RESUMEN
BACKGROUND: Changes in protein glycosylation are widely observed in tumor cells. N-glycan branching through adding ß1,6-linked N-acetylglucosamine (ß1,6-GlcNAc) to an α1,6-linked mannose, which is catalyzed by the N-acetylglucosaminyltransferase V (MGAT5 or GnT-V), is one of the most frequently observed tumor-associated glycan structure formed. Increased levels of this branching structure play a pro-tumoral role in various ways, for example, through the stabilization of growth factor receptors, the destabilization of intercellular adhesion, or the acquisition of a migratory phenotype. CONCLUSION: In this review, we provide an updated and comprehensive summary of the physiological and pathophysiological roles of MGAT5 and ß1,6-GlcNAc branched N-glycans, including their regulatory mechanisms. Specific emphasis is given to the role of MGAT5 and ß1,6-GlcNAc branched N-glycans in cellular mechanisms that contribute to the development and progression of solid tumors. We also provide insight into possible future clinical implications, such as the use of MGAT5 as a prognostic biomarker.
Asunto(s)
Acetilglucosamina , Neoplasias , Humanos , Glicosilación , Fenotipo , Polisacáridos , N-AcetilglucosaminiltransferasasRESUMEN
The drug, 5-fluorouracil (5FU) is a standard first-line treatment for colorectal cancer (CRC) patients. However, drug resistance acquisition remains an important challenge for effective clinical outcomes. Here, we established a long-term drug-resistant CRC model and explored the cellular events underlying 5FU resistance. We showed that 5FU-treated cells (HCT-116 5FUR) using a prolonged treatment protocol were significantly more resistant than parental cells. Likewise, cell viability and IC50 values were also observed to increase in HCT-116 5FUR cells when treated with increasing doses of oxaliplatin, indicating a cross-resistance mechanism to other cytotoxic agents. Moreover, HCT-116 5FUR cells exhibited metabolic and molecular changes, as evidenced by increased thymidylate synthase levels and upregulated mRNA levels of ABCB1. HCT-116 5FUR cells were able to overcome S phase arrest and evade apoptosis, as well as activate autophagy, as indicated by increased LC3B levels. Cells treated with low and high doses displayed epithelial-mesenchymal transition (EMT) features, as observed by decreased E-cadherin and claudin-3 levels, increased vimentin protein levels, and increased SLUG, ZEB2 and TWIST1 mRNA levels. Furthermore, HCT-116 5FUR cells displayed enhanced migration and invasion capabilities. Interestingly, we found that the 5FU drug-resistance gene signature is positively associated with the mesenchymal signature in CRC samples, and that ABCB1 and ZEB2 co-expressed at high levels could predict poor outcomes in CRC patients. Overall, the 5FU long-term drug-resistance model established here induced various cellular events, and highlighted the importance of further efforts to identify promising targets involved in more than one cellular event to successfully overcome drug-resistance.
