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Rhabdomyolysis is a disorder characterized by acute damage of the sarcolemma of the skeletal muscle leading to release of potentially toxic muscle cell components into the circulation, most notably creatine phosphokinase (CK) and myoglobulin, and is frequently accompanied by myoglobinuria. In the present work, we evaluated the toxicity of p-phenylenediamine (PPD), a main component of hair dyes which is reported to induce rhabdomyolysis. We studied the metabolic effect of this compound in vivo with Wistar rats and in vitro with C2C12 muscle cells. To this aim we have combined multi-omic experimental measurements with computational approaches using model-driven methods. The integrative study presented here has unveiled the metabolic disorders associated to PPD exposure that may underlay the aberrant metabolism observed in rhabdomyolys disease. Animals treated with lower doses of PPD (10 and 20 mg/kg) showed depressed activity and myoglobinuria after 10 h of treatment. We measured the serum levels of aspartate aminotransferase (AST), alanine aminotransferase (ALT), and creatine kinase (CK) in rats after 24, 48, and 72 h of PPD exposure. At all times, treatment with PPD at higher doses (40 and 60 mg/kg) showed an increase of AST and ALT, and also an increase of lactate dehydrogenase (LDH) and CK after 24 h. Blood packed cell volume and hemoglobin levels, as well as organs weight at 48 and 72 h, were also measured. No significant differences were observed in these parameters under any condition. PPD induce cell cycle arrest in S phase and apoptosis (40% or early apoptotic cells) on mus musculus mouse C2C12 cells after 24 h of treatment. Incubation of mus musculus mouse C2C12 cells with [1,2-13C2]-glucose during 24 h, subsequent quantification of 13C isotopologues distribution in key metabolites of glucose metabolic network and a computational fluxomic analysis using in-house developed software (Isodyn) showed that PPD is inhibiting glycolysis, non-oxidative pentose phosphate pathway, glycogen turnover, and ATPAse reaction leading to a reduction in ATP synthesis. These findings unveil the glucose metabolism collapse, which is consistent with a decrease in cell viability observed in PPD-treated C2C12 cells and with the myoglubinuria and other effects observed in Wistar Rats treated with PPD. These findings shed new light on muscle dysfunction associated to PPD exposure, opening new avenues for cost-effective therapies in Rhabdomyolysis disease. ; This work has received financial support from the government of Catalunya: ITT program of the Work Community of PyreneesAgència de Gestió d'Ajuts Universitaris i de Recerca (AGAUR)—Generalitat de Catalunya (2014SGR1017). MC acknowledges the support received through the prize "ICREA Academia" for excellence in research, funded by ICREA foundation—Generalitat de Catalunya. IM and RT acknowledge the support received through the European Research Council under the European Union's Seventh Framework Programme (FP/2007–2013)/ERC Grant Agreement no. 320737. ; Peer reviewed ; Peer Reviewed

Metabolic reprogramming, a crucial cancer hallmark, shifts metabolic pathways such as glycolysis, tricarboxylic acid cycle or lipogenesis, to enable the growth characteristics of cancer cells. Here, we provide evidence that transketolase-like 1 (TKTL1) orchestrates aerobic glycolysis, fatty acid and nucleic acid synthesis, glutamine metabolism, protection against oxidative stress and cell proliferation. Furthermore, silencing of TKTL1 reduced the levels of sphingolipids such as lactosylceramide (a sphingolipid regulating cell survival, proliferation and angiogenesis) and phosphatidylinositol (which activates PI3K/Akt/mTOR signaling). Thus, in addition to its well-known roles in glucose and amino acid metabolism, TKTL1 also regulates lipid metabolism. In conclusion, our study provides unprecedented evidence that TKTL1 plays central roles in major metabolic processes subject to reprogramming in cancer cells and thus identifies TKTL1 as a promising target for new anti-cancer therapies. ; This work was supported by grants to M.C. (Agència de Gestió d'Ajuts Universitaris i de Recerca (AGAUR) – Generalitat de Catalunya, 2014SGR1017; ICREA Academia from the ICREA foundation; and MINECO-European Commission FEDER – Una manera de hacer Europa, SAF2014-56059-R), T.M.T. (MINECO-European Commission FEDER – Una manera de hacer Europa, SAF2015-66984-C2-1-R; and Xarxa de Referència en Biotecnologia) and P.C. (Research Foundation-Flanders (FWO), Belgian Science Policy grant (IUAP P7/03); long-term structural Methusalem funding by the Flemish Government; European Research Council (ERC) Advanced Research Grant (EU-ERC269073); and AXA Research Fund). ; Peer Reviewed

