RESUMEN
BACKGROUND: Oral squamous cell carcinoma has high recurrence and cisplatin resistance. As cancer stem cells, autophagy, and sphingolipids have been appointed as associated with chemotherapy resistance, we tested combined treatments targeting autophagy and/or sphingolipid metabolism with paclitaxel using cisplatin-resistant oral squamous cell carcinoma cells. METHODS: Cisplatin-resistant oral squamous cell carcinoma cells were maintained under exposition to FTY720 and chloroquine combined with paclitaxel and submitted to viability, clonogenicity, and spheres formation assays. The xenograft tumor model using cisplatin-resistant CAL27 cells was adopted to examine the drug combinations' potential antitumoral efficacy. Using an animal model, sphingolipids profiles from plasma and tissue samples were obtained by liquid chromatography coupled to mass spectrometry to identify potential lipids associated with drug response. RESULTS AND DISCUSSION: Our results showed higher autophagic flux in cisplatin-resistant Ooral squamous cell carcinoma (CAL27 and SCC9) cells than in parental cells. The combinations of an autophagy inhibitor (chloroquine) or an autophagy inducer/sphingosine kinase 1 antagonist (FTY720) with paclitaxel (PTX) had a synergistic antitumor effect. Treated CisR cells lost clonogenicity and tumor sphere abilities and reduced proteins associated with proliferation, survival, and cancer stem cells. FTY720 plus PTX had higher antitumor efficacy than PTX against CAL27 CisR xenograft tumor formation. Additionally, increases in glucosylceramide, dehydroglucosylceramide, and sphingomyelin were presented in responsive tumors. CONCLUSION: FTY720 sensitizes cisplatin-resistant oral squamous cell carcinoma cells for paclitaxel.
Asunto(s)
Carcinoma de Células Escamosas , Neoplasias de Cabeza y Cuello , Neoplasias de la Boca , Animales , Humanos , Cisplatino/farmacología , Paclitaxel/farmacología , Clorhidrato de Fingolimod/farmacología , Clorhidrato de Fingolimod/uso terapéutico , Carcinoma de Células Escamosas/tratamiento farmacológico , Carcinoma de Células Escamosas de Cabeza y Cuello , Apoptosis , Neoplasias de la Boca/tratamiento farmacológico , Esfingolípidos/farmacología , Cloroquina/farmacología , Línea Celular Tumoral , Resistencia a AntineoplásicosRESUMEN
The SET gene has four transcripts reported in NCBI, coding two isoforms of SET proteins. The most known function of SET protein is inhibiting protein phosphatase 2A, a tumor suppressor, which has been associated with different biological processes. In this review, our focus was on exploring the other SET functions related to epigenetic mechanisms, which impact cellular migration, cell cycle and apoptosis.
Asunto(s)
Apoptosis , Epigénesis Genética , Humanos , Procesamiento Proteico-Postraduccional , Isoformas de ProteínasRESUMEN
Bisphenol A (BPA) is an endocrine-disrupting chemical with a potential role in endocrine cancers. However, the effects of BPA on the salivary glands have been barely explored. We investigated the impact of in vivo sub-chronic exposure to BPA and its in vitro effects on human salivary gland mucoepidermoid carcinoma cell lines. Male and female mice were exposed to BPA (30 mg/kg/day). Sublingual and submandibular salivary glands from an estrogen-deficiency model were also analyzed. BPA concentration in salivary glands was evaluated by gas chromatography coupled to ion trap mass spectrometry. Immunohistochemical analysis using anti-p63 and anti-α-SMA antibodies was performed on mouse salivary gland tissues. Gene expression of estrogen receptors alpha and beta, P63 and α-SMA was quantified in mouse salivary gland and/or mucoepidermoid (UM-HMC-1 and UM-HMC-3A) cell lines. Cell viability, p63 and Ki-67 immunostaining were evaluated in vitro. BPA disrupted the tissue architecture of the submandibular and sublingual glands, particularly in female mice, and increased the expression of estrogen receptors and p63, effects that were accompanied by significant BPA accumulation in these tissues. Conversely, ovariectomy slightly impacted BPA-induced morphological changes. In vitro, BPA did not affect the proliferation of neoplastic cells, but augmented the expression of p63 and estrogen receptors. The present data highlight a potential harmful effect of BPA on salivary gland tissues, particularly in female mice, and salivary gland tumor cells. Our findings suggest that estrogen-dependent pathways may orchestrate the effects of BPA in salivary glands.
