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International audience; Metabolic reprogramming is a common hallmark of cancer, but a large variability in tumor bioenergetics exists between patients. Using high-resolution respirometry on fresh biopsies of human lung adenocarcinoma, we identified 2 subgroups reflected in the histologically normal, paired, cancer-adjacent tissue: high (OX+) mitochondrial respiration and low (OX-) mitochondrial respiration. The OX+ tumors poorly incorporated [F-18]fluorodeoxy-glucose and showed increased expression of the mitochondrial trifunctional fatty acid oxidation enzyme (MTP; HADHA) compared with the paired adjacent tissue. Genetic inhibition of MTP altered OX+ tumor growth in vivo. Trimetazidine, an approved drug inhibitor of MTP used in cardiology, also reduced tumor growth and induced disruption of the physical interaction between the MTP and respiratory chain complex I, leading to a cellular redox and energy crisis. MTP expression in tumors was assessed using histology scoring methods and varied in negative correlation with [F-18]fluorodeoxy-glucose incorporation. These findings provide proof-of-concept data for preclinical, precision, bioenergetic medicine in oxidative lung carcinomas.

International audience; Human γδ T cells contribute to tissue homeostasis and participate in epithelial stress surveillance through mechanisms that are not well understood. Here, we identified ephrin type-A receptor 2 (EphA2) as a stress antigen recognized by a human Vγ9Vδ1 TCR. EphA2 is recognized coordinately by ephrin A to enable γδ TCR activation. We identified a putative TCR binding site on the ligand-binding domain of EphA2 that was distinct from the ephrin A binding site. Expression of EphA2 was up-regulated upon AMP-activated protein kinase (AMPK)-dependent metabolic reprogramming of cancer cells, and coexpression of EphA2 and active AMPK in tumors was associated with higher CD3 T cell infiltration in human colorectal cancer tissue. These results highlight the potential of the human γδ TCR to cooperate with a co-receptor to recognize non-MHC-encoded proteins as signals of cellular dysregulation, potentially allowing γδ T cells to sense metabolic energy changes associated with either viral infection or cancer.

Mitochondrial dysfunction in the spinal cord is a hallmark of amyotrophic lateral sclerosis (ALS), but the neurometabolic alterations during early stages of the disease remain unknown. Here, we investigated the bioenergetic and proteomic changes in ALS mouse motor neurons and patients’ skin fibroblasts. We first observed that SODG93A mice presymptomatic motor neurons display alterations in the coupling efficiency of oxidative phosphorylation, along with fragmentation of the mitochondrial network. The proteome of presymptomatic ALS mice motor neurons also revealed a peculiar metabolic signature with upregulation of most energy-transducing enzymes, including the fatty acid oxidation (FAO) and the ketogenic components HADHA and ACAT2, respectively. Accordingly, FAO inhibition altered cell viability specifically in ALS mice motor neurons, while uncoupling protein 2 (UCP2) inhibition recovered cellular ATP levels and mitochondrial network morphology. These findings suggest a novel hypothesis of ALS bioenergetics linking FAO and UCP2. Lastly, we provide a unique set of data comparing the molecular alterations found in human ALS patients’ skin fibroblasts and SODG93A mouse motor neurons, revealing conserved changes in protein translation, folding and assembly, tRNA aminoacylation and cell adhesion processes.

