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1.
Cancer Cell ; 41(3): 602-619.e11, 2023 03 13.
Artigo em Inglês | MEDLINE | ID: mdl-36868226

RESUMO

Tumor cells promote the recruitment of immunosuppressive neutrophils, a subset of myeloid cells driving immune suppression, tumor proliferation, and treatment resistance. Physiologically, neutrophils are known to have a short half-life. Here, we report the identification of a subset of neutrophils that have upregulated expression of cellular senescence markers and persist in the tumor microenvironment. Senescent-like neutrophils express the triggering receptor expressed on myeloid cells 2 (TREM2) and are more immunosuppressive and tumor-promoting than canonical immunosuppressive neutrophils. Genetic and pharmacological elimination of senescent-like neutrophils decreases tumor progression in different mouse models of prostate cancer. Mechanistically, we have found that apolipoprotein E (APOE) secreted by prostate tumor cells binds TREM2 on neutrophils, promoting their senescence. APOE and TREM2 expression increases in prostate cancers and correlates with poor prognosis. Collectively, these results reveal an alternative mechanism of tumor immune evasion and support the development of immune senolytics targeting senescent-like neutrophils for cancer therapy.


Assuntos
Apolipoproteínas E , Neoplasias da Próstata , Animais , Humanos , Masculino , Camundongos , Apolipoproteínas E/metabolismo , Senescência Celular/genética , Glicoproteínas de Membrana/genética , Células Mieloides/metabolismo , Neoplasias da Próstata/metabolismo , Receptores Imunológicos/metabolismo , Microambiente Tumoral
2.
Blood ; 139(5): 732-747, 2022 02 03.
Artigo em Inglês | MEDLINE | ID: mdl-34653238

RESUMO

Splenic marginal zone B-cell lymphoma (SMZL) is a heterogeneous clinico-biological entity. The clinical course is variable, multiple genes are mutated with no unifying mechanism, and essential regulatory pathways and surrounding microenvironments are diverse. We sought to clarify the heterogeneity of SMZL by resolving different subgroups and their underlying genomic abnormalities, pathway signatures, and microenvironment compositions to uncover biomarkers and therapeutic vulnerabilities. We studied 303 SMZL spleen samples collected through the IELSG46 multicenter international study (NCT02945319) by using a multiplatform approach. We carried out genetic and phenotypic analyses, defined self-organized signatures, validated the findings in independent primary tumor metadata and in genetically modified mouse models, and determined correlations with outcome data. We identified 2 prominent genetic clusters in SMZL, termed NNK (58% of cases, harboring NF-κB, NOTCH, and KLF2 modules) and DMT (32% of cases, with DNA-damage response, MAPK, and TLR modules). Genetic aberrations in multiple genes as well as cytogenetic and immunogenetic features distinguished NNK- from DMT-SMZLs. These genetic clusters not only have distinct underpinning biology, as judged by differences in gene-expression signatures, but also different outcomes, with inferior survival in NNK-SMZLs. Digital cytometry and in situ profiling segregated 2 basic types of SMZL immune microenvironments termed immune-suppressive SMZL (50% of cases, associated with inflammatory cells and immune checkpoint activation) and immune-silent SMZL (50% of cases, associated with an immune-excluded phenotype) with distinct mutational and clinical connotations. In summary, we propose a nosology of SMZL that can implement its classification and also aid in the development of rationally targeted treatments.


Assuntos
Linfoma de Zona Marginal Tipo Células B , Neoplasias Esplênicas , Idoso , Animais , Feminino , Humanos , Masculino , Camundongos , Pessoa de Meia-Idade , Aberrações Cromossômicas , Imunofenotipagem , Linfoma de Zona Marginal Tipo Células B/diagnóstico , Linfoma de Zona Marginal Tipo Células B/genética , Família Multigênica , Mutação , Baço/patologia , Neoplasias Esplênicas/diagnóstico , Neoplasias Esplênicas/genética , Transcriptoma , Microambiente Tumoral
3.
Nat Commun ; 12(1): 7033, 2021 12 02.
Artigo em Inglês | MEDLINE | ID: mdl-34857732

RESUMO

Comprehensive genomic studies have delineated key driver mutations linked to disease progression for most cancers. However, corresponding transcriptional changes remain largely elusive because of the bias associated with cross-study analysis. Here, we overcome these hurdles and generate a comprehensive prostate cancer transcriptome atlas that describes the roadmap to tumor progression in a qualitative and quantitative manner. Most cancers follow a uniform trajectory characterized by upregulation of polycomb-repressive-complex-2, G2-M checkpoints, and M2 macrophage polarization. Using patient-derived xenograft models, we functionally validate our observations and add single-cell resolution. Thereby, we show that tumor progression occurs through transcriptional adaption rather than a selection of pre-existing cancer cell clusters. Moreover, we determine at the single-cell level how inhibition of EZH2 - the top upregulated gene along the trajectory - reverts tumor progression and macrophage polarization. Finally, a user-friendly web-resource is provided enabling the investigation of dynamic transcriptional perturbations linked to disease progression.


