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1.
J Neurooncol ; 139(2): 281-291, 2018 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-29730815

RESUMO

BACKGROUND: PD-L1 expression has been evaluated as a predictive biomarker for immunotherapy in numerous tumor types. However, very limited data are available in pediatric brain tumors. The aim of this study was to characterize PD-1 and PD-L1 expressions of four pediatric malignant brain tumors and gene expression profile. METHODS: This study included 89 pediatric patients receiving standard treatment at Seoul National University Children's Hospital and Seoul National University Bundang Hospital between 1990 and 2014: atypical teratoid/rhabdoid tumor (AT/RT) 20; ependymoma (EPN) 20; high grade glioma (HGG) 21; and medulloblastoma (MBL) 28. We performed immunohistochemistry assays for PD-1 and PD-L1. To characterize the gene expression, a custom immune-response focused gene panel was used. RESULTS: PD-1 expression was positive in 7 (35%) AT/RT, 7 (35%) EPN, 4 (19%) HGG, and 3 (11%) MBL patients. PD-L1 expression was positive in 8 (40%) AT/RT, 4 (20%) EPN, and 4 (19%) HGG; negative in all MBL patients. There was no statistically significant difference in the overall survival of PD-L1 positive patients. The gene expression analysis demonstrated differences in two clustering functional categories: cell-cell signaling and antigen presentation pathway. CONCLUSIONS: AT/RT, EPN, and HGG showed a relatively higher expression rate of PD-L1 (19-40%). This suggests these tumor types might be good candidates for PD-1 checkpoint blockade. We determined that gene expression may potentially serve as a molecular tool in predicting which patients will respond to immunotherapy. Further investigation is required to better understand the predictive and prognostic role of PD-L1 in pediatric brain tumors.


Assuntos
Antígeno B7-H1/metabolismo , Neoplasias Encefálicas/imunologia , Receptor de Morte Celular Programada 1/metabolismo , Adolescente , Biomarcadores Tumorais/metabolismo , Encéfalo/imunologia , Encéfalo/patologia , Neoplasias Encefálicas/mortalidade , Neoplasias Encefálicas/patologia , Neoplasias Encefálicas/terapia , Criança , Pré-Escolar , Ependimoma/imunologia , Ependimoma/mortalidade , Ependimoma/patologia , Ependimoma/terapia , Feminino , Seguimentos , Regulação Neoplásica da Expressão Gênica , Glioma/imunologia , Glioma/mortalidade , Glioma/patologia , Glioma/terapia , Humanos , Lactente , Masculino , RNA Mensageiro/metabolismo , Estudos Retrospectivos , Tumor Rabdoide/imunologia , Tumor Rabdoide/mortalidade , Tumor Rabdoide/patologia , Tumor Rabdoide/terapia , Análise de Sobrevida , Teratoma/imunologia , Teratoma/mortalidade , Teratoma/patologia , Teratoma/terapia
2.
Mol Cancer Ther ; 21(2): 282-293, 2022 02.
Artigo em Inglês | MEDLINE | ID: mdl-34815361

RESUMO

The innate immune agonist STING (STimulator of INterferon Genes) binds its natural ligand 2'3'-cGAMP (cyclic guanosine-adenosine monophosphate) and initiates type I IFN production. This promotes systemic antigen-specific CD8+ T-cell priming that eventually provides potent antitumor activity. To exploit this mechanism, we synthesized a novel STING agonist, MSA-1, that activates both mouse and human STING with higher in vitro potency than cGAMP. Following intratumoral administration of MSA-1 to a panel of syngeneic mouse tumors on immune-competent mice, cytokine upregulation and its exposure were detected in plasma, other tissues, injected tumors, and noninjected tumors. This was accompanied by effective antitumor activity. Mechanistic studies in immune-deficient mice suggested that antitumor activity of intratumorally dosed STING agonists is in part due to necrosis and/or innate immune responses such as TNF-α activity, but development of a robust adaptive antitumor immunity is necessary for complete tumor elimination. Combination with PD-1 blockade in anti-PD-1-resistant murine models showed that MSA-1 may synergize with checkpoint inhibitors but can also provide superior tumor control as a single agent. We show for the first time that potent cyclic dinucleotides can promote a rapid and stronger induction of the same genes eventually regulated by PD-1 blockade. This may have contributed to the relatively early tumor control observed with MSA-1. Taken together, these data strongly support the development of STING agonists as therapy for patients with aggressive tumors that are partially responsive or nonresponsive to single-agent anti-PD-1 treatment by enhancing the anti-PD-1 immune profile.


