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1.
Bioengineered ; 10(1): 282-291, 2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31311401

RESUMO

Transforming growth factor (TGF)-ß1 plays a crucial role in the epithelial-to-mesenchymal transition (EMT) in many cancer types and in thyroid cancers. Epigallocatechin-3-gallate (EGCG), the most important ingredient in the green tea, has been reported to possess antioxidant and anticancer activities. However, the cellular and molecular mechanisms explaining its action have not been completely understood. In this study, we found that EGCG significantly suppresses EMT, invasion and migration in anaplastic thyroid carcinoma (ATC) 8505C cells in vitro by regulating the TGF-ß/Smad signaling pathways. EGCG significantly inhibited TGF-ß1-induced expression of EMT markers (E-cadherin reduction and vimentin induction) in 8505C cells in vitro. Treatment with EGCG completely blocked the phosphorylation of Smad2/3, translocation of Smad4. Taken together, these results suggest that EGCG suppresses EMT and invasion and migration by blocking TGFß/Smad signaling pathways.


Assuntos
Antineoplásicos Fitogênicos/farmacologia , Catequina/análogos & derivados , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica , Transdução de Sinais/efeitos dos fármacos , Células Epiteliais da Tireoide/efeitos dos fármacos , Fator de Crescimento Transformador beta1/antagonistas & inibidores , Antígenos CD/genética , Antígenos CD/metabolismo , Caderinas/antagonistas & inibidores , Caderinas/genética , Caderinas/metabolismo , Catequina/farmacologia , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Transição Epitelial-Mesenquimal/genética , Humanos , Fosforilação/efeitos dos fármacos , Transporte Proteico/efeitos dos fármacos , Proteína Smad2/genética , Proteína Smad2/metabolismo , Proteína Smad3/genética , Proteína Smad3/metabolismo , Proteína Smad4/genética , Proteína Smad4/metabolismo , Células Epiteliais da Tireoide/metabolismo , Células Epiteliais da Tireoide/patologia , Fator de Crescimento Transformador beta1/farmacologia , Vimentina/agonistas , Vimentina/genética , Vimentina/metabolismo
2.
BMC Cancer ; 19(1): 634, 2019 Jun 27.
Artigo em Inglês | MEDLINE | ID: mdl-31248373

RESUMO

BACKGROUND: Metastasis is a leading cause of breast cancer mortality. The induction of epithelial-to-mesenchymal transition (EMT) and complex oncogenic signaling is a vital step in the evolution of highly metastatic and therapeutically-intractable breast cancer; necessitating novel target discovery or development of therapeutics that target metastatic breast cells (MBCs). METHODS: To achieve this, this study employs a combination of in silico bioinformatics analyses, protein and transcript analyses, drug sensitivity assays, functional assays and animal studies. RESULTS: The present study identified CDH11 as an inductor and/or facilitator of metastatic signaling, and biomarker of poor prognosis in MBCs. Furthermore, we showed that in the presence of CDH11-rich cancer-associated fibroblasts (CAFs), MCF7 and MDA-MB-231 MBC cell lines acquired enhanced metastatic phenotype with increased CDH11, ß-catenin, vimentin, and fibronectin (FN) expression. We also demonstrated, for the first time to the best of our knowledge that exposure to anti-CDH11 antibody suppresses metastasis, reduces CDH11, FN and ß-catenin expression, and abrogate the cancer stem cell (CSC)-like traits of MBC cells. Interestingly, ectopic expression of miR-335 suppressed CDH11, ß-catenin and vimentin expression, in concert with attenuated metastatic and CSC potentials of the MBC cells; conversely, inhibition of miR-335 resulted in increased metastatic potential. Finally, corroborating the in silica and in vitro findings, in vivo assays showed that the administration of anti-CDH11 antibody or miR-335 mimic suppressed tumorigenesis and inhibited cancer metastasis. CONCLUSIONS: These findings validate our hypotheses that miR-335 mediates anti-CDH11 antibody therapy response and that an enhanced miR-335/CDH11 ratio elicits marked suppression of the MBC CSC-like and metastatic phenotypes, thus revealing a therapeutically-exploitable inverse correlation between CDH11-enhanced CSC-like and metastatic phenotype and miR-335 expression in MBCs. Thus, we highlight the therapeutic promise of humanized anti-CDH11 antibodies or miR-335-mimic, making a case for their clinical application as efficacious therapeutic option in patients with MBC.


Assuntos
Neoplasias da Mama/patologia , Caderinas/antagonistas & inibidores , Transição Epitelial-Mesenquimal , Regulação Neoplásica da Expressão Gênica , MicroRNAs/genética , Células-Tronco Neoplásicas/metabolismo , Animais , Anticorpos Monoclonais/farmacologia , Biomarcadores Tumorais/antagonistas & inibidores , Biomarcadores Tumorais/genética , Neoplasias da Mama/genética , Caderinas/genética , Linhagem Celular Tumoral , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Feminino , Humanos , Camundongos Endogâmicos NOD , Camundongos SCID , MicroRNAs/farmacologia , Metástase Neoplásica , Prognóstico , RNA Interferente Pequeno/genética , Ensaios Antitumorais Modelo de Xenoenxerto
3.
Cancer Res ; 79(6): 1113-1123, 2019 03 15.
Artigo em Inglês | MEDLINE | ID: mdl-30674537