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Neoplasias del Colon , Neoplasias Colorrectales , Apoptosis , Autofagia , Cadherinas/genética , Línea Celular Tumoral , Proliferación Celular , Claudina-3 , Neoplasias del Colon/tratamiento farmacológico , Neoplasias Colorrectales/tratamiento farmacológico , Neoplasias Colorrectales/genética , Neoplasias Colorrectales/metabolismo , Citotoxinas , Resistencia a Antineoplásicos , Transición Epitelial-Mesenquimal , Fluorouracilo/farmacología , Humanos , Oxaliplatino/farmacología , ARN Mensajero , Timidilato Sintasa , VimentinaRESUMEN
Mesenchymal stromal cells (MSCs) were first used as a source for cell therapy in 1995; however, despite their versatility and unambiguous demonstration of efficacy and safety in preclinical/phase I studies, the positive effect of MSCs in human phase III studies did not resemble the success obtained in mouse models of disease. This dissonance highlights the need to more thoroughly study the immunobiology of MSCs to make better use of these cells. Thus, we aimed to study the immunobiology of MSCs by using chip array analysis as a method for general screening to obtain a global picture in our model study and found IFNy and IL-17 signaling as the first two "top canonical pathways" involved in MSCs immunomodulation. The role of IFNy in triggering the immunosuppressive properties of MSCs is well recognized by many groups; however, the role of IL-17 in this process remains uncertain. Interestingly, in contrast to IFNy, which actively improved the MSCs-mediated immunosuppression, IL-17 did not improve directly the MSCs-mediated immunosuppression. Instead, IL-17 signaling induced the migration of MSCs and inflammatory cells, bringing these cell types together and increasing the likelihood of the lymphocytes sensing the immunosuppressive molecules produced by the MSCs. These effects also correlated with high levels of cytokine/chemokine production and metalloprotease activation by MSCs. Importantly, this treatment maintained the MSCs safety profile by not inducing the expression of molecules related to antigen presentation. In this way, our findings highlight the possibility of using IL-17, in combination with IFNy, to prime MSCs for cell therapy to improve their biological properties and thus their therapeutic efficacy. Finally, the use of preactivated MSCs may also minimize variations among MSCs to produce more uniform therapeutic products. In the not-so-distant future, we envisage a portfolio of MSCs activated by different cocktails specifically designed to target and treat specific diseases. Graphical abstract.
Asunto(s)
Movimiento Celular , Terapia de Inmunosupresión , Interferón gamma/metabolismo , Interleucina-17/metabolismo , Células Madre Mesenquimatosas/metabolismo , Movimiento Celular/genética , Citocinas/metabolismo , Regulación de la Expresión Génica , Humanos , Mediadores de Inflamación/metabolismo , Prueba de Cultivo Mixto de Linfocitos , Células Madre Mesenquimatosas/inmunología , Fenotipo , Transducción de SeñalRESUMEN
Citral is a natural compound that has shown cytotoxic and antiproliferative effects on breast and hematopoietic cancer cells; however, there are few studies on melanoma cells. Oxidative stress is known to be involved in all stages of melanoma development and is able to modulate intracellular pathways related to cellular proliferation and death. In this study, we hypothesize that citral exerts its cytotoxic effect on melanoma cells by the modulation of cellular oxidative status and/or intracellular signaling. To test this hypothesis, we investigated the antiproliferative and cytotoxic effects of citral on B16F10 murine melanoma cells evaluating its effects on cellular oxidative stress, DNA damage, cell death, and important signaling pathways, as these pathways, namely, extracellular signal-regulated kinases 1/2 (ERK1/2), AKT, and phosphatidylinositol-3 kinase, are involved in cell proliferation and differentiation. The p53 and nuclear factor kappa B were also investigated due to their ability to respond to intracellular stress. We observed that citral exerted antiproliferative and cytotoxic effects in B16F10; induced oxidative stress, DNA lesions, and p53 nuclear translocation; and reduced nitric oxide levels and nuclear factor kappa B, ERK1/2, and AKT. To investigate citral specificity, we used non-neoplastic human and murine cells, HaCaT (human skin keratinocytes) and NIH-3T3 cells (murine fibroblasts), and observed that although citral effects were not specific for cancer cells, non-neoplastic cells were more resistant to citral than B16F10. These findings highlight the potential clinical utility of citral in melanoma, with a mechanism of action involving the oxidative stress generation, nitric oxide depletion, and interference in signaling pathways related to cell proliferation.