Assuntos
Imunidade Inata/imunologia , Imunoterapia/métodos , Interferons/metabolismo , Neoplasias/imunologia , Animais , Linhagem Celular Tumoral , Feminino , Humanos , Camundongos
3.
Mol Cancer Ther ; 21(3): 427-439, 2022 03 01.
Artigo em Inglês | MEDLINE | ID: mdl-34965960

RESUMO

Targeting the programmed death 1/programmed death ligand 1 (PD-1/PD-L1) pathway with immunotherapy has revolutionized the treatment of many cancers. Somatic tumor mutational burden (TMB) and T-cell-inflamed gene expression profile (GEP) are clinically validated pan-tumor genomic biomarkers that can predict responsiveness to anti-PD-1/PD-L1 monotherapy in many tumor types. We analyzed the association between these biomarkers and the efficacy of PD-1 inhibitor in 11 commonly used preclinical syngeneic tumor mouse models using murinized rat anti-mouse PD-1 DX400 antibody muDX400, a surrogate for pembrolizumab. Response to muDX400 treatment was broadly classified into three categories: highly responsive, partially responsive, and intrinsically resistant to therapy. Molecular and cellular profiling validated differences in immune cell infiltration and activation in the tumor microenvironment of muDX400-responsive tumors. Baseline and on-treatment genomic analysis showed an association between TMB, murine T-cell-inflamed gene expression profile (murine-GEP), and response to muDX400 treatment. We extended our analysis to investigate a canonical set of cancer and immune biology-related gene signatures, including signatures of angiogenesis, myeloid-derived suppressor cells, and stromal/epithelial-to-mesenchymal transition/TGFß biology previously shown to be inversely associated with the clinical efficacy of immune checkpoint blockade. Finally, we evaluated the association between murine-GEP and preclinical efficacy with standard-of-care chemotherapy or antiangiogenic agents that previously demonstrated promising clinical activity, in combination with muDX400. Our profiling studies begin to elucidate the underlying biological mechanisms of response and resistance to PD-1/PD-L1 blockade represented by these models, thereby providing insight into which models are most appropriate for the evaluation of orthogonal combination strategies.


Assuntos
Antígeno B7-H1 , Imunoterapia , Neoplasias , Receptor de Morte Celular Programada 1 , Animais , Antígeno B7-H1/antagonistas & inibidores , Biomarcadores Tumorais/genética , Linhagem Celular Tumoral , Modelos Animais de Doenças , Humanos , Inibidores de Checkpoint Imunológico , Camundongos , Neoplasias/tratamento farmacológico , Neoplasias/genética , Receptor de Morte Celular Programada 1/antagonistas & inibidores , Microambiente Tumoral
4.
J Immunol ; 182(9): 5748-56, 2009 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-19380822

RESUMO

IL-27 is secreted by APCs in response to inflammatory stimuli and exerts a proinflammatory Th1-enhancing activity but also has significant anti-inflammatory functions. We examined the molecular mechanism by which IL-27 regulates TGFbeta plus IL-6- or IL-23-dependent Th17 development in the mouse and human systems. IL-27 inhibited the production of IL-17A and IL-17F in naive T cells by suppressing, in a STAT1-dependent manner, the expression of the Th17-specific transcription factor RORgamma t. The in vivo significance of the role of IL-27 was addressed in delayed-type hypersensitivity response and experimental autoimmune encephalomyelitis (EAE). By generating mice deficient for the p28 subunit of IL-27, we showed that IL-27 regulated the severity of delayed-type hypersensitivity response and EAE through its effects on Th17 cells. Furthermore, up-regulation of IL-10 in the CNS, which usually occurs late after EAE onset and plays a role in the resolution of the disease, was notably absent in IL-27p28(-/-) mice. These results show that IL-27 acts as a negative regulator of the developing IL-17A response in vivo, suggesting a potential therapeutic role for IL-27 in autoimmune diseases.


Assuntos
Linhagem da Célula/imunologia , Inibidores do Crescimento/fisiologia , Interleucina-17/antagonistas & inibidores , Interleucinas/fisiologia , Receptores do Ácido Retinoico/antagonistas & inibidores , Receptores dos Hormônios Tireóideos/antagonistas & inibidores , Linfócitos T Auxiliares-Indutores/imunologia , Animais , Diferenciação Celular/genética , Diferenciação Celular/imunologia , Linhagem da Célula/genética , Células Cultivadas , Encefalomielite Autoimune Experimental/genética , Encefalomielite Autoimune Experimental/imunologia , Encefalomielite Autoimune Experimental/patologia , Regulação da Expressão Gênica/imunologia , Predisposição Genética para Doença , Inibidores do Crescimento/deficiência , Inibidores do Crescimento/genética , Humanos , Interleucina-17/biossíntese , Interleucinas/deficiência , Interleucinas/genética , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Membro 3 do Grupo F da Subfamília 1 de Receptores Nucleares , Subunidades Proteicas/deficiência , Subunidades Proteicas/genética , Subunidades Proteicas/fisiologia , Receptores do Ácido Retinoico/biossíntese , Receptores do Ácido Retinoico/genética , Receptores dos Hormônios Tireóideos/biossíntese , Receptores dos Hormônios Tireóideos/genética , Linfócitos T Auxiliares-Indutores/metabolismo , Linfócitos T Auxiliares-Indutores/patologia
5.
Front Immunol ; 12: 752348, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34912335

