RESUMEN
Cancer cells adapt their metabolic processes to support rapid proliferation, but less is known about how cancer cells alter metabolism to promote cell survival in a poorly vascularized tumour microenvironment. Here we identify a key role for serine and glycine metabolism in the survival of brain cancer cells within the ischaemic zones of gliomas. In human glioblastoma multiforme, mitochondrial serine hydroxymethyltransferase (SHMT2) and glycine decarboxylase (GLDC) are highly expressed in the pseudopalisading cells that surround necrotic foci. We find that SHMT2 activity limits that of pyruvate kinase (PKM2) and reduces oxygen consumption, eliciting a metabolic state that confers a profound survival advantage to cells in poorly vascularized tumour regions. GLDC inhibition impairs cells with high SHMT2 levels as the excess glycine not metabolized by GLDC can be converted to the toxic molecules aminoacetone and methylglyoxal. Thus, SHMT2 is required for cancer cells to adapt to the tumour environment, but also renders these cells sensitive to glycine cleavage system inhibition.
Asunto(s)
Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/patología , Glioblastoma/metabolismo , Glioblastoma/patología , Glicina Hidroximetiltransferasa/metabolismo , Glicina/metabolismo , Isquemia/metabolismo , Acetona/análogos & derivados , Acetona/metabolismo , Acetona/toxicidad , Animales , Neoplasias Encefálicas/irrigación sanguínea , Neoplasias Encefálicas/enzimología , Hipoxia de la Célula , Línea Celular Tumoral , Supervivencia Celular , Femenino , Glioblastoma/irrigación sanguínea , Glioblastoma/enzimología , Glicina-Deshidrogenasa (Descarboxilante)/antagonistas & inhibidores , Glicina-Deshidrogenasa (Descarboxilante)/metabolismo , Humanos , Isquemia/enzimología , Isquemia/patología , Ratones , Necrosis , Consumo de Oxígeno , Piruvaldehído/metabolismo , Piruvaldehído/toxicidad , Piruvato Quinasa/metabolismo , Microambiente Tumoral , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Succinate is known to act as an inflammatory signal in classically activated macrophages through stabilization of HIF-1α leading to IL-1ß production. Relevant to this, hypoxia is known to drive succinate accumulation and release into the extracellular milieu. The metabolic alterations associated with succinate release during inflammation and under hypoxia are poorly understood. Data are presented showing that Mycoplasma arginini infection of VM-M3 cancer cells enhances the Warburg effect associated with succinate production in mitochondria and eventual release into the extracellular milieu. We investigated how succinate production and release was related to the changes of other soluble metabolites, including itaconate and 2-HG. Furthermore, we found that hypoxia alone could induce succinate release from the VM-M3 cells and that this could occur in the absence of glucose-driven lactate production. Our results elucidate metabolic pathways responsible for succinate accumulation and release in cancer cells, thus identifying potential targets involved in both inflammation and hypoxia. This article is part of a Special Issue entitled 20th European Bioenergetics Conference, edited by László Zimányi and László Tretter.
Asunto(s)
Neoplasias Encefálicas/patología , Glioblastoma/patología , Hipoxia/complicaciones , Inflamación/complicaciones , Infecciones por Mycoplasma/complicaciones , Mycoplasma/patogenicidad , Succinatos/metabolismo , Animales , Neoplasias Encefálicas/etiología , Neoplasias Encefálicas/metabolismo , Metabolismo Energético , Glioblastoma/etiología , Glioblastoma/metabolismo , Metaboloma , Ratones , Células Tumorales CultivadasRESUMEN
Malignant brain tumors are a significant health problem in children and adults. Conventional therapeutic approaches have been largely unsuccessful in providing long-term management. As primarily a metabolic disease, malignant brain cancer can be managed through changes in metabolic environment. In contrast to normal neurons and glia, which readily transition to ketone bodies (ß-hydroxybutyrate) for energy under reduced glucose, malignant brain tumors are strongly dependent on glycolysis for energy. The transition from glucose to ketone bodies as a major energy source is an evolutionary conserved adaptation to food deprivation that permits the survival of normal cells during extreme shifts in nutritional environment. Only those cells with a flexible genome and normal mitochondria can effectively transition from one energy state to another. Mutations restrict genomic and metabolic flexibility thus making tumor cells more vulnerable to energy stress than normal cells. We propose an alternative approach to brain cancer management that exploits the metabolic flexibility of normal cells at the expense of the genetically defective and metabolically challenged tumor cells. This approach to brain cancer management is supported from recent studies in mice and humans treated with calorie restriction and the ketogenic diet. Issues of implementation and use protocols are presented for the metabolic management of brain cancer.
