Autonomous metabolic reprogramming and oxidative stress characterize endothelial dysfunction in acute myocardial infarction.

Compelling evidence has accumulated on the role of oxidative stress on the endothelial cell (EC) dysfunction in acute coronary syndrome. Unveiling the underlying metabolic determinants has been hampered by the scarcity of appropriate cell models to address cell-autonomous mechanisms of EC dysfunction. We have generated endothelial cells derived from thrombectomy specimens from patients affected with acute myocardial infarction (AMI) and conducted phenotypical and metabolic characterizations. AMI-derived endothelial cells (AMIECs) display impaired growth, migration, and tubulogenesis. Metabolically, AMIECs displayed augmented ROS and glutathione intracellular content, with a diminished glucose consumption coupled to high lactate production. In AMIECs, while PFKFB3 protein levels of were downregulated, PFKFB4 levels were upregulated, suggesting a shunting of glycolysis towards the pentose phosphate pathway, supported by upregulation of G6PD. Furthermore, the glutaminolytic enzyme GLS was upregulated in AMIECs, providing an explanation for the increase in glutathione content. Finally, AMIECs displayed a significantly higher mitochondrial membrane potential than control ECs, which, together with high ROS levels, suggests a coupled mitochondrial activity. We suggest that high mitochondrial proton coupling underlies the high production of ROS, balanced by PPP- and glutaminolysis-driven synthesis of glutathione, as a primary, cell-autonomous abnormality driving EC dysfunction in AMI.

Integrating systemic and molecular levels to infer key drivers sustaining metabolic adaptations.

Metabolic adaptations to complex perturbations, like the response to pharmacological treatments in multifactorial diseases such as cancer, can be described through measurements of part of the fluxes and concentrations at the systemic level and individual transporter and enzyme activities at the molecular level. In the framework of Metabolic Control Analysis (MCA), ensembles of linear constraints can be built integrating these measurements at both systemic and molecular levels, which are expressed as relative differences or changes produced in the metabolic adaptation. Here, combining MCA with Linear Programming, an efficient computational strategy is developed to infer additional non-measured changes at the molecular level that are required to satisfy these constraints. An application of this strategy is illustrated by using a set of fluxes, concentrations, and differentially expressed genes that characterize the response to cyclin-dependent kinases 4 and 6 inhibition in colon cancer cells. Decreases and increases in transporter and enzyme individual activities required to reprogram the measured changes in fluxes and concentrations are compared with down-regulated and up-regulated metabolic genes to unveil those that are key molecular drivers of the metabolic response.

Quantitative Proteomic Approach Reveals Altered Metabolic Pathways in Response to the Inhibition of Lysine Deacetylases in A549 Cells under Normoxia and Hypoxia.

Growing evidence is showing that acetylation plays an essential role in cancer, but studies on the impact of KDAC inhibition (KDACi) on the metabolic profile are still in their infancy. Here, we analyzed, by using an iTRAQ-based quantitative proteomics approach, the changes in the proteome of KRAS-mutated non-small cell lung cancer (NSCLC) A549 cells in response to trichostatin-A (TSA) and nicotinamide (NAM) under normoxia and hypoxia. Part of this response was further validated by molecular and biochemical analyses and correlated with the proliferation rates, apoptotic cell death, and activation of ROS scavenging mechanisms in opposition to the ROS production. Despite the differences among the KDAC inhibitors, up-regulation of glycolysis, TCA cycle, oxidative phosphorylation and fatty acid synthesis emerged as a common metabolic response underlying KDACi. We also observed that some of the KDACi effects at metabolic levels are enhanced under hypoxia. Furthermore, we used a drug repositioning machine learning approach to list candidate metabolic therapeutic agents for KRAS mutated NSCLC. Together, these results allow us to better understand the metabolic regulations underlying KDACi in NSCLC, taking into account the microenvironment of tumors related to hypoxia, and bring new insights for the future rational design of new therapies.

Targeting the Metabolic Adaptation of Metastatic Cancer.