Assuntos
Proteína Potenciadora do Homólogo 2 de Zeste/genética , Proteínas de Neoplasias/genética , Neoplasias da Próstata/genética , Transcriptoma , Animais , Atlas como Assunto , Linhagem Celular Tumoral , Progressão da Doença , Proteína Potenciadora do Homólogo 2 de Zeste/metabolismo , Pontos de Checagem da Fase G2 do Ciclo Celular/genética , Perfilação da Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Xenoenxertos , Humanos , Macrófagos/metabolismo , Macrófagos/patologia , Masculino , Camundongos , Proteínas de Neoplasias/metabolismo , Complexo Repressor Polycomb 2/genética , Complexo Repressor Polycomb 2/metabolismo , Análise de Componente Principal , Próstata/metabolismo , Próstata/patologia , Neoplasias da Próstata/metabolismo , Neoplasias da Próstata/patologia , Transdução de Sinais , Análise de Célula Única
4.
Science ; 374(6564): 216-224, 2021 Oct 08.
Artigo em Inglês | MEDLINE | ID: mdl-34618582

RESUMO

The microbiota comprises the microorganisms that live in close contact with the host, with mutual benefit for both counterparts. The contribution of the gut microbiota to the emergence of castration-resistant prostate cancer (CRPC) has not yet been addressed. We found that androgen deprivation in mice and humans promotes the expansion of defined commensal microbiota that contributes to the onset of castration resistance in mice. Specifically, the intestinal microbial community in mice and patients with CRPC was enriched for species capable of converting androgen precursors into active androgens. Ablation of the gut microbiota by antibiotic therapy delayed the emergence of castration resistance even in immunodeficient mice. Fecal microbiota transplantation (FMT) from CRPC mice and patients rendered mice harboring prostate cancer resistant to castration. In contrast, tumor growth was controlled by FMT from hormone-sensitive prostate cancer patients and Prevotella stercorea administration. These results reveal that the commensal gut microbiota contributes to endocrine resistance in CRPC by providing an alternative source of androgens.


Assuntos
Androgênios/biossíntese , Bactérias/metabolismo , Microbioma Gastrointestinal/fisiologia , Interações entre Hospedeiro e Microrganismos , Neoplasias de Próstata Resistentes à Castração/metabolismo , Neoplasias de Próstata Resistentes à Castração/microbiologia , Idoso , Idoso de 80 Anos ou mais , Antagonistas de Androgênios/uso terapêutico , Animais , Antibacterianos/farmacologia , Bactérias/efeitos dos fármacos , Bactérias/genética , Linhagem Celular Tumoral , Transplante de Microbiota Fecal , Microbioma Gastrointestinal/efeitos dos fármacos , Microbioma Gastrointestinal/genética , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos NOD , Camundongos SCID , Pessoa de Meia-Idade , Neoplasias Experimentais , Prevotella/metabolismo , Neoplasias de Próstata Resistentes à Castração/tratamento farmacológico , Simbiose , Ensaios Antitumorais Modelo de Xenoenxerto
5.
Swiss Med Wkly ; 151: w20483, 2021 03 15.
Artigo em Inglês | MEDLINE | ID: mdl-33872378

RESUMO

Tumour-infiltrating myeloid-derived suppressor cells (MDSCs) are a heterogeneous population of myeloid cells. The main feature of MDSCs is their ability to suppress T-cell activation and function, which leads to immunosuppressive activity in the tumour microenvironment. Higher numbers of circulating and tumour-infiltrating MDSCs have been observed in a large number of patients with various types of tumour, and are linked to poor prognosis, especially in hormone-driven tumours. Recently, it has been demonstrated that the recruitment of MDSCs in prostate cancer confers resistance to canonical endocrine therapies, opening a new approach to the treatment of hormone-driven cancer patients.


Assuntos
Células Supressoras Mieloides , Neoplasias da Próstata , Hormônios , Humanos , Masculino , Células Mieloides , Microambiente Tumoral
6.
Cancer Cell ; 39(1): 68-82.e9, 2021 01 11.
Artigo em Inglês | MEDLINE | ID: mdl-33186519

RESUMO

Metastases account for most cancer-related deaths, yet the mechanisms underlying metastatic spread remain poorly understood. Recent evidence demonstrates that senescent cells, while initially restricting tumorigenesis, can induce tumor progression. Here, we identify the metalloproteinase inhibitor TIMP1 as a molecular switch that determines the effects of senescence in prostate cancer. Senescence driven either by PTEN deficiency or chemotherapy limits the progression of prostate cancer in mice. TIMP1 deletion allows senescence to promote metastasis, and elimination of senescent cells with a senolytic BCL-2 inhibitor impairs metastasis. Mechanistically, TIMP1 loss reprograms the senescence-associated secretory phenotype (SASP) of senescent tumor cells through activation of matrix metalloproteinases (MMPs). Loss of PTEN and TIMP1 in prostate cancer is frequent and correlates with resistance to docetaxel and worst clinical outcomes in patients treated in an adjuvant setting. Altogether, these findings provide insights into the dual roles of tumor-associated senescence and can potentially impact the treatment of prostate cancer.