The search for new drugs capable of blocking the metabolic vulnerabilities of human tumors has now entered the clinical evaluation stage, but several projects already failed in phase I or phase II. In particular, very promising in vitro studies could not be translated in vivo at preclinical stage and beyond. This was the case for most glycolysis inhibitors that demonstrated systemic toxicity. A more recent example is the inhibition of glutamine catabolism in lung adenocarcinoma that failed in vivo despite a strong addiction of several cancer cell lines to glutamine in vitro. Such contradictory findings raised several questions concerning the optimization of drug discovery strategies in the field of cancer metabolism. For instance, the cell culture models in 2D or 3D might already show strong limitations to mimic the tumor micro- and macro-environment. The microenvironment of tumors is composed of cancer cells of variegated metabolic profiles, supporting local metabolic exchanges and symbiosis, but also of immune cells and stroma that further interact with and reshape cancer cell metabolism. The macroenvironment includes the different tissues of the organism, capable of exchanging signals and fueling the tumor 'a distance'. Moreover, most metabolic targets were identified from their increased expression in tumor transcriptomic studies, or from targeted analyses looking at the metabolic impact of particular oncogenes or tumor suppressors on selected metabolic pathways. Still, very few targets were identified from in vivo analyses of tumor metabolism in patients because such studies are difficult and adequate imaging methods are only currently being developed for that purpose. For instance, perfusion of patients with [13C]-glucose allows deciphering the metabolomics of tumors and opens a new area in the search for effective targets. Metabolic imaging with positron emission tomography and other techniques that do not involve [13C] can also be used to evaluate tumor metabolism and to follow the efficiency of a treatment at a preclinical or clinical stage. Relevant descriptors of tumor metabolism are now required to better stratify patients for the development of personalized metabolic medicine. In this review, we discuss the current limitations in basic research and drug discovery in the field of cancer metabolism to foster the need for more clinically relevant target identification and validation. We discuss the design of adapted drug screening assays and compound efficacy evaluation methods for the discovery of innovative anti-cancer therapeutic approaches at the level of tumor energetics. This article is part of a Special Issue entitled Mitochondria in Cancer, edited by Giuseppe Gasparre, Rodrigue Rossignol and Pierre Sonveaux.

In this review we discuss the structure and functions of the aspartate/glutamate carriers (AGC1-aralar and AGC2-citrin). Those proteins supply the aspartate synthesized within mitochondrial matrix to the cytosol in exchange for glutamate and a proton. A structure of an AGC carrier is not available yet but comparative 3D models were proposed. Moreover, transport assays performed by using the recombinant AGC1 and AGC2, reconstituted into liposome vesicles, allowed to explore the kinetics of those carriers and to reveal their specific transport properties. AGCs participate to a wide range of cellular functions, as the control of mitochondrial respiration, calcium signaling and antioxydant defenses. AGC1 might also play peculiar tissue-specific functions, as it was found to participate to cell-to-cell metabolic symbiosis in the retina. On the other hand, AGC1 is involved in the glutamate-mediated excitotoxicity in neurons and AGC gene or protein alterations were discovered in rare human diseases. Accordingly, a mice model of AGC1 gene knock-out presented with growth delay and generalized tremor, with myelinisation defects. More recently, AGC was proposed to play a crucial role in tumor metabolism as observed from metabolomic studies showing that the asparate exported from the mitochondrion by AGC1 is employed in the regeneration of cytosolic glutathione. Therefore, given the central role of AGCs in cell metabolism and human pathology, drug screening are now being developed to identify pharmacological modulators of those carriers. This article is part of a Special Issue entitled: Mitochondrial Channels edited by Pierre Sonveaux, Pierre Maechler and Jean-Claude Martinou.

Resume Les cytopathies mitochondriales sont des maladies rares pour lesquelles les enfants atteints sont soumis a des anesthesies repetees, dans le cadre d’une prise en charge de leur pathologie ou de l’urgence. Les agents de l’anesthesie presentent de nombreuses interactions avec le metabolisme mitochondrial, notamment les anesthesiques locaux, les agents halogenes et le propofol. De ce fait, il parait essentiel de parfaitement connaitre les malades suspects des la consultation d’anesthesie afin d’optimiser les protocoles d’anesthesie et de prevenir les complications perioperatoire. Les decompensations metaboliques a type d’acidose lactique, cardiorespiratoires et neurologiques sont a surveiller.

