RESUMEN
The risk of cancer and associated mortality increases substantially in humans from the age of 65 years onwards1-6. Nonetheless, our understanding of the complex relationship between age and cancer is still in its infancy2,3,7,8. For decades, this link has largely been attributed to increased exposure time to mutagens in older individuals. However, this view does not account for the established role of diet, exercise and small molecules that target the pace of metabolic ageing9-12. Here we show that metabolic alterations that occur with age can produce a systemic environment that favours the progression and aggressiveness of tumours. Specifically, we show that methylmalonic acid (MMA), a by-product of propionate metabolism, is upregulated in the serum of older people and functions as a mediator of tumour progression. We traced this to the ability of MMA to induce SOX4 expression and consequently to elicit transcriptional reprogramming that can endow cancer cells with aggressive properties. Thus, the accumulation of MMA represents a link between ageing and cancer progression, suggesting that MMA is a promising therapeutic target for advanced carcinomas.
Asunto(s)
Envejecimiento/metabolismo , Progresión de la Enfermedad , Ácido Metilmalónico/metabolismo , Invasividad Neoplásica , Metástasis de la Neoplasia , Neoplasias/patología , Adulto , Anciano , Envejecimiento/sangre , Envejecimiento/genética , Animales , Línea Celular Tumoral , Femenino , Regulación Neoplásica de la Expresión Génica , Humanos , Masculino , Ácido Metilmalónico/sangre , Ratones , Persona de Mediana Edad , Invasividad Neoplásica/genética , Invasividad Neoplásica/patología , Metástasis de la Neoplasia/genética , Metástasis de la Neoplasia/patología , Neoplasias/sangre , Neoplasias/genética , Factores de Transcripción SOXC/metabolismo , Transducción de Señal , Transcriptoma/genética , Factor de Crecimiento Transformador beta/metabolismoRESUMEN
Cancer cells reprogram their metabolism to support growth and to mitigate cellular stressors. The serine synthesis pathway has been identified as a metabolic pathway frequently altered in cancers and there has been considerable interest in developing pharmacological agents to target this pathway. Here, we report a series of indole amides that inhibit human 3-phosphoglycerate dehydrogenase (PHGDH), the enzyme that catalyzes the first committed step of the serine synthesis pathway. Using X-ray crystallography, we show that the indole amides bind the NAD+ pocket of PHGDH. Through structure-based optimization we were able to develop compounds with low nanomolar affinities for PHGDH in an enzymatic IC50 assay. In cellular assays, the most potent compounds inhibited de novo serine synthesis with low micromolar to sub-micromolar activities and these compounds successfully abrogated the proliferation of cancer cells in serine free media. The indole amide series reported here represent an important improvement over previously published PHGDH inhibitors as they are markedly more potent and their mechanism of action is better defined.
Asunto(s)
Amidas/química , Inhibidores Enzimáticos/química , Indoles/química , Fosfoglicerato-Deshidrogenasa/antagonistas & inhibidores , Serina/biosíntesis , Amidas/metabolismo , Amidas/farmacología , Sitios de Unión , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Cristalografía por Rayos X , Inhibidores Enzimáticos/metabolismo , Inhibidores Enzimáticos/farmacología , Humanos , Simulación de Dinámica Molecular , Fosfoglicerato-Deshidrogenasa/metabolismo , Estructura Terciaria de Proteína , Relación Estructura-ActividadRESUMEN
Cancer cells have metabolic dependencies that distinguish them from their normal counterparts. Among these dependencies is an increased use of the amino acid glutamine to fuel anabolic processes. Indeed, the spectrum of glutamine-dependent tumours and the mechanisms whereby glutamine supports cancer metabolism remain areas of active investigation. Here we report the identification of a non-canonical pathway of glutamine use in human pancreatic ductal adenocarcinoma (PDAC) cells that is required for tumour growth. Whereas most cells use glutamate dehydrogenase (GLUD1) to convert glutamine-derived glutamate into α-ketoglutarate in the mitochondria to fuel the tricarboxylic acid cycle, PDAC relies on a distinct pathway in which glutamine-derived aspartate is transported into the cytoplasm where it can be converted into oxaloacetate by aspartate transaminase (GOT1). Subsequently, this oxaloacetate is converted into malate and then pyruvate, ostensibly increasing the NADPH/NADP(+) ratio which can potentially maintain the cellular redox state. Importantly, PDAC cells are strongly dependent on this series of reactions, as glutamine deprivation or genetic inhibition of any enzyme in this pathway leads to an increase in reactive oxygen species and a reduction in reduced glutathione. Moreover, knockdown of any component enzyme in this series of reactions also results in a pronounced suppression of PDAC growth in vitro and in vivo. Furthermore, we establish that the reprogramming of glutamine metabolism is mediated by oncogenic KRAS, the signature genetic alteration in PDAC, through the transcriptional upregulation and repression of key metabolic enzymes in this pathway. The essentiality of this pathway in PDAC and the fact that it is dispensable in normal cells may provide novel therapeutic approaches to treat these refractory tumours.
