RESUMEN
Secretory cells in glands and the nervous system frequently package and store proteins destined for regulated secretion in dense-core granules (DCGs), which disperse when released from the cell surface. Despite the relevance of this dynamic process to diseases such as diabetes and human neurodegenerative disorders, our mechanistic understanding is relatively limited, because of the lack of good cell models to follow the nanoscale events involved. Here, we employ the prostate-like secondary cells (SCs) of the Drosophila male accessory gland to dissect the cell biology and genetics of DCG biogenesis. These cells contain unusually enlarged DCGs, which are assembled in compartments that also form secreted nanovesicles called exosomes. We demonstrate that known conserved regulators of DCG biogenesis, including the small G-protein Arf1 and the coatomer complex AP-1, play key roles in making SC DCGs. Using real-time imaging, we find that the aggregation events driving DCG biogenesis are accompanied by a change in the membrane-associated small Rab GTPases which are major regulators of membrane and protein trafficking in the secretory and endosomal systems. Indeed, a transition from trans-Golgi Rab6 to recycling endosomal protein Rab11, which requires conserved DCG regulators like AP-1, is essential for DCG and exosome biogenesis. Our data allow us to develop a model for DCG biogenesis that brings together several previously disparate observations concerning this process and highlights the importance of communication between the secretory and endosomal systems in controlling regulated secretion.
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Proteínas de Drosophila , Exosomas , Animales , Humanos , Masculino , Vesículas de Núcleo Denso , Drosophila , Proteínas de Drosophila/genética , Exosomas/genética , Proteínas , Proteínas de Unión al GTP rab/genética , Factor de Transcripción AP-1RESUMEN
Deregulated metabolism is one of the hallmarks of cancer. It is well-known that tumour cells tend to metabolize glucose via glycolysis even when oxygen is available and mitochondrial respiration is functional. However, the lower energy efficiency of aerobic glycolysis with respect to mitochondrial respiration makes this behaviour, namely the Warburg effect, counter-intuitive, although it has now been recognized as source of anabolic precursors. On the other hand, there is evidence that oxygenated tumour cells could be fuelled by exogenous lactate produced from glycolysis. We employed a multi-scale approach that integrates multi-agent modelling, diffusion-reaction, stoichiometric equations, and Boolean networks to study metabolic cooperation between hypoxic and oxygenated cells exposed to varying oxygen, nutrient, and inhibitor concentrations. The results show that the cooperation reduces the depletion of environmental glucose, resulting in an overall advantage of using aerobic glycolysis. In addition, the oxygen level was found to be decreased by symbiosis, promoting a further shift towards anaerobic glycolysis. However, the oxygenated and hypoxic populations may gradually reach quasi-equilibrium. A sensitivity analysis using Latin hypercube sampling and partial rank correlation shows that the symbiotic dynamics depends on properties of the specific cell such as the minimum glucose level needed for glycolysis. Our results suggest that strategies that block glucose transporters may be more effective to reduce tumour growth than those blocking lactate intake transporters.
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Neoplasias , Simbiosis , Humanos , Glucólisis , Ácido Láctico/metabolismo , Neoplasias/metabolismo , Glucosa/metabolismo , Hipoxia , OxígenoRESUMEN
Radiotherapy and chemotherapy are effective treatment methods for many types of cancer, but resistance is common. Recent findings indicate that antiviral type I interferon (IFN) signaling is induced by these treatments. However, the underlying mechanisms still need to be elucidated. Expression of a set of IFN-stimulated genes comprises an IFN-related DNA damage resistance signature (IRDS), which correlates strongly with resistance to radiotherapy and chemotherapy across different tumors. Classically, during viral infection, the presence of foreign DNA in the cytoplasm of host cells can initiate type I IFN signaling. Here, we demonstrate that DNA-damaging modalities used during cancer therapy lead to the release of ssDNA fragments from the cell nucleus into the cytosol, engaging this innate immune response. We found that the factors that control DNA end resection during double-strand break repair, including the Bloom syndrome (BLM) helicase and exonuclease 1 (EXO1), play a major role in generating these DNA fragments and that the cytoplasmic 3'-5' exonuclease Trex1 is required for their degradation. Analysis of mRNA expression profiles in breast tumors demonstrates that those with lower Trex1 and higher BLM and EXO1 expression levels are associated with poor prognosis. Targeting BLM and EXO1 could therefore represent a novel approach for circumventing the IRDS produced in response to cancer therapeutics.