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Proliferación Celular/efectos de los fármacos , Melanoma Experimental/tratamiento farmacológico , Melanoma/tratamiento farmacológico , Monoterpenos/administración & dosificación , Estrés Oxidativo/efectos de los fármacos , Monoterpenos Acíclicos , Animales , Apoptosis/efectos de los fármacos , Ciclo Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Daño del ADN/efectos de los fármacos , Humanos , Melanoma/metabolismo , Melanoma/patología , Melanoma Experimental/genética , Melanoma Experimental/patología , Ratones , FN-kappa B/genética , Células 3T3 NIH , Óxido Nítrico/metabolismo , Proteína p53 Supresora de Tumor/genéticaRESUMEN
Radiotherapy is widely used for advanced rectal tumors. However, tumor recurrence after this treatment tends to be more aggressive and is associated with a poor prognosis. Uncovering the molecular mechanism that controls this recurrence is essential for developing new therapeutic applications. In the present study, we demonstrated that radiation increases the EphA4 activation level of the survivor progeny of colorectal cancer cells submitted to this treatment and that such activation promoted the internalization of a complex E-cadherin-EphA4, inducing cell-cell adhesion disruption. Moreover, EphA4 knockdown in the progeny of irradiated cells reduced the migratory and invasive potentials and metalloprotease activity induced by irradiation. Finally, we demonstrated that the cell migration and invasion potential were regulated by AKT and ERK1/2 signaling, with the ERK1/2 activity being dependent on EphA4. In summary, our study demonstrates that these signaling pathways could be responsible for the therapeutic failure, thereby promoting local invasion and metastasis in rectal cancer after radiotherapy. We also postulate that EphA4 is a potential therapeutic target for colorectal cancer treatment.
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Neoplasias Colorrectales/radioterapia , Receptor EphA4/fisiología , Transducción de Señal/fisiología , Antígenos CD , Cadherinas/análisis , Neoplasias Colorrectales/patología , Doxazosina/farmacología , Quinasas MAP Reguladas por Señal Extracelular/fisiología , Células HT29 , Humanos , Invasividad Neoplásica , Proteínas Proto-Oncogénicas c-akt/fisiologíaRESUMEN
Changes in glycosylation, which is one of the most common protein post-translational modifications, are considered to be a hallmark of cancer. N-glycans can modulate cell migration, cell-cell adhesion, cell signaling, growth and metastasis. The colorectal cancer (CRC) is a leading cause of cancer-related mortality and the correlation between CRC progression and changes in the pattern of expression of N-glycans is being considered in the search for new biomarkers. Here, we review the role of N-glycans in CRC cell biology. The perspectives on emerging N-glycan-related anticancer therapies, along with new insights and challenges, are also discussed.
Asunto(s)
Biomarcadores de Tumor/metabolismo , Neoplasias Colorrectales/metabolismo , N-Acetilglucosaminiltransferasas/metabolismo , Polisacáridos/metabolismo , Animales , Antineoplásicos/uso terapéutico , Biomarcadores de Tumor/antagonistas & inhibidores , Neoplasias Colorrectales/tratamiento farmacológico , Neoplasias Colorrectales/patología , Progresión de la Enfermedad , Glicosilación/efectos de los fármacos , Humanos , Modelos Biológicos , N-Acetilglucosaminiltransferasas/antagonistas & inhibidores , Polisacáridos/antagonistas & inhibidores , Polisacáridos/químicaRESUMEN
The participation of oxidative stress in the mechanism of metformin action in breast cancer remains unclear. We investigated the effects of clinical (6 and 30 µM) and experimental concentrations of metformin (1000 and 5000 µM) in MCF-7 and in MDA-MB-231 cells, verifying cytotoxicity, oxidative stress, DNA damage, and intracellular pathways related to cell growth and survival after 24 h of drug exposure. Clinical concentrations of metformin decreased metabolic activity of MCF-7 cells in the MTT assay, which showed increased oxidative stress and DNA damage, although cell death and impairment in the proliferative capacity were observed only at higher concentrations. The reduction in metabolic activity and proliferation in MDA-MB-231 cells was present only at experimental concentrations after 24 h of drug exposition. Oxidative stress and DNA damage were induced in this cell line at experimental concentrations. The drug decreased cytoplasmic extracellular signal-regulated kinases 1 and 2 (ERK1/2) and AKT and increased nuclear p53 and cytoplasmic transforming growth factor ß1 (TGF-ß1) in both cell lines. These findings suggest that metformin reduces cell survival by increasing reactive oxygen species, which induce DNA damage and apoptosis. A relationship between the increase in TGF-ß1 and p53 levels and the decrease in ERK1/2 and AKT was also observed. These findings suggest the mechanism of action of metformin in both breast cancer cell lineages, whereas cell line specific undergoes redox changes in the cells in which proliferation and survival signaling are modified. Taken together, these results highlight the potential clinical utility of metformin as an adjuvant during the treatment of luminal and triple-negative breast cancer.