Assuntos
Docetaxel/administração & dosagem , Deleção de Genes , PTEN Fosfo-Hidrolase/genética , Neoplasias da Próstata/patologia , Inibidor Tecidual de Metaloproteinase-1/genética , Animais , Senescência Celular/efeitos dos fármacos , Docetaxel/farmacologia , Regulação Neoplásica da Expressão Gênica , Humanos , Masculino , Metaloproteinases da Matriz/metabolismo , Camundongos , Metástase Neoplásica , Transplante de Neoplasias , Células PC-3 , Neoplasias da Próstata/tratamento farmacológico , Neoplasias da Próstata/genética , Neoplasias da Próstata/metabolismo
7.
Clin Cancer Res ; 26(23): 6387-6398, 2020 12 01.
Artigo em Inglês | MEDLINE | ID: mdl-32928793

RESUMO

PURPOSE: In search of novel strategies to improve the outcome of advanced prostate cancer, we considered that prostate cancer cells rearrange iron homeostasis, favoring iron uptake and proliferation. We exploited this adaptation by exposing prostate cancer preclinical models to high-dose iron to induce toxicity and disrupt adaptation to androgen starvation. EXPERIMENTAL DESIGN: We analyzed markers of cell viability and mechanisms underlying iron toxicity in androgen receptor-positive VCaP and LNCaP, castration-resistant DU-145 and PC-3, and murine TRAMP-C2 cells treated with iron and/or the antiandrogen bicalutamide. We validated the results in vivo in VCaP and PC-3 xenografts and in TRAMP-C2 injected mice treated with iron and/or bicalutamide. RESULTS: Iron was toxic for all prostate cancer cells. In particular, VCaP, LNCaP, and TRAMP-C2 were highly iron sensitive. Toxicity was mediated by oxidative stress, which primarily affected lipids, promoting ferroptosis. In highly sensitive cells, iron additionally caused protein damage. High-basal iron content and oxidative status defined high iron sensitivity. Bicalutamide-iron combination exacerbated oxidative damage and cell death, triggering protein oxidation also in poorly iron-sensitive DU-145 and PC-3 cells.In vivo, iron reduced tumor growth in TRAMP-C2 and VCaP mice. In PC-3 xenografts, bicalutamide-iron combination caused protein oxidation and successfully impaired tumor expansion while single compounds were ineffective. Macrophages influenced body iron distribution but did not limit the iron effect on tumor expansion. CONCLUSIONS: Our models allow us to dissect the direct iron effect on cancer cells. We demonstrate the proof of principle that iron toxicity inhibits prostate cancer cell proliferation, proposing a novel tool to strengthen antiandrogen treatment efficacy.


Assuntos
Antagonistas de Androgênios/farmacologia , Anilidas/farmacologia , Apoptose , Sinergismo Farmacológico , Ferro/farmacologia , Nitrilas/farmacologia , Neoplasias da Próstata/tratamento farmacológico , Compostos de Tosil/farmacologia , Animais , Proliferação de Células , Humanos , Masculino , Camundongos , Neoplasias da Próstata/patologia , Células Tumorais Cultivadas , Ensaios Antitumorais Modelo de Xenoenxerto
8.
Front Immunol ; 9: 1786, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30108594

RESUMO

Prostate adenocarcinoma (PCa) and melanoma are paradigmatic examples of tumors that are either poorly or highly sensitive to therapies based on monoclonal antibodies directed against regulatory pathways in T lymphocytes [i.e., immune checkpoint blockade (ICB)]. Yet, approximately 40% of melanoma patients are resistant or acquire resistance to ICB. What characterize the microenvironment of PCa and ICB-resistant melanoma are a scanty cytotoxic T cell infiltrate and a strong immune suppression, respectively. Here, we compare the tumor microenvironment in these two subgroups of cancer patients, focusing on some among the most represented immune checkpoint molecules: cytotoxic T lymphocyte-associated antigen-4, programmed death-1, lymphocyte activation gene-3, and T cell immunoglobulin and mucin-domain containing-3. We also report on several examples of crosstalk between cancer and immune cells that are mediated by inhibitory immune checkpoints and identify promising strategies aimed at overcoming ICB resistance both in PCa and melanoma.


Assuntos
Antineoplásicos Imunológicos/imunologia , Neoplasias/imunologia , Neoplasias da Próstata/imunologia , Linfócitos T Citotóxicos/imunologia , Adenocarcinoma/imunologia , Adenocarcinoma/patologia , Adenocarcinoma/terapia , Antineoplásicos Imunológicos/uso terapêutico , Antígeno B7-H1/antagonistas & inibidores , Antígeno B7-H1/imunologia , Antígeno B7-H1/metabolismo , Antígeno CTLA-4/antagonistas & inibidores , Antígeno CTLA-4/imunologia , Antígeno CTLA-4/metabolismo , Humanos , Imunoterapia/métodos , Masculino , Melanoma/imunologia , Melanoma/patologia , Melanoma/terapia , Neoplasias/patologia , Neoplasias/terapia , Receptor de Morte Celular Programada 1/antagonistas & inibidores , Receptor de Morte Celular Programada 1/imunologia , Receptor de Morte Celular Programada 1/metabolismo , Neoplasias da Próstata/patologia , Neoplasias da Próstata/terapia , Linfócitos T Citotóxicos/citologia
9.
Immunotherapy ; 10(9): 753-767, 2018 07.
Artigo em Inglês | MEDLINE | ID: mdl-30008257