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Daño del ADN , Exodesoxirribonucleasas/metabolismo , Inmunidad Innata/genética , Interferones/metabolismo , Fosfoproteínas/metabolismo , Animales , Neoplasias de la Mama/genética , Neoplasias de la Mama/mortalidad , Neoplasias de la Mama/terapia , Línea Celular Tumoral , Citoplasma/enzimología , Citoplasma/inmunología , Citoplasma/metabolismo , Daño del ADN/efectos de los fármacos , ADN de Cadena Simple/inmunología , ADN de Cadena Simple/metabolismo , Perfilación de la Expresión Génica , Regulación Neoplásica de la Expresión Génica , Humanos , Células MCF-7 , Ratones , Mutágenos/uso terapéutico , Mutágenos/toxicidad , Tolerancia a Radiación/inmunología , Radiación Ionizante , Especies Reactivas de Oxígeno , Transducción de SeñalRESUMEN
Exosomes are secreted extracellular vesicles carrying diverse molecular cargos, which can modulate recipient cell behaviour. They are thought to derive from intraluminal vesicles formed in late endosomal multivesicular bodies (MVBs). An alternate exosome formation mechanism, which is conserved from fly to human, is described here, with exosomes carrying unique cargos, including the GTPase Rab11, generated in Rab11-positive recycling endosomal MVBs. Release of Rab11-positive exosomes from cancer cells is increased relative to late endosomal exosomes by reducing growth regulatory Akt/mechanistic Target of Rapamycin Complex 1 (mTORC1) signalling or depleting the key metabolic substrate glutamine, which diverts membrane flux through recycling endosomes. Vesicles produced under these conditions promote tumour cell proliferation and turnover and modulate blood vessel networks in xenograft mouse models in vivo. Their growth-promoting activity, which is also observed in vitro, is Rab11a-dependent, involves ERK-MAPK-signalling and is inhibited by antibodies against amphiregulin, an EGFR ligand concentrated on these vesicles. Therefore, glutamine depletion or mTORC1 inhibition stimulates release from Rab11a compartments of exosomes with pro-tumorigenic functions, which we propose promote stress-induced tumour adaptation.
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Proliferación Celular , Exosomas , Glutamina/deficiencia , Sistema de Señalización de MAP Quinasas , Neoplasias , Animales , Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Drosophila melanogaster , Exosomas/genética , Exosomas/metabolismo , Exosomas/patología , Diana Mecanicista del Complejo 1 de la Rapamicina/genética , Diana Mecanicista del Complejo 1 de la Rapamicina/metabolismo , Neoplasias/genética , Neoplasias/metabolismo , Neoplasias/patología , Proteínas de Unión al GTP rab/genética , Proteínas de Unión al GTP rab/metabolismoRESUMEN
Angiogenesis, the fundamental process by which new blood vessels form from existing ones, depends on precise spatial and temporal gene expression within specific compartments of the endothelium. However, the molecular links between proangiogenic signals and downstream gene expression remain unclear. During sprouting angiogenesis, the specification of endothelial cells into the tip cells that lead new blood vessel sprouts is coordinated by vascular endothelial growth factor A (VEGFA) and Delta-like ligand 4 (Dll4)/Notch signaling and requires high levels of Notch ligand DLL4. Here, we identify MEF2 transcription factors as crucial regulators of sprouting angiogenesis directly downstream from VEGFA. Through the characterization of a Dll4 enhancer directing expression to endothelial cells at the angiogenic front, we found that MEF2 factors directly transcriptionally activate the expression of Dll4 and many other key genes up-regulated during sprouting angiogenesis in both physiological and tumor vascularization. Unlike ETS-mediated regulation, MEF2-binding motifs are not ubiquitous to all endothelial gene enhancers and promoters but are instead overrepresented around genes associated with sprouting angiogenesis. MEF2 target gene activation is directly linked to VEGFA-induced release of repressive histone deacetylases and concurrent recruitment of the histone acetyltransferase EP300 to MEF2 target gene regulatory elements, thus establishing MEF2 factors as the transcriptional effectors of VEGFA signaling during angiogenesis.