RESUMO

Identifying controlling features of responsiveness to checkpoint blockade therapies is an urgent goal in oncology research. Our group and others have previously shown melanoma tumors resistant to checkpoint blockade display features of mesenchymal transition, including E-cadherin loss. Here, we present the first in vivo evidence that E-cadherin from tumor cells facilitate immune attack, using a B16F10 melanoma mouse model in which E-cadherin is exogenously expressed (B16.Ecad). We find, compared with vector control, B16.Ecad exhibits delayed tumor growth, reduced metastatic potential, and increased overall survival in vivo. Transplantation of B16.Ecad into Rag1-/- and CD103-/- mice abrogated the tumor growth delay. This indicates the anti-melanoma response against B16.Ecad is both immune and CD103+ mediated. Moreover, B16.Ecad showed increased responsiveness to combination immune checkpoint blockade (ICB) compared with vector control. This work establishes a rationale for ICB responses observed in high E-cadherin-expressing tumors and suggests therapeutic advancement through amplifying CD103+ immune cell subsets.Significance: These findings identify the mechanism behind checkpoint blockade resistance observed in melanoma that has undergone mesenchymal transition and suggest activation of CD103+ immune cells as a therapeutic strategy against other E-cadherin-expressing malignancies.Graphical Abstract: http://cancerres.aacrjournals.org/content/canres/79/6/1113/F1.large.jpg.


Assuntos
Antígenos CD/metabolismo , Antineoplásicos Imunológicos/farmacologia , Caderinas/metabolismo , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Cadeias alfa de Integrinas/metabolismo , Neoplasias Pulmonares/secundário , Melanoma Experimental/patologia , Animais , Antígenos CD/genética , Apoptose , Linfócitos T CD8-Positivos/efeitos dos fármacos , Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/patologia , Caderinas/antagonistas & inibidores , Caderinas/genética , Proliferação de Células , Regulação Neoplásica da Expressão Gênica , Cadeias alfa de Integrinas/genética , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/imunologia , Neoplasias Pulmonares/metabolismo , Melanoma Experimental/tratamento farmacológico , Melanoma Experimental/imunologia , Melanoma Experimental/metabolismo , Camundongos , Células Tumorais Cultivadas , Microambiente Tumoral
4.
Oncogene ; 38(4): 455-468, 2019 01.
Artigo em Inglês | MEDLINE | ID: mdl-30111817

RESUMO

Epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor (TKI) is a major advance in treating NSCLC with EGFR-activating mutations. However, acquired resistance, due partially to secondary mutations limits their use. Here we report that NSCLC cells with acquired resistance to gefitinib or osimertinib (AZD9291) exhibit EMT features, with a decrease in E-cadherin, and increases in vimentin and stemness, without possessing any EGFR secondary mutations. Knockdown of E-cadherin in parental cells increased gefitinib resistance and stemness, while knockdown of vimentin in resistant cells resulted in opposite effects. Src activation and Hakai upregulation were found in gefitinib-resistant cells. Knockdown of Hakai elevated E-cadherin expression, attenuated stemness, and resensitized the cells to gefitinib. Clinical cancer specimens with acquired gefitinib resistance also showed a decrease in E-cadherin and an increase in Hakai expression. The dual HDAC and HMGR inhibitor JMF3086 inhibited the Src/Hakai and Hakai/E-cadherin interaction to reverse E-cadherin expression, and attenuated vimentin and stemness to restore gefitinib sensitivity. The EMT features of AZD9291-resistant H1975 cells were related to the upregulation of Zeb1. Both gefitinib and AZD9291 sensitivity was restored by JMF3086 through reversing EMT. Our study not only revealed a common mechanism of EMT in both gefitinib and AZD9291 resistance beyond EGFR mutations per se, but also provides a new strategy to overcome it.


Assuntos
Antineoplásicos/farmacologia , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Resistencia a Medicamentos Antineoplásicos/genética , Transição Epitelial-Mesenquimal/fisiologia , Gefitinibe/farmacologia , Neoplasias Pulmonares/tratamento farmacológico , Proteínas de Neoplasias/fisiologia , Piperazinas/farmacologia , Inibidores de Proteínas Quinases/farmacologia , Acrilamidas , Compostos de Anilina , Animais , Antígenos CD/biossíntese , Antígenos CD/genética , Caderinas/antagonistas & inibidores , Caderinas/biossíntese , Caderinas/genética , Carcinoma Pulmonar de Células não Pequenas/genética , Carcinoma Pulmonar de Células não Pequenas/patologia , Linhagem Celular Tumoral , Resistencia a Medicamentos Antineoplásicos/fisiologia , Ensaios de Seleção de Medicamentos Antitumorais , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Transição Epitelial-Mesenquimal/genética , Receptores ErbB/genética , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Inibidores de Histona Desacetilases/farmacologia , Humanos , Inibidores de Hidroximetilglutaril-CoA Redutases/farmacologia , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Proteínas de Neoplasias/antagonistas & inibidores , Proteínas de Neoplasias/genética , Células-Tronco Neoplásicas/efeitos dos fármacos , Células-Tronco Neoplásicas/patologia , Interferência de RNA , RNA Interferente Pequeno/farmacologia , Organismos Livres de Patógenos Específicos , Ubiquitina-Proteína Ligases/antagonistas & inibidores , Ubiquitina-Proteína Ligases/biossíntese , Ubiquitina-Proteína Ligases/genética , Vimentina/antagonistas & inibidores , Vimentina/biossíntese , Vimentina/genética , Homeobox 1 de Ligação a E-box em Dedo de Zinco/biossíntese , Homeobox 1 de Ligação a E-box em Dedo de Zinco/genética
5.
Carcinogenesis ; 40(1): 15-26, 2019 03 12.
Artigo em Inglês | MEDLINE | ID: mdl-30508037