RESUMO

Allogeneic hematopoietic cell transplantation (allo-HCT) is an adoptive immunotherapy strategy whose effectiveness relies on graft-versus-tumor (GVT) effect. We explored the feasibility of enhancing GVT after allo-HCT by peptide vaccination. Two myeloma patients were transplanted with a fludarabine-total body irradiation conditioning regimen and vaccinated with an HLA-A*0201-restricted modified survivin nonapeptide, plus montanide as adjuvant. At time of first vaccination, one patient had just attained serological remission despite documented relapse after transplant, while the other patient was in stable disease. Both patients had an immune response to vaccination: the frequency of survivin-specific CD8+ T cells increased between second and sixth vaccination and accounted for 0.5-0.8% of CD8+ cells; CD8+ cells were functional in ELISPOT assay. The first patient persists in complete remission with a follow-up of >5 years, while the second patient did not have a clinical response and vaccination was halted. We analyzed the T-cell receptor (TCR) repertoire of the first patient by spectratyping and found that vaccination did not affect the diversity of TCR profile, indicating that survivin clonotypes were probably spread in multiple TCR families. We generated a limited number (n = 4) of survivin-specific T cell clones: three were reactive only against the modified peptide, whereas one clone recognized also the naive peptide. Peptide vaccination is safe and applicable after allo-HCT and elicits an efficient antigen-specific T cell response without causing graft-versus-host disease.


Assuntos
Neoplasias Ósseas/terapia , Linfócitos T CD8-Positivos/imunologia , Doença Enxerto-Hospedeiro/prevenção & controle , Efeito Enxerto vs Tumor/imunologia , Transplante de Células-Tronco Hematopoéticas , Mieloma Múltiplo/terapia , Peptídeos/imunologia , Survivina/imunologia , Neoplasias Ósseas/secundário , Células Clonais , Citotoxicidade Imunológica , ELISPOT , Evolução Fatal , Feminino , Humanos , Imunidade Celular , Masculino , Mieloma Múltiplo/patologia , Recidiva Local de Neoplasia , Indução de Remissão , Transplante Homólogo , Vacinação
10.
Oncoimmunology ; 7(4): e1341032, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29632712

RESUMO

Many patients with B-cell malignancies can be successfully treated, although tumor eradication is rarely achieved. T-cell-directed killing of tumor cells using engineered T-cells or bispecific antibodies is a promising approach for the treatment of hematologic malignancies. We investigated the efficacy of CD19xCD3 DART bispecific antibody in a broad panel of human primary B-cell malignancies. The CD19xCD3 DART identified 2 distinct subsets of patients, in which the neoplastic lymphocytes were eliminated with rapid or slow kinetics. Delayed responses were always overcome by a prolonged or repeated DART exposure. Both CD4 and CD8 effector cytotoxic cells were generated, and DART-mediated killing of CD4+ cells into cytotoxic effectors required the presence of CD8+ cells. Serial exposures to DART led to the exponential expansion of CD4 + and CD8 + cells and to the sequential ablation of neoplastic cells in absence of a PD-L1-mediated exhaustion. Lastly, patient-derived neoplastic B-cells (B-Acute Lymphoblast Leukemia and Diffuse Large B Cell Lymphoma) could be proficiently eradicated in a xenograft mouse model by DART-armed cytokine induced killer (CIK) cells. Collectively, patient tailored DART exposures can result in the effective elimination of CD19 positive leukemia and B-cell lymphoma and the association of bispecific antibodies with unmatched CIK cells represents an effective modality for the treatment of CD19 positive leukemia/lymphoma.

11.
Clin Cancer Res ; 24(9): 2171-2181, 2018 05 01.
Artigo em Inglês | MEDLINE | ID: mdl-29490991

RESUMO

Purpose: Irregular blood flow and endothelial cell anergy, which characterize many solid tumors, hinder tumor infiltration by cytotoxic T lymphocytes (CTL). This confers resistance to cancer immunotherapy with monoclonal antibodies directed against regulatory pathways in T lymphocytes (i.e., immune checkpoint blockade, ICB). We investigated whether NGR-TNF, a TNF derivative capable of targeting the tumor vasculature, and improving intratumor infiltration by activated CTLs, could sensitize tumors to ICB with antibodies specific for the PD-1 and CTLA-4 receptors.Experimental Design: Transgenic adenocarcinoma of the mouse prostate (TRAMP) mice with autochthonous prostate cancer and C57BL/6 mice with orthotopic B16 melanoma were treated with NGR-TNF, adoptive T-cell therapy (ACT), and ICB, and monitored for immune surveillance and disease progression.Results: The combination of ACT, NGR-TNF, and ICB was the most effective in delaying disease progression, and in improving overall survival of mice bearing ICB-resistant prostate cancer or melanoma. Mechanistically, the therapeutic effects were associated with potent tumor infiltration, especially by endogenous but also by adoptively transferred PD-1+, granzyme B+, and interferon-γ+ CTLs. The therapeutic effects were also associated with favorable T-effector/regulatory T cell ratios.Conclusions: Targeting the tumor vasculature with low-dose TNF in association with ACT may represent a novel strategy for enhancing T-cell infiltration in tumors and overcoming resistance to immune checkpoint blockers. Clin Cancer Res; 24(9); 2171-81. ©2018 AACR.