Asunto(s)
Neoplasias de las Glándulas Salivales , Glándulas Salivales , Humanos , Animales , Ratones , Masculino , Femenino , Estrógenos , Receptores de Estrógenos/genética , Receptores de Estrógenos/metabolismo , Línea Celular Tumoral , Neoplasias de las Glándulas Salivales/inducido químicamenteRESUMEN
Pericytes are perivascular cells related to vessel structure and angiogenesis that can interact with neoplastic cells, interfering with cancer progression and outcomes. This study focused on the characterization of pericytes in oral squamous cell carcinoma (OSCC) using clinical samples and a transgenic mouse model of oral carcinogenesis. Nestin-/NG2+ (type-1) and nestin+/NG2+ (type-2) pericytes were analyzed by direct fluorescence after induction of oral carcinogenesis (4-nitroquinoline-1-oxide). Gene expression of neuron glial antigen-2 (NG2), platelet-derived growth factor receptor beta (PDGFR-ß), and cluster of differentiation 31 (CD31) was examined in human OSCC tissues. The protein expression of von Willebrand factor and NG2 was assessed in oral leukoplakia (i.e., oral potentially malignant disorders) and OSCC samples. Additionally, clinicopathological aspects and survival data were correlated and validated by bioinformatics using The Cancer Genome Atlas (TCGA). Induction of carcinogenesis in mice produced an increase in both NG2+ pericyte subsets. In human OSCC, advanced-stage tumors showed a significant reduction in CD31 mRNA and von Willebrand factor-positive vessels. Low PDGFR-ß expression was related to a shorter disease-free survival time, while NG2 mRNA overexpression was associated with a reduction in overall survival, consistent with the TCGA data. Herein, oral carcinogenesis resulted in an increase in NG2+ pericytes, which negatively affected survival outcomes.
Asunto(s)
Carcinoma de Células Escamosas , Neoplasias de Cabeza y Cuello , Neoplasias de la Boca , Ratones , Humanos , Animales , Pericitos/metabolismo , Carcinoma de Células Escamosas/metabolismo , Nestina/metabolismo , Neoplasias de la Boca/metabolismo , Carcinoma de Células Escamosas de Cabeza y Cuello/patología , Factor de von Willebrand/genética , Factor de von Willebrand/metabolismo , Ratones Transgénicos , Receptor beta de Factor de Crecimiento Derivado de Plaquetas/genética , Receptor beta de Factor de Crecimiento Derivado de Plaquetas/metabolismo , Carcinogénesis/patología , Neoplasias de Cabeza y Cuello/patología , ARN Mensajero/metabolismoRESUMEN
CONTEXT: Nicotinamide nucleotide transhydrogenase (NNT) acts as an antioxidant defense mechanism. NNT mutations cause familial glucocorticoid deficiency (FGD). How impaired oxidative stress disrupts adrenal steroidogenesis remains poorly understood. OBJECTIVE: To ascertain the role played by NNT in adrenal steroidogenesis. METHODS: The genotype-phenotype association of a novel pathogenic NNT variant was evaluated in a boy with FGD. Under basal and oxidative stress (OS) induced conditions, transient cell cultures of the patient's and controls' wild-type (WT) mononuclear blood cells were used to evaluate antioxidant mechanisms and mitochondrial parameters (reactive oxygen species [ROS] production, reduced glutathione [GSH], and mitochondrial mass). Using CRISPR/Cas9, a stable NNT gene knockdown model was built in H295R adrenocortical carcinoma cells to determine the role played by NNT in mitochondrial parameters and steroidogenesis. NNT immunohistochemistry was assessed in fetal and postnatal human adrenals. RESULTS: The homozygous NNT p.G866D variant segregated with the FGD phenotype. Under basal and OS conditions, p.G866D homozygous mononuclear blood cells exhibited increased ROS production, and decreased GSH levels and mitochondrial mass than WT NNT cells. In line H295R, NNT knocked down cells presented impaired NNT protein expression, increased ROS production, decreased the mitochondrial mass, as well as the size and the density of cholesterol lipid droplets. NNT knockdown affected steroidogenic enzyme expression, impairing cortisol and aldosterone secretion. In human adrenals, NNT is abundantly expressed in the transition fetal zone and in zona fasciculata. CONCLUSION: Together, these studies demonstrate the essential role of NNT in adrenal redox homeostasis and steroidogenesis.
