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1.
iScience ; 25(10): 105149, 2022 Oct 21.
Artigo em Inglês | MEDLINE | ID: mdl-36185376

RESUMO

Age-related diseases are major concern in developed countries. To avoid disabilities that accompany increased lifespan, pharmaceutical approaches are considered. Therefore, appropriate animal models are required for a better understanding of aging processes and potential in vivo assays to evaluate the impact of molecules that may delay the occurrence of age-related diseases. Few mouse models exhibiting pathological aging exist, but currently, none of them reproducibly mimics human diseases like osteoporosis, cognitive dysfunctions or sarcopenia that can be seen in some, but not all, elders. Here, we describe the premature aging phenotypes of Dicer-deficient mature animals, which exhibit an overall deterioration of many organs and tissues (skin, heart, and adipose tissue) ultimately leading to a significant reduction of their lifespan. Molecular characterization of transcriptional responses focused on the adipose tissue suggested that both canonical and non-canonical functions of DICER are involved in this process and highlight potential actionable pathways to revert it.

2.
Matrix Biol ; 108: 20-38, 2022 04.
Artigo em Inglês | MEDLINE | ID: mdl-35227929

RESUMO

The extracellular matrix molecule Tenascin-C (TNC) promotes cancer and chronic inflammation by multiple mechanisms. Recently, TNC was shown to promote an immune suppressive tumor microenvironment (TME) through binding soluble chemoattracting factors, thus retaining leukocytes in the stroma. TNC also binds to fibronectin (FN) and other molecules, raising the question of a potential common TNC binding mechanism. By sequence comparison of two TNC-interacting domains in FN, the fifth (FN5) and thirteenth (FN13) fibronectin type III domains we identified a MAtrix REgulating MOtif "MAREMO" or M-motif that is highly conserved amongst vertebrates. By sequence analysis, structural modeling and functional analysis we found also putative M-motifs in TNC itself. We showed by negative staining electron microscopic imaging that the M-motif in FN mediates interactions with FN as well as with TNC. We generated two M-motif mimetic peptides P5 and P13 resembling the M-motif in FN5 and FN13, respectively. By using structural information we modelled binding of these M-motif mimetics revealing a putative MAREMO binding site MBS in FN5 and TN3, respectively overlapping with the M-motif. We further demonstrated that the M-motif mimetic peptides blocked several functions of TNC, such as binding of TNC to FN, cell rounding on a mixed FN/TNC substratum, FN matrix expression and subsequent assembly, TNC-induced signaling and gene expression, TNC chemokine binding and dendritic cell retention, thus providing novel opportunities to inhibit TNC actions. Our results suggest that targeting the MAREMO/MBS interaction could be exploited for reducing inflammation and matrix functions in cancer and fibrosis.


Assuntos
Neoplasias , Tenascina , Animais , Matriz Extracelular/metabolismo , Inflamação , Neoplasias/genética , Peptídeos , Tenascina/genética , Tenascina/metabolismo , Microambiente Tumoral
3.
EMBO Mol Med ; 13(6): e13270, 2021 06 07.
Artigo em Inglês | MEDLINE | ID: mdl-33988305

RESUMO

Immune checkpoint therapy, where CD8 tumor infiltrating T lymphocytes (TIL) are reactivated, is a promising anti-cancer treatment approach, yet with low response rates. The extracellular matrix, in particular tenascin-C, may generate barriers for TIL. To investigate this possibility, we used a MMTV-NeuNT and syngeneic mammary gland grafting model derived thereof with engineered tenascin-C levels and observed accumulation of CD8 TIL in tenascin-C-rich stroma. Inhibition studies revealed that tenascin-C induced CXCL12 through TLR4. By binding CXCL12, tenascin-C retained CD8 TIL in the stroma. Blockade of CXCR4, the receptor of CXCL12, enhanced macrophage and CD8 TIL infiltration and reduced tumor growth and subsequent metastasis. Retention of CD8 TIL by tenascin-C/CXCL12 was also observed in human breast cancer by tissue staining. Moreover, whereas high CD8 TIL numbers correlated with longer metastasis-free survival, this was not the case when also tenascin-C and CXCL12 levels were high. Altogether, these results may be useful for improving tumor immunity as diagnostic tool and to formulate a future "TIL-matrix-release-and-reactivate" strategy.


Assuntos
Linfócitos do Interstício Tumoral , Neoplasias , Linfócitos T CD8-Positivos , Quimiocina CXCL12 , Matriz Extracelular , Humanos , Tenascina
4.
Front Immunol ; 12: 635166, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33790905