Asunto(s)
Neoplasias Encefálicas/terapia , Metabolismo Energético/fisiología , Glioblastoma/terapia , Adulto , Animales , Neoplasias Encefálicas/complicaciones , Neoplasias Encefálicas/dietoterapia , Neoplasias Encefálicas/metabolismo , Restricción Calórica , Niño , Dieta Cetogénica , Modelos Animales de Enfermedad , Glioblastoma/complicaciones , Glioblastoma/dietoterapia , Glioblastoma/metabolismo , Humanos , Ratones , Enfermedades Mitocondriales/complicaciones , Enfermedades Mitocondriales/metabolismo , Enfermedades Mitocondriales/terapia , Modelos BiológicosRESUMEN
Metastatic cancer is a major cause of morbidity and mortality. Current therapeutic options consist of chemotherapy, radiation or targeted therapies. However, these therapies are often toxic, effective over a small range of cancer types or result in drug resistance. Therefore, a more global, less toxic strategy for the management of metastatic cancer is required. Although most cancers display increased glucose metabolism, glutamine is also a major energy substrate for many cancers. We evaluated the antimetastatic potential of 6-diazo-5-oxo-L-norleucine (DON), a glutamine analog, using the new VM mouse model of systemic metastasis. We found that primary tumor growth was â¼20-fold less in DON-treated mice than in untreated control mice. We also found that DON treatment inhibited metastasis to liver, lung and kidney as detected by bioluminescence imaging and histology. Our findings provide proof of concept that metabolic therapies targeting glutamine metabolism can manage systemic metastatic cancer.
Asunto(s)
Antimetabolitos Antineoplásicos/farmacología , Neoplasias Encefálicas/tratamiento farmacológico , Diazooxonorleucina/farmacología , Glutamina/metabolismo , Animales , Glucemia/metabolismo , Peso Corporal , Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/patología , Restricción Calórica , Procesos de Crecimiento Celular/efectos de los fármacos , Línea Celular Tumoral , Cerebro/metabolismo , Cerebro/patología , Sistemas de Liberación de Medicamentos , Femenino , Glucosa/deficiencia , Glucosa/metabolismo , Glutamina/deficiencia , Masculino , Ratones , Metástasis de la NeoplasiaRESUMEN
We recently identified a new tumor (VM-M3), which arose spontaneously in the brain of an inbred VM mouse. When grown outside the brain, the VM-M3 tumor expresses all major biological processes of metastasis to include local invasion, intravasation, immune system survival, extravasation, and secondary tumor formation involving lung, liver, kidney, spleen and brain. The VM-M3 tumor also expresses multiple properties of macrophage-like cells similar to those described previously in numerous human metastatic cancers suggesting that the VM-M3 model will be useful for studying most types of metastatic cancer, regardless of tissue origin. VM-M3 tumor cells, expressing firefly luciferase (VM-M3/Fluc), were grown subcutaneously in the immunocompetent and syngeneic VM mouse host. The antimetastatic effects of methotrexate (MTX; 25 mg/kg) and cisplatin (10-15 mg/kg) were evaluated following i.p. injections administered once/wk for 3 weeks. Bioluminescent imaging was used to measure VM-M3/Fluc growth and metastasis. All (12/12) control mice developed systemic cancer within 21 days of subcutaneous VM-M3/Fluc implantation. Although methotrexate did not inhibit VM-M3/Fluc primary tumor growth, it reduced lung and liver metastasis by 50% and completely inhibited metastasis to kidneys, spleen and brain. Cisplatin significantly reduced primary tumor growth, blocked metastasis to lung, liver, kidneys, spleen and brain, and significantly increased survival in all treated animals. Our findings show that the response of the VM-M3/Fluc tumor to MTX and cisplatin is similar to that reported in humans with metastatic disease. These findings indicate that the VM-M3/Fluc tumor is a reliable preclinical model for evaluating antimetastatic cancer therapies and underlying control pathways.
