immunocompetent mice, compared to immunocompromised (29 vs. 24 days, p = 0.0374). Flow cytometry and RT-qPCR analyses highlight that [131I]ICF01012 induces adaptive and innate immune cell recruitment in the tumor microenvironment. [131I]ICF01012 combination with ICIs (anti-CTLA-4, anti-PD-1, anti-PD-L1) has shown that tolerance is a main immune escape mechanism, whereas exhaustion is not present after TRT. Furthermore, [131I]ICF01012 and ICI combination has systematically resulted in a prolonged survival (p < 0.0001) compared to TRT alone. Specifically, [131I]ICF01012 + anti-CTLA-4 combination significantly increases survival compared to anti-CTLA-4 alone (41 vs. 26 days; p = 0.0011), without toxicity. This work represents the first global characterization of TRT-induced modifications of the antitumor immune response, demonstrating that tolerance is a main immune escape mechanism and that combining TRT and ICI is promising.


Assuntos
Antineoplásicos Imunológicos/uso terapêutico , Imunoterapia/métodos , Radioisótopos do Iodo/uso terapêutico , Melanoma Experimental/imunologia , Melanoma Experimental/terapia , Tolerância a Radiação/efeitos dos fármacos , Animais , Terapia Combinada , Melanoma Experimental/patologia , Camundongos , Células Tumorais Cultivadas
12.
Oncogene ; 39(22): 4344-4357, 2020 05.
Artigo em Inglês | MEDLINE | ID: mdl-32335582

RESUMO

We explore a novel strategy of activating immune signaling through increased micronuclei formation utilizing a cell cycle checkpoint inhibitor to drive cell cycle progression following ionizing radiation. The Chk1/2 inhibitor AZD7762 is used to abrogate radiation therapy (RT)-induced G2/M cell cycle arrest in multiple cell lines and, we find that this therapeutic combination promotes increased micronuclei formation in vitro and subsequently drives increased type I interferon signaling and cytotoxic T-cell activation. In vivo studies using B16-F10 melanoma cancer cells implanted in C57/BL6 mice demonstrate improved rates of tumor control at the abscopal (unirradiated) site, located outside of the radiation field, only in the AZD7762 + RT group, with a corresponding reduction in mean tumor volume, increase in the CD8 T-cell population, and immune activated gene signaling. Our results demonstrate that targeted inhibition of cell cycle checkpoint activation following ionizing radiation drives increased production of immunogenic micronuclei, leading to systemic tumor response with potential future clinical benefit.


Assuntos
Núcleo Celular/efeitos dos fármacos , Núcleo Celular/efeitos da radiação , Quinase 1 do Ponto de Checagem/antagonistas & inibidores , Quinase do Ponto de Checagem 2/antagonistas & inibidores , Pontos de Checagem da Fase G2 do Ciclo Celular/efeitos dos fármacos , Pontos de Checagem da Fase G2 do Ciclo Celular/efeitos da radiação , Melanoma Experimental/imunologia , Proteínas de Neoplasias/antagonistas & inibidores , Tiofenos/farmacologia , Ureia/análogos & derivados , Animais , Linhagem Celular Tumoral , Feminino , Humanos , Interferon beta/biossíntese , Interferon beta/genética , Melanoma Experimental/tratamento farmacológico , Melanoma Experimental/patologia , Melanoma Experimental/radioterapia , Proteínas de Membrana/biossíntese , Proteínas de Membrana/genética , Camundongos Endogâmicos C57BL , Testes para Micronúcleos , Proteínas de Neoplasias/biossíntese , Proteínas de Neoplasias/genética , Interferência de RNA , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/farmacologia , Fator de Transcrição STAT1/biossíntese , Fator de Transcrição STAT1/genética , Carga Tumoral/efeitos dos fármacos , Carga Tumoral/efeitos da radiação , Ureia/farmacologia , Ensaios Antitumorais Modelo de Xenoenxerto
13.
Immunity ; 52(4): 668-682.e7, 2020 04 14.
Artigo em Inglês | MEDLINE | ID: mdl-32294407