RESUMO

Histone modification plays important molecular roles in development and progression of cancers. Dysregulation of histone H3 arginine (R) methylation is still unknown in primary cancer, including gastric cancer (GC). Although PRMT6 contributes to asymmetric dimethylation at H3R2 (H3R2me2as) in cancer cells, its molecular functions are poorly understood in GC. In this study, we assessed H3R2me2as and PRMT6 expression levels in 133 primary GC tissues by immunohistochemistry. Increased H3R2me2as was found in 68 GC (51.1%) cases and independently related to poor prognosis. PRMT6 was overexpressed in 70 GC (52.6%) and strongly correlated with the global H3R2me2as levels (P < 0.001). By analyzing biological functions of PRMT6 in GC cell lines by lentivirus-based systems, PRMT6 overexpression enhanced global H3R2me2as and invasiveness in vitro, while PRMT6 knockout (PRMT6-KO) suppressed these effects and tumorigenicity in vivo. ChIP and microarray assays demonstrated that PRMT6-KO GC cells decreased the enrichments of H3R2me2as at the promoter regions of PCDH7, SCD and IGFBP5, resulting in upregulation of their gene expression. PRMT6 was recruited to the promoter regions of PCDH7 and SCD in the PRMT6-overexpressed cells. Knockdown of tumor suppressor PCDH7 in the PRMT6-KO GC cells elevated cell migration and invasion. PRMT6 expression inversely correlated with PCDH7 expression in primary GC (P = 0.021). Collectively, our findings strongly indicate that H3R2me2as is a strong prognostic indicator of GC patients, and PRMT6-overexpressing GC cells may acquire invasiveness through direct transcriptional inhibition of PCDH7 by increasing H3R2me2as level. Thus, inhibition of the PRMT6-H3R2me2as pathway could be a promising new therapeutic strategy in GC.


Assuntos
Histonas/metabolismo , Proteínas Nucleares/fisiologia , Proteína-Arginina N-Metiltransferases/fisiologia , Neoplasias Gástricas/metabolismo , Animais , Arginina/metabolismo , Caderinas/antagonistas & inibidores , Caderinas/fisiologia , Linhagem Celular Tumoral , Progressão da Doença , Feminino , Humanos , Masculino , Metilação , Camundongos , Neoplasias Gástricas/patologia
6.
Biomed Pharmacother ; 109: 2293-2304, 2019 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-30551487

RESUMO

EndMT plays an important role in the relationship between endothelial dysfunction and atherosclerosis. This work will elucidate the biofunction induced by miR-449a and lipid rafts in EndMT and development of atherosclerosis. The differential miRNA expression between atherosclerotic plaques and normal arteries were analyzed. The luciferase activities of AdipoR2 3' UTR treated with miR-449a were determined. ECs were dealt with miR-449a mimics or inhibitors, then cell proliferation and migration were assessed. Moreover, the expression of AdipoR2 and mesenchymal cell markers were analyzed. The influences of lipid rafts related to reciprocity between E-cadherin and AdipoR2 on TNF-α-induced damage in ECs were investigated. ApoE KO diabetic mice were used to explore the potential roles of miR-449a on atherosclerosis. Our results indicated that compared with normal arteries, 17 miRNAs were upregulated and 3 miRNAs were down-regulated in atherosclerotic plaques. The relative expression of miR-449a in plaques was significantly higher than that in normal arteries. MiR-449a suppressed AdipoR2 expression, additionally its interaction protein E-cadherin in ECs. MiR-449a enhanced expression of mesenchymal cell markers, induced cell proliferation and migration of ECs, regulated the interaction between E-cadherin and AdipoR2 interceded by lipid rafts. The miR-449a antagomir could protect against the development process of atherosclerosis in ApoE KO diabetic mice. In conclusion, miR-449a targeted to AdipoR2, and was a crucial mediator of EndMT and atherosclerosis in ECs through regulating E-cadherin bindability with AdipoR2 in lipid rafts. These results suggested that aim to lipid rafts and miR-449a in chronic EC inflammation response, was a feasible therapy strategy for atherosclerosis.


Assuntos
Aterosclerose/metabolismo , Caderinas/metabolismo , Transição Epitelial-Mesenquimal/fisiologia , Microdomínios da Membrana/metabolismo , MicroRNAs/biossíntese , Receptores de Adiponectina/biossíntese , Animais , Aterosclerose/patologia , Caderinas/antagonistas & inibidores , Diabetes Mellitus Experimental/metabolismo , Diabetes Mellitus Experimental/patologia , Humanos , Masculino , Microdomínios da Membrana/patologia , Camundongos , Camundongos Knockout , Receptores de Adiponectina/antagonistas & inibidores
7.
Mol Vis ; 24: 759-766, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30581282

RESUMO

Purpose: To identify retinal protein changes that mediate beneficial effects of intravitreal bevacizumab in experimental branch retinal vein occlusion (BRVO). Methods: In six Danish Landrace pigs, BRVO was induced with argon laser in both eyes. After BRVO was induced, the right eye of each animal was given an intravitreal injection of bevacizumab while the left eye was treated with saline water. The retinas were collected 15 days after BRVO, and differentially expressed proteins were analyzed with tandem mass tags-based mass spectrometry. Validation of statistically significantly changed proteins was performed with immunohistochemistry and western blotting. Results: Fluorescein angiography showed no recanalization of the occluded vessels. A total of 4,013 proteins were successfully identified and quantified. Nine proteins were statistically significantly changed following bevacizumab intervention. In experimental BRVO, bevacizumab treatment resulted in upregulation of transthyretin (TTR) and pantothenate kinase 3. Bevacizumab downregulated protocadherin 7, protein FAM192A, and ATP synthase protein 8. Immunohistochemistry revealed that TTR was highly abundant in the choroid following bevacizumab intervention. Conclusions: Bevacizumab intervention in experimental BRVO resulted in an increased level of TTR. This is the second study in which we showed an increased retinal level of TTR following anti-vascular endothelial growth factor (VEGF) intervention in experimental BRVO. We hypothesize that there is an interaction between TTR and VEGF and that bevacizumab may exert a beneficial effect on the retina by upregulating TTR.