5 pages, 1 table. ; Natural polyphenolic compounds from plants and fruits have been reported to be potent antioxidants and to present antitumoral properties. The difficulty to clearly identify structure/antitumoral activity of this family of compounds makes difficult to optimize the formulation of combinations of polyphenolic extracts from different plants or fruits to achieve the best antitumoral potency. Recent advances have been achieved in this field from the exhaustive characterization of polyphenolic extracts with different degree of polymerization and percentage of galloylation. Here we review the results obtained in this field from our team (Lizarraga et al., 2007; Lozano et al., 2006; Lozano et al., 2005; Matito et al., 2003; Tourino et al., 2008; Tourino et al., 2005) that relate degree of polymerization and galloylation to radical scavenging capacity, tumor cell proliferation inhibition and induction of apoptosis. From this review we concluded that natural polyphenolic extract with high degree of polymerization and galloylation will present the higher antiproliferative and induction of apoptosis capacities. ; This work was supported by Spanish Government and the European Union FEDER funds (SAF2005-01627 and AGL2004-07579-C04-03/ALI) and ISCIII-RTICC (RD06/0020/0046); Generalitat de Catalunya (2005SGR00204; 2006ITT-10007). ; Peer reviewed

[Background] Genome-scale metabolic models (GSMM) integrating transcriptomics have been widely used to study cancer metabolism. This integration is achieved through logical rules that describe the association between genes, proteins, and reactions (GPRs). However, current gene-to-reaction formulation lacks the stoichiometry describing the transcript copies necessary to generate an active catalytic unit, which limits our understanding of how genes modulate metabolism. The present work introduces a new state-of-the-art GPR formulation that considers the stoichiometry of the transcripts (S-GPR). As case of concept, this novel gene-to-reaction formulation was applied to investigate the metabolic effects of the chronic exposure to Aldrin, an endocrine disruptor, on DU145 prostate cancer cells. To this aim we integrated the transcriptomic data from Aldrin-exposed and non-exposed DU145 cells through S-GPR or GPR into a human GSMM by applying different constraint-based-methods. ; [Results] Our study revealed a significant improvement of metabolite consumption/production predictions when S-GPRs are implemented. Furthermore, our computational analysis unveiled important alterations in carnitine shuttle and prostaglandine biosynthesis in Aldrin-exposed DU145 cells that is supported by bibliographic evidences of enhanced malignant phenotype. ; [Conclusions] The method developed in this work enables a more accurate integration of gene expression data into model-driven methods. Thus, the presented approach is conceptually new and paves the way for more in-depth studies of aberrant cancer metabolism and other diseases with strong metabolic component with important environmental and clinical implications. ; The research leading to these results has received funding from the European Research Council under the European Union's Seventh Framework Programme (FP/2007–2013) / ERC Grant Agreement n. 320737 and from the Novo Nordisk Foundation (NNF10CC1016517 and NNF14OC0009473). MC is funded by ICREA Academia programme-2015 (Icrea Fundation), AGAUR-Generalitat de Catalunya (2017SGR-1033), MINECO European Commission FEDER funds (SAF2017-89673-R) and Instituto de Salud Carlos III (CIBEREHD CB17/04/00023). ; Peer reviewed

