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UNLABELLED: Emerging evidence suggests that epithelial-mesenchymal transitions (EMTs) play important roles in tumor metastasis and recurrence. Understanding molecular mechanisms that regulate the EMT process is crucial for improving treatment of hepatocellular carcinoma (HCC). MicroRNAs (miRNAs) play important roles in HCC; however, the mechanisms by which miRNAs target the EMT and their therapeutic potential remains largely unknown. To better explore the roles of miRNAs in the EMT process, we established an EMT model in HCC cells by transforming growth factor beta 1 treatment and found that several tumor-related miRNAs were significantly decreased. Among these miRNAs, miR-125b expression was most strongly suppressed. We also found down-regulation of miR-125b in most HCC cells and clinical specimens, which correlated with cellular differentiation in HCC patients. We then demonstrated that miR-125b overexpression attenuated EMT phenotype in HCC cancer cells, whereas knockdown of miR-125b promoted the EMT phenotype in vitro and in vivo. Moreover, we found that miR-125b attenuated EMT-associated traits, including chemoresistance, migration, and stemness in HCC cells, and negatively correlated with EMT and cancer stem cell (CSC) marker expressions in HCC specimens. miR-125b overexpression could inhibit CSC generation and decrease tumor incidence in the mouse xenograft model. Mechanistically, our data revealed that miR-125b suppressed EMT and EMT-associated traits of HCC cells by targeting small mothers against decapentaplegic (SMAD)2 and 4. Most important, the therapeutic delivery of synthetic miR-125b mimics decreased the target molecule of CSC and inhibited metastasis in the mice model. These findings suggest a potential therapeutic treatment of miR-125b for liver cancer. CONCLUSION: miR-125b exerts inhibitory effects on EMT and EMT-associated traits in HCC by SMAD2 and 4. Ectopic expression of miR-125b provides a promising strategy to treat HCC.
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Transición Epitelial-Mesenquimal/genética , Regulación Neoplásica de la Expresión Génica , MicroARNs/genética , Células Madre Neoplásicas/patología , Proteína Smad2/metabolismo , Proteína Smad4/metabolismo , Animales , Carcinoma Hepatocelular/metabolismo , Carcinoma Hepatocelular/patología , Modelos Animales de Enfermedad , Regulación hacia Abajo , Humanos , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/patología , Masculino , Ratones , Ratones Desnudos , MicroARNs/metabolismo , Células Madre Neoplásicas/metabolismo , Distribución Aleatoria , Sensibilidad y Especificidad , Transfección , Células Tumorales CultivadasRESUMEN
UNLABELLED: Cancer-associated mesenchymal stem cells (MSCs) play a pivotal role in modulating tumor progression. However, the interactions between liver cancer-associated MSCs (LC-MSCs) and hepatocellular carcinoma (HCC) remain unreported. Here, we identified the presence of MSCs in HCC tissues. We also showed that LC-MSCs significantly enhanced tumor growth in vivo and promoted tumor sphere formation in vitro. LC-MSCs also promoted HCC metastasis in an orthotopic liver transplantation model. Complementary DNA (cDNA) microarray analysis showed that S100A4 expression was significantly higher in LC-MSCs compared with liver normal MSCs (LN-MSCs) from adjacent cancer-free tissues. Importantly, the inhibition of S100A4 led to a reduction of proliferation and invasion of HCC cells, while exogenous S100A4 expression in HCC cells resulted in heavier tumors and more metastasis sites. Our results indicate that S100A4 secreted from LC-MSCs can promote HCC cell proliferation and invasion. We then found the expression of oncogenic microRNA (miR)-155 in HCC cells was significantly up-regulated by coculture with LC-MSCs and by S100A4 ectopic overexpression. The invasion-promoting effects of S100A4 were significantly attenuated by a miR-155 inhibitor. These results suggest that S100A4 exerts its effects through the regulation of miR-155 expression in HCC cells. We demonstrate that S100A4 secreted from LC-MSCs promotes the expression of miR-155, which mediates the down-regulation of suppressor of cytokine signaling 1, leading to the subsequent activation of STAT3 signaling. This promotes the expression of matrix metalloproteinases 9, which results in increased tumor invasiveness. CONCLUSION: S100A4 secreted from LC-MSCs is involved in the modulation of HCC progression, and may be a potential therapeutic target. (HEPATOLOGY 2013).