Assuntos
Inibidores da Angiogênese/farmacologia , Bevacizumab/farmacologia , Regulação da Expressão Gênica/efeitos dos fármacos , Pré-Albumina/genética , Retina/efeitos dos fármacos , Oclusão da Veia Retiniana/tratamento farmacológico , Animais , Caderinas/antagonistas & inibidores , Caderinas/genética , Caderinas/metabolismo , Corioide/irrigação sanguínea , Corioide/diagnóstico por imagem , Corioide/efeitos dos fármacos , Corioide/metabolismo , Angiofluoresceinografia , Perfilação da Expressão Gênica , Humanos , Cadeias gama de Imunoglobulina/genética , Cadeias gama de Imunoglobulina/metabolismo , Cadeias kappa de Imunoglobulina/genética , Cadeias kappa de Imunoglobulina/metabolismo , Injeções Intravítreas , ATPases Mitocondriais Próton-Translocadoras/antagonistas & inibidores , ATPases Mitocondriais Próton-Translocadoras/genética , ATPases Mitocondriais Próton-Translocadoras/metabolismo , Fosfotransferases (Aceptor do Grupo Álcool)/genética , Fosfotransferases (Aceptor do Grupo Álcool)/metabolismo , Pré-Albumina/agonistas , Pré-Albumina/metabolismo , Retina/diagnóstico por imagem , Retina/metabolismo , Retina/patologia , Oclusão da Veia Retiniana/diagnóstico por imagem , Oclusão da Veia Retiniana/genética , Oclusão da Veia Retiniana/patologia , Suínos
8.
Science ; 361(6402): 599-603, 2018 08 10.
Artigo em Inglês | MEDLINE | ID: mdl-30093598

RESUMO

Excess dietary lipid uptake causes obesity, a major global health problem. Enterocyte-absorbed lipids are packaged into chylomicrons, which enter the bloodstream through intestinal lymphatic vessels called lacteals. Here, we show that preventing lacteal chylomicron uptake by inducible endothelial genetic deletion of Neuropilin1 (Nrp1) and Vascular endothelial growth factor receptor 1 (Vegfr1; also known as Flt1) renders mice resistant to diet-induced obesity. Absence of NRP1 and FLT1 receptors increased VEGF-A bioavailability and signaling through VEGFR2, inducing lacteal junction zippering and chylomicron malabsorption. Restoring permeable lacteal junctions by VEGFR2 and vascular endothelial (VE)-cadherin signaling inhibition rescued chylomicron transport in the mutant mice. Zippering of lacteal junctions by disassembly of cytoskeletal VE-cadherin anchors prevented chylomicron uptake in wild-type mice. These data suggest that lacteal junctions may be targets for preventing dietary fat uptake.


Assuntos
Quilomícrons/metabolismo , Dieta Hiperlipídica/efeitos adversos , Gorduras na Dieta/metabolismo , Neuropilina-1/genética , Obesidade/etiologia , Obesidade/genética , Receptor 1 de Fatores de Crescimento do Endotélio Vascular/genética , Animais , Antígenos CD/metabolismo , Caderinas/antagonistas & inibidores , Caderinas/metabolismo , Quilomícrons/efeitos adversos , Gorduras na Dieta/efeitos adversos , Enterócitos/metabolismo , Deleção de Genes , Absorção Intestinal/genética , Absorção Intestinal/fisiologia , Masculino , Camundongos , Camundongos Knockout , Transdução de Sinais , Fator A de Crescimento do Endotélio Vascular/antagonistas & inibidores , Fator A de Crescimento do Endotélio Vascular/metabolismo , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/antagonistas & inibidores , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/metabolismo
9.
Biochemistry ; 57(32): 4867-4879, 2018 08 14.
Artigo em Inglês | MEDLINE | ID: mdl-30001488

RESUMO

Despite the well-established biophysical principle of adhesion-guided in vitro morphogenesis, there are few single synthetic molecular species that can rapidly enable morphogenesis (e.g., a cell monolayer to cell spheroids) in a cell culture because adhesion inherently involves many signals. Here we show the use of adaptive multifunctional supramolecular assemblies of glycopeptides, consisting of cell adhesion sequence and saccharide, to induce cell spheroids rapidly from a monolayer of cells. Having a general architecture of N-terminal capping, glycosylation, and an integrin-binding sequence, the glycopeptides self-assemble to form a dynamic continuum of nanostructures (i.e., from nanoparticles to nanofibers) to affect the interactions of integrins, E-selectin, and cadherins with their natural ligands and to act adaptively according to the cellular environment. Such adaptive (i.e., context-dependent) interactions weaken cell-substratum adhesion and enhance intercellular interactions, which rapidly and transiently induce cell spheroids. This work illustrates the use of supramolecular assemblies of simple glycopeptides to modulate biophysical conditions for regulating cell functions, which is a new approach for developing biomaterials.