RESUMO

Programmed cell death-1 (PD-1) blockade has a profound effect on the ability of the immune system to eliminate tumors, but many questions remain about the cell types involved and the underlying mechanisms of immune activation. To shed some light on this, the cellular and molecular events following inhibition of PD-1 signaling was investigated in the MC-38 colon carcinoma model using constitutive (PD-1 KO) and conditional (PD1cKO) mice and in wild-type mice treated with PD-1 antibody. The impact on both tumor growth and the development of tumor immunity was assessed. In the PD-1cKO mice, a complete deletion of Pdcd1 in tumor-infiltrating T cells (TILs) after tamoxifen treatment led to the inhibition of tumor growth of both small and large tumors. Extensive immune phenotypic analysis of the TILs by flow and mass cytometry identified 20-different T cell subsets of which specifically 5-CD8 positive ones expanded in all three models after PD-1 blockade. All five subsets expressed granzyme B and interferon gamma (IFNγ). Gene expression analysis of the tumor further supported the phenotypic analysis in both PD-1cKO- and PD-1 Ab-treated mice and showed an upregulation of pathways related to CD4 and CD8 T-cell activation, enhanced signaling through costimulatory molecules and IFNγ, and non-T-cell processes. Altogether, using PD-1cKO mice, we define the intrinsic nature of PD-1 suppression of CD8 T-cell responses in tumor immunity.


Assuntos
Linfócitos T CD8-Positivos/imunologia , Linfócitos do Interstício Tumoral/imunologia , Neoplasias Experimentais/imunologia , Receptor de Morte Celular Programada 1/imunologia , Subpopulações de Linfócitos T/imunologia , Animais , Linfócitos T CD8-Positivos/efeitos dos fármacos , Feminino , Inibidores de Checkpoint Imunológico/farmacologia , Linfócitos do Interstício Tumoral/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Receptor de Morte Celular Programada 1/deficiência
6.
Infect Immun ; 78(3): 1353-63, 2010 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-20065029

RESUMO

Paired immunoglobulin-like type 2 receptors (PILRs) inhibitory PILRalpha and activating PILRbeta are predominantly expressed on myeloid cells. Their functions in host defense and inflammation are largely unknown, and in this study, we evaluated their roles in an acute Staphylococcus aureus pneumonia model. Compared to their respective controls, Pilrb(-/-) mice or mice in which PILRalpha was activated with an agonistic antibody showed improved clearance of pulmonary staphylococci and improved survival. These mice had reduced serum or bronchoalveolar lavage fluid levels of interleukin-1beta (IL-1beta), tumor necrosis factor alpha (TNF-alpha), and IL-6 and elevated levels of gamma interferon (IFN-gamma), IL-12, and IL-10. In contrast, mice in which PILRbeta was activated had increased lung bacterial burdens and higher mortality coupled with an intense proinflammatory response with highly elevated levels of IL-1beta, TNF-alpha, and IL-6. Treatment groups with reduced bacterial burdens had higher levels of Keratinocyte-derived chemokine (KC), macrophage inflammatory protein 2 (MIP-2), and MIP-1alpha in bronchoalveolar lavage fluid and an increased influx of neutrophils and macrophages to the lungs. Consistent with our in vivo findings, bone marrow-derived macrophages from Pilrb(-/-) mice released significantly less IL-1beta and TNF-alpha and more IFN-gamma and IL-12 than did the wild-type macrophages when directly stimulated with heat-killed S. aureus. To our knowledge, this is the first evidence that S. aureus directly interacts with PILRbeta. It provides a mechanism by which manipulating the balance in favor of an inhibitory PILR signal, by activation of PILRalpha or deletion of PILRbeta, helps to control acute S. aureus-mediated pneumonia and attenuate the inflammatory response. These results highlight the importance of PILRs in innate immunity and the control of inflammation.