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Carcinoma Hepatocelular/metabolismo , Neoplasias Hepáticas/metabolismo , Células Madre Mesenquimatosas/metabolismo , MicroARNs/metabolismo , Proteínas S100/metabolismo , Animales , Carcinoma Hepatocelular/patología , Proliferación Celular , Progresión de la Enfermedad , Humanos , Neoplasias Hepáticas/patología , Masculino , Metaloproteinasa 9 de la Matriz/metabolismo , Células Madre Mesenquimatosas/patología , Ratones , Ratones Desnudos , Invasividad Neoplásica , Metástasis de la Neoplasia , Proteína de Unión al Calcio S100A4 , Factor de Transcripción STAT3/metabolismo , Proteína 1 Supresora de la Señalización de Citocinas , Proteínas Supresoras de la Señalización de Citocinas/metabolismoRESUMEN
The enzymatic activities of Furin, Transmembrane serine proteinase 2 (TMPRSS2), Cathepsin L (CTSL), and Angiotensin-converting enzyme 2 (ACE2) receptor binding are necessary for the entry of coronaviruses into host cells. Precise inhibition of these key proteases in ACE2+ lung cells during a viral infection cycle shall prevent viral Spike (S) protein activation and its fusion with a host cell membrane, consequently averting virus entry to the cells. In this study, dual-drug-combined (TMPRSS2 inhibitor Camostat and CTSL inhibitor E-64d) nanocarriers (NCs) are constructed conjugated with an anti-human ACE2 (hACE2) antibody and employ Red Blood Cell (RBC)-hitchhiking, termed "Nanoengineered RBCs," for targeting lung cells. The significant therapeutic efficacy of the dual-drug-loaded nanoengineered RBCs in pseudovirus-infected K18-hACE2 transgenic mice is reported. Notably, the modular nanoengineered RBCs (anti-receptor antibody+NCs+RBCs) precisely target key proteases of host cells in the lungs to block the entry of Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), regardless of virus variations. These findings are anticipated to benefit the development of a series of novel and safe host-cell-protecting antiviral therapies.
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COVID-19 , Catepsina L , SARS-CoV-2 , Inhibidores de Serina Proteinasa , Animales , Ratones , Enzima Convertidora de Angiotensina 2/metabolismo , Catepsina L/antagonistas & inhibidores , Catepsina L/metabolismo , COVID-19/prevención & control , COVID-19/virología , Eritrocitos , Pulmón/metabolismo , Péptido Hidrolasas/metabolismo , SARS-CoV-2/metabolismo , SARS-CoV-2/patogenicidad , Serina Endopeptidasas/metabolismo , Inhibidores de Serina Proteinasa/farmacología , Inhibidores de Serina Proteinasa/uso terapéuticoRESUMEN
Mesenchymal stem cells (MSCs) play a critical role in promoting cancer progression. However, it is not clear whether MSCs are located in breast cancer tissues and correlated with tumor proliferation. The aim of this study was to investigate the presence of MSCs in breast cancer tissues and evaluate their interactions with cancer cells. We successfully isolated and identified MSCs from primary breast cancer tissues. Breast cancer-associated MSCs (BC-MSCs) showed homogenous immunophenotype, and possessed tri-lineage differentiation potential (osteoblast, adipocyte, and chondrocyte). When co-transplanted with cancer cells in a xenograft model in vivo, BC-MSCs significantly increased the volume and weight of tumors. We observed that BC-MSCs stimulated mammosphere formation in the transwell co-culture system in vitro. This effect was significantly suppressed by the EGF receptor inhibitor. We verified that BC-MSCs could secrete EGF and activate cancer cell's EGF receptors. Furthermore, our data showed that EGF derived from BC-MSCs could promote mammosphere formation via the PI3K/Akt signaling pathway. Our results confirmed the presence of MSC in primary breast cancer tissues, and they could provide a favorable microenvironment for tumor cell growth in vivo, partially enhance mammosphere formation via the EGF/EGFR/Akt pathway.