Asunto(s)
Neoplasias de la Corteza Suprarrenal , NADP Transhidrogenasas , Masculino , Recién Nacido , Humanos , NADP Transhidrogenasas/genética , NADP Transhidrogenasas/metabolismo , Antioxidantes , Especies Reactivas de Oxígeno/metabolismo , Mitocondrias/metabolismo , Neoplasias de la Corteza Suprarrenal/genéticaRESUMEN
BACKGROUND: Chemoresistance is associated with recurrence and metastasis in oral squamous cell carcinoma (OSCC). The cancer stem cell (CSC) subpopulation is highly resistant to therapy, and they are regulated by epigenetic mechanisms. HDACs are histone deacetylase enzymes that epigenetically regulate gene expression. HDAC6 acts on several physiological processes, including oxidative stress, autophagy and DNA damage response, and its accumulation is associated with cancer. Here, we investigate the role of HDAC6 in CSC-mediated chemoresistance in oral carcinoma in addition to its application as a therapeutic target to reverse chemoresistance. METHODS: Wild-type oral carcinoma cell lines (CAL27 WT and SCC9 WT), cisplatin-resistant (CAL27 CisR and SCC9 CisR), and the subpopulations of cancer stem cells (CSC+) and non-stem (CSC-) derived from CisR cells were investigated. HDAC6 accumulation was analyzed by Western blot and immunofluorescence; DNA damage was evaluated by immunofluorescence of phospho-H2A.X; the qPCR for PRDX2, PRDX6, SOD2, and TXN and ROS assay assessed oxidative stress. Apoptosis and CSC accumulation were investigated by flow cytometry. RESULTS: We identified the accumulation of HDAC6 in CisR cell lines and CSC. Cisplatin-resistant cell lines and CSC demonstrated a reduction in DNA damage and ROS and elevated expression of PRDX2. The administration of tubastatin A (a specific HDAC6 inhibitor) increased oxidative stress and DNA damage and decreased PRDX2. Tubastatin A as a monotherapy induced apoptosis in CisR and CSC and reduced the stemness phenotype. CONCLUSION: High levels of HDAC6 sustain CSC subpopulation and chemoresistance in OSCC, suggesting HDAC6 as a pharmacological target to overcome resistance and perhaps prevent recurrence in OSCC.
Asunto(s)
Carcinoma de Células Escamosas , Neoplasias de Cabeza y Cuello , Neoplasias de la Boca , Carcinoma de Células Escamosas de Cabeza y Cuello , Carcinoma de Células Escamosas/tratamiento farmacológico , Carcinoma de Células Escamosas/metabolismo , Línea Celular Tumoral , Cisplatino/farmacología , Cisplatino/uso terapéutico , Resistencia a Antineoplásicos/efectos de los fármacos , Neoplasias de Cabeza y Cuello/patología , Histona Desacetilasa 6/antagonistas & inhibidores , Histona Desacetilasa 6/metabolismo , Histona Desacetilasas/metabolismo , Humanos , Neoplasias de la Boca/patología , Células Madre Neoplásicas/efectos de los fármacos , Células Madre Neoplásicas/metabolismo , Células Madre Neoplásicas/patología , Especies Reactivas de Oxígeno/metabolismo , Carcinoma de Células Escamosas de Cabeza y Cuello/tratamiento farmacológico , Carcinoma de Células Escamosas de Cabeza y Cuello/patologíaRESUMEN
BACKGROUND: Ischemic stroke produces a large health impact worldwide, with scarce therapeutic options. OBJECTIVE: This study aimed to reveal the role of NADPH oxidase and neuroinflammatory genes in the cerebral anti-ischemic effects of C-Phycocyanin (C-PC), the chief biliprotein of Spirulina platensis. METHODS: Rats with either focal cerebral ischemia/reperfusion (I/R) or acute brain hypoperfusion, received C-PC at different doses, or a vehicle, for up to 6 h post-stroke. Neurological, behavioral and histochemical parameters were assessed in I/R rats at 24 h. Cerebral gene expression and hippocampal neuron viability were evaluated in hypoperfused rats at acute (24 h) or chronic phases (30 days), respectively. A molecular docking analysis of NOX2 and C-PC-derived Phycocyanobilin (PCB) was also performed. RESULTS: C-PC, obtained with a purity of 4.342, significantly reduced the infarct volume and neurological deficit in a dose-dependent manner, and improved the exploratory activity of I/R rats. This biliprotein inhibited NOX2 expression, a crucial NADPH oxidase isoform in the brain, and the superoxide increase produced by the ischemic event. Moreover, C-PC-derived PCB showed a high binding affinity in silico with NOX2. C-PC downregulated the expression of pro-inflammatory genes (IFN-γ, IL-6, IL-17A, CD74, CCL12) and upregulated immune suppressive genes (Foxp3, IL-4, TGF-ß) in hypoperfused brain areas. This compound also decreased chronic neuronal death in the hippocampus of hypoperfused rats. CONCLUSION: These results suggest that the inhibition of cerebral NADPH oxidase and the improvement of neuroinflammation are key mechanisms mediating the neuroprotective actions of C-PC against brain ischemia.