RESUMO

The extracellular matrix (ECM) molecule Tenascin-C (TNC) is well-known to promote tumor progression by multiple mechanisms. However, reliable TNC detection in tissues of tumor banks remains limited. Therefore, we generated dromedary single-domain nanobodies Nb3 and Nb4 highly specific for human TNC (hTNC) and characterized the interaction with TNC by several approaches including ELISA, western blot, isothermal fluorescence titration and negative electron microscopic imaging. Our results revealed binding of both nanobodies to distinct sequences within fibronectin type III repeats of hTNC. By immunofluroescence and immunohistochemical imaging we observed that both nanobodies detected TNC expression in PFA and paraffin embedded human tissue from ulcerative colitis, solid tumors and liver metastasis. As TNC impairs cell adhesion to fibronectin we determined whether the nanobodies abolished this TNC function. Indeed, Nb3 and Nb4 restored adhesion of tumor and mesangial cells on a fibronectin/TNC substratum. We recently showed that TNC orchestrates the immune-suppressive tumor microenvironment involving chemoretention, causing tethering of CD11c+ myeloid/dendritic cells in the stroma. Here, we document that immobilization of DC2.4 dendritic cells by a CCL21 adsorbed TNC substratum was blocked by both nanobodies. Altogether, our novel TNC specific nanobodies could offer valuable tools for detection of TNC in the clinical practice and may be useful to inhibit the immune-suppressive and other functions of TNC in cancer and other diseases.


Assuntos
Anticorpos Neutralizantes/imunologia , Camelus/imunologia , Anticorpos de Domínio Único/imunologia , Tenascina/antagonistas & inibidores , Animais , Anticorpos Neutralizantes/farmacologia , Especificidade de Anticorpos , Sítios de Ligação de Anticorpos , Adesão Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Colite Ulcerativa/imunologia , Colo/imunologia , Ensaio de Imunoadsorção Enzimática , Células HEK293 , Humanos , Imuno-Histoquímica , Neoplasias Hepáticas/imunologia , Neoplasias Hepáticas/secundário , Ligação Proteica , Anticorpos de Domínio Único/farmacologia , Tenascina/administração & dosagem , Tenascina/imunologia
5.
Cancer Immunol Res ; 8(9): 1122-1138, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32665262

RESUMO

Inherent immune suppression represents a major challenge in the treatment of human cancer. The extracellular matrix molecule tenascin-C promotes cancer by multiple mechanisms, yet the roles of tenascin-C in tumor immunity are incompletely understood. Using a 4NQO-induced oral squamous cell carcinoma (OSCC) model with abundant and absent tenascin-C, we demonstrated that tenascin-C enforced an immune-suppressive lymphoid stroma via CCL21/CCR7 signaling, leading to increased metastatic tumors. Through TLR4, tenascin-C increased expression of CCR7 in CD11c+ myeloid cells. By inducing CCL21 in lymphatic endothelial cells via integrin α9ß1 and binding to CCL21, tenascin-C immobilized CD11c+ cells in the stroma. Inversion of the lymph node-to-tumor CCL21 gradient, recruitment of T regulatory cells, high expression of anti-inflammatory cytokines, and matrisomal components were hallmarks of the tenascin-C-instructed lymphoid stroma. Ablation of tenascin-C or CCR7 blockade inhibited the lymphoid immune-suppressive stromal properties, reducing tumor growth, progression, and metastasis. Thus, targeting CCR7 could be relevant in human head and neck tumors, as high tenascin-C expression and an immune-suppressive stroma correlate to poor patient survival.


Assuntos
Neoplasias Bucais/imunologia , Carcinoma de Células Escamosas de Cabeça e Pescoço/imunologia , Tenascina/imunologia , Animais , Quimiocina CCL21/imunologia , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Neoplasias Bucais/patologia , Receptores CCR7/imunologia , Proteínas Recombinantes/farmacologia , Linfócitos T Reguladores/imunologia , Tenascina/farmacologia , Microambiente Tumoral/imunologia
6.
Biochem Biophys Res Commun ; 530(2): 471-478, 2020 09 17.
Artigo em Inglês | MEDLINE | ID: mdl-32593416

RESUMO

Tenascin-C (TNC) and tenascin-W (TNW), large hexameric glycoproteins overexpressed in the tumor microenvironment, are useful tumor biomarkers for theranostic applications. For now, polyclonal and monoclonal antibodies, as well as aptamers targeting TNC and TNW have been developed. However, the immunostaining sensitivity of antibodies is very heterogenous. The main aim of this study was to generate antibodies in dromedary that detect TNC and TNW, respectively. We show that immune sera from immunized dromedaries are able to specifically bind native TNC and TNW by ELISA and also to detect TNC and TNW in matrix tracks of mammary tumors by immunostaining. Furthermore, we demonstrate that purified IgG subtypes are able to interact specifically with TNC or TNW by ELISA and immunostaining. These camelid antibodies are a good basis to develop tools for the detection of TNC and TNW in the tumor microenvironment and could potentially have a broader application for early diagnosis of solid cancers.