Asunto(s)
Antineoplásicos/farmacología , Cisplatino/farmacología , Modelos Animales de Enfermedad , Metotrexato/farmacología , Metástasis de la Neoplasia/patología , Animales , Progresión de la Enfermedad , Ratones , Ratones Endogámicos , Análisis de SupervivenciaRESUMEN
Glioblastoma multiforme (GBM) is a rapidly progressive disease of morbidity and mortality and is the most common form of primary brain cancer in adults. Lack of appropriate in vivo models has been a major roadblock to developing effective therapies for GBM. A new highly invasive in vivo GBM model is described that was derived from a spontaneous brain tumor (VM-M3) in the VM mouse strain. Highly invasive tumor cells could be identified histologically on the hemisphere contralateral to the hemisphere implanted with tumor cells or tissue. Tumor cells were highly expressive for the chemokine receptor CXCR4 and the proliferation marker Ki-67 and could be identified invading through the pia mater, the vascular system, the ventricular system, around neurons, and over white matter tracts including the corpus callosum. In addition, the brain tumor cells were labeled with the firefly luciferase gene, allowing for non-invasive detection and quantitation through bioluminescent imaging. The VM-M3 tumor has a short incubation time with mortality occurring in 100% of the animals within approximately 15 days. The VM-M3 brain tumor model therefore can be used in a pre-clinical setting for the rapid evaluation of novel anti-invasive therapies.
Asunto(s)
Astrocitoma/patología , Neoplasias Encefálicas/patología , Movimiento Celular , Proliferación Celular , Modelos Animales de Enfermedad , Neuronas/patología , Animales , Astrocitoma/genética , Astrocitoma/metabolismo , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/metabolismo , Adhesión Celular , Técnicas para Inmunoenzimas , Proteína 2 de Unión a Factor de Crecimiento Similar a la Insulina/genética , Proteína 2 de Unión a Factor de Crecimiento Similar a la Insulina/metabolismo , Ratones , Ratones Endogámicos C57BL , Invasividad Neoplásica , Neuronas/metabolismo , ARN Mensajero/genética , Receptores CXCR4/genética , Receptores CXCR4/metabolismo , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Tasa de Supervivencia , Células Tumorales CultivadasRESUMEN
Metastasis is the process by which cancer cells disseminate from the primary neoplasm and invade surrounding tissue and distant organs, and is the primary cause of morbidity and mortality for cancer patients. Most conventional cancer therapies are ineffective in managing tumor metastasis. This has been due in large part to the absence of in vivo metastatic models that represent the full spectrum of metastatic disease. Here we identify 3 new spontaneously arising tumors in the inbred VM mouse strain, which has a relatively high incidence of CNS tumors. Two of the tumors (VM-M2 and VM-M3) reliably expressed all of the major biological processes of metastasis to include local invasion, intravasation, immune system survival, extravasation and secondary tumor formation involving liver, kidney, spleen, lung and brain. Metastasis was assessed through visual organ inspection, histology, immunohistochemistry and bioluminescence imaging. The metastatic VM tumor cells also expressed multiple properties of macrophages including morphological appearance, surface adhesion, phagocytosis, total lipid composition (glycosphingolipids and phospholipids) and gene expression (CD11b, Iba1, F4/80, CD68, CD45 and CXCR4). The third tumor (VM-NM1) grew rapidly and expressed properties of neural stem/progenitor cells, but was neither invasive nor metastatic. Our data indicate that spontaneous brain tumors can arise from different cell types in VM mice and that metastatic cancer can represent a disease of macrophage-like cells similar to those described in several human metastatic cancers. The new VM tumor model will be useful for defining the biological processes of cancer metastasis and for evaluating potential therapies for tumor management.