Asunto(s)
Isquemia Encefálica , Fármacos Neuroprotectores , Daño por Reperfusión , Animales , Isquemia Encefálica/tratamiento farmacológico , Isquemia Encefálica/metabolismo , Modelos Animales de Enfermedad , Simulación del Acoplamiento Molecular , NADPH Oxidasas/metabolismo , Fármacos Neuroprotectores/farmacología , Fármacos Neuroprotectores/uso terapéutico , Ficocianina/farmacología , Ficocianina/uso terapéutico , Ratas , Daño por Reperfusión/tratamiento farmacológicoAsunto(s)
Histonas , Esfingolípidos , Acetilación , Metilación de ADN , Epigénesis Genética , Expresión Génica , Histonas/metabolismo , HumanosRESUMEN
Enzymes related to sphingolipids metabolism has been suggested as altered in oral squamous cell carcinoma (OSCC). However, clinical relevance of diverse sphingolipids in OSCC is not fully known. Here, we evaluated sphingolipidomics in plasma and tumor tissues as a tool for diagnosis/prognosis in OSCC patients. Plasma was obtained from 58 controls and 56 OSCC patients, and paired tumor and surgical margin tissues (n = 42). The levels of 28 sphingolipids molecules were obtained by mass spectrometry. Furthermore, sphingolipids were analyzed with clinical and pathological characteristics to search the potential for diagnosis and prognosis. Lower levels of 17 sphingolipids was found in the plasma of OSCC patients compared to controls while four were elevated in tumor tissues. C18:0 dyhidroceramide and C24:0 lactosylceramide in plasma were associated with perineural invasion, while tissue levels of ceramide and dyhidroceramide were associated with advanced tumor stage and perineural invasion. High plasma levels of C24:0 ceramide (HR = 0.10, p = 0.0036) and C24:1 glucosylceramide (HR = 6.62, p = 0.0023), and tissue levels of C24:0 dyhidroceramide (HR = 3.95, p = 0.032) were identified as independent prognostic factors. Moreover, we identified signatures composed by i) sphinganine-1-phosphate and C16 ceramide-1-phosphate in plasma with significant diagnostic accuracy, while ii) C24:0 ceramide, C24:0 dyhidroceramide, and C24:1 glucosylceramide plasma levels, and iii) C24:0 dyhidrosphingomyelin and C24:0 ceramide tissue levels showed value to predict survival in patients aged 60 years or older. We proposed the sphingolipids signatures in plasma and tumor tissues as biomarkers candidates to diagnosis and prognosis in OSCC.
Asunto(s)
Metabolismo de los Lípidos/genética , Pronóstico , Esfingolípidos/sangre , Carcinoma de Células Escamosas de Cabeza y Cuello/genética , Adulto , Anciano , Ceramidas/sangre , Femenino , Regulación Neoplásica de la Expresión Génica/genética , Glucosilceramidas/sangre , Humanos , Masculino , Persona de Mediana Edad , Esfingolípidos/genética , Carcinoma de Células Escamosas de Cabeza y Cuello/sangre , Carcinoma de Células Escamosas de Cabeza y Cuello/patología , Transcriptoma/genéticaRESUMEN
BACKGROUND: Sphingolipids are bioactive lipids that play a role in cancer development. However, the clinical role of sphingolipid (SPL)-related genes in oral cancer (OC) remains not fully understood. OBJECTIVE: This study, aimed to examine the mRNA expression of 14 sphingolipid-related genes in oral cancer patients and their implication with clinicopathological features and prognosis. METHODS: qPCR analysis was performed in 50 OC tissues and their matched surgical margins. Next, Kaplan-Meier, Cox regression, and Receiver operating characteristics (ROC) analysis were applied to evaluate the impact of sphingolipid-related genes expression on the prognosis of OC. RESULTS: The genes SET, ACER3, SK1 and S1PR5 were predominantly up-regulated, while ABCG2, S1PR1, ABCB1 and SPNS2 were down-regulated in OC patients. Analyzing the Cancer Genome Atlas Head-Neck Squamous Cell Carcinoma (TCGA-HNSC) data, which are predominantly composed of OC samples, these genes displayed a similar profile. In OC patients, high levels of SK1 were associated with lymph node metastasis, extracapsular invasion, desmoplasia, locoregional relapse, and disease status. Low levels of SPNS2 were associated with lymph node metastasis, perineural invasion, and disease status. Furthermore, OC and HNSC patients with higher SK1 expression demonstrated shorter disease-free survival (p= 0.0037; p= 0.0087), whereas those with lower SPNS2 expression exhibited shorter overall survival (p= 0.051; p= 0.0012). High levels of ACER3 and low levels of S1PR1 were associated with shorter disease-free and overall survival in HNSC patients. CONCLUSION: Several sphingolipid-related genes are deregulated in OC at the mRNA level and are associated with clinicopathological features and presented potencial for the prediction of poor prognosis in OC patients.