Assuntos
Glicopeptídeos/química , Caderinas/antagonistas & inibidores , Caderinas/metabolismo , Adesão Celular/efeitos dos fármacos , Técnicas de Cultura de Células , Selectina E/antagonistas & inibidores , Selectina E/metabolismo , Ácidos Graxos Monoinsaturados/farmacologia , Fibronectinas/antagonistas & inibidores , Fibronectinas/metabolismo , Glicopeptídeos/farmacologia , Humanos , Integrinas/antagonistas & inibidores , Integrinas/metabolismo , Morfogênese/efeitos dos fármacos , Nanoestruturas/química
10.
Chem Biol Interact ; 292: 24-29, 2018 Aug 25.
Artigo em Inglês | MEDLINE | ID: mdl-29932878

RESUMO

Elevated cyclooxygenase-2 (COX-2) closely associates with tumor progression and distant metastasis in various human cancers. However, the role of COX-2 in epithelial ovarian cancer (EOC), and its mechanistic details, remain poorly understood. In the present study, we tested hypothesis that COX-2 induces loss of expression of E-cadherin, with resulting promotion of cancer cells' invasiveness in ovarian cancer. First, we observed an inverse relationship between COX-2 and E-cadherin expression as COX-2 was enhanced but E-cadherin was decreased in surgically-resected specimens of EOC. Depletion of COX-2, by celecoxib treatment, resulted in attenuated nuclear translocation of Snail, and, in turn, significantly increased E-cadherin in EOC cell line SKOV3, which was established to be due to the reduced binding of Snail onto E-cadherin promoter. Such COX-2 inhibition resulted in reduced invasion of EOC cells, similar to what was achieved through Snail silencing in SKOV as well as ES-2 EOC cells. These results suggest that COX-2-Snail signaling plays a critical role in regulation of E-cadherin and might provide insights into mechanisms for paracrine inflammation-mediated aggressiveness in EOC.


Assuntos
Caderinas/genética , Caderinas/metabolismo , Celecoxib/farmacologia , Ciclo-Oxigenase 2/metabolismo , Neoplasias Epiteliais e Glandulares/fisiopatologia , Neoplasias Ovarianas/fisiopatologia , Fatores de Transcrição da Família Snail/metabolismo , Western Blotting , Caderinas/antagonistas & inibidores , Carcinoma Epitelial do Ovário , Linhagem Celular Tumoral , Regulação para Baixo , Feminino , Humanos , Inflamação/tratamento farmacológico , Neoplasias Epiteliais e Glandulares/tratamento farmacológico , Neoplasias Ovarianas/tratamento farmacológico , Reação em Cadeia da Polimerase , Regiões Promotoras Genéticas , Transdução de Sinais/efeitos dos fármacos
11.
12.
Toxicol Lett ; 294: 135-144, 2018 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-29778911

RESUMO

Di(2-ethylhexyl) phthalate (DEHP) is a widely used plasticizer that is metabolized to mono(2-ethylhexyl) phthalate (MEHP). Inhalation is an important exposure route for both phthalates, and their effects on lungs include inflammation, alteration of postnatal maturation (alveolarization), enlarged airspaces and cell differentiation changes, suggesting that alveolar epithelial cells-2 (AEC) are targets of phthalates. This study evaluated the cell progression, epithelial and mesenchymal markers, including surfactant secretion in A549 cells (AEC) that were exposed to DEHP (1-100 µM) or MEHP (1-50 µM) for 24-72 h. The results showed an increased cell proliferation at all concentrations of each phthalate at 24 and 48 h. Cell migration showed a concentration-dependent increase at 24 and 48 h of exposure to either phthalate and enlarged structures were seen. Decreased levels of both surfactants (SP-B/SP-C) were observed after the exposure to either phthalate at 48 h, and of SP-C positive cells exposed to MEHP, suggesting a loss of the epithelial phenotype. While a decrease in the epithelial marker E-cadherin and an increase in the mesenchymal marker fibronectin were observed following exposure to either phthalate. Our results showed that DEHP and MEHP altered the structure and migration of A549 cells and promoted the loss of the epithelial phenotype.


Assuntos
Células Epiteliais Alveolares/efeitos dos fármacos , Desdiferenciação Celular/efeitos dos fármacos , Dietilexilftalato/análogos & derivados , Dietilexilftalato/toxicidade , Plastificantes/toxicidade , Proteína B Associada a Surfactante Pulmonar/antagonistas & inibidores , Proteína C Associada a Surfactante Pulmonar/antagonistas & inibidores , Células A549 , Células Epiteliais Alveolares/citologia , Células Epiteliais Alveolares/metabolismo , Antígenos CD , Biomarcadores/metabolismo , Caderinas/antagonistas & inibidores , Caderinas/metabolismo , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Forma Celular/efeitos dos fármacos , Tamanho Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Fibronectinas/agonistas , Fibronectinas/metabolismo , Humanos , Cinética , Proteína B Associada a Surfactante Pulmonar/metabolismo , Proteína C Associada a Surfactante Pulmonar/metabolismo
13.
Cell Death Differ ; 25(11): 2023-2036, 2018 11.
Artigo em Inglês | MEDLINE | ID: mdl-29666468

RESUMO

During tissue repair, the injury site releases various bioactive molecules as damage signals to actively recruit stem cells to the damaged region. Despite convincing evidence that mesenchymal stem cells (MSCs) can sense damage signals and promote repair processes, the identity of these signals and how these signals regulate stem cell-mediated tissue repair remain unknown. Glycyl tRNA synthetase (GRS) is a ubiquitously expressed enzyme that catalyzes the first step of protein synthesis in all organisms. In addition to this canonical function, we identified for the first time that GRS is released by damaged tissues or cells in response to various injury signals and may function as a damage signal that activates the proliferative, differentiation, and migratory potential of MSCs, possibly through its identified receptor, cadherin-6 (CDH-6). Binding between GRS and CDH-6 activates survival signals, such as those of the PI3K/Akt and/or FAK/ERK1/2 pathways. More importantly, we also found that MSCs stimulated with GRS show significantly improved homing and differentiation potential and subsequent in vivo therapeutic effects, in a liver fibrosis animal model. Collectively, our findings provide compelling evidence for a novel function of GRS in enhancing the multiple beneficial functions of stem cells via a non-canonical mechanism as a damage signal.