Assuntos
Pneumonia Estafilocócica/imunologia , Pneumonia Estafilocócica/patologia , Receptores Imunológicos/metabolismo , Transdução de Sinais , Staphylococcus aureus/imunologia , Animais , Análise Química do Sangue , Líquido da Lavagem Broncoalveolar/química , Contagem de Colônia Microbiana , Citocinas/análise , Citocinas/sangue , Feminino , Pulmão/microbiologia , Macrófagos/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Receptores Imunológicos/deficiência , Receptores Imunológicos/imunologia , Análise de Sobrevida
7.
Breast Cancer Res ; 12(3): R32, 2010.
Artigo em Inglês | MEDLINE | ID: mdl-20525379

RESUMO

INTRODUCTION: Anterior-gradient 2 (AGR2) is an estrogen-responsive secreted protein. Its upregulation has been well documented in a number of cancers, particularly breast cancer, for which mixed data exist on the prognostic implications of AGR2 expression. Although emerging evidence indicates that AGR2 is associated with poor prognosis, its function and impact on cancer-relevant pathways have not been elucidated in breast cancer. METHODS: To investigate the biologic role of AGR2 in breast cancer, AGR2 was transiently knocked down, by using siRNA, in T47 D and ZR-75-1 (estrogen receptor-alpha (ER)-positive) and MDA-MB-231 and SK-BR-3 (ER-negative) human breast cancer cell lines. The impact of silencing AGR2 was evaluated in both anchorage-dependent and anchorage-independent growth (soft agar, spheroid) assays. Cell-cycle profiles in ER-positive cell lines were determined with BrdU incorporation, and cell death was measured with Annexin V, JC-1, and F7-26 staining. After transiently silencing AGR2 or stimulating with recombinant AGR2, modulation of key regulators of growth and survival pathways was assessed with Western blot. Combination studies of AGR2 knockdown with the antiestrogens tamoxifen and fulvestrant were carried out and assessed at the level of anchorage-dependent growth inhibition and target modulation (cyclin D1, ER). RESULTS: AGR2 knockdown inhibited growth in anchorage-dependent and anchorage-independent assays, with a more-pronounced effect in ER-positive cell lines. Cyclin D1 levels and BrdU incorporation were reduced with AGR2 knockdown. Conversely, cyclin D1 was induced with recombinant AGR2. AGR2 knockdown induced cell death in ZR-75-1 and T47 D cells, and also downregulated survivin and c-Myc. Evidence of AGR2-ER crosstalk was demonstrated by a reduction of ER at the protein level after transiently silencing AGR2. AGR2 knockdown in combination with fulvestrant or tamoxifen did not preclude the efficacy of the antiestrogens, but enhanced it. In addition, p-Src, implicated in tamoxifen resistance, was downregulated with AGR2 knockdown. CONCLUSIONS: Transiently silencing AGR2 in ER-positive breast cancer cell lines inhibited cell growth and cell-cycle progression and induced cell death. Breast cancer drivers (ER and cyclin D1) as well as cancer-signaling nodes (pSrc, c-Myc, and survivin) were demonstrated to be downstream of AGR2. Collectively, the data presented support the utility of anti-AGR2 therapy in ER-positive breast cancers because of its impact on cancer-relevant pathways.


Assuntos
Neoplasias da Mama/metabolismo , Proliferação de Células , Ciclina D1/metabolismo , Receptor alfa de Estrogênio/metabolismo , Proteínas Associadas aos Microtúbulos/metabolismo , Proteínas/fisiologia , Animais , Apoptose/efeitos dos fármacos , Western Blotting , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/genética , Adesão Celular/efeitos dos fármacos , Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Ensaio de Unidades Formadoras de Colônias , Ciclina D1/genética , Estradiol/análogos & derivados , Estradiol/farmacologia , Antagonistas de Estrogênios/farmacologia , Receptor alfa de Estrogênio/genética , Feminino , Citometria de Fluxo , Fulvestranto , Humanos , Imunização , Proteínas Inibidoras de Apoptose , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Proteínas Associadas aos Microtúbulos/genética , Mucoproteínas , Proteínas Oncogênicas , Proteínas/antagonistas & inibidores , RNA Mensageiro/genética , RNA Interferente Pequeno/farmacologia , Ratos , Ratos Endogâmicos Lew , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Survivina , Tamoxifeno/farmacologia
8.
Mol Ther ; 17(3): 508-15, 2009 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-19127252