Asunto(s)
Glutamina/metabolismo , Redes y Vías Metabólicas , Proteína Oncogénica p21(ras)/metabolismo , Neoplasias Pancreáticas/metabolismo , Neoplasias Pancreáticas/patología , Proteínas Proto-Oncogénicas/metabolismo , Proteínas ras/metabolismo , Adenocarcinoma/genética , Adenocarcinoma/metabolismo , Adenocarcinoma/patología , Aspartato Aminotransferasas/deficiencia , Aspartato Aminotransferasas/genética , Aspartato Aminotransferasas/metabolismo , Línea Celular Tumoral , Proliferación Celular , Ciclo del Ácido Cítrico , Glutamato Deshidrogenasa/metabolismo , Homeostasis , Humanos , Ácidos Cetoglutáricos/metabolismo , Proteína Oncogénica p21(ras)/genética , Oncogenes/genética , Oxidación-Reducción , Neoplasias Pancreáticas/genética , Proteínas Proto-Oncogénicas/genética , Proteínas Proto-Oncogénicas p21(ras) , Especies Reactivas de Oxígeno/metabolismo , Proteínas ras/genéticaRESUMEN
Cancer cells reprogram their metabolism to promote growth and proliferation. The genetic evidence pointing to the importance of the amino acid serine in tumorigenesis is striking. The gene encoding the enzyme 3-phosphoglycerate dehydrogenase (PHGDH), which catalyzes the first committed step of serine biosynthesis, is overexpressed in tumors and cancer cell lines via focal amplification and nuclear factor erythroid-2-related factor 2 (NRF2)-mediated up-regulation. PHGDH-overexpressing cells are exquisitely sensitive to genetic ablation of the pathway. Here, we report the discovery of a selective small molecule inhibitor of PHGDH, CBR-5884, identified by screening a library of 800,000 drug-like compounds. CBR-5884 inhibited de novo serine synthesis in cancer cells and was selectively toxic to cancer cell lines with high serine biosynthetic activity. Biochemical characterization of the inhibitor revealed that it was a noncompetitive inhibitor that showed a time-dependent onset of inhibition and disrupted the oligomerization state of PHGDH. The identification of a small molecule inhibitor of PHGDH not only enables thorough preclinical evaluation of PHGDH as a target in cancers, but also provides a tool with which to study serine metabolism.