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Neoplasias de la Mama/patología , Carcinoma Ductal de Mama/patología , Factor de Crecimiento Epidérmico/fisiología , Células Madre Mesenquimatosas/metabolismo , Células Madre Neoplásicas/metabolismo , Esferoides Celulares/metabolismo , Animales , Antígenos de Diferenciación/metabolismo , Diferenciación Celular , Proliferación Celular , Forma de la Célula , Técnicas de Cocultivo , Factor de Crecimiento Epidérmico/metabolismo , Receptores ErbB/metabolismo , Femenino , Humanos , Ratones , Ratones Desnudos , Trasplante de Neoplasias , Fosfatidilinositol 3-Quinasas/metabolismo , Proteínas Proto-Oncogénicas c-akt/metabolismo , Transducción de Señal , Carga Tumoral , Células Tumorales CultivadasRESUMEN
UNLABELLED: The high incidence rate of hepatocellular carcinoma (HCC) is mainly the result of frequent metastasis and tumor recurrence. Unfortunately, the underlying molecular mechanisms driving HCC metastasis are still not fully understood. It has been demonstrated that tumor stroma cells contribute to primary tumor growth and metastasis. Within the HCC environment, activated hepatic stellate cells (HSCs) can release a number of molecules and enhance cancer cell proliferation and invasiveness in a paracrine manner. Here, for the first time, we demonstrate that epimorphin (EPM; also called syntaxin-2), an extracellular protein, is strongly elevated in activated HSCs within tumor stroma. We show that knockdown of EPM expression in HSCs substantially abolishes their effects on cancer cell invasion and metastasis. Ectopic expression of EPM in HCC cancer cells enhances their invasiveness; we demonstrate that the cells expressing EPM have markedly increased metastasis potential. Furthermore, EPM-mediated invasion and metastasis of cancer cells is found to require up-regulation of matrix metalloproteinase-9 (MMP-9) through the activation of focal adhesion kinase (FAK)/extracellular signal-regulated kinase (ERK) axis. CONCLUSION: Our results show that EPM, secreted by activated HSCs within HCC stroma, promotes invasion and metastasis of cancer cells by activating MMP-9 expression through the FAK-ERK pathway.
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Carcinoma Hepatocelular/metabolismo , Carcinoma Hepatocelular/secundario , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/patología , Sistema de Señalización de MAP Quinasas/fisiología , Sintaxina 1/metabolismo , División Celular/fisiología , Movimiento Celular/fisiología , Quinasas MAP Reguladas por Señal Extracelular/metabolismo , Quinasa 1 de Adhesión Focal/metabolismo , Células Hep G2 , Células Estrelladas Hepáticas/metabolismo , Células Estrelladas Hepáticas/patología , Humanos , Hígado/metabolismo , Hígado/patología , Metaloproteinasa 9 de la Matriz/metabolismo , Invasividad NeoplásicaRESUMEN
A novel chemical functionalization of guar gum (GG) by benzoic acid (BA) via nucleophilic substitution reaction in aqueous solution has been reported. BA moieties are chosen due to coordination chemistry of carboxylic acid moieties, hydrophobicity and intermolecular interaction of aromatic rings. The presence of conjugated BA on guar gum-benzoic acid (GG-BA) with grafting density of 5.5% is confirmed by 1H NMR. Amorphous GG-BA with irregular morphology has been studied by UV-Vis, FTIR, XRD, SEM, TEM, TGA, computational chemistry and contact angle measurement. GG-BA in a concentration range from 0 to 4000 µg mL-1 has good biocompatibility to mouse embryonic fibroblasts (MEF), human mammary epithelial cells (MCF-10A) after 48 and 72 h of treatment using WST-1 assay. GG-BA shows great potential for the development of biomaterials such as bioadhesives, hydrogels, and coacervates.