Metabolic adaptation is emerging as an important hallmark of cancer and metastasis. In the last decade, increasing evidence has shown the importance of metabolic alterations underlying the metastatic process, especially in breast cancer metastasis but also in colorectal cancer metastasis. Being the main cause of cancer-related deaths, it is of great importance to developing new therapeutic strategies that specifically target metastatic cells. In this regard, targeting metabolic pathways of metastatic cells is one of the more promising windows for new therapies of metastatic colorectal cancer, where still there are no approved inhibitors against metabolic targets. In this study, we review the recent advances in the field of metabolic adaptation of cancer metastasis, focusing our attention on colorectal cancer. In addition, we also review the current status of metabolic inhibitors for cancer treatment.

Cysteine and Folate Metabolism Are Targetable Vulnerabilities of Metastatic Colorectal Cancer.

With most cancer-related deaths resulting from metastasis, the development of new therapeutic approaches against metastatic colorectal cancer (mCRC) is essential to increasing patient survival. The metabolic adaptations that support mCRC remain undefined and their elucidation is crucial to identify potential therapeutic targets. Here, we employed a strategy for the rational identification of targetable metabolic vulnerabilities. This strategy involved first a thorough metabolic characterisation of same-patient-derived cell lines from primary colon adenocarcinoma (SW480), its lymph node metastasis (SW620) and a liver metastatic derivative (SW620-LiM2), and second, using a novel multi-omics integration workflow, identification of metabolic vulnerabilities specific to the metastatic cell lines. We discovered that the metastatic cell lines are selectively vulnerable to the inhibition of cystine import and folate metabolism, two key pathways in redox homeostasis. Specifically, we identified the system xCT and MTHFD1 genes as potential therapeutic targets, both individually and combined, for combating mCRC.

Software Supporting a Workflow of Quantitative Dynamic Flux Maps Estimation in Central Metabolism from SIRM Experimental Data.

Stable isotope-resolved metabolomics (SIRM), based on the analysis of biological samples from living cells incubated with artificial isotope enriched substrates, enables mapping the rates of biochemical reactions (metabolic fluxes). We developed software supporting a workflow of analysis of SIRM data obtained with mass spectrometry (MS). The evaluation of fluxes starting from raw MS recordings requires at least three steps of computer support: first, extraction of mass spectra of metabolites of interest, then correction of the spectra for natural isotope abundance, and finally, evaluation of fluxes by simulation of the corrected spectra using a corresponding mathematical model. A kinetic model based on ordinary differential equations (ODEs) for isotopomers of metabolites of the corresponding biochemical network supports the final part of the analysis, which provides a dynamic flux map.

De novo MYC addiction as an adaptive response of cancer cells to CDK4/6 inhibition.

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 HIF-1α-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.

Methylseleninic acid promotes antitumour effects via nuclear FOXO3a translocation through Akt inhibition.

Selenium supplement has been shown in clinical trials to reduce the risk of different cancers including lung carcinoma. Previous studies reported that the antiproliferative and pro-apoptotic activities of methylseleninic acid (MSA) in cancer cells could be mediated by inhibition of the PI3K pathway. A better understanding of the downstream cellular targets of MSA will provide information on its mechanism of action and will help to optimize its use in combination therapies with PI3K inhibitors. For this study, the effects of MSA on viability, cell cycle, metabolism, apoptosis, protein and mRNA expression, and reactive oxygen species production were analysed in A549 cells. FOXO3a subcellular localization was examined in A549 cells and in stably transfected human osteosarcoma U2foxRELOC cells. Our results demonstrate that MSA induces FOXO3a nuclear translocation in A549 cells and in U2OS cells that stably express GFP-FOXO3a. Interestingly, sodium selenite, another selenium compound, did not induce any significant effects on FOXO3a translocation despite inducing apoptosis. Single strand break of DNA, disruption of tumour cell metabolic adaptations, decrease in ROS production, and cell cycle arrest in G1 accompanied by induction of apoptosis are late events occurring after 24h of MSA treatment in A549 cells. Our findings suggest that FOXO3a is a relevant mediator of the antiproliferative effects of MSA. This new evidence on the mechanistic action of MSA can open new avenues in exploiting its antitumour properties and in the optimal design of novel combination therapies. We present MSA as a promising chemotherapeutic agent with synergistic antiproliferative effects with cisplatin.