Asunto(s)
Neoplasias Encefálicas/química , Neoplasias Encefálicas/patología , Proteínas de Unión al Calcio/análisis , Macrófagos , Metástasis de la Neoplasia , Animales , Western Blotting , Adhesión Celular , Cromatografía Líquida de Alta Presión , Modelos Animales de Enfermedad , Femenino , Regulación Neoplásica de la Expresión Génica , Neoplasias Renales/química , Neoplasias Renales/secundario , Lípidos/análisis , Neoplasias Hepáticas/química , Neoplasias Hepáticas/secundario , Masculino , Ratones , Proteínas de Microfilamentos , Trasplante de Neoplasias , Fagocitosis , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Neoplasias del Bazo/química , Neoplasias del Bazo/secundario , Transducción GenéticaAsunto(s)
Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/terapia , Metabolismo Energético/efectos de los fármacos , Metabolismo Energético/efectos de la radiación , Glioblastoma/metabolismo , Glioblastoma/terapia , Antineoplásicos/efectos adversos , Humanos , Radioterapia/efectos adversosRESUMEN
Intratumoral hypoxia stimulates enrichment of breast cancer stem cells (BCSC), which are critical for metastasis and patient mortality. Here we report a metabolic adaptation that is required for hypoxia-induced BCSC enrichment and metastasis. Hypoxia-inducible factors coordinately regulate expression of genes encoding phosphoglycerate dehydrogenase (PHGDH) and five downstream enzymes in the serine synthesis pathway and mitochondrial one-carbon (folate) cycle. RNAi-mediated silencing of PHGDH expression in both estrogen receptor-positive and negative breast cancer cells led to decreased NADPH levels, disturbed mitochondrial redox homeostasis, and increased apoptosis, which abrogated BCSC enrichment under hypoxic conditions. PHGDH-deficient cells exhibited increased oxidant levels and apoptosis, as well as loss of BCSC enrichment, in response to treatment with carboplatin or doxorubicin. PHGDH-deficient cells were relatively weakly tumorigenic and tumors that did form were deficient in BCSCs, abolishing metastatic capacity. Our findings highlight a role for PHGDH in the formation of secondary (recurrent or metastatic) tumors, with potential implications for therapeutic targeting of advanced cancers. Cancer Res; 76(15); 4430-42. ©2016 AACR.
Asunto(s)
Neoplasias de la Mama/genética , Neoplasias Pulmonares/secundario , Mitocondrias/metabolismo , Células Madre Neoplásicas/metabolismo , Fosfoglicerato-Deshidrogenasa/metabolismo , Neoplasias de la Mama/patología , Femenino , Humanos , Metástasis de la Neoplasia , Células Madre Neoplásicas/patología , TransfecciónRESUMEN
Mammary epithelial cells transition between periods of proliferation and quiescence during development, menstrual cycles, and pregnancy, and as a result of oncogenic transformation. Utilizing an organotypic 3D tissue culture model coupled with quantitative metabolomics and proteomics, we identified significant differences in glutamate utilization between proliferating and quiescent cells. Relative to quiescent cells, proliferating cells catabolized more glutamate via transaminases to couple non-essential amino acid (NEAA) synthesis to α-ketoglutarate generation and tricarboxylic acid (TCA) cycle anaplerosis. As cells transitioned to quiescence, glutamine consumption and transaminase expression were reduced, while glutamate dehydrogenase (GLUD) was induced, leading to decreased NEAA synthesis. Highly proliferative human tumors display high transaminase and low GLUD expression, suggesting that proliferating cancer cells couple glutamine consumption to NEAA synthesis to promote biosynthesis. These findings describe a competitive and partially redundant relationship between transaminases and GLUD, and they reveal how coupling of glutamate-derived carbon and nitrogen metabolism can be regulated to support cell proliferation.