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Células Endoteliales/citología , Células Endoteliales/fisiología , Regulación del Desarrollo de la Expresión Génica , Factores de Transcripción MEF2/metabolismo , Neovascularización Fisiológica/genética , Animales , Células Cultivadas , Embrión no Mamífero , Células Endoteliales/enzimología , Elementos de Facilitación Genéticos/genética , Histona Desacetilasas/genética , Histona Desacetilasas/metabolismo , Humanos , Péptidos y Proteínas de Señalización Intracelular/genética , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Factores de Transcripción MEF2/química , Factores de Transcripción MEF2/genética , Proteínas de la Membrana/genética , Proteínas de la Membrana/metabolismo , Ratones , Neovascularización Patológica/genética , Dominios y Motivos de Interacción de Proteínas , Retina/embriología , Transducción de Señal , Factor A de Crecimiento Endotelial Vascular/metabolismo , Pez CebraRESUMEN
Obesity-related cancers account for 40% of the cancer cases observed in the USA and obesity is overtaking smoking as the most widespread modifiable risk factor for carcinogenesis. Here, we use the hallmarks of cancer framework to delineate how obesity might influence the carcinogenic hallmarks in somatic cells. We discuss the effects of obesity on (a) sustaining proliferative signaling; (b) evading growth suppressors; (c) resisting cell death; (d) enabling replicative immortality; (e) inducing angiogenesis; (f) activating invasion and metastasis; (g) reprogramming energy metabolism; and (h) avoiding immune destruction, together with its effects on genome instability and tumour-promoting inflammation. We present the current understanding and controversies in this evolving field, and highlight some areas in need of further cross-disciplinary focus. For instance, the relative importance of the many potentially causative obesity-related factors is unclear for each type of malignancy. Even within a single tumour type, it is currently unknown whether one obesity-related factor consistently plays a predominant role, or if this varies between patients or, even in a single patient with time. Clarifying how the hallmarks are affected by obesity may lead to novel prevention and treatment strategies for the increasingly obese population.
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Carcinogénesis , Neoplasias , Humanos , Neoplasias/metabolismo , Neovascularización Patológica/patología , Obesidad/complicaciones , Transducción de SeñalRESUMEN
Over the past 15 years, there has been great interest in the potential to repurpose the diabetes drug, metformin, as a cancer treatment. However, despite considerable efforts being made to investigate its efficacy in a number of large randomised clinical trials in different tumour types, results have been disappointing to date. This perspective article summarises how interest initially developed in the oncological potential of metformin and the diverse clinical programme of work to date including our contribution to establishing the intra-tumoral pharmacodynamic effects of metformin in the clinic. We also discuss the lessons that can be learnt from this experience and whether a further clinical investigation of metformin in cancer is warranted.
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Metformina , Neoplasias , Humanos , Metformina/uso terapéutico , Reposicionamiento de Medicamentos , Hipoglucemiantes/uso terapéutico , Neoplasias/tratamiento farmacológicoRESUMEN
Decades of research have shown that rare highly penetrant mutations can promote tumorigenesis, but it is still unclear whether variants observed at high-frequency in the broader population could modulate the risk of developing cancer. Genome-wide Association Studies (GWAS) have generated a wealth of data linking single nucleotide polymorphisms (SNPs) to increased cancer risk, but the effect of these mutations are usually subtle, leaving most of cancer heritability unexplained. Understanding the role of high-frequency mutations in cancer can provide new intervention points for early diagnostics, patient stratification and treatment in malignancies with high prevalence, such as breast cancer. Here we review state-of-the-art methods to study cancer heritability using GWAS data and provide an updated map of breast cancer susceptibility loci at the SNP and gene level.