Assuntos
Neoplasias/etiologia , Neoplasias/metabolismo , Neovascularização Patológica/imunologia , Neovascularização Patológica/metabolismo , Linfócitos T/imunologia , Linfócitos T/metabolismo , Fator de Necrose Tumoral alfa/metabolismo , Animais , Antineoplásicos Imunológicos/farmacologia , Biomarcadores Tumorais , Linhagem Celular Tumoral , Terapia Combinada , Modelos Animais de Doenças , Imunomodulação/efeitos dos fármacos , Imunofenotipagem , Imunoterapia Adotiva , Linfócitos do Interstício Tumoral/efeitos dos fármacos , Linfócitos do Interstício Tumoral/imunologia , Linfócitos do Interstício Tumoral/metabolismo , Masculino , Melanoma Experimental , Camundongos , Camundongos Knockout , Neoplasias/patologia , Neoplasias/terapia , Linfócitos T/efeitos dos fármacos , Linfócitos T Citotóxicos/imunologia , Linfócitos T Citotóxicos/metabolismo
12.
Cytokine Growth Factor Rev ; 36: 17-24, 2017 08.
Artigo em Inglês | MEDLINE | ID: mdl-28606732

RESUMO

Cancer immunotherapy with monoclonal antibodies directed against regulatory pathways in T lymphocytes has been revolutionizing medical oncology, and the clinical success of monoclonal antibodies targeting either cytotoxic T lymphocyte antigen-4 (CTLA-4) or program death-1 (PD-1) in patients affected by melanoma, Hodgkin's lymphoma, Merkel cell carcinoma, and head and neck, bladder, renal cell or non-small cell lung cancer is way beyond the most optimistic expectation. However, immune checkpoint blockade (ICB) has failed to arrest progression in a consistent amount of patients affected by those tumors, and various histological types, including breast, colon and prostate cancer, are less sensitive to this therapeutic approach. Such clinical findings have fueled massive research efforts in the attempt to identify pre-existing and acquired mechanisms of resistance to ICB. Here we focus on evidences emerging from studies in humans on how tumor cells and the tumor microenvironment contribute to the heterogeneous clinical responses, and we propose strategies stemming from pre-clinical models that might improve clinical outcomes for patients.


Assuntos
Anticorpos Monoclonais/uso terapêutico , Resistencia a Medicamentos Antineoplásicos , Imunoterapia , Neoplasias/terapia , Microambiente Tumoral , Animais , Antígeno CTLA-4/imunologia , Carcinoma Pulmonar de Células não Pequenas/imunologia , Carcinoma Pulmonar de Células não Pequenas/terapia , Terapia Combinada , Humanos , Neoplasias Pulmonares/imunologia , Neoplasias Pulmonares/terapia , Melanoma/terapia , Camundongos , Neoplasias/imunologia , Receptor de Morte Celular Programada 1/imunologia , Linfócitos T/imunologia
13.
Cancer Res ; 76(17): 5151-62, 2016 09 01.
Artigo em Inglês | MEDLINE | ID: mdl-27328731

RESUMO

miR-214 and miR-148b have been proposed to antagonize the effects of each other in enabling or blocking metastasis, respectively. In this study, we provide evidence deepening their role and interrelationship in the process of metastatic dissemination. Depleting miR-214 or elevating miR-148b blocked the dissemination of melanoma or breast cancer cells, an effect that could be accentuated by dual alteration. Mechanistic investigations indicated that dual alteration suppressed passage of malignant cells through the blood vessel endothelium by reducing expression of the cell adhesion molecules ITGA5 and ALCAM. Notably, transendothelial migration in vitro and extravasation in vivo impaired by singly alternating miR-214 or miR-148b could be overridden by overexpression of ITGA5 or ALCAM in the same tumor cells. In clinical specimens of primary breast cancer or metastatic melanoma, we found a positive correlation between miR-214 and ITGA5 or ALCAM along with an inverse correlation of miR-214 and miR-148b in the same specimens. Our findings define an antagonistic relationship of miR-214 and miR-148b in determining the dissemination of cancer cells via tumor-endothelial cell interactions, with possible implications for microRNA-mediated therapeutic interventions aimed at blocking cancer extravasation. Cancer Res; 76(17); 5151-62. ©2016 AACR.


Assuntos
Neoplasias da Mama/patologia , Melanoma/patologia , MicroRNAs/genética , Invasividade Neoplásica/genética , Animais , Antígenos CD/biossíntese , Neoplasias da Mama/genética , Moléculas de Adesão Celular/biossíntese , Moléculas de Adesão Celular Neuronais/biossíntese , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células/genética , Células Endoteliais/patologia , Feminino , Proteínas Fetais/biossíntese , Regulação Neoplásica da Expressão Gênica/genética , Xenoenxertos , Humanos , Immunoblotting , Melanoma/genética , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID
14.
Clin Immunol ; 152(1-2): 152-63, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-24632064

RESUMO

In eosinophilic granulomatosis with polyangiitis (EGPA) clonally expanded T cells might concur in granuloma formation and vascular injury. The TCR ß-variable (BV) chain repertoire and third complementarity determining region (CDR3) of peripheral CD4+ and CD8+ cells in EGPA patients and age-matched controls and the expression of cytokines and chemokine receptors were investigated. The CD8+ lymphocytes of EGPA patients showed an increased frequency of BV expansions with a skewed profile of BV CDR3 lengths, increased CCR5 and CXCR3 expression and increased INFγ and TNFα production. In two patients, the TCR CDR3 cDNA sequences of the expanded BV family were identified. The CD4+ lymphocytes of EGPA patients revealed a higher expression of CRTH2 and increased production of IL-5. In conclusion, CD4+ T cells display a Th2 profile and CD8+ T cells are clonally expanded in EGPA and have a proinflammatory phenotype, suggesting their pathogenic role in vasculitic damage.