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Metformina/administración & dosificación , Estrés Oxidativo/efectos de los fármacos , Factor de Crecimiento Transformador beta1/biosíntesis , Neoplasias de la Mama Triple Negativas/tratamiento farmacológico , Apoptosis/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Daño del ADN/efectos de los fármacos , Femenino , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Humanos , Células MCF-7 , Especies Reactivas de Oxígeno/metabolismo , Transducción de Señal , Factor de Crecimiento Transformador beta1/genética , Neoplasias de la Mama Triple Negativas/genética , Neoplasias de la Mama Triple Negativas/patología , Proteína p53 Supresora de Tumor/biosíntesis , Proteína p53 Supresora de Tumor/genéticaRESUMEN
Burkitt lymphoma is a highly aggressive non-Hodgkin lymphoma that is characterized by MYC deregulation. Recently, the PI3K pathway has emerged as a cooperative prosurvival mechanism in Burkitt lymphoma. Despite the highly successful results of treatment that use high-dose chemotherapy regimens in pediatric Burkitt lymphoma patients, the survival rate of pediatric patients with progressive or recurrent disease is low. PI3Ks are also known to regulate cell migration, and abnormal cell migration may contribute to cancer progression and dissemination in Burkitt lymphoma. Little is known about Burkitt lymphoma cell migration, but the cooperation between MYC and PI3K in Burkitt lymphoma pathogenesis suggests that a drug combination could be used to target the different steps involved in Burkitt lymphoma cell dissemination and disease progression. The aim of this study was to investigate the effects of the histone deacetylase inhibitor suberoylanilide hydroxamic acid combined with the PI3K inhibitor LY294002 on Burkitt lymphoma cell growth and migration. The combination enhanced the cell growth inhibition and cell-cycle arrest induced by the PI3K inhibitor or histone deacetylase inhibitor individually. Moreover, histone deacetylase inhibitor/PI3K inhibitor cotreatment suppressed Burkitt lymphoma cell migration and decreased cell polarization, Akt and ERK1/2 phosphorylation, and leads to RhoB induction. In summary, the histone deacetylase inhibitor/PI3Ki combination inhibits cell proliferation and migration via alterations in PI3K signaling and histone deacetylase activity, which is involved in the acetylation of α-tubulin and the regulation of RhoB expression.
Asunto(s)
Linfoma de Burkitt/enzimología , Movimiento Celular/efectos de los fármacos , Cromonas/farmacología , Inhibidores de Histona Desacetilasas/farmacología , Sistema de Señalización de MAP Quinasas/efectos de los fármacos , Morfolinas/farmacología , Inhibidores de las Quinasa Fosfoinosítidos-3 , Linfoma de Burkitt/patología , Línea Celular Tumoral , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Histona Desacetilasas/metabolismo , Humanos , Proteína Quinasa 1 Activada por Mitógenos/biosíntesis , Proteína Quinasa 3 Activada por Mitógenos/biosíntesis , Proteínas Proto-Oncogénicas c-akt/biosíntesis , Proteínas Proto-Oncogénicas c-myc/biosíntesis , Proteína de Unión al GTP rhoB/biosíntesisRESUMEN
Radiotherapy remains a major approach to adjuvant therapy for patients with advanced colorectal cancer, however, the fractionation schedules frequently allow for the repopulation of surviving tumors cells, neoplastic progression, and subsequent metastasis. The aim of the present study was to analyze the transgenerational effects induced by radiation and evaluate whether it could increase the malignant features on the progeny derived from irradiated parental colorectal cancer cells, Caco-2, HT-29, and HCT-116. The progeny of these cells displayed a differential radioresistance as seen by clonogenic and caspase activation assay and had a direct correlation with survivin expression as observed by immunoblotting. Immunofluorescence showed that the most radioresistant progenies had an aberrant morphology, disturbance of the cell-cell adhesion contacts, disorganization of the actin cytoskeleton, and vimentin filaments. Only the progeny derived from intermediary radioresistant cells, HT-29, reduced the E-cadherin expression and overexpressed ß-catenin and vimentin with increased cell migration, invasion, and metalloprotease activation as seen by immunoblotting, wound healing, invasion, and metalloprotease activity assay. We also observed that this most aggressive progeny increased the Wnt/ß-catenin-dependent TCF/LEF activity and underwent an upregulation of mesenchymal markers and downregulation of E-cadherin, as determined by qRT-PCR. Our results showed that the intermediate radioresistant cells can generate more aggressive cellular progeny with the EMT-like phenotype. The Wnt/ß-catenin pathway may constitute an important target for new adjuvant treatment schedules with radiotherapy, with the goal of reducing the migratory and invasive potential of the remaining cells after treatment.