Assuntos
Glicina-tRNA Ligase/metabolismo , Células-Tronco Mesenquimais/metabolismo , Tecido Adiposo/citologia , Caderinas/antagonistas & inibidores , Caderinas/genética , Caderinas/metabolismo , Tetracloreto de Carbono/toxicidade , Diferenciação Celular/efeitos dos fármacos , Movimento Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Quinase 1 de Adesão Focal/metabolismo , Glicina-tRNA Ligase/genética , Glicina-tRNA Ligase/farmacologia , Humanos , Falência Hepática Aguda/induzido quimicamente , Falência Hepática Aguda/metabolismo , Falência Hepática Aguda/patologia , Sistema de Sinalização das MAP Quinases , Metaloproteinase 2 da Matriz/metabolismo , Células-Tronco Mesenquimais/citologia , Proteína Homeobox Nanog/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Interferência de RNA , RNA Interferente Pequeno/metabolismo , Transdução de Sinais/efeitos dos fármacos
14.
Biochim Biophys Acta Rev Cancer ; 1869(2): 321-332, 2018 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-29673969

RESUMO

We propose a new cadherin family classification comprising epithelial cadherins (cadherin 17 [CDH17], cadherin 16, VE-cadherin, cadherin 6 and cadherin 20) containing RGD motifs within their sequences. Expression of some RGD cadherins is associated with aggressive forms of cancer during the late stages of metastasis, and CDH17 and VE-cadherin have emerged as critical actors in cancer metastasis. After binding to α2ß1 integrin, these cadherins promote integrin ß1 activation, and thereby cell adhesion, invasion and proliferation, in liver and lung metastasis. Activation of α2ß1 integrin provokes an affinity increase for type IV collagen, a major component of the basement membrane and a critical partner for cell anchoring in liver and other metastatic organs. Activation of α2ß1 integrin by RGD motifs breaks an old paradigm of integrin classification and supports an important role of this integrin in cancer metastasis. Recently, synthetic peptides containing the RGD motif of CDH17 elicited highly specific and selective antibodies that block the ability of CDH17 RGD to activate α2ß1 integrin. These monoclonal antibodies inhibit metastatic colonization in orthotopic mouse models of liver and lung metastasis for colorectal cancer and melanoma, respectively. Hopefully, blocking the cadherin RGD ligand capacity will give us control over the integrin activity in solid tumors metastasis, paving the way for development of new agents of cancer treatment.


Assuntos
Caderinas/metabolismo , Movimento Celular , Integrina alfa2beta1/metabolismo , Neoplasias/metabolismo , Oligopeptídeos/metabolismo , Receptores Imunológicos/metabolismo , Receptores de Peptídeos/metabolismo , Animais , Anticorpos Monoclonais/uso terapêutico , Antineoplásicos Imunológicos/uso terapêutico , Sítios de Ligação , Caderinas/antagonistas & inibidores , Caderinas/imunologia , Adesão Celular , Movimento Celular/efeitos dos fármacos , Proliferação de Células , Humanos , Integrina alfa2beta1/antagonistas & inibidores , Integrina alfa2beta1/imunologia , Metástase Neoplásica , Neoplasias/tratamento farmacológico , Neoplasias/patologia , Ligação Proteica , Receptores Imunológicos/antagonistas & inibidores , Receptores Imunológicos/imunologia , Receptores de Peptídeos/antagonistas & inibidores , Receptores de Peptídeos/imunologia , Transdução de Sinais
15.
Cell Physiol Biochem ; 46(2): 482-491, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29614512

RESUMO

BACKGROUND/AIMS: An adequate matrix production of nucleus pulposus (NP) cells is an important tissue engineering-based strategy to regenerate degenerative discs. Here, we mainly aimed to investigate the effects and mechanism of mechanical compression (i.e., static compression vs. dynamic compression) on the matrix synthesis of three-dimensional (3D) cultured NP cells in vitro. METHODS: Rat NP cells seeded on small intestinal submucosa (SIS) cryogel scaffolds were cultured in the chambers of a self-developed, mechanically active bioreactor for 10 days. Meanwhile, the NP cells were subjected to compression (static compression or dynamic compression at a 10% scaffold deformation) for 6 hours once per day. Unloaded NP cells were used as controls. The cellular phenotype and matrix biosynthesis of NP cells were investigated by real-time PCR and Western blotting assays. Lentivirus-mediated N-cadherin (N-CDH) knockdown and an inhibitor, LY294002, were used to further investigate the role of N-CDH and the PI3K/Akt pathway in this process. RESULTS: Dynamic compression better maintained the expression of cell-specific markers (keratin-19, FOXF1 and PAX1) and matrix macromolecules (aggrecan and collagen II), as well as N-CDH expression and the activity of the PI3K/Akt pathway, in the 3D-cultured NP cells compared with those expression levels and activity in the cells grown under static compression. Further analysis showed that the N-CDH knockdown significantly down-regulated the expression of NP cell-specific markers and matrix macromolecules and inhibited the activation of the PI3K/Akt pathway under dynamic compression. However, inhibition of the PI3K/Akt pathway had no effects on N-CDH expression but down-regulated the expression of NP cell-specific markers and matrix macromolecules under dynamic compression. CONCLUSION: Dynamic compression increases the matrix synthesis of 3D-cultured NP cells compared with that of the cells under static compression, and the N-CDH-PI3K/Akt pathway is involved in this regulatory process. This study provides a promising strategy to promote the matrix deposition of tissue-engineered NP tissue in vitro prior to clinical transplantation.