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Ácido Benzoico/química , Materiales Biocompatibles/química , Galactanos/química , Mananos/química , Gomas de Plantas/química , Animales , Ácido Benzoico/síntesis química , Materiales Biocompatibles/síntesis química , Técnicas de Química Sintética , Humanos , Ratones , Modelos Moleculares , Estructura Molecular , Análisis Espectral , TermodinámicaRESUMEN
Cancer stem cells (CSCs) are reported to play essential roles in chemoresistance and metastasis. Pathways regulating CSC self-renewal and proliferation, such as Hedgehog, Notch, Wnt/ß-catenin, TGF-ß, and Myc, may be potential therapeutic targets. Here, a functional screening from the focused library with 365 compounds is performed by a step-by-step strategy. Among these candidate molecules, phenyl-2-pyrimidinyl ketone 4-allyl-3-amino selenourea (CU27) is chosen for further identification because it proves to be the most effective compound over others on CSC inhibition. Through ingenuity pathway analysis, it is shown CU27 may inhibit CSC through a well-known stemness-related transcription factor c-Myc. Gene set enrichment analysis, dual-luciferase reporter assays, expression levels of typical c-Myc targets, molecular docking, surface plasmon resonance, immunoprecipitation, and chromatin immunoprecipitation are conducted. These results together suggest CU27 binds c-Myc bHLH/LZ domains, inhibits c-Myc-Max complex formation, and prevents its occupancy on target gene promoters. In mouse models, CU27 significantly sensitizes sorafenib-resistant tumor to sorafenib, reduces the primary tumor size, and inhibits CSC generation, showing a dramatic anti-metastasis potential. Taken together, CU27 exerts inhibitory effects on CSC and CSC-associated traits in hepatocellular carcinoma (HCC) via c-Myc transcription activity inhibition. CU27 may be a promising therapeutic to treat sorafenib-resistant HCC.
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Carcinoma Hepatocelular , Neoplasias Hepáticas , Compuestos de Selenio , Selenio , Animales , Carcinoma Hepatocelular/tratamiento farmacológico , Carcinoma Hepatocelular/genética , Línea Celular Tumoral , Detección Precoz del Cáncer , Neoplasias Hepáticas/tratamiento farmacológico , Neoplasias Hepáticas/genética , Ratones , Simulación del Acoplamiento Molecular , Células Madre Neoplásicas/metabolismo , Células Madre Neoplásicas/patología , Selenio/metabolismo , Selenio/farmacología , Compuestos de Selenio/metabolismo , Compuestos de Selenio/farmacología , Sorafenib/metabolismo , Sorafenib/farmacologíaRESUMEN
Hepatocellular carcinoma (HCC) is the most common liver cancer with high mortality. Here, we found that hnRNPU is overexpressed in HCC tissues and is correlated with the poor prognosis of HCC patients. Besides, hnRNPU is of high significance in regulating the proliferation, apoptosis, self-renewal, and tumorigenic potential of HCC cells. Mechanismly, c-Myc regulates hnRNPU expression at the transcriptional level, and meanwhile, hnRNPU stabilizes the mRNA of c-MYC. We found that the hnRNPU and c-Myc regulatory loop exerts a synergistic effect on the proliferation and self-renewal of HCC, and promotes the HCC progression. Taken together, hnRNPU functions as a novel transcriptional target of c-Myc and promotes HCC progression, which may become a promising target for the treatment of c-Myc-driven HCC.