Asunto(s)
Biomarcadores de Tumor/metabolismo , Carcinoma de Células Escamosas/genética , Metástasis Linfática/genética , Neoplasias de la Boca/genética , Esfingolípidos/metabolismo , Humanos , Persona de Mediana Edad , PronósticoRESUMEN
HnRNP K protein is a heterogeneous nuclear ribonucleoprotein which has been proposed to be involved in the leukemogenesis of acute promyelocytic leukemia (APL), as well as in differentiation induced by all-trans retinoic acid (ATRA). We previously demonstrated a connection between SET and hnRNP K function in head and neck squamous cell carcinoma (HNSCC) cells related to splicing processing. The objective of this study was to characterize the participation of hnRNP K and SET proteins in ATRA-induced differentiation in APL. We observed higher (5- to 40-fold) levels of hnRNP K and SET mRNA in APL patients at the diagnosis phase compared with induction and maintenance phases. hnRNP K knockdown using short-hairpin RNA led to cell death in ATRA-sensitive NB4 and resistant NB4-R2 cells by apoptosis with SET cleavage. In addition, hnRNP K knockdown increased granulocytic differentiation in APL cells, mainly in NB4-R2 with ATRA. hnRNP K knockdown had an effect similar to that of treatment with U0126 (an meiosis-specific serine/threonine protein kinase/ERK inhibitor), mainly in NB4-R2 cells. SET knockdown in APL cells revealed that apoptosis induction in cells with hnRNP K knockdown occurred by SET cleavage rather than by reduction in SET protein. Transplantation of NB4-R2 cells into nude mice confirmed that arsenic trioxide (ATO) combined with U0126 has higher potential against tumor progression when compared to ATO. Therefore, hnRNP K/SET and ERK are potential therapeutic targets for both antineoplastic leukemia therapy and relapsed APL patients with ATRA resistance.
Asunto(s)
Leucemia Promielocítica Aguda , Animales , Trióxido de Arsénico/metabolismo , Trióxido de Arsénico/uso terapéutico , Ribonucleoproteína Heterogénea-Nuclear Grupo K/genética , Humanos , Leucemia Promielocítica Aguda/tratamiento farmacológico , Leucemia Promielocítica Aguda/genética , Leucemia Promielocítica Aguda/metabolismo , Ratones , Ratones Desnudos , Tretinoina/metabolismo , Tretinoina/farmacologíaRESUMEN
ERK1/2 inhibitors have attracted special attention concerning the ability of circumventing cases of innate or log-term acquired resistance to RAF and MEK kinase inhibitors. Based on the 4-aminoquinazoline pharmacophore of kinases, herein we describe the synthesis of 4-aminoquinazoline derivatives bearing a 1,2,3-triazole stable core to bridge different aromatic and heterocyclic rings using copper-catalysed azide-alkyne cycloaddition reaction (CuAAC) as a Click Chemistry strategy. The initial screening of twelve derivatives in tumoral cells (CAL-27, HN13, HGC-27, and BT-20) revealed that the most active in BT-20 cells (25a, IC50 24.6 µM and a SI of 3.25) contains a more polar side chain (sulfone). Furthermore, compound 25a promoted a significant release of lactate dehydrogenase (LDH), suggesting the induction of cell death by necrosis. In addition, this compound induced G0/G1 stalling in BT-20 cells, which was accompanied by a decrease in the S phase. Western blot analysis of the levels of p-STAT3, p-ERK, PARP, p53 and cleaved caspase-3 revealed p-ERK1/2 and p-STA3 were drastically decreased in BT-20 cells under 25a incubation, suggesting the involvement of these two kinases in the mechanisms underlying 25a-induced cell cycle arrest, besides loss of proliferation and viability of the breast cancer cell. Molecular docking simulations using the ERK-ulixertinib crystallographic complex showed compound 25a could potentially compete with ATP for binding to ERK in a slightly higher affinity than the reference ERK1/2 inhibitor. Further in silico analyses showed comparable toxicity and pharmacokinetic profiles for compound 25a in relation to ulixertinib.