Assuntos
Caderinas/metabolismo , Força Compressiva/fisiologia , Matriz Extracelular/metabolismo , Animais , Caderinas/antagonistas & inibidores , Caderinas/genética , Células Cultivadas , Cromonas/farmacologia , Fatores de Transcrição Forkhead/genética , Fatores de Transcrição Forkhead/metabolismo , Géis/química , Regulação da Expressão Gênica , Queratina-19/genética , Masculino , Morfolinas/farmacologia , Núcleo Pulposo/citologia , Núcleo Pulposo/metabolismo , Fenótipo , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Interferência de RNA , Ratos , Ratos Sprague-Dawley , Transdução de Sinais/efeitos dos fármacos , Tecidos Suporte/química
16.
Cell Physiol Biochem ; 45(5): 1966-1974, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29518783

RESUMO

BACKGROUND/AIMS: Studies have shown that miR-194 functions as a tumour suppressor and is associated with tumour growth and metastasis. This study intends to uncover the mechanism of tumour suppression by miR-194. The expression of miR-194 in osteosarcoma cell lines and tissues were monitored by real-time PCR. METHODS: The proliferation ability was examined by MTT assay. Migration and apoptosis of cells were monitored by migration assay and flow cytometry, respectively. The regulation of miR-194 on CDH2 was determined by luciferase assays and western blot assays. RESULTS: The results showed that miR-194 was significantly reduced in osteosarcoma compared with that in normal bone tissue. Overexpression of miR-194 significantly attenuated the proliferation and migration and induced the apoptosis of osteosarcoma cells. Furthermore, we demonstrated that miR-194 has inhibited the malignant behaviour of osteosarcoma by downregulating CDH2 expression. CONCLUSIONS: These findings suggested that miR-194 may act as a tumour suppressor in osteosarcoma. miR-194/CDH2 may be a novel therapeutic target in the treatment of osteosarcoma.


Assuntos
Antígenos CD/metabolismo , Apoptose , Neoplasias Ósseas/patologia , Caderinas/metabolismo , Proliferação de Células , MicroRNAs/metabolismo , Osteossarcoma/patologia , Regiões 3' não Traduzidas , Antagomirs/metabolismo , Antígenos CD/genética , Sequência de Bases , Neoplasias Ósseas/genética , Neoplasias Ósseas/metabolismo , Caderinas/antagonistas & inibidores , Caderinas/genética , Linhagem Celular Tumoral , Movimento Celular , Regulação para Baixo , Humanos , MicroRNAs/antagonistas & inibidores , MicroRNAs/genética , Osteossarcoma/genética , Osteossarcoma/metabolismo , Alinhamento de Sequência
17.
J Med Chem ; 61(7): 2989-3007, 2018 04 12.
Artigo em Inglês | MEDLINE | ID: mdl-29566337

RESUMO

Structure-based optimization was conducted to improve the potency, selectivity, and cell-based activities of ß-catenin/B-cell lymphoma 9 (BCL9) inhibitors based on the 4'-fluoro- N-phenyl-[1,1'-biphenyl]-3-carboxamide scaffold, which was designed to mimic the side chains of the hydrophobic α-helical hot spots at positions i, i + 3, and i + 7. Compound 29 was found to disrupt the ß-catenin/BCL9 protein-protein interaction (PPI) with a Ki of 0.47 µM and >1900-fold selectivity for ß-catenin/BCL9 over ß-catenin/E-cadherin PPIs. The proposed binding mode of new inhibitors was consistent with the results of site-directed mutagenesis and structure-activity relationship studies. Cell-based studies indicated that 29 disrupted the ß-catenin/BCL9 interaction without affecting the ß-catenin/E-cadherin interaction, selectively suppressed transactivation of Wnt/ß-catenin signaling, downregulated expression of Wnt target genes, and inhibited viability of Wnt/ß-catenin-dependent cancer cells in dose-dependent manners. A comparison of the biochemical and cell-based assay results offered the directions for future inhibitor optimization.


Assuntos
Antineoplásicos/síntese química , Antineoplásicos/farmacologia , Proteínas de Neoplasias/antagonistas & inibidores , Caderinas/antagonistas & inibidores , Linhagem Celular Tumoral , Relação Dose-Resposta a Droga , Desenho de Drogas , Humanos , Conformação Molecular , Mutagênese Sítio-Dirigida , Proteínas de Neoplasias/genética , Reação em Cadeia da Polimerase , Bibliotecas de Moléculas Pequenas , Relação Estrutura-Atividade , Fatores de Transcrição , Via de Sinalização Wnt/efeitos dos fármacos , beta Catenina/efeitos dos fármacos
18.
Artigo em Inglês | MEDLINE | ID: mdl-29474162

RESUMO

Cadherin-17 (CDH17) is highly expressed in gastric cancer and is thus considered to be a good target for antibody therapy. CDH17 is classified as a nonclassical cadherin, in that it is composed of seven extracellular cadherin domains. We generated anti-CDH17 monoclonal antibodies (mAbs) which recognize the extracellular domain of CDH17. Competitive assay using AGS, a gastric cancer cell line, cells revealed that five selected anti-CDH17 mAbs recognize different epitopes on CDH17. As AGS cells were shown to exhibit broad expression pattern of CDH17 by flow cytometry, we separated three clones with a low (10,000/cell), medium (50,000/cell), and high (200,000/cell) expression level, designating them as AGSlow, AGSmed, and AGShigh, respectively. The mAbs, coupled with saporin, exhibited effective cytotoxicity to AGShigh, but poor cytotoxicity to AGSlow. By contrast, the immunotoxin cocktail using the three clones D2101, D2005, and D2008, which recognize different epitopes, exhibited efficient cytotoxicity, even to the AGSlow group. The effect of the immunotoxin cocktail is synergistic, as the combination index was demonstrated to be below 1.0, as calculated by the method of Chou and Talalay using CalcuSyn software. These results suggest that the immunotoxin cocktail targeted to multiple epitopes has synergistic effects on low expression level cells, which expand the applicable range of immunotoxin therapy for cancer.