Asunto(s)
Neoplasias/metabolismo , Fosfoglicerato-Deshidrogenasa/antagonistas & inhibidores , Serina/biosíntesis , Línea Celular Tumoral , Proliferación Celular , Humanos , Neoplasias/patologíaRESUMEN
Pancreatic cancer cells are characterized by deregulated metabolic programs that facilitate growth and resistance to oxidative stress. Among these programs, pancreatic cancers preferentially utilize a metabolic pathway through the enzyme aspartate aminotransferase 1 [also known as glutamate oxaloacetate transaminase 1 (GOT1)] to support cellular redox homeostasis. As such, small molecule inhibitors that target GOT1 could serve as starting points for the development of new therapies for pancreatic cancer. We ran a high-throughput screen for inhibitors of GOT1 and identified a small molecule, iGOT1-01, with in vitro GOT1 inhibitor activity. Application in pancreatic cancer cells revealed metabolic and growth inhibitory activity reflecting a promiscuous inhibitory profile. We then performed an in silico docking analysis to study inhibitor-GOT1 interactions with iGOT1-01 analogues that possess improved solubility and potency properties. These results suggested that the GOT1 inhibitor competed for binding to the pyridoxal 5-phosphate (PLP) cofactor site of GOT1. To analyze how the GOT1 inhibitor bound to GOT1, a series of GOT1 mutant enzymes that abolished PLP binding were generated. Application of the mutants in X-ray crystallography and thermal shift assays again suggested but were unable to formally conclude that the GOT1 inhibitor bound to the PLP site. Mutational studies revealed the relationship between PLP binding and the thermal stability of GOT1 while highlighting the essential nature of several residues for GOT1 catalytic activity. Insight into the mode of action of GOT1 inhibitors may provide leads to the development of drugs that target redox balance in pancreatic cancer.
Asunto(s)
Aspartato Aminotransferasa Citoplasmática/antagonistas & inhibidores , Proliferación Celular/efectos de los fármacos , Neoplasias del Colon/patología , Inhibidores Enzimáticos/farmacología , Mutación , Neoplasias Pancreáticas/patología , Aspartato Aminotransferasa Citoplasmática/genética , Aspartato Aminotransferasa Citoplasmática/metabolismo , Sitios de Unión , Neoplasias del Colon/tratamiento farmacológico , Neoplasias del Colon/metabolismo , Cristalografía por Rayos X , Inhibidores Enzimáticos/química , Ensayos Analíticos de Alto Rendimiento , Humanos , Metabolómica , Modelos Moleculares , Neoplasias Pancreáticas/tratamiento farmacológico , Neoplasias Pancreáticas/metabolismo , Conformación Proteica , Relación Estructura-Actividad , Células Tumorales CultivadasRESUMEN
Pancreatic ductal adenocarcinoma (PDAC) is a lethal malignancy that is extremely refractory to the therapeutic approaches that have been evaluated to date. Recently, it has been demonstrated that PDAC tumors are dependent upon a metabolic pathway involving aspartate aminotransferase 1, also known as glutamate-oxaloacetate transaminase 1 (GOT1), for the maintenance of redox homeostasis and sustained proliferation. As such, small molecule inhibitors targeting this metabolic pathway may provide a novel therapeutic approach for the treatment of this devastating disease. To this end, from a high throughput screen of â¼800,000 molecules, 4-(1H-indol-4-yl)-N-phenylpiperazine-1-carboxamide was identified as an inhibitor of GOT1. Mouse pharmacokinetic studies revealed that potency, rather than inherent metabolic instability, would limit immediate cell- and rodent xenograft-based experiments aimed at validating this potential cancer metabolism-related target. Medicinal chemistry-based optimization resulted in the identification of multiple derivatives with >10-fold improvements in potency, as well as the identification of a tryptamine-based series of GOT1 inhibitors.
Asunto(s)
Aspartato Aminotransferasas/antagonistas & inhibidores , Inhibidores Enzimáticos/uso terapéutico , Indoles/uso terapéutico , Compuestos de Fenilurea/uso terapéutico , Piperazinas/uso terapéutico , Transaminasas/antagonistas & inhibidores , Animales , Aspartato Aminotransferasa Citoplasmática , Carcinoma Ductal Pancreático/tratamiento farmacológico , Descubrimiento de Drogas , Estabilidad de Medicamentos , Inhibidores Enzimáticos/síntesis química , Inhibidores Enzimáticos/química , Inhibidores Enzimáticos/farmacocinética , Femenino , Indoles/síntesis química , Indoles/química , Indoles/farmacocinética , Ratones , Compuestos de Fenilurea/síntesis química , Compuestos de Fenilurea/química , Compuestos de Fenilurea/farmacocinética , Piperazinas/síntesis química , Piperazinas/química , Piperazinas/farmacocinética , Relación Estructura-ActividadRESUMEN
The amphimedosides, discovered in 2006, are the first examples of naturally occurring glycosylated alkoxyamines. We report syntheses of amphimedosides A-C that feature a stereoselective oxyamine neoglycosylation and found that these alkaloids display modest cytotoxicity toward seven diverse human cancer cell lines, exhibiting IC(50) values ranging from 3.0 µM to greater than 100 µM.