RESUMO

The primary mechanisms supporting immunoregulatory polarization of myeloid cells upon infiltration into tumors remain largely unexplored. Elucidation of these signals could enable better strategies to restore protective anti-tumor immunity. Here, we investigated the role of the intrinsic activation of the PKR-like endoplasmic reticulum (ER) kinase (PERK) in the immunoinhibitory actions of tumor-associated myeloid-derived suppressor cells (tumor-MDSCs). PERK signaling increased in tumor-MDSCs, and its deletion transformed MDSCs into myeloid cells that activated CD8+ T cell-mediated immunity against cancer. Tumor-MDSCs lacking PERK exhibited disrupted NRF2-driven antioxidant capacity and impaired mitochondrial respiratory homeostasis. Moreover, reduced NRF2 signaling in PERK-deficient MDSCs elicited cytosolic mitochondrial DNA elevation and, consequently, STING-dependent expression of anti-tumor type I interferon. Reactivation of NRF2 signaling, conditional deletion of STING, or blockade of type I interferon receptor I restored the immunoinhibitory potential of PERK-ablated MDSCs. Our findings demonstrate the pivotal role of PERK in tumor-MDSC functionality and unveil strategies to reprogram immunosuppressive myelopoiesis in tumors to boost cancer immunotherapy.


Assuntos
Carcinoma Pulmonar de Lewis/imunologia , Carcinoma Epitelial do Ovário/imunologia , Regulação Neoplásica da Expressão Gênica , Melanoma Experimental/imunologia , Proteínas de Membrana/imunologia , Neoplasias Cutâneas/imunologia , eIF-2 Quinase/imunologia , Animais , Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/patologia , Carcinoma Pulmonar de Lewis/genética , Carcinoma Pulmonar de Lewis/metabolismo , Carcinoma Pulmonar de Lewis/patologia , Carcinoma Epitelial do Ovário/genética , Carcinoma Epitelial do Ovário/metabolismo , Carcinoma Epitelial do Ovário/patologia , Feminino , Humanos , Imunossupressão , Interferon-alfa/genética , Interferon-alfa/imunologia , Interferon beta/genética , Interferon beta/imunologia , Masculino , Melanoma Experimental/genética , Melanoma Experimental/metabolismo , Melanoma Experimental/patologia , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Mitocôndrias/imunologia , Mitocôndrias/metabolismo , Células Supressoras Mieloides/imunologia , Células Supressoras Mieloides/patologia , Fator 2 Relacionado a NF-E2/genética , Fator 2 Relacionado a NF-E2/imunologia , Receptores de Interferon/genética , Receptores de Interferon/imunologia , Transdução de Sinais , Neoplasias Cutâneas/genética , Neoplasias Cutâneas/metabolismo , Neoplasias Cutâneas/patologia , Resposta a Proteínas não Dobradas/imunologia , eIF-2 Quinase/deficiência , eIF-2 Quinase/genética
14.
Cancer Immunol Immunother ; 69(8): 1461-1475, 2020 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-32285171

RESUMO

Interleukin-33 (IL-33) regulates innate and acquired immune response to pathogens, self-antigens and tumors. IL-33 effects on tumors depend on the dose and mode of administration along with the type of malignancy. We studied the effects of IL-33 on the development of primary and metastatic melanoma induced by B16-F1 cell line in C57BL/6 mice. Intraperitoneally applied IL-33 restricts primary tumor growth. When administered intranasally 3 days prior to the intravenous injection of the tumor cells, IL-33 promoted growth of B16-F1 melanoma metastases, while B16-F10 gave massive metastases independently of IL-33. To mimic natural dissemination, we next used a limited number (5 × 104) of B16-F1 cells intravenously followed by application of IL-33 intraperitoneally. IL-33 increased the size of metastases (10.96 ± 3.96 mm2) when compared to the control group (0.86 ± 0.39 mm2), without changing incidence and number of metastases. IL-33 increased expression of ST2 on both tumor and immune cells in metastases. Also, IL-33 enhanced eosinophils and anti-tumor NK cells in the lung. The striking finding was reduced cytotoxicity of CD8+ T cells derived from metastatic lung of IL-33 injected mice. IL-33 reduced the percentage of TNF-α+ and IFN-γ+ CD8+ T cells while increasing the frequency of CD8+ T cells that express inhibitory molecules (PD-1, KLRG-1 and CTLA-4). There was a significant accumulation of CD11b+Gr-1+ myeloid suppressor cells and FoxP3+, IL-10+ and CTLA-4+ regulatory T cells in the metastatic lung of IL-33 injected mice. The relevance of IL-33 for melanoma metastases was also documented in a significantly increased level of serum IL-33 in stage III melanoma patients.