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Biomarcadores de Tumor/genética , Neoplasias de la Mama/epidemiología , Neoplasias de la Mama/genética , Predisposición Genética a la Enfermedad , Estudio de Asociación del Genoma Completo , Modelos Estadísticos , Polimorfismo de Nucleótido Simple , Neoplasias de la Mama/patología , Femenino , Humanos , PronósticoRESUMEN
NEAT1 is a highly abundant nuclear architectural long non-coding RNA. There are two overlapping NEAT1 isoforms, NEAT1_1 and NEAT1_2, of which the latter is an essential scaffold for the assembly of a class of nuclear ribonucleoprotein bodies called paraspeckles. Paraspeckle formation is elevated by a wide variety of cellular stressors and in certain developmental processes, either through transcriptional upregulation of the NEAT1 gene or through a switch from NEAT1_1 to NEAT1_2 isoform production. In such conditions, paraspeckles modulate cellular processes by sequestering proteins or RNA molecules. NEAT1 is abnormally expressed in many cancers and a growing body of evidence suggests that, in many cases, high NEAT1 levels are associated with therapy resistance and poor clinical outcome. Here we review the current knowledge of NEAT1 expression and functions in breast cancer, highlighting its established role in postnatal mammary gland development. We will discuss possible isoform-specific roles of NEAT1_1 and NEAT1_2 in different breast cancer subtypes, which critically needs to be considered when studying NEAT1 and breast cancer.
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Empalme Alternativo , Neoplasias de la Mama/genética , ARN Largo no Codificante/genética , Neoplasias de la Mama/metabolismo , Resistencia a Antineoplásicos , Femenino , Regulación Neoplásica de la Expresión Génica , Humanos , Paraspeckles/metabolismo , Regulación hacia ArribaRESUMEN
BACKGROUND: Height and other anthropometric measures are consistently found to associate with differential cancer risk. However, both genetic and mechanistic insights into these epidemiological associations are notably lacking. Conversely, inherited genetic variants in tumour suppressors and oncogenes increase cancer risk, but little is known about their influence on anthropometric traits. METHODS: By integrating inherited and somatic cancer genetic data from the Genome-Wide Association Study Catalog, expression Quantitative Trait Loci databases and the Cancer Gene Census, we identify SNPs that associate with different cancer types and differential gene expression in at least one tissue type, and explore the potential pleiotropic associations of these SNPs with anthropometric traits through SNP-wise association in a cohort of 500,000 individuals. RESULTS: We identify three regulatory SNPs for three important cancer genes, FANCA, MAP3K1 and TP53 that associate with both anthropometric traits and cancer risk. Of particular interest, we identify a previously unrecognised strong association between the rs78378222[C] SNP in the 3' untranslated region (3'-UTR) of TP53 and both increased risk for developing non-melanomatous skin cancer (OR=1.36 (95% 1.31 to 1.41), adjusted p=7.62E-63), brain malignancy (OR=3.12 (2.22 to 4.37), adjusted p=1.43E-12) and increased standing height (adjusted p=2.18E-24, beta=0.073±0.007), lean body mass (adjusted p=8.34E-37, beta=0.073±0.005) and basal metabolic rate (adjusted p=1.13E-31, beta=0.076±0.006), thus offering a novel genetic link between these anthropometric traits and cancer risk. CONCLUSION: Our results clearly demonstrate that heritable variants in key cancer genes can associate with both differential cancer risk and anthropometric traits in the general population, thereby lending support for a genetic basis for linking these human phenotypes.