Asunto(s)
Alcaloides/química , Amino Azúcares/síntesis química , Antineoplásicos/síntesis química , Alcaloides/farmacología , Alcaloides/toxicidad , Amino Azúcares/química , Amino Azúcares/farmacología , Antineoplásicos/química , Antineoplásicos/farmacología , Línea Celular , Línea Celular Tumoral , Ensayos de Selección de Medicamentos Antitumorales , Glicosilación , Humanos , Concentración 50 Inhibidora , EstereoisomerismoRESUMEN
The alteration of metabolic pathways is a critical strategy for cancer cells to attain the traits necessary for metastasis in disease progression. Here, we find that dysregulation of propionate metabolism produces a pro-aggressive signature in breast and lung cancer cells, increasing their metastatic potential. This occurs through the downregulation of methylmalonyl coenzyme A epimerase (MCEE), mediated by an extracellular signal-regulated kinase 2-driven transcription factor Sp1/early growth response protein 1 transcriptional switch driven by metastatic signalling at its promoter level. The loss of MCEE results in reduced propionate-driven anaplerotic flux and intracellular and intratumoral accumulation of methylmalonic acid, a by-product of propionate metabolism that promotes cancer cell invasiveness. Altogether, we present a previously uncharacterized dysregulation of propionate metabolism as an important contributor to cancer and a valuable potential target in the therapeutic treatment of metastatic carcinomas.
Asunto(s)
Neoplasias , Propionatos , Humanos , Ácido Metilmalónico/metabolismo , Fenotipo , Propionatos/farmacología , Transducción de SeñalRESUMEN
A significant increase in dietary fructose consumption has been implicated as a potential driver of cancer. Metabolic adaptation of cancer cells to utilize fructose confers advantages for their malignant growth, but compelling therapeutic targets have not been identified. Here, we show that fructose metabolism of leukemic cells can be inhibited by targeting the de novo serine synthesis pathway (SSP). Leukemic cells, unlike their normal counterparts, become significantly dependent on the SSP in fructose-rich conditions as compared to glucose-rich conditions. This metabolic program is mediated by the ratio of redox cofactors, NAD+/NADH, and the increased SSP flux is beneficial for generating alpha-ketoglutarate from glutamine, which allows leukemic cells to proliferate even in the absence of glucose. Inhibition of PHGDH, a rate-limiting enzyme in the SSP, dramatically reduces leukemia engraftment in mice in the presence of high fructose, confirming the essential role of the SSP in the metabolic plasticity of leukemic cells.
Asunto(s)
Fructosa/metabolismo , Leucemia Mieloide Aguda/metabolismo , Serina/biosíntesis , Animales , Humanos , Leucemia Mieloide Aguda/patología , Ratones , Ratones Congénicos , Ratones Endogámicos NOD , Células Tumorales CultivadasRESUMEN
Metastasis is the leading cause of cancer mortality. Chromatin remodeling provides the foundation for the cellular reprogramming necessary to drive metastasis. However, little is known about the nature of this remodeling and its regulation. Here, we show that metastasis-inducing pathways regulate histone chaperones to reduce canonical histone incorporation into chromatin, triggering deposition of H3.3 variant at the promoters of poor-prognosis genes and metastasis-inducing transcription factors. This specific incorporation of H3.3 into chromatin is both necessary and sufficient for the induction of aggressive traits that allow for metastasis formation. Together, our data clearly show incorporation of histone variant H3.3 into chromatin as a major regulator of cell fate during tumorigenesis, and histone chaperones as valuable therapeutic targets for invasive carcinomas.