Assuntos
Anticorpos/imunologia , Camelus/imunologia , Tenascina/imunologia , Animais , Anticorpos/análise , Biomarcadores Tumorais/análise , Biomarcadores Tumorais/imunologia , Neoplasias da Mama/imunologia , Neoplasias da Mama/patologia , Ensaio de Imunoadsorção Enzimática , Feminino , Células HEK293 , Humanos , Imunização , Camundongos , Microscopia de Fluorescência , Tenascina/análise , Microambiente Tumoral
7.
Cancer Immunol Res ; 8(3): 368-382, 2020 03.
Artigo em Inglês | MEDLINE | ID: mdl-31941671

RESUMO

The interplay between cancer cells and immune cells is a key determinant of tumor survival. Here, we uncovered how tumors exploit the immunomodulatory properties of the extracellular matrix to create a microenvironment that enables their escape from immune surveillance. Using orthotopic grafting of mammary tumor cells in immunocompetent mice and autochthonous models of breast cancer, we discovered how tenascin-C, a matrix molecule absent from most healthy adult tissues but expressed at high levels and associated with poor patient prognosis in many solid cancers, controls the immune status of the tumor microenvironment. We found that, although host-derived tenascin-C promoted immunity via recruitment of proinflammatory, antitumoral macrophages, tumor-derived tenascin-C subverted host defense by polarizing tumor-associated macrophages toward a pathogenic, immune-suppressive phenotype. Therapeutic monoclonal antibodies that blocked tenascin-C activation of Toll-like receptor 4 reversed this phenotypic switch in vitro and reduced tumor growth and lung metastasis in vivo, providing enhanced benefit in combination with anti-PD-L1 over either treatment alone. Combined tenascin-C:macrophage gene-expression signatures delineated a significant survival benefit in people with breast cancer. These data revealed a new approach to targeting tumor-specific macrophage polarization that may be effective in controlling the growth and spread of breast tumors.


Assuntos
Antineoplásicos Imunológicos/farmacologia , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/imunologia , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/imunologia , Macrófagos/imunologia , Animais , Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Matriz Extracelular/efeitos dos fármacos , Matriz Extracelular/imunologia , Feminino , Humanos , Vigilância Imunológica , Imunoterapia/métodos , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/secundário , Ativação de Macrófagos/efeitos dos fármacos , Ativação de Macrófagos/imunologia , Macrófagos/efeitos dos fármacos , Camundongos , Fenótipo , Tenascina/imunologia , Células Tumorais Cultivadas , Microambiente Tumoral/efeitos dos fármacos , Microambiente Tumoral/imunologia
8.
Matrix Biol ; 83: 26-47, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31288084

RESUMO

Metastasis is a major cause of death in cancer patients. The extracellular matrix molecule tenascin-C is a known promoter of metastasis, however the underlying mechanisms are not well understood. To further analyze the impact of tenascin-C on cancer progression we generated MMTV-NeuNT mice that develop spontaneous mammary tumors, on a tenascin-C knockout background. We also developed a syngeneic orthotopic model in which tumor cells derived from a MMTV-NeuNT tumor. Tumor cells were transfected with control shRNA or with shRNA to knockdown tenascin-C expression and, were grafted into the mammary gland of immune competent, wildtype or tenascin-C knockout mice. We show that stromal-derived tenascin-C increases metastasis by reducing apoptosis and inducing the cellular plasticity of cancer cells located in pulmonary blood vessels invasions (BVI), before extravasation. We characterized BVI as organized structures of tightly packed aggregates of proliferating tumor cells with epithelial characteristics, surrounded by Fsp1+ cells, internally located platelets and, a luminal monolayer of endothelial cells. We found extracellular matrix, in particular, tenascin-C, between the stromal cells and the tumor cell cluster. In mice lacking stromal-derived tenascin-C, the organization of pulmonary BVI was significantly affected, revealing novel functions of host-derived tenascin-C in supporting the integrity of the endothelial cell coat, increasing platelet abundance, tumor cell survival, epithelial plasticity, thereby promoting overall lung metastasis. Many effects of tenascin-C observed in BVI including enhancement of cellular plasticity, survival and migration, could be explained by activation of TGF-ß signaling. Finally, in several human cancers, we also observed BVI to be surrounded by an endothelial monolayer and to express tenascin-C. Expression of tenascin-C is specific to BVI and is not observed in lymphatic vascular invasions frequent in breast cancer, which lack an endothelial lining. Given that BVI have prognostic significance for many tumor types, such as shorter cancer patient survival, increased metastasis, vessel occlusion, and organ failure, our data revealing a novel mechanism by which stromal tenascin-C promotes metastasis in human cancer, may have potential for diagnosis and therapy.


Assuntos
Vasos Sanguíneos/patologia , Neoplasias Pulmonares/patologia , Neoplasias Pulmonares/secundário , Neoplasias Mamárias Experimentais/patologia , Receptor ErbB-2/genética , Tenascina/genética , Animais , Vasos Sanguíneos/metabolismo , Linhagem Celular Tumoral , Feminino , Técnicas de Inativação de Genes , Humanos , Neoplasias Pulmonares/irrigação sanguínea , Neoplasias Pulmonares/genética , Neoplasias Mamárias Experimentais/irrigação sanguínea , Neoplasias Mamárias Experimentais/genética , Neoplasias Mamárias Experimentais/metabolismo , Camundongos , Camundongos Transgênicos , Ratos , Transdução de Sinais , Células Estromais , Tenascina/metabolismo , Fator de Crescimento Transformador beta/metabolismo
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