Asunto(s)
Antineoplásicos/farmacología , Quinasas MAP Reguladas por Señal Extracelular/antagonistas & inhibidores , Simulación del Acoplamiento Molecular , Inhibidores de Proteínas Quinasas/farmacología , Quinazolinas/farmacología , Triazoles/farmacología , Antineoplásicos/síntesis química , Antineoplásicos/química , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Ensayos de Selección de Medicamentos Antitumorales , Quinasas MAP Reguladas por Señal Extracelular/metabolismo , Humanos , Estructura Molecular , Inhibidores de Proteínas Quinasas/síntesis química , Inhibidores de Proteínas Quinasas/química , Quinazolinas/química , Relación Estructura-Actividad , Triazoles/químicaRESUMEN
Bisphenol A (BPA) is an endocrine disrupting chemical able to promote hormone-responsive tumors. The major route of BPA contamination being oral, the aim of the present study was to investigate BPA effects on oral cells. Here, we evaluated the impact of sub-chronic in vivo exposure to BPA and its in vitro effects on neoplastic and non-neoplastic oral cells. We evaluated the oral mucosa of mice chronically exposed to BPA (200 mg/L). The response of keratinocytes (NOK-SI) and Head and Neck (HN) Squamous Cell Carcinoma (SCC), HN12 and HN13 cell lines to BPA was examined. In vivo, BPA accumulated in oral tissues and caused an increase in epithelial proliferative activity. BPA disrupted the function of keratinocytes by altering pro-survival and proliferative pathways and the secretion of cytokines and growth factors. In tumor cells, BPA induced proliferative, invasive, pro-angiogenic, and epigenetic paths. Our data highlight the harmful effects of BPA on oral mucosa and, tumorigenic and non-tumorigenic cells. Additionally, BPA may be a modifier of oral cancer cell behavior by prompting a functional shift to a more aggressive phenotype.
Asunto(s)
Disruptores Endocrinos , Neoplasias de la Boca , Animales , Compuestos de Bencidrilo/toxicidad , Disruptores Endocrinos/toxicidad , Ratones , Mucosa Bucal , Neoplasias de la Boca/inducido químicamente , Fenoles/toxicidadRESUMEN
In vitro 3D culture models have emerged in the cancer field due to their ability to recapitulate characteristics of the in vivo tumor. Herein, we described the establishment and characterization of 3D multicellular spheroids using ovarian cancer cells (SKOV-3) in co-culture with mesenchymal cells (MUC-9) or fibroblasts (CCD27-Sk). We demonstrated that SKOV-3 cells in co-culture were able to form regular and compact spheroids with diameters ranging from 300 to 400 µm and with a roundness close to 1.0 regardless of the type of stromal cell used. In the 3D culture an increase was not observed in spheroid diameter nor was there significant cell growth. What is more, the 3D co-cultures presented an up regulation of genes related to tumorigenesis, angiogenesis and metastases (MMP2, VEGFA, SNAI1, ZEB1 and VIM) when compared with 2D and 3D monoculture. As expected, both 3D cultures (mono and co-cultures) exhibited a higher Paclitaxel chemoresistance when compared to 2D condition. Although we did not observe differences in the Paclitaxel resistance between the 3D mono and co-cultures, the gene expression results indicate that the presence of mesenchymal cells and fibroblasts better recapitulate the in vivo tumor microenvironment, being able, therefore, to more accurately evaluate drug efficacy for ovarian cancer therapy.