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Apoptosis/fisiología , Carcinoma Hepatocelular/patología , Ribonucleoproteína Heterogénea-Nuclear Grupo U/fisiología , Neoplasias Hepáticas/patología , Proteínas Proto-Oncogénicas c-myc/metabolismo , Transcripción Genética , Animales , Línea Celular Tumoral , Humanos , Ratones Endogámicos NOD , Ratones SCID , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Protein kinase A (PKA) activation has recently been reported to inhibit epithelial-mesenchymal transition (EMT) and cancer stem cell (CSC) ability, which is considered to be responsible for chemoresistance and tumor recurrence in patients. While current studies mainly focus on gene manipulation of the EMT process, the direct delivery of PKA enzymes to cancer cells has never been investigated. Here, we utilize the commercial Lipofectamine CRISPRMAX reagent to directly deliver PKAs to breast cancer cells and evaluate its effects on EMT regulation. We optimized the delivery parameters with fluorescent-labeled bovine serum albumin, and successfully delivered fluorescent PKAs through CRISPRMAX into breast cancer cells. Then, we evaluated the biological effects by immunofluorescence, flow cytometry, mammosphere assay, and chemoresistance assay. Our data showed the expression of EMT-related markers, α-smooth muscle actin and N-cadherin, was downregulated after CRISPRMAX-PKA treatment. Although the CD44+/CD24- population did not change considerably, the size of mammospheres significantly decreased. In paclitaxel and doxorubicin chemoresistance assays, we noticed PKA delivery significantly inhibited paclitaxel resistance rather than doxorubicin resistance. Taken together, these results suggest our direct enzyme delivery can be a potential strategy for inhibiting EMT/CSC-associated traits, providing a safer approach and having more clinical translational efficacy than gene manipulation. This strategy will also facilitate the direct testing of other target enzymes/proteins on their biological functions.
RESUMEN
Liver cancer stem cells (L-CSCs) are considered to be an important therapeutic target for hepatocellular carcinoma (HCC). This study provides a new in vitro long-term culture model for a specific subpopulation of L-CSCs enriched by cell surface markers. We combined CD13, CD133 and EpCAM to selectively enrich L-CSCs, which we then cultured in modified chemically defined medium. The enriched L-CSCs exhibited enhanced proliferation, self-renewal and long-term clonal maintenance ability as compared with non-CSCs. Compared with wild-type hepatocellular carcinoma, the expression of stemness surface markers, oncogenes, drug resistance and tumorigenicity in enriched L-CSCs was significantly increased. In summary, the subpopulation of L-CSCs still maintains cancer stem cell-related phenotypes after 14 days of culture.
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Antígeno AC133/metabolismo , Biomarcadores de Tumor/metabolismo , Antígenos CD13/metabolismo , Carcinoma Hepatocelular/patología , Molécula de Adhesión Celular Epitelial/metabolismo , Neoplasias Hepáticas/patología , Células Madre Neoplásicas/patología , Carcinoma Hepatocelular/metabolismo , Humanos , Neoplasias Hepáticas/metabolismo , Células Madre Neoplásicas/metabolismo , Células Tumorales CultivadasRESUMEN
PURPOSE: A better understanding of the underlying molecular mechanisms in treatment failure of bevacizumab (BEV) for malignant glioma would contribute to overcome therapeutic resistance. METHODS: Here, we used a quantitative proteomic method to identify molecular signatures of glioblastoma cell after BEV treatment by two-dimensional liquid chromatography-tandem mass spectrometry analysis and 6-plex iTRAQ quantification. Next, the function of cold-inducible RNA-binding protein (CIRP), one of the most significantly affected proteins by drug treatment, was evaluated in drug resistance of glioma cells by invasion assays and animal xenograft assays. Target molecules bound by CIRP were determined using RNA-binding protein immunoprecipitation and microarray analysis. Then, these mRNAs were identified by quantitative real-time PCR. RESULTS: Eighty-seven proteins were identified with significant fold changes. The biological functional analysis indicated that most of the proteins were involved in the process of cellular signal transduction, cell adhesion, and protein transport. The expression of CIRP greatly decreased after BEV treatment, and ectopic expression of CIRP abolished cell migration in BEV-treated glioma cells. In addition, CIRP could bind mRNA of CXCL12 and inhibit BEV-induced increase of CXCL12 in glioma cells. CONCLUSION: These data suggested that CIRP may take part in BEV-induced migration of gliomas by binding of migration-relative RNAs.