Asunto(s)
Movimiento Celular/efectos de la radiación , Transición Epitelial-Mesenquimal/efectos de la radiación , Vía de Señalización Wnt , Citoesqueleto de Actina/metabolismo , Antígenos CD , Apoptosis , Células CACO-2 , Cadherinas/metabolismo , Caspasas/metabolismo , Forma de la Célula , Neoplasias Colorrectales , Células HT29 , Humanos , Proteínas Inhibidoras de la Apoptosis/metabolismo , Invasividad Neoplásica , Tolerancia a Radiación , Survivin , Vimentina/metabolismo , beta Catenina/metabolismoRESUMEN
Studies have reported that Na,K-ATPase interacts with E-cadherin to stabilize (AJs) and regulate the expression of claudins, the main proteins present in the tight junction (TJ) in epithelial cells containing caveolae. However, the role of this ATPase in the regulation of the AJ and TJ proteins in colorectal cancer cells as well as the molecular events underlying this event in a caveolae-independent system remain undefined. In the present study, we used ouabain, a classic drug known to inhibit Na,K-ATPase, and Caco-2 cells, which are a well-established human colorectal cancer model that does not exhibit caveolae. We demonstrated that ouabain treatment resulted in a reduction of the ß1 Na,K-ATPase protein and cell redistribution of the AJ proteins E-cadherin and ß-catenin, as well as the α1 Na,K-ATPase subunit. Furthermore, ouabain increased claudin-3 protein levels, impaired the TJ barrier function and increased cell viability and proliferation during the early stages of treatment. Additionally, the observed ouabain-induced events were dependent on the activation of ERK1/2 signaling; but in contrast to previous studies, this signaling cascade was caveolae-independent. In conclusion, our findings strongly suggest that α1 and ß1 Na,K-ATPase downregulation and ERK1/2 activation induced by ouabain are interlinked events that play an important role during cell-cell adhesion loss, which is an important step during the tumor progression of colorectal carcinomas.
Asunto(s)
Caveolas/metabolismo , Neoplasias Colorrectales/metabolismo , Proteína Quinasa 1 Activada por Mitógenos/metabolismo , Proteína Quinasa 3 Activada por Mitógenos/metabolismo , Ouabaína/farmacología , ATPasa Intercambiadora de Sodio-Potasio/metabolismo , Uniones Adherentes/efectos de los fármacos , Uniones Adherentes/metabolismo , Cadherinas/metabolismo , Línea Celular Tumoral , Proliferación Celular , Claudina-3 , Neoplasias Colorrectales/genética , Humanos , Transducción de Señal , Uniones Estrechas/efectos de los fármacos , Uniones Estrechas/metabolismoRESUMEN
Changes in glycosylation are considered a hallmark of cancer, and one of the key targets of glycosylation modifications is E-cadherin. We and others have previously demonstrated that E-cadherin has a role in the regulation of bisecting GlcNAc N-glycans expression, remaining to be determined the E-cadherin-dependent signaling pathway involved in this N-glycans expression regulation. In this study, we analysed the impact of E-cadherin expression in the activation profile of receptor tyrosine kinases such as insulin receptor (IR) and IGF-I receptor (IGF-IR). We demonstrated that exogenous E-cadherin expression inhibits IR, IGF-IR and ERK 1/2 phosphorylation. Stimulation with insulin and IGF-I in MDA-MD-435 cancer cells overexpressing E-cadherin induces a decrease of bisecting GlcNAc N-glycans that was accompanied with alterations on E-cadherin cellular localization. Concomitantly, IR/IGF-IR signaling activation induced a mesenchymal-like phenotype of cancer cells together with an increased tumor cell invasion capability. Altogether, these results demonstrate an interplay between E-cadherin and IR/IGF-IR signaling as major networking players in the regulation of bisecting N-glycans expression, with important effects in the modulation of epithelial characteristics and tumor cell invasion. Here we provide new insights into the role that Insulin/IGF-I signaling play during cancer progression through glycosylation modifications.