Asunto(s)
Detección Precoz del Cáncer , Neoplasias Ováricas , Línea Celular Tumoral , Técnicas de Cocultivo , Evaluación Preclínica de Medicamentos , Femenino , Humanos , Neoplasias Ováricas/tratamiento farmacológico , Esferoides Celulares , Microambiente TumoralRESUMEN
Dehydroepiandrosterone (DHEA) is a steroid hormone secreted by the adrenal glands, gonads and brain. It is a precursor to sex hormones and also is known to have immune modulatory activity. However, little is known about the relationship between DHEA and neutrophils and thus our study evaluates the influence of DHEA in the effector functions of neutrophils. Human neutrophils were treated in vitro with DHEA and further infected with Salmonella enterica serovar Typhimurium. The treatment of neutrophils with 0.01 µM of DHEA increased the phagocytosis of Salmonella independent of TLR4 as the treatment did not modulate the TLR4 expression. Additionally, DHEA caused a decrease in ROS (reactive oxygen species) production and did not influence the formation of the neutrophil extracellular trap (NET). Steroid treated neutrophils, infected or stimulated with LPS (lipopolysaccharide), showed reduced production of IL-8, compared to untreated cells. Also, the protein levels of p-NFκB were decreased in neutrophils treated with DHEA, and this reduction could explain the reduced levels of IL-8. These results led us to conclude that the steroid hormone DHEA has important modulatory functions in neutrophils
Asunto(s)
Humanos , Masculino , Adulto , Técnicas In Vitro , Deshidroepiandrosterona/análisis , Neutrófilos/metabolismo , Fagocitosis/genética , Hormonas Esteroides Gonadales/farmacología , Glándulas Suprarrenales/metabolismo , Salmonella enterica/clasificaciónRESUMEN
As SET protein is overexpressed and PP2A activity is reduced in oral squamous cell carcinoma (OSCC), this study aimed to assess the effects induced by OP449, a PP2A activator/SET inhibitor, on OSCC cells in vitro, and its potential either isolated or combined with FTY720, a PP2A activator/sphingosine kinase 1 antagonist, as antitumoral therapy in vivo. SET protein was analyzed in cells by immunoblotting and cancer stem cells by aldehyde dehydrogenase 1 assay (ALDH1). The cytotoxicity of OP449 was determined in five OSCC lineages by resazurin assay. Molecular actions of OP449 in SET targets were determined by immunoblotting. The coefficient of drug interaction (CDI) was used to characterize the synergism of OP449 and FTY720. The xenograft HN12 tumor model in nude mice was used to assess the antitumoral effect of OP449 and/or FTY720. HN12 (metastatic) cells showed higher SET and ALDH1 levels, and together with SCC9 cells were selected for molecular analysis. OP449 altered several SET functions/targets, such as histone H3 acetylation and NFkB. A synergism in cytotoxicity was observed when HN12 and SCC9 cells were pre-treated with 2 µM OP449 in combination with 15 µM FTY720 (CDI = 0.27 ± 0.088). Nude mice bearing xenograft HN12 tumors treated with OP449 and FTY720 showed reduced tumor mass. Moreover, NFkB was reduced in tumors after treatment. OP449 targets several SET functions, not only PP2A inhibition. Besides, OP449 plus FTY720 has a synergistic antitumoral effect on OSCC. Our results suggest new combined therapies and highlight SET and NFκB signaling as targets for OSCC therapy.
Asunto(s)
Antineoplásicos/uso terapéutico , Carcinoma de Células Escamosas/tratamiento farmacológico , Clorhidrato de Fingolimod/uso terapéutico , Neoplasias de la Boca/tratamiento farmacológico , Péptidos/uso terapéutico , Moduladores de los Receptores de fosfatos y esfingosina 1/uso terapéutico , Animales , Antineoplásicos/farmacología , Carcinoma de Células Escamosas/metabolismo , Línea Celular , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Proteínas de Unión al ADN/metabolismo , Sinergismo Farmacológico , Clorhidrato de Fingolimod/farmacología , Chaperonas de Histonas/metabolismo , Humanos , Masculino , Ratones Endogámicos BALB C , Ratones Desnudos , Neoplasias de la Boca/metabolismo , FN-kappa B/metabolismo , Péptidos/farmacología , Proteína Fosfatasa 2/metabolismo , Moduladores de los Receptores de fosfatos y esfingosina 1/farmacologíaRESUMEN
Non-T cell activation linker (NTAL) is a lipid raft-membrane protein expressed by normal and leukemic cells and involved in cell signaling. In acute promyelocytic leukemia (APL), NTAL depletion from lipid rafts decreases cell viability through regulation of the Akt/PI3K pathway. The role of NTAL in APL cell processes, and its association with clinical outcome, has not, however, been established. Here, we show that reduced levels of NTAL were associated with increased all-trans retinoic acid (ATRA)-induced differentiation, generation of reactive oxygen species, and mitochondrial dysfunction. Additionally, NTAL-knockdown (NTAL-KD) in APL cell lines led to activation of Ras, inhibition of Akt/mTOR pathways, and increased expression of autophagy markers, leading to an increased apoptosis rate following arsenic trioxide treatment. Furthermore, NTAL-KD in NB4 cells decreased the tumor burden in (NOD scid gamma) NSG mice, suggesting its implication in tumor growth. A retrospective analysis of NTAL expression in a cohort of patients treated with ATRA and anthracyclines, revealed that NTAL overexpression was associated with a high leukocyte count (P = 0.007) and was independently associated with shorter overall survival (Hazard Ratio: 3.6; 95% Confidence Interval: 1.17-11.28; P = 0.026). Taken together, our data highlights the importance of NTAL in APL cell survival and response to treatment.
