Resting natural killer cell homeostasis relies on tryptophan/NAD+^{+} metabolism and HIF-1α

Natural killer (NK) cells are forced to cope with different oxygen environments even under resting conditions. The adaptation to low oxygen is regulated by oxygen-sensitive transcription factors, the hypoxia-inducible factors (HIFs). The function of HIFs for NK cell activation and metabolic rewiring remains controversial. Activated NK cells are predominantly glycolytic, but the metabolic programs that ensure the maintenance of resting NK cells are enigmatic. By combining in situ metabolomic and transcriptomic analyses in resting murine NK cells, our study defines HIF-1α as a regulator of tryptophan metabolism and cellular nicotinamide adenine dinucleotide (NAD$^{+}$ ) levels. The HIF-1α/NAD$^{+}$ axis prevents ROS production during oxidative phosphorylation (OxPhos) and thereby blocks DNA damage and NK cell apoptosis under steady-state conditions. In contrast, in activated NK cells under hypoxia, HIF-1α is required for glycolysis, and forced HIF-1α expression boosts glycolysis and NK cell performance in vitro and in vivo. Our data highlight two distinct pathways by which HIF-1α interferes with NK cell metabolism. While HIF-1α-driven glycolysis is essential for NK cell activation, resting NK cell homeostasis relies on HIF-1α-dependent tryptophan/NAD$^{+}$ metabolism

Vaccination-based immunotherapy to target profibrotic cells in liver and lung

Fibrosis is the final path of nearly every form of chronic disease, regardless of the pathogenesis. Upon chronic injury, activated, fibrogenic fibroblasts deposit excess extracellular matrix, and severe tissue fibrosis can occur in virtually any organ. However, antifibrotic therapies that target fibrogenic cells, while sparing homeostatic fibroblasts in healthy tissues, are limited. We tested whether specific immunization against endogenous proteins, strongly expressed in fibrogenic cells but highly restricted in quiescent fibroblasts, can elicit an antigen-specific cytotoxic T cell response to ameliorate organ fibrosis. In silico epitope prediction revealed that activation of the genes Adam12 and Gli1 in profibrotic cells and the resulting “self-peptides” can be exploited for T cell vaccines to ablate fibrogenic cells. We demonstrate the efficacy of a vaccination approach to mount CD8+ T cell responses that reduce fibroblasts and fibrosis in the liver and lungs in mice. These results provide proof of principle for vaccination-based immunotherapies to treat fibrosis

Hypoxia, Metabolism and Immune Cell Function

Hypoxia is a hallmark of inflamed, infected or damaged tissue, and the adaptation to inadequate tissue oxygenation is regulated by hypoxia-inducible factors (HIFs). HIFs are key mediators of the cellular response to hypoxia, but they are also associated with pathological stress such as inflammation, bacteriological infection or cancer. In addition, HIFs are central regulators of many innate and adaptive immunological functions, including migration, antigen presentation, production of cytokines and antimicrobial peptides, phagocytosis as well as cellular metabolic reprogramming. A characteristic feature of immune cells is their ability to infiltrate and operate in tissues with low level of nutrients and oxygen. The objective of this article is to discuss the role of HIFs in the function of innate and adaptive immune cells in hypoxia, with a focus on how hypoxia modulates immunometabolism

Etude du métabolisme cellulaire de la tumeur et sa relation avec l'immunothérapie médiée par les cellules NK