Asunto(s)
Acetilglucosamina/metabolismo , Cadherinas/metabolismo , Factor I del Crecimiento Similar a la Insulina/metabolismo , Insulina/metabolismo , Invasividad Neoplásica , Transducción de Señal , Western Blotting , Línea Celular Tumoral , Citoplasma/metabolismo , Técnica del Anticuerpo Fluorescente , Glicosilación , Humanos , Fosforilación , Polisacáridos/metabolismo , Reacción en Cadena en Tiempo Real de la Polimerasa , beta Catenina/metabolismoRESUMEN
In many gut chronic inflammatory conditions, intestinal epithelium (IE) is deprived of the protection of the mucus secreted by IE-specialized cells. In these events, bleeding and subsequent lysis of erythrocytes are common. This may lead to the release of high amounts of heme in the intestinal lumen, which interacts with IE. Previous works from our group have shown that heme itself is a proinflammatory molecule, activating a number of phlogistic signaling events in a nicotinamide adenine dinucleotide phosphate oxidase (NADPHox)-dependent manner. In this study, we aim to evaluate the effects of heme upon a well-established nontransformed small intestine epithelial cell lineage (IEC 6). Our results show that free heme evokes intracellular reactive oxygen species (ROS) production by IEC 6 cells, which is inhibited both by pharmacological inhibition with diphenyleneiodonium (10 µM), a NADPHox inhibitor, and small interfering RNA-mediated suppression of NOX1, a constitutive NADPHox isoform present in intestinal epithelial cells. Focal adhesion kinase phosphorylation and actin cytoskeleton polymerization are also induced by heme in a NADPHox-dependent manner. Heme increases monolayer permeability and redistributes key modulators of cell-cell adhesion as zona occludens-1 and E-cadherin proteins via NADPHox signaling. Heme promotes IEC 6 cell migration and proliferation, phenomena also regulated by NADPHox-derived ROS. Heme, in NADPHox-activating concentrations, is able to induce mRNA expression of IL-6, a cytokine implicated in inflammatory and tumorigenic responses. These data indicate a prominent role for heme-derived signaling in the pathophysiology of intestinal mucosa dysfunction and address an important role of NADPHox activity on the pathogenesis of intestinal inflammatory conditions.