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Pesos y Medidas Corporales , Neoplasias/genética , Oncogenes , Polimorfismo de Nucleótido Simple , Adulto , Anciano , Antropometría , Estudios de Cohortes , Femenino , Ligamiento Genético , Pleiotropía Genética , Predisposición Genética a la Enfermedad , Estudio de Asociación del Genoma Completo , Humanos , Masculino , Persona de Mediana Edad , Sitios de Carácter Cuantitativo , Carácter Cuantitativo Heredable , Medición de RiesgoRESUMEN
Tumor hypoxia is associated with poor patient outcomes in estrogen receptor-α-positive (ERα+) breast cancer. Hypoxia is known to affect tumor growth by reprogramming metabolism and regulating amino acid (AA) uptake. Here, we show that the glutamine transporter, SNAT2, is the AA transporter most frequently induced by hypoxia in breast cancer, and is regulated by hypoxia both in vitro and in vivo in xenografts. SNAT2 induction in MCF7 cells was also regulated by ERα, but it became predominantly a hypoxia-inducible factor 1α (HIF-1α)-dependent gene under hypoxia. Relevant to this, binding sites for both HIF-1α and ERα overlap in SNAT2's cis-regulatory elements. In addition, the down-regulation of SNAT2 by the ER antagonist fulvestrant was reverted in hypoxia. Overexpression of SNAT2 in vitro to recapitulate the levels induced by hypoxia caused enhanced growth, particularly after ERα inhibition, in hypoxia, or when glutamine levels were low. SNAT2 up-regulation in vivo caused complete resistance to antiestrogen and, partially, anti-VEGF therapies. Finally, high SNAT2 expression levels correlated with hypoxia profiles and worse outcome in patients given antiestrogen therapies. Our findings show a switch in the regulation of SNAT2 between ERα and HIF-1α, leading to endocrine resistance in hypoxia. Development of drugs targeting SNAT2 may be of value for a subset of hormone-resistant breast cancer.
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Sistema de Transporte de Aminoácidos A/metabolismo , Antineoplásicos Hormonales/uso terapéutico , Neoplasias de la Mama/patología , Hipoxia de la Célula , Resistencia a Antineoplásicos , Moduladores de los Receptores de Estrógeno/uso terapéutico , Animales , Neoplasias de la Mama/tratamiento farmacológico , Neoplasias de la Mama/metabolismo , Línea Celular Tumoral , Receptor alfa de Estrógeno/metabolismo , Femenino , Xenoinjertos , Humanos , Ratones , Microambiente TumoralRESUMEN
BACKGROUND: Glutamine (Gln) is an abundant nutrient used by cancer cells. Breast cancers cells and particularly triple-receptor negative breast cancer (TNBC) are reported to be dependent on Gln to produce the energy required for survival and proliferation. Despite intense research on the role of the intracellular Gln pathway, few reports have focussed on Gln transporters in breast cancer and TNBC. METHODS: The role and localisation of the Gln transporter SLC38A2/SNAT2 in response to Gln deprivation or pharmacological stresses was examined in a panel of breast cancer cell lines. Subsequently, the effect of SLC38A2 knockdown in Gln-sensitive cell lines was analysed. The prognostic value of SLC38A2 in a cohort of breast cancer was determined by immunohistochemistry. RESULTS: SLC38A2 was identified as a strongly expressed amino acid transporter in six breast cancer cell lines. We confirmed an autophagic route of degradation for SLC38A2. SLC38A2 knockdown decreased Gln consumption, inhibited cell growth, induced autophagy and led to ROS production in a subgroup of Gln-sensitive cell lines. High expression of SLC38A2 protein was associated with poor breast cancer specific survival in a large cohort of patients (p = 0.004), particularly in TNBC (p = 0.02). CONCLUSIONS: These results position SLC38A2 as a selective target for inhibiting growth of Gln-dependent breast cancer cell lines.