Assuntos
Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/imunologia , Síndrome de Churg-Strauss/imunologia , Receptores de Antígenos de Linfócitos T alfa-beta/imunologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Biomarcadores , Células Cultivadas , Síndrome de Churg-Strauss/sangue , Regiões Determinantes de Complementaridade , Feminino , Granuloma/imunologia , Humanos , Switching de Imunoglobulina/imunologia , Inflamação/imunologia , Interferon gama/biossíntese , Interleucina-5/biossíntese , Masculino , Pessoa de Meia-Idade , Receptores de Antígenos de Linfócitos T alfa-beta/genética , Receptores CCR5/biossíntese , Receptores CXCR3/biossíntese , Receptores Imunológicos/biossíntese , Receptores de Prostaglandina/biossíntese , Fator de Necrose Tumoral alfa/biossíntese
15.
PLoS One ; 9(1): e84859, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-24400121

RESUMO

MicroRNAs are single-stranded non-coding RNAs that simultaneously down-modulate the expression of multiple genes post-transcriptionally by binding to the 3'UTRs of target mRNAs. Here we used computational methods to predict microRNAs relevant in breast cancer progression. Specifically, we applied different microRNA target prediction algorithms to various groups of differentially expressed protein-coding genes obtained from four breast cancer datasets. Six potential candidates were identified, among them miR-223, previously described to be highly expressed in the tumor microenvironment and known to be actively transferred into breast cancer cells. To investigate the function of miR-223 in tumorigenesis and to define its molecular mechanism, we overexpressed miR-223 in breast cancer cells in a transient or stable manner. Alternatively we overexpressed miR-223 in mouse embryonic fibroblasts or HEK293 cells and used their conditioned medium to treat tumor cells. With both approaches, we obtained elevated levels of miR-223 in tumor cells and observed decreased migration, increased cell death in anoikis conditions and augmented sensitivity to chemotherapy but no effect on adhesion and proliferation. The analysis of miR-223 predicted targets revealed enrichment in cell death and survival-related genes and in pathways frequently altered in breast cancer. Among these genes, we showed that protein levels for STAT5A, ITGA3 and NRAS were modulated by miR-223. In addition, we proved that STAT5A is a direct miR-223 target and highlighted a possible correlation between miR-223 and STAT5A in migration and chemotherapy response. Our investigation revealed that a computational analysis of cancer gene expression datasets can be a relevant tool to identify microRNAs involved in cancer progression and that miR-223 has a prominent role in breast malignancy that could potentially be exploited therapeutically.


Assuntos
Neoplasias da Mama/genética , Neoplasias da Mama/patologia , MicroRNAs/genética , Regiões 3' não Traduzidas , Anoikis/genética , Antineoplásicos/farmacologia , Pareamento de Bases , Sequência de Bases , Neoplasias da Mama/metabolismo , Morte Celular/efeitos dos fármacos , Morte Celular/genética , Linhagem Celular Tumoral , Movimento Celular/genética , Biologia Computacional , Bases de Dados Factuais , Progressão da Doença , Feminino , Perfilação da Expressão Gênica , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , MicroRNAs/química , MicroRNAs/metabolismo , Processamento Pós-Transcricional do RNA , Fator de Transcrição STAT5/química , Fator de Transcrição STAT5/genética , Fator de Transcrição STAT5/metabolismo , Transdução de Sinais/efeitos dos fármacos
16.
Stem Cells ; 31(7): 1422-33, 2013 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-23592522

RESUMO

It has recently been proposed that defective differentiation of mammary luminal progenitors predisposes to basal-like breast cancer. However, the molecular and cellular mechanisms involved are still unclear. Here, we describe that the adaptor protein p130Cas is a crucial regulator of mouse mammary epithelial cell (MMEC) differentiation. Using a transgenic mouse model, we show that forced p130Cas overexpression in the luminal progenitor cell compartment results in the expansion of luminal cells, which aberrantly display basal cell features and reduced differentiation in response to lactogenic stimuli. Interestingly, MMECs overexpressing p130Cas exhibit hyperactivation of the tyrosine kinase receptor c-Kit. In addition, we demonstrate that the constitutive c-Kit activation alone mimics p130Cas overexpression, whereas c-Kit downregulation is sufficient to re-establish proper differentiation of p130Cas overexpressing cells. Overall, our data indicate that high levels of p130Cas, via abnormal c-Kit activation, promote mammary luminal cell plasticity, thus providing the conditions for the development of basal-like breast cancer. Consistently, p130Cas is overexpressed in human triple-negative breast cancer, further suggesting that p130Cas upregulation may be a priming event for the onset of basal-like breast cancer.