Asunto(s)
Carcinoma/patología , Cromatina/metabolismo , Regulación Neoplásica de la Expresión Génica , Histonas/metabolismo , Metástasis de la Neoplasia/genética , Animales , Carcinogénesis/genética , Carcinoma/genética , Línea Celular Tumoral , Cromatina/genética , Factor 1 de Ensamblaje de la Cromatina/genética , Factor 1 de Ensamblaje de la Cromatina/metabolismo , Progresión de la Enfermedad , Epigénesis Genética , Transición Epitelial-Mesenquimal/genética , Femenino , Histonas/genética , Humanos , Masculino , Ratones , Regiones Promotoras Genéticas/genética , RNA-Seq , Factores de Transcripción/genética , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Given the putative role of PHGDH in cancer, development of inhibitors is required to explore its function. In this context, we established and validated a straightforward enzymatic assay suitable for high-throughput screening and we identified inhibitors with similar chemical scaffolds. Through a convergent pharmacophore approach, we synthesized α-ketothioamides that exhibit interesting in vitro PHGDH inhibition and encouraging cellular results. These novel probes may be used to understand the emerging biology of this metabolic target.
Asunto(s)
Inhibidores Enzimáticos/química , Inhibidores Enzimáticos/farmacología , Fosfoglicerato-Deshidrogenasa/antagonistas & inhibidores , Tioamidas/química , Tioamidas/farmacología , Antineoplásicos/química , Antineoplásicos/farmacología , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Evaluación Preclínica de Medicamentos , Ensayos Analíticos de Alto Rendimiento , Humanos , Neoplasias/tratamiento farmacológico , Neoplasias/enzimología , Fosfoglicerato-Deshidrogenasa/metabolismoRESUMEN
Serine metabolism is likely to play a critical role in cancer cell growth. A recent study reports the identification of a novel small-molecule inhibitor of serine synthesis that targets 3-phosphoglycerate dehydrogenase (PHGDH), the first enzyme of the serine synthesis pathway, and selectively abrogates the proliferation of PHGDH overexpressing breast cancer cells.
RESUMEN
Tumors have high energetic and anabolic needs for rapid cell growth and proliferation, and the serine biosynthetic pathway was recently identified as an important source of metabolic intermediates for these processes. We integrated metabolic tracing and transcriptional profiling of a large panel of non-small cell lung cancer (NSCLC) cell lines to characterize the activity and regulation of the serine/glycine biosynthetic pathway in NSCLC. Here we show that the activity of this pathway is highly heterogeneous and is regulated by NRF2, a transcription factor frequently deregulated in NSCLC. We found that NRF2 controls the expression of the key serine/glycine biosynthesis enzyme genes PHGDH, PSAT1 and SHMT2 via ATF4 to support glutathione and nucleotide production. Moreover, we show that expression of these genes confers poor prognosis in human NSCLC. Thus, a substantial fraction of human NSCLCs activates an NRF2-dependent transcriptional program that regulates serine and glycine metabolism and is linked to clinical aggressiveness.