Assuntos
Linfócitos T CD8-Positivos/imunologia , Interleucina-33/administração & dosagem , Interleucina-33/sangue , Neoplasias Pulmonares/secundário , Melanoma Experimental/patologia , Linfócitos T Citotóxicos/imunologia , Linfócitos T Reguladores/imunologia , Animais , Apoptose , Biomarcadores Tumorais/sangue , Linfócitos T CD8-Positivos/efeitos dos fármacos , Estudos de Casos e Controles , Proliferação de Células , Regulação Neoplásica da Expressão Gênica , Humanos , Neoplasias Pulmonares/imunologia , Neoplasias Pulmonares/metabolismo , Melanoma Experimental/imunologia , Melanoma Experimental/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Prognóstico , Linfócitos T Citotóxicos/efeitos dos fármacos , Linfócitos T Reguladores/efeitos dos fármacos , Células Tumorais Cultivadas
15.
Cancer Res ; 80(13): 2874-2888, 2020 07 01.
Artigo em Inglês | MEDLINE | ID: mdl-32265223

RESUMO

Myeloid-derived suppressor cells (MDSC) include immature monocytic (M-MDSC) and granulocytic (PMN-MDSC) cells that share the ability to suppress adaptive immunity and to hinder the effectiveness of anticancer treatments. Of note, in response to IFNγ, M-MDSCs release the tumor-promoting and immunosuppressive molecule nitric oxide (NO), whereas macrophages largely express antitumor properties. Investigating these opposing activities, we found that tumor-derived prostaglandin E2 (PGE2) induces nuclear accumulation of p50 NF-κB in M-MDSCs, diverting their response to IFNγ toward NO-mediated immunosuppression and reducing TNFα expression. At the genome level, p50 NF-κB promoted binding of STAT1 to regulatory regions of selected IFNγ-dependent genes, including inducible nitric oxide synthase (Nos2). In agreement, ablation of p50 as well as pharmacologic inhibition of either the PGE2 receptor EP2 or NO production reprogrammed M-MDSCs toward a NOS2low/TNFαhigh phenotype, restoring the in vivo antitumor activity of IFNγ. Our results indicate that inhibition of the PGE2/p50/NO axis prevents MDSC-suppressive functions and restores the efficacy of anticancer immunotherapy. SIGNIFICANCE: Tumor-derived PGE2-mediated induction of nuclear p50 NF-κB epigenetically reprograms the response of monocytic cells to IFNγ toward an immunosuppressive phenotype, thus retrieving the anticancer properties of IFNγ. GRAPHICAL ABSTRACT: http://cancerres.aacrjournals.org/content/canres/80/13/2874/F1.large.jpg.


Assuntos
Diferenciação Celular , Neoplasias Colorretais/patologia , Dinoprostona/farmacologia , Monócitos/patologia , Células Supressoras Mieloides/patologia , Subunidade p50 de NF-kappa B/metabolismo , Neoplasias Pancreáticas/patologia , Animais , Apoptose , Proliferação de Células , Neoplasias Colorretais/tratamento farmacológico , Neoplasias Colorretais/imunologia , Neoplasias Colorretais/metabolismo , Humanos , Tolerância Imunológica , Interferon gama/metabolismo , Melanoma Experimental/tratamento farmacológico , Melanoma Experimental/imunologia , Melanoma Experimental/metabolismo , Melanoma Experimental/patologia , Camundongos , Monócitos/efeitos dos fármacos , Monócitos/imunologia , Monócitos/metabolismo , Células Supressoras Mieloides/efeitos dos fármacos , Células Supressoras Mieloides/imunologia , Células Supressoras Mieloides/metabolismo , Subunidade p50 de NF-kappa B/genética , Óxido Nítrico/metabolismo , Ocitócicos/farmacologia , Neoplasias Pancreáticas/tratamento farmacológico , Neoplasias Pancreáticas/imunologia , Neoplasias Pancreáticas/metabolismo , Células Tumorais Cultivadas
16.
J Immunother Cancer ; 8(1)2020 03.
Artigo em Inglês | MEDLINE | ID: mdl-32152220