Carbohydrate response element-binding protein (ChREBP) is a transcription factor that mediates glucose signaling in mammalian liver, leading to the expression of different glycolytic and lipogenic genes, such as pyruvate kinase (L-PK) and fatty acid synthase (FAS). The current model for ChREBP activation in response to sugar phosphates holds that glucose metabolization to xylulose 5-phosphate (X-5-P) triggers the activation of protein phosphatase 2A, which dephosphorylates ChREBP and leads to its nuclear translocation and activation. However, evidence indicates that glucose 6-phosphate (G-6-P) is the most likely signal metabolite for the glucose-induced transcription of these genes. The glucose derivative that is responsible for carbohydrate-dependent gene expression remains to be identified. The difficulties in measuring G-6-P and X-5-P concentrations simultaneously and in changing them independently have hindered such identification. To discriminate between these possibilities, we adapted a liquid chromatography mass spectrometry method to identify and quantify sugar phosphates in human hepatocarcinoma cells (Hep G2) and rat hepatocytes in response to different carbon sources and in the presence/absence of a glucose-6-phosphate dehydrogenase inhibitor. We also used this method to demonstrate that these cells could not metabolize 2-deoxyglucose beyond 2-deoxyglucose-6-phosphate. The simultaneous quantification of sugar phosphates and FAS and L-PK expression levels demonstrated that both X-5-P and G-6-P play a role in the modulation of gene expression. In conclusion, this report presents for the first time a single mechanism that incorporates the effects of X-5-P and G-6-P on the enhancement of the expression of carbohydrate-responsive genes. © 2012 the American Physiological Society. ; This study was supported by the projects SAF2009-12602 and SAF2011-25726 and by RD06/0020/0046 and RD06/0014/0025 from Red Temática de Investigación Cooperativa en Cáncer and Red de Centros FIS-RECAVA, respectively, from the Instituto de Salud Carlos III, both funded by the Ministerio de Ciencia e Innovación-Spanish government and European Regional Development Funds "Una manera de hacer Europa." It has also received financial support from the European Union-funded project ETHERPATHS (FP7-KBBE-222639) (http://www.etherpaths.org/) and from the Agència de Gestió d'Ajuts Universitaris i de Recerca-Generalitat de Catalunya (2009SGR01308, 2006ITT-10007, and 2009CTP-00026). ; Peer Reviewed

Cyclin-dependent kinases (CDK) are rational cancer therapeutic targets fraught with the development of acquired resistance by tumor cells. Through metabolic and transcriptomic analyses, we show that the inhibition of CDK4/6 leads to a metabolic reprogramming associated with gene networks orchestrated by the MYC transcription factor. Upon inhibition of CDK4/6, an accumulation of MYC protein ensues which explains an increased glutamine metabolism, activation of the mTOR pathway and blunting of HIF1a-mediated responses to hypoxia. These MYC-driven adaptations to CDK4/6 inhibition render cancer cells highly sensitive to inhibitors of MYC, glutaminase or mTOR and to hypoxia, demonstrating that metabolic adaptations to antiproliferative drugs unveil new vulnerabilities that can be exploited to overcome acquired drug tolerance and resistance by cancer cells ; This work was supported by grants to M.C. from Agència de Gestió d'Ajuts Universitaris i de Recerca (AGAUR)—Generalitat de Catalunya, (2014SGR1017), ICREA Foundation (ICREA Academia) and MINECO from the Spanish Government and FEDER funds (SAF2014-56059-R, SAF2015-70270- REDT, European Commission FEDER—Una manera de hacer Europa), and to T.M.T. from MINECO (SAF2012-40017-C02-01 and SAF2015-66984-C2-1-R, European Commission FEDER—Una manera de hacer Europa) and Xarxa de Referència en Biotecnologia. This work was also supported by the Francis Crick Institute which receives its core funding from Cancer Research UK (FC001223), the UK Medical Research Council (FC001223), and the Wellcome Trust (FC001223). The CRG/UPF Proteomics Unit is part of the "Plataforma de Recursos Biomoleculares y Bioinformáticos (ProteoRed)" supported by grant PT13/ 0001 of ISCIII and Spanish Ministry of Economy and Competitiveness. ; Peer reviewed