La formation et le développement d'une tumeur sont provoqués par une série de défauts qui se produisent à l'intérieur de la cellule cancéreuse et dans son microenvironnement. Ces anomalies permettent à la cellule de développer ses propres stratégies de croissance, de prolifération, de différenciation et de métabolisme. Toutes ces adaptations, ainsi que la création d'un micro-environnement unique favorisent la croissance de la tumeur et inhibent la réponse immunitaire anti-tumorale. Le métabolisme des cellules cancéreuses et l'évasion immunitaire sont des points très sensibles dans le développement des cancers et peuvent être utilisés en clinique. Les études récentes suggèrent que ces deux phénomènes sont liés, et que le métabolisme des cellules cancéreuses peut amener à l'échappement immunitaire par la tumeur. Le métabolisme des cellules tumorales a tendance à éviter l'activité mitochondriale et la phosphorylation oxydative, et est principalement basée sur la glycolyse pour la production d'énergie (effet Warburg). Mon travail de thèse est divisé en deux parties. Dans la première partie nous avons proposé un concept thérapeutique novateur avec une nouvelle thérapie combinatoire pour le traitement de cancers hématologiques. Cette thérapie est basée sur l'induction de changements métaboliques par le dichloroacétate (DCA), et elle est associée avec la chimiothérapie conventionnelle (doxorubicine, vincristine) pour réactiver les fonctions de p53. Les tumeurs avec p53 mutantes sont résistantes à cette combinaison. Dans ce cas, nous avons constaté que le DCA peut coopère avec 17-AAG (l'inhibiteur de Hsp90) pour éliminer spécifiquement les cellules cancéreuses. En conséquence, une meilleure compréhension des signaux et des mécanismes par lesquels le DCA sensibilise les cellules tumorales à la chimiothérapie est nécessaire pour en comprendre le mode d'action. En outre, l'identification de ce mécanisme permettra d'élucider les voies métaboliques impliquées dans la survie des cellules cancéreuses. La deuxième partie de ma thèse se concentre sur la biologie des cellules NK. Les cellules NK sont des lymphocytes du système immunitaire inné et possèdent une cytotoxicité naturelle contre les cibles, c'est à dire les cellules tumorales. L'utilisation optimale des cellules NK en clinique nécessite leur expansion et leur activation in vitro. Les cellules NK s'activent en présence de cytokines ou par le contact avec les cellules cibles. L'activation des cellules NK induit la prolifération, mais celle-ci dépend aussi de la présence d'autres cellules immunitaires. L'activation, par les cytokines et par les cellules cibles, induit un différent ARNm/microARN profil d'expression. L'analyse détaillée des isoformes de la protéine tyrosine phosphatase CD45 a permis de caractériser de nouvelles populations de cellules NK anti-tumorales humaines. L'identification de différentes populations de cellules NK est très importante pour la compréhension de leur physiologie et pour l'amélioration de leur utilisation en immunothérapie clinique. Cela peut également donner des informations précieuses sur l'état physiologique de l'hôte. En effet, l'augmentation des cellules CD45RAdim et CD45RO + dans le compartiment des cellules NK matures identifie clairement les patients avec des hémopathies malignes. Nous pensons que leur détection peut être utilisée comme un outil de diagnostic et également pour évaluer l'efficacité des traitements anti-tumoraux, car ces populations de cellules NK spécifiques devraient diminuer lors de l'élimination de cellules tumorales cibles. Dans l'avenir, nous voulons combiner le traitement du métabolisme de la tumeur avec la thérapie anti-tumorale basée sur les cellules NK. Sur la base de nos données préliminaires, nous pouvons proposer le traitement des cellules cancéreuses par des médicaments métaboliques pour augmenter la sensibilité et la reconnaissance par les cellules NK activées.Tumor formation and development are caused by a range of defects that occur inside the cancer cell and in the external cellular microenvironment. These abnormalities allow developing tumors to establish their own strategies of growth, proliferation, differentiation and metabolism. All these adaptations, as well as the creation of a unique microenvironment, promote tumor growth and suppress the anti-cancer immune response. Tumor cell metabolism and immune evasion are sensitive points of cancer development that can be targeted in clinic. Recent studies suggest that these two phenomena are related and that cancer cell metabolism may propel tumor immune escape. Tumor cell metabolism tends to avoid mitochondrial activity and oxidative phosphorylation (OXPHOS), and largely relies on glycolysis to produce energy (Warburg effect). My thesis work is divided into two parts. The first one proposes an innovative therapeutic strategy, which is the use of different combinatorial therapy depending on the p53 status for the treatment of hematological cancers. This is based on the induction of metabolic changes by dichloroacetate (DCA), combined with conventional chemotherapy (doxorubicin, vincristine) to reactivate wild type p53 functions. Mutant p53 tumors are resistant to this combination approach. However, we found that DCA synergized with the Hsp90 inhibitor 17-AAG to specifically eliminate these cells. Therefore, a clearer understanding of the signals and mechanisms by which DCA sensitize cancer cells to chemotherapy was needed to understand its mode of action. We uncovered it in our work. In addition, identification of this mechanism will help to elucidate metabolic pathways involved in cancer cell survival.The second part of my thesis is focused on the study of NK cell biology. NK cell is an innate immune system lymphocyte lineage with natural cytotoxicity against targets, i.e. tumor cells. Its optimal use in the clinic requires in vitro expansion and activation. Cytokines and the encounter with target cells activate NK cells, induce their proliferation, and cause clearly different mRNA/miRNA expression profile. Detailed analysis of the leucocyte-specific phosphatase CD45 isoforms allowed us to characterize new human anti-tumor NK cell populations. The identification of the different NK cell populations is important for understanding their physiology and for improving their therapeutic use in the clinic. It can also give valuable information about the host physiological status. Indeed, the increase of CD45RAdim and CD45RO+ cells in the mature NK cell compartment clearly identifies patients with hematological malignancies. We thus hypothesize that their detection could be used as a diagnostic tool, and also to assess the efficacy of antitumor treatments, because these specific NK cell populations should decrease upon removal of the targeted tumor cells. Our future goal is to use a novel combinatorial therapy in hematological cancers that will combine metabolic drugs and NK cell-based therapy. Based on our preliminary data, we propose that the treatment of cancer cells with metabolic drugs could increase their sensitivity and recognition by activated NK cells

Hypoxia, Metabolism and Immune Cell Function

Hypoxia is a hallmark of inflamed, infected or damaged tissue, and the adaptation to inadequate tissue oxygenation is regulated by hypoxia-inducible factors (HIFs). HIFs are key mediators of the cellular response to hypoxia, but they are also associated with pathological stress such as inflammation, bacteriological infection or cancer. In addition, HIFs are central regulators of many innate and adaptive immunological functions, including migration, antigen presentation, production of cytokines and antimicrobial peptides, phagocytosis as well as cellular metabolic reprogramming. A characteristic feature of immune cells is their ability to infiltrate and operate in tissues with low level of nutrients and oxygen. The objective of this article is to discuss the role of HIFs in the function of innate and adaptive immune cells in hypoxia, with a focus on how hypoxia modulates immunometabolism

From tumor cell metabolism to tumor immune escape.