RESUMO

BACKGROUND: Interferon-α (IFN-α) plays a pivotal role in host antitumor immunity, and the evasion of IFN-α signaling pathway can lead to IFN-α resistance during the treatment of cancer. Although the interplay between IFN-α and tumor cells has been extensively investigated in differentiated tumor cells, much less attention has been directed to tumor-repopulating cells (TRCs). METHODS: Three-dimentional soft fibrin matrix was used to select and grow highly malignant and tumorigenic melanoma TRCs. The regulation of integrin ß3 (ITGB3)-c-SRC-STAT signaling pathway in melanoma TRCs was investigated both in vitro and in vivo. The relevant mRNA and protein expression levels were analyzed by qRT-PCR and western blot analysis. Immunoprecipitation and chromatin immunoprecipitation (ChIP) followed by qPCR (ChIP-qPCR) assays were performed to detect protein-protein and protein-DNA interactions. The clinical impacts of retinoic acid inducible gene-I (RIG-I) were assessed in melanoma datasets obtained from The Cancer Genome Atlas and Gene Expression Omnibus profiles. RESULTS: IFN-α-induced apoptosis was decreased in melanoma TRCs. Compared with conventional flask-cultured cells, IFN-α-mediated STAT1 activation was diminished in melanoma TRCs. Decreased expression of RIG-I in melanoma TRCs led to diminished activation of STAT1 via enhancing the interaction between Src homology region 2 domain-containing phosphatase-1 and STAT1. In addition, low expression levels of RIG-I correlated with poor prognosis in patients with melanoma. STAT3 was highly phosphorylated in TRCs and knockdown of STAT3 reversed the downregulation of RIG-I in TRCs. Knockdown of STAT3 resulted in STAT1 activation and increased expression of the pro-apoptosis genes in IFN-α-treated TRCs. Combined treatment of STAT3 inhibitor and IFN-α increased the apoptosis rate of TRCs. Disruption of ITGB3/c-SRC/STAT3 signaling pathway significantly elevated the efficiency of IFN-α-induced apoptosis of TRCs. CONCLUSIONS: In melanoma TRCs, ITGB3-c-SRC-STAT3 pathway caused RIG-I repression and then affect STAT1 activation to cause resistance to IFN-α-induced apoptosis. RIG-I is a prognostic marker in patients with melanoma. Combination of STAT3 inhibitor and IFN-α could enhance the efficacy of melanoma treatment. Our findings may provide a new concept of combinatorial treatment for future immunotherapy.


Assuntos
Proteína DEAD-box 58/metabolismo , Integrina beta3/metabolismo , Interferon-alfa/farmacologia , Melanoma Experimental/tratamento farmacológico , Melanoma/tratamento farmacológico , Proteínas Proto-Oncogênicas pp60(c-src)/metabolismo , Fator de Transcrição STAT3/metabolismo , Animais , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Proteína DEAD-box 58/antagonistas & inibidores , Proteína DEAD-box 58/genética , Regulação para Baixo , Feminino , Células Hep G2 , Humanos , Fatores Imunológicos/farmacologia , Melanoma/imunologia , Melanoma/metabolismo , Melanoma/patologia , Melanoma Experimental/imunologia , Melanoma Experimental/metabolismo , Melanoma Experimental/patologia , Camundongos , Camundongos Endogâmicos C57BL , Prognóstico , Fator de Transcrição STAT1/metabolismo , Transdução de Sinais , Taxa de Sobrevida
17.
Proc Natl Acad Sci U S A ; 117(11): 6047-6055, 2020 03 17.
Artigo em Inglês | MEDLINE | ID: mdl-32123114

RESUMO

Interleukin (IL)-2 and IL-21 dichotomously shape CD8+ T cell differentiation. IL-2 drives terminal differentiation, generating cells that are poorly effective against tumors, whereas IL-21 promotes stem cell memory T cells (TSCM) and antitumor responses. Here we investigated the role of metabolic programming in the developmental differences induced by these cytokines. IL-2 promoted effector-like metabolism and aerobic glycolysis, robustly inducing lactate dehydrogenase (LDH) and lactate production, whereas IL-21 maintained a metabolically quiescent state dependent on oxidative phosphorylation. LDH inhibition rewired IL-2-induced effects, promoting pyruvate entry into the tricarboxylic acid cycle and inhibiting terminal effector and exhaustion programs, including mRNA expression of members of the NR4A family of nuclear receptors, as well as Prdm1 and Xbp1 While deletion of Ldha prevented development of cells with antitumor effector function, transient LDH inhibition enhanced the generation of memory cells capable of triggering robust antitumor responses after adoptive transfer. LDH inhibition did not significantly affect IL-21-induced metabolism but caused major transcriptomic changes, including the suppression of IL-21-induced exhaustion markers LAG3, PD1, 2B4, and TIM3. LDH inhibition combined with IL-21 increased the formation of TSCM cells, resulting in more profound antitumor responses and prolonged host survival. These findings indicate a pivotal role for LDH in modulating cytokine-mediated T cell differentiation and underscore the therapeutic potential of transiently inhibiting LDH during adoptive T cell-based immunotherapy, with an unanticipated cooperative antitumor effect of LDH inhibition and IL-21.