El pdf del artículo es la versión post-print. ; Macrophages play a relevant role ininnate and adaptive immunity depending on the balance of the stimuli received. From an analytical and functional point of view, macrophage stimulation can be segregated into three main modes, as follows: innate, classic, and alternative pathways. These differential activations result in the expression of specific sets of genes involved in the release of pro-or anti-inflammatory stimuli. In the present work, we have analyzed whether specific metabolic patterns depend on the signaling pathway activated. A[1,2-13C2]glucose tracer-based metabolomics approach has been used to characterize the metabolic flux distributions in macrophages stimulated through the classic, innate, and alternative pathways. Using this methodology combined with mass isotopomer distribution analysis of the new formed metabolites, the data show that activated macrophages are essentially glycolytic cells, and a clear cut off between the classic/innate activation and the alternative pathway exists. Interestingly, macrophage activation through LPS/IFN-γ or TLR-2, -3, -4, and -9 results in similar flux distribution patterns regardless of the pathway activated. However, stimulation through the alternative pathway has minor metabolic effects. Themolecular basis of the differences between these two types of behavior involves a switch in the expression of 6-phosphofructo-2-kinase/fructose-2,6- bisphosphatase (PFK2) from the liver type-PFK2 to the more active ubiquitous PFK2 isoenzyme, which responds to Hif-1α activation and increases fructose-2,6-bisphosphate concentration and the glycolytic flux. However, using macrophages targeted for Hif-1α, the switch of PFK2 isoenzymes still occurs in LPS/IFN-γ-activated macrophages, suggesting that this pathway regulates ubiquitous PFK2 expression through Hif-1a-independent mechanisms. Copyright © 2010 by The American Association of Immunologists, Inc. ; This work was supported by European Commission (FP7) Etherpath KBBE-Grant Agreement no. 222639, Grants SAF2005-01627, SAF2008-00164, and BFU2008- 02161 from Ministerio de Ciencia e Innovacio´n of the Spanish Government, 2005SGR00204 from the Generalitat de Catalunya, S-BIO-0283/2006 from Comunidad de Madrid, Red Temática de Investigación Sobre Cáncer (RETICC RD6/0020/ 0046) and Red Temática de Investigación Sobre Enfermedades Cardiovasculares (RECAVA RD6/0014/006), and FIS-RECAVA RD06/0014/0025. RECAVA and Ciberehd are funded by Instituto de Salud Carlos III. J.-C.R.-P. is supported by a fellowship from the University of Barcelona. ; Peer Reviewed

[Abstract] Epithelial-mesenchymal-transition promotes intra-tumoral heterogeneity, by enhancing tumor cell invasiveness and promoting drug resistance. We integrated transcriptomic data for two clonal subpopulations from a prostate cancer cell line (PC-3) into a genome-scale metabolic network model to explore their metabolic differences and potential vulnerabilities. In this dual cell model, PC-3/S cells express Epithelial-mesenchymal transition markers and display high invasiveness and low metastatic potential, while PC-3/M cells present the opposite phenotype and higher proliferative rate. Model-driven analysis and experimental validations unveiled a marked metabolic reprogramming in long-chain fatty acids metabolism. While PC-3/M cells showed an enhanced entry of long-chain fatty acids into the mitochondria, PC-3/S cells used long-chain fatty acids as precursors of eicosanoid metabolism. We suggest that this metabolic reprogramming endows PC-3/M cells with augmented energy metabolism for fast proliferation and PC-3/S cells with increased eicosanoid production impacting angiogenesis, cell adhesion and invasion. PC-3/S metabolism also promotes the accumulation of docosahexaenoic acid, a long-chain fatty acid with antiproliferative effects. The potential therapeutic significance of our model was supported by a differential sensitivity of PC-3/M cells to etomoxir, an inhibitor of long-chain fatty acid transport to the mitochondria ; [Author summary] The coexistence within the same tumor of a variety of subpopulations, featuring different phenotypes (intra-tumoral heterogeneity) represents a challenge for diagnosis, prognosis and targeted therapies. In this work, we have explored the metabolic differences underlying tumor heterogeneity by building cell-type-specific genome-scale metabolic models that integrate transcriptome and metabolome data of two clonal subpopulations derived from the same prostate cancer cell line (PC-3). These subpopulations display either highly