Tumours display different cell populations with distinct metabolic phenotypes. Thus, subpopulations can adjust to different environments, particularly with regard to oxygen and nutrient availability. Our results indicate that progression to metastasis requires mitochondrial function. Our research, centered on cell lines that display increasing degrees of malignancy, focused on metabolic events, especially those involving mitochondria, which could reveal which stages are mechanistically associated with metastasis. Melanocytes were subjected to several cycles of adhesion impairment, producing stable cell lines exhibiting phenotypes representing a progression from non-tumorigenic to metastatic cells. Metastatic cells (4C11+) released the highest amounts of lactate, part of which was derived from glutamine catabolism. The 4C11+ cells also displayed an increased oxidative metabolism, accompanied by enhanced rates of oxygen consumption coupled to ATP synthesis. Enhanced mitochondrial function could not be explained by an increase in mitochondrial content or mitochondrial biogenesis. Furthermore, 4C11+ cells had a higher ATP content, and increased succinate oxidation (complex II activity) and fatty acid oxidation. In addition, 4C11+ cells exhibited a 2-fold increase in mitochondrial membrane potential (ΔΨmit). Consistently, functional assays showed that the migration of cells depended on glutaminase activity. Metabolomic analysis revealed that 4C11+ cells could be grouped as a subpopulation with a profile that was quite distinct from the other cells investigated in the present study. The results presented here have centred on how the multiple metabolic inputs of tumour cells may converge to compose the so-called metastatic phenotype.

Human tissues differ by the type and the amount of energy substrate that they use and store, as dictated by their genetically programmed “metabolic rigidity” which defines the expression of tissue-specific catabolic and anabolic enzymes. For instance, adipocytes and neurons cannot perform fat oxidation, while cardiomyocytes utilize mainly fatty acids for ATP synthesis. These programs evolve during embryogenesis, with a typical burst of oxidative metabolism at birth. Bioenergetic homeostasis is regulated at the cellular level by effectors such as insulin and at the organismal level by the metabolic coupling between “energy donor” tissues, as the adipose tissue and the liver, and “receivers” and the muscle and the brain. In contrast to normal tissues, most tumors develop a dramatic “metabolic flexibility,” which allows them to consume whatever energy substrate is available. Yet, recent findings suggest that this flexibility is restricted by specific anabolic needs, so that alternative energy-transducing pathways are not selected randomly. Cancer cells also avoid the metabolic control by insulin, growth factors, and energy substrate starvation or excess by typical mutations in the MAPK, PI3K, RAS, and related pathways. Metabolic flexibility is made possible by the molecular rewiring of several metabolic pathways, which undergo branching, truncation, and reversal, as extensively documented for the Krebs cycle in the last decade. Like normal tissues, tumors can take advantage of metabolic coupling with other tissues, although this phenomenon remains poorly described. A related bioenergetic peculiarity of cancer cells concerns the metabolic coupling which occurs inside the tumors, between hypoxic-glycolytic and oxygenated-oxidative cancer cells. Undoubtedly, the fundamental studies on metabolic rigidity in normal tissues and on metabolic flexibility in cancer cells have revealed novel means of bioenergetic control, such as the essential signaling activity of the oncometabolites succinate, fumarate, oxoglutarate, or acetyl-CoA on HIF1α and NRF2 transcription factors, but also histone acetylases, respectively. The well-known regulation of OXPHOS by citrate, NADH, F1,6BP, Ca2+, and ATP on glycolytic enzymes IDH, PDH, and CIV also operates in cancer cells. Besides energy homeostasis, the study of cancer cell metabolic remodeling has shed light on the close links between the modalities of energy transduction and the capacity to synthesize amino acids or lipids necessary for tumor growth. Metabolic remodeling is not restricted to cancer and has been investigated in genetically inherited metabolic disorders as mitochondrial diseases or in the multifactorial metabolic syndrome. The results demonstrated the existence of alternative pathways of ATP synthesis and the effect of oxidative stress on TCA truncation via aconitase cleavage. Nowadays, large-scale analyses of comprehensive proteomics, transcriptomics, and metabolomics, potentially supported by computer modeling of metabolism and genetic approaches of subpathways validation, allow the deciphering of the fine changes in cell metabolism, both in physiology and pathology. Patterns of metabolic deviations and the corresponding genetic signatures can be confirmed by flux analyses using 13C-labeled isotopes of relevant metabolites. Analysis of deviant metabolism can identify potential targets so that therapeutic strategies could be derived from this knowledge, and ongoing preclinical studies aim to alter the alternative metabolic pathways in cancer.