Asunto(s)
Adenocarcinoma/metabolismo , Biomarcadores de Tumor/genética , Carcinoma de Pulmón de Células no Pequeñas/metabolismo , Regulación Neoplásica de la Expresión Génica , Neoplasias Pulmonares/metabolismo , Factor 2 Relacionado con NF-E2/genética , Serina/biosíntesis , Adenocarcinoma/genética , Adenocarcinoma/mortalidad , Adenocarcinoma/secundario , Animales , Apoptosis , Biomarcadores de Tumor/metabolismo , Western Blotting , Carcinoma de Pulmón de Células no Pequeñas/genética , Carcinoma de Pulmón de Células no Pequeñas/mortalidad , Carcinoma de Pulmón de Células no Pequeñas/secundario , Proliferación Celular , Humanos , Técnicas para Inmunoenzimas , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/mortalidad , Neoplasias Pulmonares/patología , Ratones , Factor 2 Relacionado con NF-E2/antagonistas & inhibidores , Factor 2 Relacionado con NF-E2/metabolismo , ARN Mensajero/genética , ARN Interferente Pequeño/genética , Reacción en Cadena en Tiempo Real de la Polimerasa , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Células Tumorales Cultivadas , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
More than half of human colorectal cancers (CRCs) carry either KRAS or BRAF mutations and are often refractory to approved targeted therapies. We found that cultured human CRC cells harboring KRAS or BRAF mutations are selectively killed when exposed to high levels of vitamin C. This effect is due to increased uptake of the oxidized form of vitamin C, dehydroascorbate (DHA), via the GLUT1 glucose transporter. Increased DHA uptake causes oxidative stress as intracellular DHA is reduced to vitamin C, depleting glutathione. Thus, reactive oxygen species accumulate and inactivate glyceraldehyde 3-phosphate dehydrogenase (GAPDH). Inhibition of GAPDH in highly glycolytic KRAS or BRAF mutant cells leads to an energetic crisis and cell death not seen in KRAS and BRAF wild-type cells. High-dose vitamin C impairs tumor growth in Apc/Kras(G12D) mutant mice. These results provide a mechanistic rationale for exploring the therapeutic use of vitamin C for CRCs with KRAS or BRAF mutations.
Asunto(s)
Ácido Ascórbico/uso terapéutico , Neoplasias Colorrectales/tratamiento farmacológico , Neoplasias Colorrectales/genética , Proteínas Proto-Oncogénicas B-raf/genética , Proteínas Proto-Oncogénicas/genética , Proteínas ras/genética , Proteína de la Poliposis Adenomatosa del Colon/genética , Animales , Ácido Ascórbico/administración & dosificación , Ácido Ascórbico/farmacología , Línea Celular Tumoral , Ácido Deshidroascórbico/metabolismo , Femenino , Transportador de Glucosa de Tipo 1/metabolismo , Gliceraldehído-3-Fosfato Deshidrogenasa (Fosforilante)/metabolismo , Glucólisis/efectos de los fármacos , Humanos , Ratones , Ratones Mutantes , Ratones Desnudos , Proteínas Proto-Oncogénicas p21(ras)/genética , Especies Reactivas de Oxígeno/metabolismo , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
The metabolic requirements of cancer cells differ from that of their normal counterparts. To support their proliferation, cancer cells switch to a fermentative metabolism that is thought to support biomass production. Instances where metabolic enzymes promote tumorigenesis remain rare. However, an enzyme involved in the de novo synthesis of serine, 3-phosphoglycerate dehydrogenase (PHGDH), was recently identified as a putative oncogene. The potential mechanisms by which PHGDH promotes cancer are discussed.
Asunto(s)
Amplificación de Genes , Glucosa/metabolismo , Melanoma/enzimología , Melanoma/genética , Fosfoglicerato-Deshidrogenasa/genética , Neoplasias Cutáneas/enzimología , Neoplasias Cutáneas/genética , Humanos , Redes y Vías MetabólicasRESUMEN
Most tumors exhibit increased glucose metabolism to lactate, however, the extent to which glucose-derived metabolic fluxes are used for alternative processes is poorly understood. Using a metabolomics approach with isotope labeling, we found that in some cancer cells a relatively large amount of glycolytic carbon is diverted into serine and glycine metabolism through phosphoglycerate dehydrogenase (PHGDH). An analysis of human cancers showed that PHGDH is recurrently amplified in a genomic region of focal copy number gain most commonly found in melanoma. Decreasing PHGDH expression impaired proliferation in amplified cell lines. Increased expression was also associated with breast cancer subtypes, and ectopic expression of PHGDH in mammary epithelial cells disrupted acinar morphogenesis and induced other phenotypic alterations that may predispose cells to transformation. Our findings show that the diversion of glycolytic flux into a specific alternate pathway can be selected during tumor development and may contribute to the pathogenesis of human cancer.