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Sistema de Transporte de Aminoácidos A/metabolismo , Glutamina/metabolismo , Estrés Oxidativo/fisiología , Neoplasias de la Mama Triple Negativas/metabolismo , Neoplasias de la Mama Triple Negativas/patología , Adulto , Anciano , Línea Celular Tumoral , Femenino , Humanos , Persona de Mediana Edad , PronósticoRESUMEN
BACKGROUND: There is limited knowledge about DCIS cellular composition and relationship with breast cancer events (BCE). METHODS: Immunofluorescence multiplexing (MxIF) was used to image and quantify 32 cellular biomarkers in FFPE DCIS tissue microarrays. Over 75,000 DCIS cells from 51 patients (median 9 years follow-up for non-BCE cases) were analysed for profiles predictive of BCE. K-means clustering was used to evaluate cellular co-expression of epithelial markers with ER and HER2. RESULTS: Only ER, PR and HER2 significantly correlated with BCE. Cluster analysis identified 6 distinct cell groups with different levels of ER, Her2, cMET and SLC7A5. Clusters 1 and 3 were not significant. Clusters 2 and 4 (high ER/low HER2 and SLC7A5/mixed cMET) significantly correlated with low BCE risk (P = 0.001 and P = 0.034), while cluster 6 (high HER2/low ER, cMET and SLC7A5) correlated with increased risk (P = 0.018). Cluster 5 (similar to cluster 6, except high SLC7A5) trended towards significance (P = 0.072). A continuous expression score (Escore) based on these 4 clusters predicted likelihood of BCE (AUC = 0.79, log-rank test P = 5E-05; LOOCV AUC = 0.74, log-rank test P = 0.006). CONCLUSION: Multiplexed spatial analysis of limited tissue is a novel method for biomarker analysis and predicting BCEs. Further validation of Escore is needed in a larger cohort.
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Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Biomarcadores de Tumor/metabolismo , Neoplasias de la Mama/patología , Carcinoma Ductal de Mama/patología , Carcinoma Intraductal no Infiltrante/patología , Mastectomía/métodos , Anciano , Neoplasias de la Mama/metabolismo , Neoplasias de la Mama/terapia , Carcinoma Ductal de Mama/metabolismo , Carcinoma Ductal de Mama/terapia , Carcinoma Intraductal no Infiltrante/metabolismo , Carcinoma Intraductal no Infiltrante/terapia , Terapia Combinada , Femenino , Estudios de Seguimiento , Humanos , Persona de Mediana Edad , Pronóstico , Estudios Retrospectivos , Tasa de SupervivenciaRESUMEN
BACKGROUND: There is a need to improve the treatment of prostate cancer (PCa) and reduce treatment side effects. Vascular-targeted photodynamic therapy (VTP) is a focal therapy for low-risk low-volume localised PCa, which rapidly disrupts targeted tumour vessels. There is interest in expanding the use of VTP to higher-risk disease. Tumour vasculature is characterised by vessel immaturity, increased permeability, aberrant branching and inefficient flow. FRT alters the tumour microenvironment and promotes transient 'vascular normalisation'. We hypothesised that multimodality therapy combining fractionated radiotherapy (FRT) and VTP could improve PCa tumour control compared against monotherapy with FRT or VTP. METHODS: We investigated whether sequential delivery of FRT followed by VTP 7 days later improves flank TRAMP-C1 PCa tumour allograft control compared to monotherapy with FRT or VTP. RESULTS: FRT induced 'vascular normalisation' changes in PCa flank tumour allografts, improving vascular function as demonstrated using dynamic contrast-enhanced magnetic resonance imaging. FRT followed by VTP significantly delayed tumour growth in flank PCa allograft pre-clinical models, compared with monotherapy with FRT or VTP, and improved overall survival. CONCLUSION: Combining FRT and VTP may be a promising multimodal approach in PCa therapy. This provides proof-of-concept for this multimodality treatment to inform early phase clinical trials.