RESUMO

Intravascular delivery (1.5 x 10(9) particles and higher) of recombinant adenovirus (rAd) induces myeloid cell mediated, self-limiting hemodynamic responses in normal mice. However, we observed anaphylactoid-type reactions and exacerbated hemodynamic events following rAd injection in mice bearing malignant 4T1 mammary carcinoma. Because 4T1 tumors induce significant CD11b(+)Gr-1(+) myeloid cell expansion and activation, we set to determine whether this causes rAd-induced exaggerated responses. When treated with a single intravenous dose (1 x 10(10) particles) of rAd, mice implanted with 4T1 carcinoma succumbed due to the anaphylactoid-type reactions. In contrast, normal mice and mice implanted with a related mammary carcinoma (66cl4) that does not induce CD11b(+)Gr-1(+) cell expansion, showed minimal responses. Depletion of phagocytic CD11b(+)Gr-1(+) cells prior to rAd delivery protected 4T1 tumor-bearing animals, whereas passive transfer of CD11b(+)Gr-1(+) cells from 4T1 tumor-bearing animals was sufficient to convey susceptibility to anaphylactoid-type reactions in normal animals. We further show that there is upregulation of nitric oxide and leukotriene signaling pathways in the 4T1 tumor-induced CD11b(+)Gr-1(+) myeloid cells and that pretreating mice with inhibitors of nitric oxide synthetase and leukotrienes can attenuate the anaphylactoid-type reactions. These data show that malignant tumor growth can alter CD11b(+)Gr-1(+) myeloid cells, rendering hosts susceptible to anaphylactoid-type reactions upon intravascular treatment with rAd.


Assuntos
Adenoviridae/imunologia , Anafilaxia/metabolismo , Antígeno CD11b/imunologia , Diferenciação Celular , Terapia Genética/efeitos adversos , Neoplasias/patologia , Receptores de Quimiocinas/metabolismo , Adenoviridae/genética , Anafilaxia/fisiopatologia , Animais , Sistema Cardiovascular/metabolismo , Linhagem Celular Tumoral , Suscetibilidade a Doenças , Feminino , Frequência Cardíaca , Humanos , Antagonistas de Leucotrienos/farmacologia , Lipossomos , Camundongos , Células Mieloides/metabolismo , Transplante de Neoplasias , Neoplasias/genética , Neoplasias/imunologia , Neoplasias/terapia , Óxido Nítrico Sintase/antagonistas & inibidores , Óxido Nítrico Sintase/metabolismo , Fagócitos , Receptores de Quimiocinas/imunologia , Receptores de Leucotrienos/metabolismo
9.
Nucleic Acids Res ; 36(11): 3802-18, 2008 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-18492722

RESUMO

STAT5A and STAT5B proteins belong to the family of signal transducers and activators of transcription. They are encoded by two separate genes with 91% identity in their amino acid sequences. Despite their high degree of conservation, STAT5A and STAT5B exert non-redundant functions, resulting at least in part from differences in target gene activation. To better characterize the differential contribution of STAT5A and STAT5B in gene regulation, we performed single or double knockdown of STAT5A and STAT5B using small interfering RNA. Subsequent gene expression profiling and RT-qPCR analyses of IL-3-stimulated Ba/F3-beta cells led to the identification of putative novel STAT5 target genes. Chromatin immunoprecipitation assays analyzing the corresponding gene loci identified unusual STAT5 binding sites compared to conventional STAT5 responsive elements. Some of the STAT5 targets identified are upregulated in several human cancers, suggesting that they might represent potential oncogenes in STAT5-associated malignancies.


Assuntos
Regulação da Expressão Gênica , Fator de Transcrição STAT5/fisiologia , Animais , Sítios de Ligação , Fosfatase 1 de Especificidade Dupla/genética , Perfilação da Expressão Gênica , Humanos , Interleucina-3/farmacologia , Camundongos , Neoplasias/genética , Interferência de RNA , Receptores de Complemento/genética , Elementos Reguladores de Transcrição , Fator de Transcrição STAT5/antagonistas & inibidores , Fator de Transcrição STAT5/genética , Ativação Transcricional , Proteína Transmembrana Ativadora e Interagente do CAML/genética
10.
J Clin Invest ; 116(5): 1310-6, 2006 May.
Artigo em Inglês | MEDLINE | ID: mdl-16670770

RESUMO

Uncontrolled mucosal immunity in the gastrointestinal tract of humans results in chronic inflammatory bowel disease (IBD), such as Crohn disease and ulcerative colitis. In early clinical trials as well as in animal models, IL-12 has been implicated as a major mediator of these diseases based on the ability of anti-p40 mAb treatment to reverse intestinal inflammation. The cytokine IL-23 shares the same p40 subunit with IL-12, and the anti-p40 mAbs used in human and mouse IBD studies neutralized the activities of both IL-12 and IL-23. IL-10-deficient mice spontaneously develop enterocolitis. To determine how IL-23 contributes to intestinal inflammation, we studied the disease susceptibility in the absence of either IL-23 or IL-12 in this model, as well as the ability of recombinant IL-23 to exacerbate IBD induced by T cell transfer. Our study shows that in these models, IL-23 is essential for manifestation of chronic intestinal inflammation, whereas IL-12 is not. A critical target of IL-23 is a unique subset of tissue-homing memory T cells, which are specifically activated by IL-23 to produce the proinflammatory mediators IL-17 and IL-6. This pathway may be responsible for chronic intestinal inflammation as well as other chronic autoimmune inflammatory diseases.