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Hepatocellular carcinoma (HCC) has a high mortality rate due to the lack of effective treatments and drugs. Arsenic trioxide (ATO), which has been proved to successfully treat acute promyelocytic leukemia (APL), was recently reported to show therapeutic potential in solid tumors including HCC. However, its anticancer mechanisms in HCC still need further investigation. In this study, we demonstrated that ATO inhibits tumorigenesis and distant metastasis in mouse models, corresponding with a prolonged mice survival time. Also, ATO was found to significantly decrease the cancer stem cell (CSC)-associated traits. Minichromosome maintenance protein (MCM) 7 was further identified to be a potential target suppressed dramatically by ATO, of which protein expression is increased in patients and significantly correlated with tumor size, cellular differentiation, portal venous emboli, and poor patient survival. Moreover, MCM7 knockdown recapitulates the effects of ATO on CSCs and metastasis, while ectopic expression of MCM7 abolishes them. Mechanistically, our results suggested that ATO suppresses MCM7 transcription by targeting serum response factor (SRF)/MCM7 complex, which functions as an important transcriptional regulator modulating MCM7 expression. Taken together, our findings highlight the importance of ATO in the treatment of solid tumors. The identification of SRF/MCM7 complex as a target of ATO provides new insights into ATO's mechanism, which may benefit the appropriate use of this agent in the treatment of HCC.
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Antineoplásicos/farmacología , Trióxido de Arsénico/farmacología , Carcinoma Hepatocelular/tratamiento farmacológico , Neoplasias Hepáticas/tratamiento farmacológico , Componente 7 del Complejo de Mantenimiento de Minicromosoma/metabolismo , Células Madre Neoplásicas/metabolismo , Factor de Respuesta Sérica/metabolismo , Animales , Antineoplásicos/uso terapéutico , Trióxido de Arsénico/uso terapéutico , Carcinoma Hepatocelular/metabolismo , Carcinoma Hepatocelular/patología , Carcinoma Hepatocelular/secundario , Línea Celular Tumoral , Progresión de la Enfermedad , Regulación hacia Abajo/genética , Regulación Neoplásica de la Expresión Génica/genética , Ontología de Genes , Humanos , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/patología , Neoplasias Hepáticas Experimentales/tratamiento farmacológico , Neoplasias Hepáticas Experimentales/metabolismo , Neoplasias Hepáticas Experimentales/mortalidad , Neoplasias Pulmonares/tratamiento farmacológico , Neoplasias Pulmonares/mortalidad , Neoplasias Pulmonares/secundario , Masculino , Ratones , Ratones Endogámicos NOD , Ratones Desnudos , Ratones SCID , Componente 7 del Complejo de Mantenimiento de Minicromosoma/genética , Células Madre Neoplásicas/efectos de los fármacos , Pronóstico , Factor de Respuesta Sérica/antagonistas & inhibidores , Factor de Respuesta Sérica/genética , Trasplante HeterólogoRESUMEN
OBJECTIVE: To compare the differentiation ability difference of hematopoietic, mesenchymal and endothelial potential between CD41⺠cells derived from the mouse aorta-gonadmesonephros (AGM) region, yolk sac (YS) and embryonic circulating blood (CB). METHODS: CD41⺠cells were sorted from AGM, YS and CB. The CD45 and c-kit expression were studied in CD41⺠cells by flow cytometry. IL-3 and bone morphogenetic protein 4 (BMP-4) treatment together with semi solid culture were used to assess hematopoietic potential difference of CD41⺠cells. Immunofluorescence staining of α-SMA was used to assess mesenchymal potential difference. The endothelial cell induction system was used to assess endothelial potential difference. RESULTS: The proportions of CD45+ cells in CD41⺠population were 51.9% (AGM), 45.8% (YS) and 22.2% (CB), respectively, while those of c-kit⺠cells were 40.0% (AGM), 39.6% (YS) and 36.2% (CB), respectively. After stimulated by IL-3 factor, the number of total colonies increased in all three groups-derived CD41⺠cells compared to that of unstimulated group[(14.1±1.9) vs (1.2±0.2), (32.4±1.1) vs (18.4±2.2) and (41.8±0.9) vs (10.4±1.8)], (P<0.01). After stimulated by BMP-4 factor, compared to unstimulated group, CFU-Mix colony number in CD41⺠cells from AGM region and YS were significantly decreased[(0.5±0.6) vs (3.2±0.8), (1.3±0.7) vs (7.4±1.7)](P<0.01), but there was no difference in CB group[(2.5±0.5) vs (3.9±1.5)](P>0.01). The mesenchymal marker α-SMA was highly expressed in CD41⺠cells from AGM region and YS, but lowly expressed in CD41⺠cells from CB. CONCLUSION: There are some differences between CD41⺠cells in AGM region, YS and CB on hematopoietic cell surface marker expression, hematopoietic colony formation with IL-3 and BMP-4 stimulation.