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Células Epiteliales/efectos de los fármacos , Hemo/farmacología , Mucosa Intestinal/efectos de los fármacos , NADPH Oxidasas/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Transducción de Señal/efectos de los fármacos , Citoesqueleto de Actina/metabolismo , Animales , Cadherinas/fisiología , Línea Celular , Movimiento Celular/efectos de los fármacos , Movimiento Celular/fisiología , Proliferación Celular/efectos de los fármacos , Duodeno/efectos de los fármacos , Duodeno/enzimología , Inhibidores Enzimáticos/farmacología , Células Epiteliales/enzimología , Proteína-Tirosina Quinasas de Adhesión Focal/metabolismo , Silenciador del Gen , Interleucina-6/biosíntesis , Mucosa Intestinal/enzimología , NADH NADPH Oxidorreductasas/antagonistas & inhibidores , NADH NADPH Oxidorreductasas/genética , NADPH Oxidasa 1 , NADPH Oxidasas/antagonistas & inhibidores , Compuestos Onio/farmacología , Permeabilidad/efectos de los fármacos , Fosforilación , Ratas , Transducción de Señal/genética , Proteína de la Zonula Occludens-1/fisiologíaRESUMEN
During malignant transformation, changes in the expression profile of glycans may be involved in a variety of events, including the loss of cell-cell and cell-matrix adhesion, migration, invasion, and evasion of apoptosis. Therefore, modulation of glycan expression with drugs has promising therapeutic potential for various cancer types. In this study, we investigated the in vitro anticancer activity of the N-glycan biosynthesis inhibitors (swainsonine and tunicamycin) in cells derived from colorectal cancer (CRC). We also examined whether these inhibitors are able to induce radiosensitization and toxicity when used in combination with cisplatin or irinotecan, two current anticancer drugs. Our results show that treatment with tunicamycin inhibits cellular mechanisms related to the malignant phenotype, such as anchorage-dependent and anchorage-independent colony formation, migration and invasion, in undifferentiated HCT-116 colon cancer cells, whereas swainsonine only inhibits cell migration. We also observed that tunicamycin, but not swainsonine, caused radiosensitivity in HCT-116 cells. Moreover, the combination of swainsonine with cisplatin or irinotecan enhanced their toxicity in HCT-116 cells, while the combination of tunicamycin with these drugs had no effect. Given these results, we suggest that the modulation of N-glycan biosynthesis appears to be a potential therapeutic tool for CRC treatment because inhibition of this process induced anticancer activity in vitro. Additionally, the inhibition of the N-glycan biosynthesis in combination with chemotherapic drugs is a promising therapeutic strategy for enhancing radiation therapy.
Asunto(s)
Neoplasias Colorrectales/metabolismo , Polisacáridos/metabolismo , Tunicamicina/farmacología , Western Blotting , Células CACO-2 , Camptotecina/análogos & derivados , Camptotecina/farmacología , Movimiento Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Cisplatino/farmacología , Citometría de Flujo , Células HCT116 , Células HT29 , Humanos , Irinotecán , Swainsonina/farmacologíaRESUMEN
PURPOSE: Aberrant protein glycosylation and disassembly of E-cadherin-mediated cell-cell adhesion are characteristics of epithelial cancer. However, the relationship between these two events in colorectal cancer remains to be defined. In this study, we analyzed whether N-glycan expression is crucial for the loss of E-cadherin-mediated cell-cell adhesion in human colorectal cancer cells. METHODS: Differentiated Caco-2 and undifferentiated HCT-116 colon cancer cells were used as models of stable and unstable adherens junctions (AJs), respectively. Complex-type N-glycans were detected using the lectins E-PHA (Phaseolus vulgaris E.) and L-PHA (Phaseolus vulgaris L.). To study E-cadherin-mediated AJ assembly, we examined the effects of swainsonine, an inhibitor of α-mannosidase II, and tunicamycin, a drug that inhibits the biosynthesis of N-glycans, via western blot, immunofluorescence, differential extraction in Triton X-100, and electron microscopy. Cell proliferation and apoptosis were examined by crystal violet staining and flow cytometry, respectively. RESULTS: We observed positive labeling for E-PHA and L-PHA lectins in both cell lines; however, HCT-116 cells had increased E-cadherin-linked complex-type N-glycans. Interestingly, tunicamycin, but not swainsonine, was able to induce functional E-cadherin-mediated cell-cell adhesion in undifferentiated HCT-116 cells, as shown by the increased association of E-cadherin with the actin cytoskeleton. Moreover, in HCT-116 cells, tunicamycin also induced the formation of tight cell-cell contacts, and it inhibited cell proliferation without triggering apoptosis. CONCLUSIONS: Collectively, our results demonstrate for the first time that altered N-glycan expression plays an important role in the loss of AJ stability in undifferentiated colorectal cancer cells and that this loss may be associated with the progression of colorectal cancer.