Asunto(s)
Proteínas Adaptadoras Transductoras de Señales/metabolismo , Protocolos de Quimioterapia Combinada Antineoplásica/farmacología , Leucemia Promielocítica Aguda/patología , Proteínas Adaptadoras Transductoras de Señales/genética , Adolescente , Adulto , Anciano , Animales , Antraciclinas/farmacología , Antraciclinas/uso terapéutico , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Médula Ósea/patología , Diferenciación Celular/efectos de los fármacos , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Supervivencia sin Enfermedad , Femenino , Técnicas de Silenciamiento del Gen , Humanos , Leucemia Promielocítica Aguda/sangre , Leucemia Promielocítica Aguda/tratamiento farmacológico , Leucemia Promielocítica Aguda/mortalidad , Recuento de Leucocitos , Masculino , Microdominios de Membrana/metabolismo , Ratones , Persona de Mediana Edad , Estudios Retrospectivos , Análisis de Supervivencia , Tretinoina/farmacología , Tretinoina/uso terapéutico , Ensayos Antitumor por Modelo de Xenoinjerto , Adulto JovenRESUMEN
Aim: Histone acetylation and methylation control gene expression. We investigated the impact of SET knockdown on histone methylation status and the consequences for the miRNAs levels in oral squamous cell carcinoma (OSCC). Methods: OSCC cells with and without SET knockdown were analyzed by quantitative real-time PCR to determine miRNA levels, and by immunoreactions to histone modifications. Results: The knockdown of SET increased the levels of histone H4K20me2 and miR-137. Still, SET protein binds to the miR-137 promoter region. The transfection of miR-137 mimic reduced the KI67 and Rb proteins and proliferation of OSCC cells. Conclusion: Our results show for the first time a relationship between SET and histone methylation associated with the control of miRNA expression and KI67 and Rb as targets of miR-137 in OSCC.
Asunto(s)
Proteínas de Unión al ADN/fisiología , Chaperonas de Histonas/fisiología , Histonas/metabolismo , MicroARNs/genética , Neoplasias de la Boca/genética , Neoplasias de la Boca/metabolismo , Carcinoma de Células Escamosas de Cabeza y Cuello/genética , Carcinoma de Células Escamosas de Cabeza y Cuello/metabolismo , Línea Celular Tumoral , Proliferación Celular , Proteínas de Unión al ADN/metabolismo , Regulación Neoplásica de la Expresión Génica , Chaperonas de Histonas/metabolismo , Humanos , Ácidos Hidroxámicos/farmacología , Antígeno Ki-67/metabolismo , Metilación , MicroARNs/metabolismo , Neoplasias de la Boca/patología , Proteína de Retinoblastoma/metabolismo , Carcinoma de Células Escamosas de Cabeza y Cuello/patologíaRESUMEN
For decades, sphingolipids have been related to several biological functions such as immune system regulation, cell survival, and proliferation. Recently, it has been reported that sphingolipids could be biomarkers in cancer and in other human disorders such as metabolic diseases. This is evidenced by the biological complexity of the sphingolipids associated with cell type-specific signaling and diverse sphingolipids molecules. As mitochondria dynamics have serious implications in homeostasis, in the present review, we focused on the relationship between sphingolipids, mainly ceramides and sphingosine-1-phosphate, and mitochondrial dynamics directed by fission, fusion, and mitophagy. There is evidence that the balances of ceramides (C18 and C16) and S1P, as well as the location of specific ceramide synthases in mitochondria, have roles in mitophagy and fission with an impact on cell fate and metabolism. However, signaling pathways controlling the sphingolipids metabolism and their location in mitochondria need to be better understood in order to propose new interventions and therapeutic strategies.
Asunto(s)
Dinámicas Mitocondriales/inmunología , Transducción de Señal/inmunología , Esfingolípidos/metabolismo , HumanosRESUMEN
Lipid rafts are ordered membrane domains, which provide an environment for the proteins participating in signal transduction. Perifosine is an alkylphospholipid (APL) that inhibits the AKT pathway, cytotoxic to neoplastic cells. We have shown that the lipid raft adaptor protein NTAL is a target of APLs in leukemic cells. Using human mantle cell lymphoma (MCL) Granta-519 cell line we showed here that perifosine decreased NTAL in lipid raft fractions reducing AKT phosphorylation before apoptosis. We also showed that the NTAL-knockdown by shRNA induced a state of reduced AKT activation. Experimental NTAL-knockdown in NSG mouse MCL xenografts reduced AKT activity, increased the basal apoptotic rate by 3-fold (n = 8) and decreased tumor weight by 2.7-fold (n = 5), indicating that NTAL participates in tumor growth. NTAL protein was detected by western blotting in circulating cells of 7 of 8 MCL patients in the leukemic phase, suggesting involvement in the progression of the disease.