Assuntos
Colite/patologia , Inflamação/patologia , Interleucina-17/fisiologia , Interleucina-6/fisiologia , Interleucinas/fisiologia , Linfócitos T/patologia , Animais , Doenças Autoimunes/patologia , Humanos , Interleucina-23 , Subunidade p19 da Interleucina-23 , Mucosa Intestinal/patologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Knockout , Linfócitos T/metabolismo
11.
Immunohorizons ; 3(7): 294-305, 2019 07 11.
Artigo em Inglês | MEDLINE | ID: mdl-31356159

RESUMO

Bromodomain and extraterminal domain (BET) proteins help direct the differentiation of helper T cell subsets, but their role in activated T cell function has not been described in detail. In this study, we investigate various consequences of epigenetic perturbation in human T lymphocytes using MK-8628, a potent and highly selective inhibitor of BET proteins. MK-8628 reduces the expression of canonical transcripts directing the proliferation, activation, and effector function of T lymphocytes. Treatment with MK-8628 abolishes the expression of key cyclins required for cell cycle progression and induces G1 cell cycle arrest in TCR-activated lymphocytes. This antiproliferative phenotype partially results from T lymphocyte apoptosis, which is exacerbated by MK-8628. In naive and memory T cell subsets, MK-8628 antagonizes T cell activation and suppresses polyfunctional cytokine production. Collectively, our results describe potent immunosuppressive effects of BET inhibition on human T cell biology. These results have important implications for immune modulatory targeting of BET proteins in the settings of T cell-driven autoimmune inflammation.


Assuntos
Acetanilidas/farmacologia , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/metabolismo , Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/metabolismo , Compostos Heterocíclicos com 3 Anéis/farmacologia , Imunossupressores/farmacologia , Proteínas/antagonistas & inibidores , Apoptose/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Citocinas/genética , Citocinas/metabolismo , Pontos de Checagem da Fase G1 do Ciclo Celular/efeitos dos fármacos , Expressão Gênica/efeitos dos fármacos , Glicólise/efeitos dos fármacos , Voluntários Saudáveis , Humanos , Ativação Linfocitária/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos
12.
J Immunol Methods ; 456: 7-14, 2018 05.
Artigo em Inglês | MEDLINE | ID: mdl-29427592

RESUMO

LAG3 is an important regulator of T cell homeostasis and studies in mouse tumor models have demonstrated that simultaneously antagonizing LAG3 and PD1 can augment tumor-specific T cell responses and induce tumor rejection. The combined use of LAG3 antagonist antibodies with established anti-PD1 therapies is currently being evaluated in human clinical trials. A functional assay for human LAG3 was developed by co-culture of a Jurkat T-cell lymphoma line overexpressing LAG3 with a Raji B-cell lymphoma line in the presence of staphylococcal enterotoxins. Reversal of LAG3 repression was measured as an increase in IL-2 production or NFAT activation in response to treatment with MK-4280, an anti-human LAG3 antagonist antibody. Changes in cytokines, chemokines, and other mRNA transcripts were in agreement with published in vitro and in vivo models for LAG3 biology which highlights the physiological relevance of the Jurkat functional assay. Additional engineering of PD1 and PDL1 components into the LAG3 assay resulted in a bi-functional assay that is capable of inducing a 10-fold response to individual antibodies blocking either PD1 or LAG3. Importantly, when MK-4280 and pembrolizumab were combined to block both pathways, a synergistic 50-fold increase in response was observed.