Assuntos
Linfócitos T CD8-Positivos/imunologia , Inibidores Enzimáticos/farmacologia , Interleucinas/metabolismo , L-Lactato Desidrogenase/antagonistas & inibidores , Melanoma Experimental/terapia , Células-Tronco/imunologia , Animais , Linfócitos T CD8-Positivos/efeitos dos fármacos , Linfócitos T CD8-Positivos/metabolismo , Linfócitos T CD8-Positivos/transplante , Diferenciação Celular/efeitos dos fármacos , Diferenciação Celular/imunologia , Linhagem Celular Tumoral/transplante , Humanos , Memória Imunológica , Imunoterapia Adotiva/métodos , Interleucina-2/imunologia , Interleucina-2/metabolismo , Interleucinas/imunologia , L-Lactato Desidrogenase/metabolismo , Melanoma Experimental/imunologia , Camundongos , Cultura Primária de Células , Células-Tronco/efeitos dos fármacos , Células-Tronco/metabolismo
18.
J Immunother Cancer ; 8(1)2020 03.
Artigo em Inglês | MEDLINE | ID: mdl-32217757

RESUMO

BACKGROUND: CD6 is a lymphocyte surface co-receptor physically associated with the T-cell receptor (TCR)/CD3 complex at the center of the immunological synapse. There, CD6 assists in cell-to-cell contact stabilization and modulation of activation/differentiation events through interaction with CD166/ALCAM (activated leukocyte cell adhesion molecule), its main reported ligand. While accumulating evidence is attracting new interest on targeting CD6 for therapeutic purposes in autoimmune disorders, little is known on its potential in cancer. In an attempt to elucidate the in vivo relevance of blocking CD6-mediated interactions in health and disease, we explored the consequences of expressing high circulating levels of a soluble form CD6 (sCD6) as a decoy receptor. METHODS: High sCD6 serum levels were achieved by using transgenic C57BL/6 mice expressing human sCD6 under the control of lymphoid-specific transcriptional elements (shCD6LckEµTg) or wild type either transduced with hepatotropic adeno-associated virus coding for mouse sCD6 or undergoing repeated infusions of recombinant human sCD6 protein. Characterization of sCD6-induced changes was performed by ex vivo flow cytometry and functional analyses of mouse lymphoid organ cells. The in vivo relevance of those changes was explored by challenging mice with subcutaneous or metastatic tumors induced by syngeneic cancer cells of different lineage origins. RESULTS: Through a combination of in vitro and in vivo studies, we show that circulating sCD6 expression induces defective regulatory T cell (Treg) generation and function, decreased CD166/ALCAM-mediated tumor cell proliferation/migration and impaired galectin-induced T-cell apoptosis, supporting the fact that sCD6 modulates antitumor lymphocyte effector function and tumorigenesis. Accordingly, sCD6 expression in vivo resulted in delayed subcutaneous tumor growth and/or reduced metastasis on challenge of mice with syngeneic cancer cells. CONCLUSIONS: Evidence is provided for the disruption of CD6 receptor-ligand interactions as a feasible immunomodulatory approach in cancer.


Assuntos
Antígenos CD/sangue , Antígenos de Diferenciação de Linfócitos T/sangue , Neoplasias Pulmonares/imunologia , Linfoma de Células T/imunologia , Melanoma Experimental/imunologia , Sarcoma Experimental/imunologia , Linfócitos T Reguladores/imunologia , Molécula de Adesão de Leucócito Ativado/imunologia , Molécula de Adesão de Leucócito Ativado/metabolismo , Animais , Antígenos CD/administração & dosagem , Antígenos CD/biossíntese , Antígenos CD/genética , Antígenos de Diferenciação de Linfócitos T/administração & dosagem , Antígenos de Diferenciação de Linfócitos T/biossíntese , Antígenos de Diferenciação de Linfócitos T/genética , Apoptose/fisiologia , Diferenciação Celular/fisiologia , Linhagem Celular Tumoral , Feminino , Humanos , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/secundário , Linfoma de Células T/metabolismo , Masculino , Melanoma Experimental/sangue , Melanoma Experimental/patologia , Melanoma Experimental/terapia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Proteínas Recombinantes/administração & dosagem , Proteínas Recombinantes/sangue , Proteínas Recombinantes/genética , Sarcoma Experimental/sangue , Sarcoma Experimental/patologia , Linfócitos T Reguladores/metabolismo
19.
J Immunother Cancer ; 8(1)2020 03.
Artigo em Inglês | MEDLINE | ID: mdl-32217759