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Neovascularización Patológica/terapia , Fotoquimioterapia/métodos , Neoplasias de la Próstata/terapia , Animales , Línea Celular Tumoral , Terapia Combinada , Fraccionamiento de la Dosis de Radiación , Células Endoteliales de la Vena Umbilical Humana , Humanos , Masculino , Ratones , Neoplasias de la Próstata/irrigación sanguínea , Análisis de Supervivencia , Microambiente Tumoral , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
BACKGROUND: In clear cell renal cell carcinoma, 80% of cases have biallelic inactivation of the VHL gene, leading to constitutive activation of both HIF1α and HIF2α. As HIF2α is the driver of the disease promoting tumour growth and metastasis, drugs targeting HIF2α have been developed. However, resistance is common, therefore new therapies are needed. METHODS: We assessed the effect of the HIF2α antagonist PT2385 in several steps of tumour development and performed RNAseq to identify genes differentially expressed upon treatment. A drug screening was used to identify drugs with antiproliferative effects on VHL-mutated HIF2α-expressing cells and could increase effectiveness of PT2385. RESULTS: PT2385 did not reduce cell proliferation or clonogenicity but, in contrast to the genetic silencing of HIF2α, it reduced in vitro cell invasion. Many HIF-inducible genes were down-regulated upon PT2385 treatment, whereas some genes involved in cell migration or extracellular matrix were up-regulated. HIF2α was associated with resistance to statins, addition to PT2385 did not increase the sensitivity. CONCLUSIONS: this study shows key differences between inhibiting a target versus knockdown, which are potentially targetable.
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Antineoplásicos/farmacología , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/antagonistas & inhibidores , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/genética , Silenciador del Gen , Neoplasias Renales/genética , Neoplasias Renales/metabolismo , Antineoplásicos/uso terapéutico , Línea Celular Tumoral , Movimiento Celular/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Reposicionamiento de Medicamentos , Resistencia a Antineoplásicos/genética , Transición Epitelial-Mesenquimal/efectos de los fármacos , Transición Epitelial-Mesenquimal/genética , Perfilación de la Expresión Génica , Humanos , Indanos/farmacología , Indanos/uso terapéutico , Neoplasias Renales/tratamiento farmacológico , Sulfonas/farmacología , Sulfonas/uso terapéutico , Activación Transcripcional , Transcriptoma , Resultado del TratamientoRESUMEN
Cancer cells are reprogrammed to consume large amounts of glucose to support anabolic biosynthetic pathways. However, blood perfusion and consequently the supply with glucose are frequently inadequate in solid cancers. PEPCK-M (PCK2), the mitochondrial isoform of phosphoenolpyruvate carboxykinase (PEPCK), has been shown by us and others to be functionally expressed and to mediate gluconeogenesis, the reverse pathway of glycolysis, in different cancer cells. Serine and ribose synthesis have been identified as downstream pathways fed by PEPCK in cancer cells. Here, we report that PEPCK-M-dependent glycerol phosphate formation from noncarbohydrate precursors (glyceroneogenesis) occurs in starved lung cancer cells and supports de novo glycerophospholipid synthesis. Using stable isotope-labeled glutamine and lactate, we show that PEPCK-M generates phosphoenolpyruvate and 3-phosphoglycerate, which are at least partially converted to glycerol phosphate and incorporated into glycerophospholipids (GPL) under glucose and serum starvation. This pathway is required to maintain levels of GPL, especially phosphatidylethanolamine (PE), as shown by stable shRNA-mediated silencing of PEPCK-M in H23 lung cancer cells. PEPCK-M shRNA led to reduced colony formation after starvation, and the effect was partially reversed by the addition of dioleyl-PE. Furthermore, PEPCK-M silencing abrogated cancer growth in a lung cancer cell xenograft model. In conclusion, glycerol phosphate formation for de novo GPL synthesis via glyceroneogenesis is a newly characterized anabolic pathway in cancer cells mediated by PEPCK-M under conditions of severe nutrient deprivation.