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Aorta/citología , Diferenciación Celular , Gónadas/citología , Mesonefro/citología , Saco Vitelino/citología , Animales , Proteína Morfogenética Ósea 4/farmacología , Interleucina-3/farmacología , Ratones , Glicoproteína IIb de Membrana Plaquetaria/metabolismo , Proteínas Proto-Oncogénicas c-kit/metabolismoRESUMEN
SPINDLIN1, a new member of the SPIN/SSTY gene family, was first identified as a gene highly expressed in ovarian cancer cells. We have previously shown that it is involved in the process of spindle organization and chromosomal stability and plays a role in the development of cancer. Nevertheless, the mechanisms underlying its oncogenic role are still largely unknown. Here, we first showed that expression of SPINDLIN1 is upregulated in clinical tumors. Ectopic expression of SPINDLIN1 promoted cancer cell proliferation and activated WNT/T-cell factor (TCF)-4 signaling. The Ser84 and Ser99 amino acids within SPINDLIN1 were further identified as the key functional sites in WNT/TCF-4 signaling activation. Mutation of these two sites of SPINDLIN1 abolished its effects on promoting WNT/TCF-4 signaling and cancer cell proliferation. We further found that Aurora-A could interact with and phosphorylate SPINDLIN1 at its key functional sites, Ser84 and Ser99, suggesting that phosphorylation of SPINDLIN1 is involved in its oncogenic function. Collectively, these results suggest that SPINDLIN1, which may be a novel substrate of the Aurora-A kinase, promotes cancer cell growth through WNT/TCF-4 signaling activation.
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Factores de Transcripción Básicos con Cremalleras de Leucinas y Motivos Hélice-Asa-Hélice/metabolismo , Proteínas de Ciclo Celular/metabolismo , Proteínas Asociadas a Microtúbulos/metabolismo , Neoplasias/patología , Fosfoproteínas/metabolismo , Factores de Transcripción/metabolismo , Vía de Señalización Wnt , Aurora Quinasas , Línea Celular Tumoral , Proliferación Celular , Femenino , Humanos , Mutación/genética , Invasividad Neoplásica , Neoplasias/metabolismo , Neoplasias Ováricas/metabolismo , Neoplasias Ováricas/patología , Fosforilación , Fosfoserina/metabolismo , Unión Proteica , Proteínas Serina-Treonina Quinasas/metabolismo , Factor de Transcripción 4RESUMEN
A recombinant phage vaccine expressing EGFR on it's capsid was constructed and used to study the anti-tumor effect. The T7 phage display system was applied to display seven xenogenic (human, chicken) epidermal growth factor receptor extracellular domain fragments. The EGFR fragment was expressed as fused protein with 10B capsid on the surface of T7 phage. The T7-EGFR phage vaccines were injected into C57BL/6J mice, and then Lewis lung cancer cells were inoculated after 4 weeks immunization. The tumor tissue was excised and weighed after 10 days to evaluate the anti-tumor effect of each experimental group. The EGFR expression of the phage vaccine was verified by western-blot analysis. The A431 cells with high expressed EGFR was used to detect the anti-EGFR antibody by flow cytometry analysis. The results showed that the A431 cell can react with the serum obtained from the mice after three-week immunization. The experimental results confirmed that special EGFR antibody could be induced by the T7-EGFR phage vaccine. The T7-EGFR phage vaccine can elicit endogenous special EGFR antibody in mice and is capable of suppressing the tumor proliferation and retarding the growth of Lewis lung cancer. This research can be used to develop an anti-tumor vaccine for the target-therapy of EGFR(+) tumor.