Asunto(s)
Células Epiteliales/citología , Células Epiteliales/metabolismo , Ácido Glutámico/metabolismo , Glándulas Mamarias Humanas/citología , Neoplasias de la Mama/enzimología , Neoplasias de la Mama/patología , Proliferación Celular , Células Cultivadas , Femenino , Glutamato Deshidrogenasa/metabolismo , Humanos , Metabolómica , Modelos Biológicos , Isótopos de Nitrógeno , Fosfatidilinositol 3-Quinasas/metabolismo , Proteómica , Proteínas Proto-Oncogénicas c-akt/metabolismo , Transducción de Señal , Serina-Treonina Quinasas TOR/metabolismo , Transaminasas/metabolismoRESUMEN
BACKGROUND: Systematic quantitative methodologies are needed to understand the heterogeneity of cell metabolism across cell types in normal physiology, disease, and treatment. Metabolic flux analysis (MFA) can be used to infer steady state fluxes, but it does not apply for transient dynamics. Kinetic flux profiling (KFP) can be used in the context of transient dynamics, and it is the current gold standard. However, KFP requires measurements at several time points, limiting its use in high-throughput applications. RESULTS: Here we propose transient MFA (tMFA) as a cost-effective methodology to quantify metabolic fluxes using metabolomics and isotope tracing. tMFA exploits the time scale separation between the dynamics of different metabolites to obtain mathematical equations relating metabolic fluxes to metabolite concentrations and isotope fractions. We show that the isotope fractions of serine and glycine are at steady state 8 h after addition of a tracer, while those of purines and glutathione are following a transient dynamics with an approximately constant turnover rate per unit of metabolite, supporting the application of tMFA to the analysis of folate metabolism. Using tMFA, we investigate the heterogeneity of folate metabolism and the response to the antifolate methotrexate in breast cancer cells. Our analysis indicates that methotrexate not only inhibits purine synthesis but also induces an increase in the AMP/ATP ratio, activation of AMP kinase (AMPK), and the inhibition of protein and glutathione synthesis. We also find that in some cancer cells, the generation of one-carbon units from serine exceeds the biosynthetic demand. CONCLUSIONS: This work validates tMFA as a cost-effective methodology to investigate cell metabolism. Using tMFA, we have shown that the effects of treatment with the antifolate methotrexate extend beyond inhibition of purine synthesis and propagate to other pathways in central metabolism.
RESUMEN
We report that a single growth factor, NM23-H1, enables serial passaging of both human ES and iPS cells in the absence of feeder cells, their conditioned media or bFGF in a fully defined xeno-free media on a novel defined, xeno-free surface. Stem cells cultured in this system show a gene expression pattern indicative of a more "naïve" state than stem cells grown in bFGF-based media. NM23-H1 and MUC1* growth factor receptor cooperate to control stem cell self-replication. By manipulating the multimerization state of NM23-H1, we override the stem cell's inherent programming that turns off pluripotency and trick the cells into continuously replicating as pluripotent stem cells. Dimeric NM23-H1 binds to and dimerizes the extra cellular domain of the MUC1* transmembrane receptor which stimulates growth and promotes pluripotency. Inhibition of the NM23-H1/MUC1* interaction accelerates differentiation and causes a spike in miR-145 expression which signals a cell's exit from pluripotency.
Asunto(s)
Nucleósido Difosfato Quinasas NM23/farmacología , Células Madre/efectos de los fármacos , Anticuerpos Monoclonales/inmunología , Anticuerpos Monoclonales/metabolismo , Unión Competitiva , Biomarcadores/metabolismo , Diferenciación Celular , Células Cultivadas , Medios de Cultivo Condicionados/farmacología , Células Madre Embrionarias/citología , Células Madre Embrionarias/efectos de los fármacos , Células Madre Embrionarias/metabolismo , Factor 2 de Crecimiento de Fibroblastos/farmacología , Fibroblastos/metabolismo , Humanos , Células Madre Pluripotentes Inducidas/citología , Células Madre Pluripotentes Inducidas/efectos de los fármacos , Células Madre Pluripotentes Inducidas/metabolismo , Ligandos , MicroARNs/genética , MicroARNs/metabolismo , Mucina-1/inmunología , Mucina-1/metabolismo , Nucleósido Difosfato Quinasas NM23/química , Nucleósido Difosfato Quinasas NM23/metabolismo , Unión Proteica/efectos de los fármacos , Multimerización de Proteína , Células Madre/citología , Células Madre/metabolismoRESUMEN
Malignant brain cancer persists as a major disease of morbidity and mortality. The failure to recognize brain cancer as a disease of energy metabolism has contributed in large part to the failure in management. As long as brain tumor cells have access to glucose and glutamine, the disease will progress. The current standard of care provides brain tumors with access to glucose and glutamine. The high fat low carbohydrate ketogenic diet (KD) will target glucose availability and possibly that of glutamine when administered in carefully restricted amounts to reduce total caloric intake and circulating levels of glucose. The restricted KD (RKD) targets major signaling pathways associated with glucose and glutamine metabolism including the IGF-1/PI3K/Akt/Hif pathway. The RKD is anti-angiogenic, anti-invasive, anti-inflammatory, and pro-apoptotic when evaluated in mice with malignant brain cancer. The therapeutic efficacy of the restricted KD can be enhanced when combined with drugs that also target glucose and glutamine. Therapeutic efficacy of the RKD was also seen against malignant gliomas in human case reports. Hence, the RKD can be an effective non-toxic therapeutic option to the current standard of care for inhibiting the growth and invasive properties of malignant brain cancer.