Assuntos
Anticorpos Monoclonais/farmacologia , Apoptose/efeitos dos fármacos , Caderinas/imunologia , Sinergismo Farmacológico , Epitopos/imunologia , Imunotoxinas/farmacologia , Neoplasias Gástricas/patologia , Biomarcadores Tumorais/metabolismo , Caderinas/antagonistas & inibidores , Caderinas/metabolismo , Humanos , Neoplasias Gástricas/imunologia , Neoplasias Gástricas/metabolismo , Células Tumorais Cultivadas
19.
Proc Natl Acad Sci U S A ; 115(5): 998-1003, 2018 01 30.
Artigo em Inglês | MEDLINE | ID: mdl-29343641

RESUMO

The F-box protein FBXO31 is a tumor suppressor that is encoded in 16q24.3, for which there is loss of heterozygosity in various solid tumors. FBXO31 serves as the substrate-recognition component of the SKP/Cullin/F-box protein class of E3 ubiquitin ligases and has been shown to direct degradation of pivotal cell-cycle regulatory proteins including cyclin D1 and the p53 antagonist MDM2. FBXO31 levels are normally low but increase substantially following genotoxic stress through a mechanism that remains to be determined. Here we show that the low levels of FBXO31 are maintained through proteasomal degradation by anaphase-promoting complex/cyclosome (APC/C). We find that the APC/C coactivators CDH1 and CDC20 bind to a destruction-box (D-box) motif present in FBXO31 to promote its polyubiquitination and degradation in a cell-cycle-regulated manner, which requires phosphorylation of FBXO31 on serine-33 by the prosurvival kinase AKT. Following genotoxic stress, phosphorylation of FBXO31 on serine-278 by another kinase, the DNA damage kinase ATM, results in disruption of its interaction with CDH1 and CDC20, thereby preventing FBXO31 degradation. Collectively, our results reveal how alterations in FBXO31 phosphorylation, mediated by AKT and ATM, underlie physiological regulation of FBXO31 levels in unstressed and genotoxically stressed cells.


Assuntos
Ciclossomo-Complexo Promotor de Anáfase/metabolismo , Proteínas Mutadas de Ataxia Telangiectasia/metabolismo , Proteínas F-Box/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Proteínas Supressoras de Tumor/metabolismo , Ciclossomo-Complexo Promotor de Anáfase/antagonistas & inibidores , Ciclossomo-Complexo Promotor de Anáfase/genética , Antígenos CD , Caderinas/antagonistas & inibidores , Caderinas/genética , Caderinas/metabolismo , Proteínas Cdc20/antagonistas & inibidores , Proteínas Cdc20/genética , Proteínas Cdc20/metabolismo , Pontos de Checagem do Ciclo Celular , Dano ao DNA , Proteínas F-Box/química , Técnicas de Silenciamento de Genes , Células HEK293 , Humanos , Modelos Biológicos , Fosforilação , Complexo de Endopeptidases do Proteassoma/metabolismo , Proteólise , RNA Interferente Pequeno/genética , Proteínas Supressoras de Tumor/química , Ubiquitinação
20.
Clin Cancer Res ; 24(2): 433-444, 2018 01 15.
Artigo em Inglês | MEDLINE | ID: mdl-28916526

RESUMO

Purpose: New targets are required for the control of advanced metastatic disease. We investigated the use of cadherin RGD motifs, which activate the α2ß1integrin pathway, as targets for the development of therapeutic monoclonal antibodies (mAb).Experimental Design: Cadherin 17 (CDH17) fragments and peptides were prepared and used for immunization and antibody development. Antibodies were tested for inhibition of ß1 integrin and cell adhesion, proliferation, and invasion assays using cell lines from different cancer types (colorectal, pancreatic, melanoma, and breast cancer). Effects of the mAbs on cell signaling were determined by Western blot analysis. Nude mice were used for survival analysis after treatment with RGD-specific mAbs and metastasis development.Results: Antibodies against full-length CDH17 failed to block the binding to α2ß1 integrin. However, CDH17 RGD peptides generated highly selective RGD mAbs that blocked CDH17 and vascular-endothelial (VE)-cadherin-mediated ß1 integrin activation in melanoma and breast, pancreatic, and colorectal cancer cells. Antibodies provoked a significant reduction in cell adhesion and proliferation of metastatic cancer cells. Treatment with mAbs impaired the integrin signaling pathway activation of FAK in colorectal cancer, of JNK and ERK kinases in colorectal and pancreatic cancers, and of JNK, ERK, Src, and AKT in melanoma and breast cancer. In vivo, RGD-specific mAbs increased mouse survival after inoculation of melanoma and colorectal cancer cell lines to cause lung and liver metastasis, respectively.Conclusions: Blocking the interaction between RGD cadherins and α2ß1 integrin with highly selective mAbs constitutes a promising therapy against advanced metastatic disease in colon cancer, melanoma, and, potentially, other cancers. Clin Cancer Res; 24(2); 433-44. ©2017 AACRSee related commentary by Marshall, p. 253.


Assuntos
Antineoplásicos Imunológicos/farmacologia , Caderinas/antagonistas & inibidores , Neoplasias Colorretais/metabolismo , Neoplasias Colorretais/patologia , Melanoma/metabolismo , Melanoma/patologia , Animais , Caderinas/metabolismo , Adesão Celular , Linhagem Celular Tumoral , Neoplasias Colorretais/mortalidade , Modelos Animais de Doenças , Humanos , Integrina beta1/metabolismo , Metástase Neoplásica , Estadiamento de Neoplasias , Transdução de Sinais , Análise de Sobrevida , Ensaios Antitumorais Modelo de Xenoenxerto
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