Assuntos
Anticorpos Bloqueadores/análise , Antígenos CD/imunologia , Engenharia Celular , Receptor de Morte Celular Programada 1/imunologia , Transdução de Sinais/imunologia , Anticorpos Bloqueadores/imunologia , Antígenos CD/genética , Citometria de Fluxo , Perfilação da Expressão Gênica , Humanos , Células Jurkat , Receptor de Morte Celular Programada 1/genética , Reação em Cadeia da Polimerase em Tempo Real , Transdução de Sinais/genética , Linfócitos T/imunologia , Células Tumorais Cultivadas , Proteína do Gene 3 de Ativação de Linfócitos
13.
J Clin Invest ; 128(2): 644-654, 2018 02 01.
Artigo em Inglês | MEDLINE | ID: mdl-29337311

RESUMO

Blockade of the checkpoint inhibitor programmed death 1 (PD1) has demonstrated remarkable success in the clinic for the treatment of cancer; however, a majority of tumors are resistant to anti-PD1 monotherapy. Numerous ongoing clinical combination therapy studies will likely reveal additional therapeutics that complement anti-PD1 blockade. Recent studies found that immunogenic cell death (ICD) improves T cell responses against different tumors, thus indicating that ICD may further augment antitumor immunity elicited by anti-PD1. Here, we observed antitumor activity following combinatorial therapy with anti-PD1 Ab and the cyclin-dependent kinase inhibitor dinaciclib in immunocompetent mouse tumor models. Dinaciclib induced a type I IFN gene signature within the tumor, leading us to hypothesize that dinaciclib potentiates the effects of anti-PD1 by eliciting ICD. Indeed, tumor cells treated with dinaciclib showed the hallmarks of ICD including surface calreticulin expression and release of high mobility group box 1 (HMGB1) and ATP. Mice treated with both anti-PD1 and dinaciclib showed increased T cell infiltration and DC activation within the tumor, indicating that this combination improves the overall quality of the immune response generated. These findings identify a potential mechanism for the observed benefit of combining dinaciclib and anti-PD1, in which dinaciclib induces ICD, thereby converting the tumor cell into an endogenous vaccine and boosting the effects of anti-PD1.


Assuntos
Adenocarcinoma/tratamento farmacológico , Compostos Bicíclicos Heterocíclicos com Pontes/farmacologia , Receptor de Morte Celular Programada 1/antagonistas & inibidores , Compostos de Piridínio/farmacologia , Trifosfato de Adenosina/química , Animais , Anticorpos Monoclonais/farmacologia , Apoptose , Linfócitos T CD8-Positivos/citologia , Morte Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Óxidos N-Cíclicos , Citocinas/metabolismo , Células Dendríticas/citologia , Sinergismo Farmacológico , Feminino , Proteína HMGB1/metabolismo , Sistema Imunitário/efeitos dos fármacos , Imunoterapia , Indolizinas , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Knockout , Transplante de Neoplasias , Fagocitose , Receptor de Morte Celular Programada 1/metabolismo , Inibidores de Proteínas Quinases/farmacologia
14.
PLoS One ; 7(3): e31680, 2012.
Artigo em Inglês | MEDLINE | ID: mdl-22479310

RESUMO

Paired immunoglobulin-like receptors beta, PILRß, and alpha, PILRα, are related to the Siglec family of receptors and are expressed primarily on cells of the myeloid lineage. PILRß is a DAP12 binding partner expressed on both human and mouse myeloid cells. The potential ligand, CD99, is found on many cell types, such as epithelial cells where it plays a role in migration of immune cells to sites of inflammation. Pilrb deficient mice were challenged with the parasite Toxoplasma gondii in two different models of infection induced inflammation; one involving the establishment of chronic encephalitis and a second mimicking inflammatory bowel disease in order to understand the potential role of this receptor in persistent inflammatory responses. It was found that in the absence of activating signals from PILRß, antigen-presenting cells (APCs) produced increased amounts of IL-27, p28 and promoted IL-10 production in effector T cells. The sustained production of IL-27 led ultimately to enhanced survival after challenge due to dampened immune pathology in the gut. Similar protection was also observed in the CNS during chronic T. gondii infection after i.p. challenge again providing evidence that PILRß is important for regulating aberrant inflammatory responses.


Assuntos
Inflamação/imunologia , Interleucinas/imunologia , Receptores Imunológicos/imunologia , Linfócitos T/imunologia , Animais , Células Cultivadas , Células Dendríticas/imunologia , Células Dendríticas/metabolismo , Encefalite/genética , Encefalite/imunologia , Encefalite/metabolismo , Feminino , Expressão Gênica , Inflamação/genética , Inflamação/metabolismo , Interferon gama/genética , Interferon gama/imunologia , Interferon gama/metabolismo , Interleucinas/genética , Interleucinas/metabolismo , Macrófagos/imunologia , Macrófagos/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos CBA , Camundongos Knockout , Microglia/imunologia , Microglia/metabolismo , Receptores Imunológicos/deficiência , Receptores Imunológicos/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Linfócitos T/metabolismo , Toxoplasma/imunologia , Toxoplasmose Animal/genética , Toxoplasmose Animal/imunologia , Toxoplasmose Animal/metabolismo
15.
Nat Med ; 18(7): 1069-76, 2012 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-22772566