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Glicerol/metabolismo , Neoplasias/metabolismo , Fosfoenolpiruvato Carboxiquinasa (ATP)/metabolismo , Fosfolípidos/metabolismo , Células A549 , Animales , Glucosa/metabolismo , Glutamina/metabolismo , Xenoinjertos , Humanos , Ácido Láctico/metabolismo , Masculino , Ratones , Ratones Desnudos , Fosfoenolpiruvato Carboxiquinasa (ATP)/genética , Fosfolípidos/químicaRESUMEN
ELTD1 is expressed in endothelial and vascular smooth muscle cells and has a role in angiogenesis. It has been classified as an adhesion GPCR, but as yet, no ligand has been identified and its function remains unknown. To establish its role, ELTD1 was overexpressed in endothelial cells. Expression and consequently ligand independent activation of ELTD1 results in endothelial-mesenchymal transistion (EndMT) with a loss of cell-cell contact, formation of stress fibres and mature focal adhesions and an increased expression of smooth muscle actin. The effect was pro-angiogenic, increasing Matrigel network formation and endothelial sprouting. RNA-Seq analysis after the cells had undergone EndMT revealed large increases in chemokines and cytokines involved in regulating immune response. Gene set enrichment analysis of the data identified a number of pathways involved in myofibroblast biology suggesting that the endothelial cells had undergone a type II EMT. This type of EMT is involved in wound repair and is closely associated with inflammation implicating ELTD1 in these processes.
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Biomarcadores/metabolismo , Células Endoteliales/patología , Transición Epitelial-Mesenquimal , Regulación de la Expresión Génica , Miofibroblastos/patología , Neovascularización Patológica , Receptores Acoplados a Proteínas G/metabolismo , Animales , Apoptosis , Movimiento Celular , Proliferación Celular , Células Cultivadas , Células Endoteliales/metabolismo , Humanos , Ratones Endogámicos C57BL , Miofibroblastos/metabolismo , Fenotipo , RNA-Seq , Receptores Acoplados a Proteínas G/genéticaRESUMEN
Angiogenesis, the formation of new blood vessels by endothelial cells, is a finely tuned process relying on the balance between promoting and repressing signalling pathways. Among these, Notch signalling is critical in ensuring appropriate response of endothelial cells to pro-angiogenic stimuli. However, the downstream targets and pathways effected by Delta-like 4 (DLL4)/Notch signalling and their subsequent contribution to angiogenesis are not fully understood. We found that the Rho GTPase, RHOQ, is induced by DLL4 signalling and that silencing RHOQ results in abnormal sprouting and blood vessel formation both in vitro and in vivo. Loss of RHOQ greatly decreased the level of Notch signalling, conversely overexpression of RHOQ promoted Notch signalling. We describe a new feed-forward mechanism regulating DLL4/Notch signalling, whereby RHOQ is induced by DLL4/Notch and is essential for the NICD nuclear translocation. In the absence of RHOQ, Notch1 becomes targeted for degradation in the autophagy pathway and NICD is sequestered from the nucleus and targeted for degradation in lysosomes.
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Proteínas Adaptadoras Transductoras de Señales/metabolismo , Proteínas de Unión al Calcio/metabolismo , Células Endoteliales de la Vena Umbilical Humana/metabolismo , Neovascularización Fisiológica , Receptores Notch/metabolismo , Transducción de Señal , Proteínas de Unión al GTP rho/metabolismo , Proteínas Adaptadoras Transductoras de Señales/genética , Proteínas de Unión al Calcio/genética , Humanos , Dominios Proteicos , Receptores Notch/genética , Proteínas de Unión al GTP rho/genéticaRESUMEN
Cancer metabolism has undergone a resurgence in the last decade, 70 years after Warburg described aerobic glycolysis as a feature of cancer cells. A wide range of techniques have elucidated the complexity and heterogeneity in preclinical models and clinical studies. What emerges are the large differences between tissues, tumour types and intratumour heterogeneity. However, synergies with inhibition of metabolic pathways have been found for many drugs and therapeutic approaches, and a critical role of window studies and translational trial design is key to success.