Assuntos
Proteína Substrato Associada a Crk/metabolismo , Glândulas Mamárias Animais/citologia , Neoplasias Mamárias Experimentais/patologia , Proteínas Proto-Oncogênicas c-kit/metabolismo , Neoplasias de Mama Triplo Negativas/patologia , Animais , Diferenciação Celular/fisiologia , Processos de Crescimento Celular/fisiologia , Células Cultivadas , Proteína Substrato Associada a Crk/biossíntese , Proteína Substrato Associada a Crk/genética , Feminino , Humanos , Imuno-Histoquímica , Glândulas Mamárias Animais/metabolismo , Neoplasias Mamárias Experimentais/genética , Neoplasias Mamárias Experimentais/metabolismo , Camundongos , Camundongos Transgênicos , Gravidez , Proteínas Proto-Oncogênicas c-kit/genética , Transdução de Sinais , Neoplasias de Mama Triplo Negativas/genética , Neoplasias de Mama Triplo Negativas/metabolismo
17.
J Immunother ; 35(7): 579-86, 2012 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-22892454

RESUMO

Cytokine-induced killer cells (CIKs) are ex vivo expanded T-NK lymphocytes capable of HLA-unrestricted antitumor activity. CIKs are promising candidates for adoptive cancer immunotherapies; they can be generated and infused in autologous settings of cancer patients, or from donors, after allogeneic hematopoietic cell transplant. Ex vivo expansion rates of CIKs are greatly variable among patients, with consequent potential clinical limitations for "poor expanders." We compared the standard expansion protocol with a new one, which included the timed addition of irradiated allogeneic peripheral blood mononuclear cells. Our hypothesis is that allogeneic stimulation might provide CIK cells with a proliferative boost and simultaneously decrease their alloreactivity versus third parties, if HLA-mismatched from the allogeneic stimulators. Allo-stimulated CIKs (AS-CIK) reached significantly higher expansion rates compared with standard controls, regardless if generated form healthy donors (131- vs. 32-fold) or cancer patients (117- vs. 14-fold). The expansion of the CD3CD56 subset was 2243-fold for AS-CIKs compared with 362 for standard CIKs. AS-CIKs efficiently killed osteosarcoma targets in vitro, results were comparable with that of standard CIKs. Standard and AS-CIKs did not show differences in phenotype and telomere length. The alloreactivity of AS-CIKs against third party HLA-mismatched peripheral blood mononuclear cells was reduced compared with standard CIKs (37% vs. 23%). In conclusion, alloreactivity of CIK cells may be exploited enhancing their final ex vivo expansion. In clinical perspective these findings may facilitate the extension of CIK-based immunotherapy to larger numbers of patients and, translated into hematopoietic cell transplant settings, contribute to reduce the risk of graft versus host disease in the hypothesis of infusions across HLA barriers.


Assuntos
Neoplasias Ósseas/terapia , Células Matadoras Induzidas por Citocinas/imunologia , Imunoterapia Adotiva , Osteossarcoma/terapia , Neoplasias Ósseas/imunologia , Neoplasias Ósseas/patologia , Complexo CD3/metabolismo , Antígeno CD56/metabolismo , Técnicas de Cultura de Células/métodos , Proliferação de Células , Células Cultivadas , Células Matadoras Induzidas por Citocinas/transplante , Citotoxicidade Imunológica , Antígenos HLA/imunologia , Humanos , Imunização , Isoantígenos/imunologia , Leucócitos Mononucleares/imunologia , Osteossarcoma/imunologia , Osteossarcoma/patologia
18.
Clin Exp Rheumatol ; 30(1 Suppl 70): S57-61, 2012.
Artigo em Inglês | MEDLINE | ID: mdl-22640649

RESUMO

OBJECTIVES: Churg-Strauss syndrome (CSS) is a necrotising vasculitis of small vessels in which oligoclonally expanded TCR Vß CD8+ effector memory T cells populations (TEM) may be involved in vasculitic damage. The aim of this study was to assess the functional role of CD8+ T cells in CSS patients by flow cytometry analysis of membrane expression of cytotoxic markers NKG2D and CD107a. METHODS: Immunostaining of peripheral T cells and effector memory lymphocytes (TEM) from CSS patients and controls was performed by gating CD28 and CD45RA in the CD8+NKG2D+ and CD4+NKG2D+ populations. CD107a expression was evaluated in both whole CD8+ and CD4+ and the TEM cells by gating CD62 and CD45RA following polyclonal stimulation. RESULTS: NKG2D expression was shifted toward the CD8+CD28- fraction of T cells in CSS patients compared to healthy controls (56.1±25.8% versus 17.2±7.3%, respectively, p=0.002). CD8+Vß+ expanded T cells showed a significantly increased expression of NKG2D compared to the whole CD8+ T cell population (91.4±1.9% versus 79.7±3.8%, respectively, p=0.015). Moreover the CD8+ population from CSS upregulates CD107a on its surface upon polyclonal stimulation in a significantly higher proportion than healthy subjects (26.2±10.8% versus 8.2±2.9%, p=0.0031) and the majority CD8+ CD107+ cells from CSS patients showed a TEM phenotype compared to controls (64.8±4.9% vs. 19.8±2.9, respectively, p<0.001). CONCLUSIONS: In CSS, CD8+ TEM lymphocytes show markers of cytotoxic activity, which suggests a role for these cells in vasculitic damage.