Asunto(s)
Neoplasias Encefálicas/dietoterapia , Restricción Calórica , Dieta Cetogénica , Metabolismo Energético/efectos de los fármacos , Glioblastoma/dietoterapia , Animales , Glucosa/metabolismo , HumanosRESUMEN
Emerging evidence indicates that impaired cellular energy metabolism is the defining characteristic of nearly all cancers regardless of cellular or tissue origin. In contrast to normal cells, which derive most of their usable energy from oxidative phosphorylation, most cancer cells become heavily dependent on substrate level phosphorylation to meet energy demands. Evidence is reviewed supporting a general hypothesis that genomic instability and essentially all hallmarks of cancer, including aerobic glycolysis (Warburg effect), can be linked to impaired mitochondrial function and energy metabolism. A view of cancer as primarily a metabolic disease will impact approaches to cancer management and prevention.
RESUMEN
GBM (glioblastoma multiforme) is the most aggressive and invasive form of primary human brain cancer. We recently developed a novel brain cancer model in the inbred VM mouse strain that shares several characteristics with human GBM. Using bioluminescence imaging, we tested the efficacy of CR (calorie restriction) for its ability to reduce tumour size and invasion. CR targets glycolysis and rapid tumour cell growth in part by lowering circulating glucose levels. The VM-M3 tumour cells were implanted intracerebrally in the syngeneic VM mouse host. Approx. 12-15 days post-implantation, brains were removed and both ipsilateral and contralateral hemispheres were imaged to measure bioluminescence of invading tumour cells. CR significantly reduced the invasion of tumour cells from the implanted ipsilateral hemisphere into the contralateral hemisphere. The total percentage of Ki-67-stained cells within the primary tumour and the total number of blood vessels was also significantly lower in the CR-treated mice than in the mice fed ad libitum, suggesting that CR is anti-proliferative and anti-angiogenic. Our findings indicate that the VM-M3 GBM model is a valuable tool for studying brain tumour cell invasion and for evaluating potential therapeutic approaches for managing invasive brain cancer. In addition, we show that CR can be effective in reducing malignant brain tumour growth and invasion.
Asunto(s)
Neoplasias Encefálicas/dietoterapia , Restricción Calórica/métodos , Modelos Animales de Enfermedad , Glioblastoma/dietoterapia , Invasividad Neoplásica , Animales , Neoplasias Encefálicas/patología , Neoplasias Encefálicas/prevención & control , Glioblastoma/patología , Glioblastoma/prevención & control , Masculino , Ratones , Ratones Endogámicos , Invasividad Neoplásica/patología , Invasividad Neoplásica/prevención & controlRESUMEN
Gangliosides are sialic acid-containing glycosphingolipids that have long been associated with tumor malignancy and metastasis. Mounting evidence suggests that gangliosides also modulate tumor angiogenesis. Tumor cells shed gangliosides into the microenvironment, which produces both autocrine and paracrine effects on tumor cells and tumor-associated host cells. In this study, we show that the simple monosialoganglioside GM3 counteracts the proangiogenic effects of vascular endothelial growth factor (VEGF) and of the complex disialoganglioside GD1a. GM3 suppressed the action of VEGF and GD1a on the proliferation of human umbilical vein endothelial cells (HUVECs) and inhibited the migration of HUVECs toward VEGF as a chemoattractant. Enrichment of added GM3 in the HUVEC membrane also reduced the phosphorylation of vascular endothelial growth factor receptor 2 (VEGFR-2) and downstream Akt. Moreover, GM3 reduced the proangiogenic effects of GD1a and growth factors in the in vivo Matrigel plug assay. Inhibition of GM3 biosynthesis with the glucosyl transferase inhibitor, N-butyldeoxynojirimycin (NB-DNJ), increased HUVEC proliferation and the phosphorylation of VEGFR-2 and Akt. The effects of NB-DNJ on HUVECs were reversed with the addition of GM3. We conclude that GM3 has antiangiogenic action and may possess therapeutic potential for reducing tumor angiogenesis.