RESUMO

BACKGROUND: Tumor-associated macrophages (TAMs) resemble M2-polarized cells with potent immunosuppressive activity and play a pivotal role in tumor growth and progression. Converting TAMs to proinflammatory M1-like phenotype is thus an attractive strategy for antitumor immunotherapy. METHODS: A mouse IgG1 (kappa) monoclonal Ab, M-860, specific to human lactoferrin (LTF) was generated by using the traditional hybridoma cell fusion technology. TAMs were generated by culturing human and mouse CD14+ monocytes in tumor-conditioned media containing a cytokine cocktail containing recombinant interleukin-4 (IL-4), interleukin-10 (IL-10) and macrophage colony stimulating factor (M-CSF). TAMs after treatment with immunocomplex (IC) between human LTF and M860 (LTF-IC) were phenotypically and functionally characterized by flow cytometry (FACS), ELISA, Q-PCR and killing assays. The antitumor effects of LTF-IC were further analyzed using in vivo experiments employing tumor-bearing human FcγRIIa-transgenic mouse models. RESULTS: Through coligation of membrane-bound CD14 and FcγRIIa, LTF-IC rendered TAMs not only M2 to M1 conversion, evidenced by increased tumor necrosis factor α production, down-regulated M2-specific markers (CD206, arginase-1 and vascular endothelial growth factor) and upregulated M1-specific markers (CD86 and HLA-DR) expression, but also potent tumoricidal activity in vitro. LTF-IC administration conferred antitumor protective efficacy and prolonged animal survival in FcγRIIa-transgenic mice, accompanied by accumulation of M1-like macrophages as well as significantly reduced infiltration of immunosuppressive myeloid-derived suppressor cells and regulatory T cells in solid tumor tissues. CONCLUSIONS: LTF-IC is a promising cancer therapeutic agent capable of converting TAMs into tumoricidal M1-like cells.


Assuntos
Anticorpos Monoclonais/farmacologia , Citocinas/imunologia , Lactoferrina/imunologia , Macrófagos/imunologia , Melanoma Experimental/imunologia , Linfócitos T Reguladores/imunologia , Animais , Anticorpos Monoclonais/imunologia , Complexo Antígeno-Anticorpo/imunologia , Linhagem Celular Tumoral , Técnicas de Cocultura , Meios de Cultivo Condicionados , Modelos Animais de Doenças , Humanos , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Melanoma Experimental/tratamento farmacológico , Melanoma Experimental/metabolismo , Melanoma Experimental/patologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Fenótipo
20.
Nat Commun ; 11(1): 1562, 2020 03 26.
Artigo em Inglês | MEDLINE | ID: mdl-32218434

RESUMO

CCL5 is a unique chemokine with distinct stage and cell-type specificities for regulating inflammation, but how these specificities are achieved and how CCL5 modulates immune responses is not well understood. Here we identify two stage-specific enhancers: the proximal enhancer mediates the constitutive CCL5 expression during the steady state, while the distal enhancer located 1.35 Mb from the promoter induces CCL5 expression in activated cells. Both enhancers are antagonized by RUNX/CBFß complexes, and SATB1 further mediates the long-distance interaction of the distal enhancer with the promoter. Deletion of the proximal enhancer decreases CCL5 expression and augments the cytotoxic activity of tissue-resident T and NK cells, which coincides with reduced melanoma metastasis in mouse models. By contrast, increased CCL5 expression resulting from RUNX3 mutation is associated with more tumor metastasis in the lung. Collectively, our results suggest that RUNX3-mediated CCL5 repression is critical for modulating anti-tumor immunity.


Assuntos
Quimiocina CCL5/genética , Subunidades alfa de Fatores de Ligação ao Core/metabolismo , Elementos Facilitadores Genéticos/genética , Regulação da Expressão Gênica , Imunidade , Animais , Antígenos CD/metabolismo , Subunidade beta de Fator de Ligação ao Core/metabolismo , Homeostase/genética , Imunidade/genética , Ativação Linfocitária/imunologia , Proteínas de Ligação à Região de Interação com a Matriz/metabolismo , Melanoma Experimental/genética , Melanoma Experimental/imunologia , Melanoma Experimental/patologia , Camundongos Endogâmicos C57BL , Camundongos Transgênicos
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