RESUMO

The spondyloarthropathies are a group of rheumatic diseases that are associated with inflammation at anatomically distal sites, particularly the tendon-bone attachments (entheses) and the aortic root. Serum concentrations of interleukin-23 (IL-23) are elevated and polymorphisms in the IL-23 receptor are associated with ankyosing spondylitis, however, it remains unclear whether IL-23 acts locally at the enthesis or distally on circulating cell populations. We show here that IL-23 is essential in enthesitis and acts on previously unidentified IL-23 receptor (IL-23R)(+), RAR-related orphan receptor γt (ROR-γt)(+)CD3(+)CD4(-)CD8(-), stem cell antigen 1 (Sca1)(+) entheseal resident T cells. These cells allow entheses to respond to IL-23 in vitro-in the absence of further cellular recruitment--and to elaborate inflammatory mediators including IL-6, IL-17, IL-22 and chemokine (C-X-C motif) ligand 1 (CXCL1). Notably, the in vivo expression of IL-23 is sufficient to phenocopy the human disease, with the specific and characteristic development of enthesitis and entheseal new bone formation in the initial complete absence of synovitis. As in the human condition, inflammation also develops in vivo at the aortic root and valve, which are structurally similar to entheses. The presence of these entheseal resident cells and their production of IL-22, which activates signal transducer and activator of transcription 3 (STAT3)-dependent osteoblast-mediated bone remodeling, explains why dysregulation of IL-23 results in inflammation at this precise anatomical site.


Assuntos
Interleucina-23/imunologia , Espondiloartropatias/imunologia , Linfócitos T/imunologia , Tendões/imunologia , Animais , Antígenos CD/metabolismo , Aorta/patologia , Artrite Experimental/complicações , Artrite Experimental/imunologia , Artrite Experimental/patologia , Remodelação Óssea , Complexo CD3/metabolismo , Antígenos CD4/metabolismo , Antígenos CD8/metabolismo , Modelos Animais de Doenças , Extremidades/patologia , Citometria de Fluxo , Humanos , Imunização Passiva , Inflamação/complicações , Inflamação/imunologia , Inflamação/patologia , Interleucina-17 , Interleucinas , Camundongos , Membro 3 do Grupo F da Subfamília 1 de Receptores Nucleares/metabolismo , Osteogênese/imunologia , Periósteo/crescimento & desenvolvimento , Receptores de Interleucina/metabolismo , Espondiloartropatias/complicações , Espondiloartropatias/patologia , Tendões/patologia , Células Th17 , Interleucina 22
16.
Cytokine ; 29(6): 256-69, 2005 Mar 21.
Artigo em Inglês | MEDLINE | ID: mdl-15749026

RESUMO

To investigate the potential molecular mediators of tissue-specific recruitment, we explored the influence of different cytokine challenges on gene expression regulation in five primary endothelial cells (ECs), representing two different phenotypes: iliac artery and aortic (macrovascular); lung, colon and dermal (microvascular). We challenged ECs with cytokines that elicit different patterns of inflammatory and immune responses in immune cells: tumor necrosis factor (TNF-alpha), interferon-gamma (IFN-gamma) or interleukin-4 (IL-4), and used microarrays containing approximately 40,000 unique cDNAs, to assess changes in differential gene expression relative to untreated cells. Five hundred and sixty three sequences changed by at least 2.5 fold in one or more of the 15 possible EC /cytokine combinations. The list included highly regulated adhesion molecules, chemokines, cytokines, metalloproteases, and IFN-gamma-induced genes. Overall, IFN-gamma caused the largest number of gene expression changes and its profile was least correlated with IL-4. In addition to clusters that were predominantly EC/cytokine specific, we also observed several clusters that were regulated by more than one cytokine across several ECs. Furthermore, we identified genes that were reciprocally expressed in response to different cytokines that could serve as markers of inflammatory and immune expression. These results confirm the importance of microenvironment in primary ECs that could have important applications in developing targeted therapies for vascular diseases.


Assuntos
Citocinas/farmacologia , Endotélio Vascular/imunologia , Regulação da Expressão Gênica , Células Cultivadas , Células Endoteliais/efeitos dos fármacos , Células Endoteliais/metabolismo , Endotélio Vascular/citologia , Perfilação da Expressão Gênica , Humanos , Mediadores da Inflamação/farmacologia , Interferon gama/farmacologia , Interleucina-4/farmacologia , Análise de Sequência com Séries de Oligonucleotídeos , Reação em Cadeia da Polimerase
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