proliferative, cancer stem cell (PC-3/M) or highly invasive, epithelial-mesenchymal-transition-like phenotypes (PC-3/S). Our model-driven analysis and experimental validations have unveiled a differential utilization of the long-chain fatty acids pool in both subpopulations. More specifically, our findings show an enhanced entry of long-chain fatty acids into the mitochondria in PC-3/M cells, while in PC-3/S cells, long-chain fatty acids are used as precursors of eicosanoid metabolism. The different utilization of long-chain fatty acids between subpopulations endows PC-3/M cells with a highly proliferative phenotype while enhances PC-3/S invasive phenotype. The present work provides a tool to unveil key metabolic nodes associated with tumor heterogeneity and highlights potential subpopulation-specific targets with important therapeutic implications ; This work was supported by the European Commission Seventh Framework Programme FP7 (METAFLUX-Marie Curie FP7-PEOPLE-2010 ITN-264780); the Spanish Government and the European Union FEDER funds (SAF2014-56059-R, SAF2015-70270-REDT and SAF2015-66984-C2-1-R); Generalitat de Catalunya-AGAUR (2014SGR1017, xarxa de referència en biotecnologia, (TMT) and the support received through the prize "ICREA Academia" for excellence in research (MC), the Lendület Program of the Hungarian Academy of Sciences (BP) and the Welcome Trust (BP) ; Peer reviewed

INTRODUCTION: Non-small cell lung cancer (NSCLC) patients diagnosed in early stage and surgically-treated have five-year mortality rate >20%. The identification of biomarkers able to predict progression and death may help to identify patients needing closer follow-up. METHODS: A retrospective cohort of early-stage surgically-treated NSCLC patients enrolled in the International Association for the Study of Lung Cancer (IASLC) Staging Project was created, and tissue Microarrays (TMAs) were constructed with tumor and non-tumor lung tissue. Pentose phosphate pathway (PPP) proteins (transketolase [TKT] and transketolase-like 1 [TKTL1]), inflammatory markers (cyclooxygenase-2 [COX-2], tumor necrosis factor alpha [TNF-α], interleukin 1 beta [IL1β], nuclear factor kappa-light-chain-enhancer of activated B cells [NFκB]-p65 and antigen Ki-67), and programmed death-ligand 1 (PDL1) were measured by immunohistochemistry. RESULTS: NSCLC patients with adenocarcinoma (ADC) or squamous cell carcinoma (SCC) were included in the study (n = 199). TKT and TKTL1 were significantly higher in ADC than in non-tumor tissue (p 30% above those of non-tumor tissue (p < 0.001). PDL1 analysis showed a higher percentage of positivity in ADC than in non-tumor tissue (p < 0.001). Multivariate Cox proportional hazards modeling confirmed that high IL1β level in tumor tissue was independently associated with 3-year mortality in NSCLC [HR = 2.05, 95% CI (1.1-3.7), p = 0.019], a relationship driven by ADC subtype. CONCLUSION: This study confirms an increase in metabolic activity and an inflammatory response in tumor tissue of early stage NSCLC, and a significant relationship between high levels of IL1β in the tumor and poor prognosis in ADC. ; This study was funded by PII Oncología Torácica from Sociedad Española de Neumología y Cirugía Torácica (SEPAR), FIS 12/02040, FIS 12/02534 and FIS 12/01310 from the Instituto de Salud Carlos III and Centro de Investigación Biomédica en Red de Enfermedades Respiratorias (CIBERES). CIBERES is an initiative of the Instituto de Salud Carlos III. This work was also supported by grants to E.M. and M.C. from Agència de Gestió d'Ajuts Universitaris i de Recerca (AGAUR)—Generalitat de Catalunya, (2014 GRC-801/2017 GRC-716 and 2014 SGR-1017/2017 SGR-1033), ICREA Foundation (ICREA Academia) and MINECO from the Spanish Government and FEDER funds (SAF2014‐56059‐R, SAF2015‐70270‐REDT, SAF2017-89673-R, European Commission FEDER—Una manera de hacer Europa), and to C.B. from AGAUR-Generalitat de Catalunya (FI-DGR2014). M.C. also received support from CB17/04/00023 from the Instituto de Salud Carlos III and Centro de Investigación Biomédica en Red de Enfermedades Hepáticas y Digestivas (CIBERHD). CIBERHD is an initiative of the Instituto de Salud Carlos III. Financial support was also provided by the European Regional Development Fund (ERDF) and CERCA Programme / Generalitat de Catalunya.