International audienceTumorigenesis implies adaptation of tumor cells to an adverse environment. First, developing tumors must acquire nutrients to ensure their rapid growth. Second, they must escape the attack from the host immune system. Recent studies suggest that these phenomena could be related and that tumor cell metabolism may propel tumor immune escape. Tumor cell metabolism tends to avoid mitochondrial activity and oxidative phosphorylation (OXPHOS), and largely relies on glycolysis to produce energy. This specific metabolism helps tumor cells to avoid the immune attack from the host by blocking or avoiding the immune attack. By changing their metabolism, tumor cells produce or sequester a variety of amino acids, lipids and chemical compounds that directly alter immune function therefore promoting immune evasion. A second group of metabolism-related modification targets the major histocompatibility complex-I (MHC-I) and related molecules. Tumor MHC-I presents tumor-associated antigens (TAAs) to cytotoxic T-cells (CTLs) and hence, sensitizes cancer cells to the cytolytic actions of the anti-tumor adaptive immune response. Blocking tumor mitochondrial activity decreases expression of MHC-I molecules at the tumor cell surface. And peroxynitrite (PNT), produced by tumor-infiltrating myeloid cells, chemically modifies MHC-I avoiding TAA expression in the plasma membrane. These evidences on the role of tumor cell metabolism on tumor immune escape open the possibility of combining drugs designed to control tumor cell metabolism with new procedures of anti-tumor immunotherapy. This article is part of a Directed Issue entitled: Bioenergetic dysfunction, adaptation and therapy

Chemical Metabolic Inhibitors For The Treatment Of Blood-Borne Cancers.

International audience: Tumor cells, including leukemic cells, remodel their bioenergetic system in favor of aerobic glycolysis. This process is called "the Warburg effect" and offers an attractive pharmacological target to preferentially eliminate malignant cells. In addition, recent results show that metabolic changes can be linked to tumor immune evasion. Mouse models demonstrate the importance of this metabolic remodeling in leukemogenesis. Some leukemias, although treatable, remain incurable and resistance to chemotherapy produces an elevated percentage of relapse in most leukemia cases. Several groups have targeted the specific metabolism of leukemia cells in preclinical and clinical studies to improve the prognosis of these patients, i.e. using L-asparaginase to treat pediatric acute lymphocytic leukemia (ALL). Additional metabolic drugs that are currently being used to treat other diseases or tumors could also be exploited for leukemia, based on preclinical studies. Finally, we discuss the potential use of several metabolic drugs in combination therapies, including immunomodulatory drugs (IMiDs) or immune cell-based therapies, to increase their efficacy and reduce side effects in the treatment of hematological cancers

NK cells in hypoxic skin mediate a trade-off between wound healing and antibacterial defence

During skin injury, immune response and repair mechanisms have to be coordinated for rapid skin regeneration and the prevention of microbial infections. Natural Killer (NK) cells infiltrate hypoxic skin lesions and Hypoxia-inducible transcription factors (HIFs) mediate adaptation to low oxygen. We demonstrate that mice lacking the Hypoxia-inducible factor (HIF)-1α isoform in NK cells show impaired release of the cytokines Interferon (IFN)-γ and Granulocyte Macrophage - Colony Stimulating Factor (GM-CSF) as part of a blunted immune response. This accelerates skin angiogenesis and wound healing. Despite rapid wound closure, bactericidal activity and the ability to restrict systemic bacterial infection are impaired. Conversely, forced activation of the HIF pathway supports cytokine release and NK cell-mediated antibacterial defence including direct killing of bacteria by NK cells despite delayed wound closure. Our results identify, HIF-1α in NK cells as a nexus that balances antimicrobial defence versus global repair in the skin

Loss of HIF-1α in natural killer cells inhibits tumour growth by stimulating non-productive angiogenesis

Productive angiogenesis, a prerequisite for tumour growth, depends on the balanced release of angiogenic and angiostatic factors by different cell types within hypoxic tumours. Natural killer (NK) cells kill cancer cells and infiltrate hypoxic tumour areas. Cellular adaptation to low oxygen is mediated by Hypoxia-inducible factors (HIFs). We found that deletion of HIF-1α in NK cells inhibited tumour growth despite impaired tumour cell killing. Tumours developing in these conditions were characterised by a high-density network of immature vessels, severe haemorrhage, increased hypoxia, and facilitated metastasis due to non-productive angiogenesis. Loss of HIF-1α in NK cells increased the bioavailability of the major angiogenic cytokine vascular endothelial growth factor (VEGF) by decreasing the infiltration of NK cells that express angiostatic soluble VEGFR-1. In summary, this identifies the hypoxic response in NK cells as an inhibitor of VEGF-driven angiogenesis, yet, this promotes tumour growth by allowing the formation of functionally improved vessels