The field of energy metabolism dramatically progressed in the last decade, owing to a large number of cancer studies, as well as fundamental investigations on related transcriptional networks and cellular interactions with the microenvironment. The concept of metabolic flexibility was clarified in studies showing the ability of cancer cells to remodel the biochemical pathways of energy transduction and linked anabolism in response to glucose, glutamine or oxygen deprivation. A clearer understanding of the large-scale bioenergetic impact of C-MYC, MYCN, KRAS and P53 was obtained, along with its modification during the course of tumor development. The metabolic dialog between different types of cancer cells, but also with the stroma, also complexified the understanding of bioenergetics and raised the concepts of metabolic symbiosis and reverse Warburg effect. Signaling studies revealed the role of respiratory chain-derived reactive oxygen species for metabolic remodeling and metastasis development. The discovery of oxidative tumors in human and mice models related to chemoresistance also changed the prevalent view of dysfunctional mitochondria in cancer cells. Likewise, the influence of energy metabolism-derived oncometabolites emerged as a new means of tumor genetic regulation. The knowledge obtained on the multi-site regulation of energy metabolism in tumors was translated to cancer preclinical studies, supported by genetic proof of concept studies targeting LDHA, HK2, PGAM1, or ACLY. Here, we review those different facets of metabolic remodeling in cancer, from its diversity in physiology and pathology, to the search of the genetic determinants, the microenvironmental regulators and pharmacological modulators.

In this chapter we discuss the relevance of mitochondrial functions and plasticity in tumor biology. In 1920, Otto Warburg first hypothesized that mitochondrial impairment is a leading cause of cancer although he recognized the existence of oxidative tumors. Likewise, Weinhouse (1951) and others found that deficient mitochondrial respiration is not an obligatory feature of cancer and Peter Vaupel suggested in the 90s that tumor oxygenation rather than OXPHOS capacity was the limiting factor of mitochondrial energy production in cancer. Recent studies now clearly indicate that mitochondria are highly functional in mice tumors and the field of oncobioenergetics identified MYC, Oct1 and RAS as pro-OXPHOS oncogenes. In addition, cancer cells adaptation to aglycemia, metabolic symbiosis between hypoxic and non-hypoxic tumor regions as well the reverse Warburg hypothesis support the crucial role of mitochondrial plasticity in the survival of a subclass of tumors. Therefore, mitochondria are now considered as potential targets for anti-cancer therapy and tentative strategies including a bioenergetic profile characterization of the tumor and the subsequent adapted bioenergetic modulation could be considered for cancer killing.

International audience; Mitophagy has been recently described as a mechanism of elimination of damaged organelles. Although the regulation of the amount of mitochondria is a core issue concerning cellular energy homeostasis, the relationship between mitochondrial degradation and energetic activity has not yet been considered. Here, we report that the stimulation of mitochondrial oxidative phosphorylation enhances mitochondrial renewal by increasing its degradation rate. Upon high oxidative phosphorylation activity, we found that the small GTPase Rheb is recruited to the mitochondrial outer membrane. This mitochondrial localization of Rheb promotes mitophagy through a physical interaction with the mitochondrial autophagic receptor Nix and the autophagosomal protein LC3-II. Thus, Rheb-dependent mitophagy contributes to the maintenance of optimal mitochondrial energy production. Our data suggest that mitochondrial degradation contributes to a bulk renewal of the organelle in order to prevent mitochondrial aging and to maintain the efficiency of oxidative phosphorylation.