Assuntos
Linfócitos T CD8-Positivos/imunologia , Síndrome de Churg-Strauss/imunologia , Memória Imunológica , Proteínas de Membrana Lisossomal/análise , Subfamília K de Receptores Semelhantes a Lectina de Células NK/análise , Adulto , Idoso , Biomarcadores/análise , Antígenos CD28/análise , Linfócitos T CD4-Positivos/imunologia , Estudos de Casos e Controles , Selectina E/análise , Feminino , Citometria de Fluxo , Humanos , Imunofenotipagem/métodos , Itália , Antígenos Comuns de Leucócito/análise , Ativação Linfocitária , Masculino , Pessoa de Meia-Idade , Paris , Fenótipo , Receptores de Antígenos de Linfócitos T alfa-beta/análise
19.
Hum Gene Ther ; 20(12): 1576-88, 2009 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-19678763

RESUMO

We investigated the possibility of introducing exogenous T cell receptor (TCR) genes into T cells by lentiviral transduction, without prior stimulation of endogenous TCR with anti-CD3. TCR transfer is used to impose tumor antigen specificity on recipient T cells, but sustained activation required for retroviral transduction may affect the clinical efficacy of engineered T cells. Cytokine stimulation makes T cells susceptible to lentiviral transduction in the absence of TCR triggering, but this advantage has never been exploited for TCR transfer. Autoimmune diseases are a source of high-affinity TCRs specific for self/tumor antigens. We selected, from a patient with vitiligo, a Mart1-specific TCR based on intrinsic interchain pairing properties and functional avidity. After lentiviral transduction of human peripheral blood mononuclear cells, preferential pairing of exogenous alpha and beta chains was observed, together with effective recognition of Mart1(+) melanoma cells. We tested transduction efficiency on various T cell subsets prestimulated with interleukin (IL)-2, IL-7, IL-15, and IL-21 (alone or in combination). Both naive and unfractionated CD8(+) T cells could be transduced without requiring endogenous TCR triggering. IL-7 plus IL-15 was the most powerful combination, allowing high levels of transgene expression without inducing T cell differentiation (34 +/- 5% Mart1-TCR(+) cells in naive CD8(+) and 16 +/- 6% in unfractionated CD8(+)). Cytokine-prestimulated, Mart1-redirected naive and unfractionated CD8(+) cells expanded better than CD3-CD28-prestimulated counterparts in response to both peptide-pulsed antigen-presenting cells and Mart1(+) melanoma cells. This strategy allows the generation of tumor-specific T cells encompassing truly naive T cells, endowed with an intact proliferative potential and a preserved differentiation stage.


Assuntos
Antígenos de Neoplasias/imunologia , Linfócitos T CD8-Positivos/imunologia , Terapia Genética/métodos , Memória Imunológica , Melanoma/terapia , Proteínas de Neoplasias/imunologia , Receptores de Antígenos de Linfócitos T/genética , Linhagem Celular Tumoral , Vetores Genéticos/genética , Humanos , Lentivirus/genética , Antígeno MART-1 , Melanoma/imunologia , Receptores de Antígenos de Linfócitos T/imunologia , Transdução Genética , Vitiligo/imunologia
20.
J Leukoc Biol ; 84(6): 1472-82, 2008 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-18725395

RESUMO

Dendritic cells (DCs) are the most potent antigen-presenting cells and fine-tune the immune response. We have investigated hypoxia's effects on the differentiation and maturation of DCs from human monocytes in vitro, and have shown that it affects DC functions. Hypoxic immature DCs (H-iDCs) significantly fail to capture antigens through down-modulation of the RhoA/Ezrin-Radixin-Moesin pathway and the expression of CD206. Moreover, H-iDCs released higher levels of CXCL1, VEGF, CCL20, CXCL8, and CXCL10 but decreased levels of CCL2 and CCL18, which predict a different ability to recruit neutrophils rather than monocytes and create a proinflammatory and proangiogenic environment. By contrast, hypoxia has no effect on DC maturation. Hypoxic mature DCs display a mature phenotype and activate both allogeneic and specific T cells like normoxic mDCs. This study provides the first demonstration that hypoxia inhibits antigen uptake by DCs and profoundly changes the DC chemokine expression profile and may have a critical role in DC differentiation, adaptation, and activation in inflamed tissues.


Assuntos
Antígenos/metabolismo , Diferenciação Celular/fisiologia , Quimiocinas/metabolismo , Células Dendríticas/citologia , Hipóxia/metabolismo , Movimento Celular , Proliferação de Células , Células Cultivadas , Citocinas/metabolismo , Proteínas de Ligação a DNA/metabolismo , Células Dendríticas/metabolismo , Regulação para Baixo , Citometria de Fluxo , Humanos , Ativação Linfocitária , Macrófagos/citologia , Macrófagos/imunologia , Macrófagos/metabolismo , Monócitos/citologia , Monócitos/imunologia , Monócitos/metabolismo , Fenótipo , Fosforilação , Linfócitos T/imunologia , Linfócitos T/metabolismo , Fatores de Transcrição/metabolismo , Proteínas rho de Ligação ao GTP/metabolismo
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