Hereditary spastic paraplegia (HSP) is considered one of the most heterogeneous groups of neurological disorders, both clinically and genetically. The disease comprises pure and complex forms that clinically include slowly progressive lower-limb spasticity resulting from degeneration of the corticospinal tract. At least 48 loci accounting for these diseases have been mapped to date, and mutations have been identified in 22 genes, most of which play a role in intracellular trafficking. Here, we identified mutations in two functionally related genes (DDHD1 and CYP2U1) in individuals with autosomal-recessive forms of HSP by using either the classical positional cloning or a combination of whole-genome linkage mapping and next-generation sequencing. Interestingly, three subjects with CYP2U1 mutations presented with a thin corpus callosum, white-matter abnormalities, and/or calcification of the basal ganglia. These genes code for two enzymes involved in fatty-acid metabolism, and we have demonstrated in human cells that the HSP pathophysiology includes alteration of mitochondrial architecture and bioenergetics with increased oxidative stress. Our combined results focus attention on lipid metabolism as a critical HSP pathway with a deleterious impact on mitochondrial bioenergetic function.

Introduction Sur modele murin, l’infiltration prolongee de levobupivacaine apres chirurgie d’exerese de tumeurs de type spinocellulaires reduit la taille des recidives tumorales [1] . L’objectif de ce travail consiste a confirmer in vitro la cytotoxicite de la levobupivacaine sur un modele cellulaire de carcinome epidermoide humain et d’etudier les mecanismes metaboliques impliques dans l’interaction entre la levobupivacaine et les cellules cancereuses. Materiel et methodes Les effets de la levobupivacaine sur des lignees de cellules humaines de carcinome epidermoide A431 ont ete compares a ceux observes sur des fibroblastes et des keratinocytes sains HacaT spontanement immortalises. Les cellules ont ete mises en culture en presence de levobupivacaine a differentes concentrations (10 μM, 50 μM, 100 μM, 500 μM et 1 mM). L’effet synergique du 5-Flououracil (50 μM) a ete recherche. La mesure de la proliferation et de la viabilite cellulaire ont ete etudiees par la methode du cristal violet apres 48 heures d’exposition. L’effet de la levobupivacaine sur le metabolisme energetique cellulaire a ete evalue par mesure de la respiration mitochondriale ( oxygen consumption rate ) et de la glycolyse ( extracellular acidification rate ) en plaques 96 puits (Seahorse Biosciences XF96) dans les 90 minutes suivant l’exposition. Les resultats ont ete exprimes en pourcentage du controle non traite et la comparaison des differents groupes a ete effectuee par le test Anova, avec une valeur p Resultats La viabilite cellulaire est significativement reduite par la levobupivacaine pour des concentrations superieures a 500 μM de facon dose dependante ( Fig. 1 a). A une concentration de 500 μM, cette cytotoxicite est accrue sur les cellules cancereuses par rapport aux cellules saines ( Fig. 1 a). Un effet additif avec le 5FU a ete retrouve apres 48 h d’exposition prolongee. Sur le plan metabolique, la levobupivacaine entraine aux concentrations les plus elevees (500 μM et 1 mM) une double inhibition des deux voies energetiques principales, la glycolyse et les OXPHOS ( Fig. 1 b). Discussion La levobupivacaine diminue la viabilite cellulaire et reduit plus specifiquement la proliferation des cellules de carcinome epidermoide vs les cellules saines. Cette cytotoxicite pourrait s’expliquer par sa capacite a inhiber partiellement les deux voies energetiques principales, glycolytique et mitochondriales. Il convient maintenant d’approfondir la comprehension de ces mecanismes et de confirmer ces resultats sur des extraits tissulaires humains.