TP53INP1 deficiency maintains murine B lymphopoiesis in aged bone marrow through redox-controlled IL-7R/STAT5 signaling.

Bone marrow (BM) produces all blood and immune cells deriving from hematopoietic stem cells (HSCs). The decrease of immune cell production during aging is one of the features of immunosenescence. The impact of redox dysregulation in BM aging is still poorly understood. Here we use TP53INP1-deficient (KO) mice endowed with chronic oxidative stress to assess the influence of aging-associated redox alterations in BM homeostasis. We show that TP53INP1 deletion has no impact on aging-related accumulation of HSCs. In contrast, the aging-related contraction of the lymphoid compartment is mitigated in TP53INP1 KO mice. B cells that accumulate in old KO BM are differentiating cells that can mature into functional B cells. Importantly, this phenotype results from B cell-intrinsic events associated with defective redox control. Finally, we show that oxidative stress in aged TP53INP1-deficient mice maintains STAT5 expression and activation in early B cells, driving high Pax5 expression, which provides a molecular mechanism for maintenance of B cell development upon aging.

Targeting Redox Metabolism in Pancreatic Cancer.

Cell metabolism is reprogrammed in cancer cells to meet their high bioenergetics and biosynthetic demands. This metabolic reprogramming is accompanied by alterations in redox metabolism, characterized by accumulation of reactive oxygen species (ROS). Elevated production of ROS, mostly by mitochondrial respiration, is counteracted by higher production of antioxidant defenses (mainly glutathione and antioxidant enzymes). Cancer cells are adapted to a high concentration of ROS, which contributes to tumorigenesis, metastasis formation, resistance to therapy and relapse. Frequent genetic alterations observed in pancreatic ductal adenocarcinoma (PDAC) affect KRAS and p53 proteins, which have a role in ROS production and control, respectively. These observations led to the proposal of the use of antioxidants to prevent PDAC development and relapse. In this review, we focus on the therapeutic strategies to further increase ROS level to induce PDAC cell death. Combining the promotion of ROS production and inhibition of antioxidant capacity is a promising avenue for pancreatic cancer therapy in the clinic.

PML hyposumoylation is responsible for the resistance of pancreatic cancer.

The dismal prognosis of pancreatic ductal adenocarcinoma (PDAC) is mainly due to its rapidly acquired resistance to all conventional treatments. Despite drug-specific mechanisms of resistance, none explains how these cells resist the stress induced by any kind of anticancer treatment. Activation of stress-response pathways relies on the post-translational modifications (PTMs) of involved proteins. Among all PTMs, those mediated by the ubiquitin family of proteins play a central role. Our aim was to identify alterations of ubiquitination, neddylation, and sumoylation associated with the multiresistant phenotype and demonstrate their implications in the survival of PDAC cells undergoing treatment. This approach pointed at an alteration of promyelocytic leukemia (PML) protein sumoylation associated with both gemcitabine and oxaliplatin resistance. We could show that this alteration of PML sumoylation is part of a general mechanism of drug resistance, which in addition involves the abnormal activation of NF-κB and cAMP response element binding pathways. Importantly, using patient-derived tumors and cell lines, we identified a correlation between the levels of PML expression and sumoylation and the sensitivity of tumors to anticancer treatments.-Swayden, M., Alzeeb, G., Masoud, R., Berthois, Y., Audebert, S., Camoin, L., Hannouche, L., Vachon, H., Gayet, O., Bigonnet, M., Roques, J., Silvy, F., Carrier, A., Dusetti, N., Iovanna, J. L., Soubeyran, P. PML hyposumoylation is responsible for the resistance of pancreatic cancer.

Thymus-specific serine protease, a protease that shapes the CD4 T cell repertoire.

The lifespan of T cells is determined by continuous interactions of their T cell receptors (TCR) with self-peptide-MHC (self-pMHC) complexes presented by different subsets of antigen-presenting cells (APC). In the thymus, developing thymocytes are positively selected through recognition of self-pMHC presented by cortical thymic epithelial cells (cTEC). They are subsequently negatively selected by medullary thymic epithelial cells (mTEC) or thymic dendritic cells (DC) presenting self-pMHC complexes. In the periphery, the homeostasis of mature T cells is likewise controlled by the interaction of their TCR with self-pMHC complexes presented by lymph node stromal cells while they may be tolerized by DC presenting tissue-derived self-antigens. To perform these tasks, the different subsets of APC are equipped with distinct combination of antigen processing enzymes and consequently present specific repertoire of self-peptides. Here, we discuss one such antigen processing enzyme, the thymus-specific serine protease (TSSP), which is predominantly expressed by thymic stromal cells. In thymic DC and TEC, TSSP edits the repertoire of peptide presented by class II molecules and thus shapes the CD4 T cell repertoire.

Neuroglobin protects astroglial cells from hydrogen peroxide-induced oxidative stress and apoptotic cell death.

Oxidative stress, resulting from accumulation of reactive oxygen species, plays a critical role in astroglial cell death occurring in diverse neuropathological conditions. Numerous studies indicate that neuroglobin (Ngb) promotes neuron survival, but nothing is known regarding the action of Ngb in astroglial cell survival. Thus, the purpose of this study was to investigate the potential glioprotective effect of Ngb on hydrogen peroxide (H2 O2 )-induced oxidative stress and apoptosis in cultured mouse astrocytes. Incubation of cells with subnanomolar concentrations of Ngb (10-14 -10-10  M) was found to prevent both H2 O2 -evoked reduction in surviving cells number and accumulation of reactive oxygen species in a concentration-dependent manner. Furthermore, Ngb treatment abolishes H2 O2 -induced increase in mitochondrial oxygen consumption rates. Concomitantly, Ngb treatment rescues H2 O2 -associated reduced expression of endogenous antioxidant enzymes (superoxide dismutases and catalase) and prevents the stimulation of the expression of pro-inflammatory genes (inducible nitric oxide synthase, cyclooxygenase-2, and interleukin (IL) IL-6 and IL-33). Moreover, Ngb blocks the stimulation of Bax (pro-apoptotic) and the inhibition of Bcl-2 (anti-apoptotic) gene expression induced by H2 O2 , which in turn abolishes caspase 3 activation. The protective effect of Ngb upon H2 O2 induced activation of caspase 3 activity and cell death can be accounted for by activation of protein kinase A and mitogen-activated protein kinase transduction cascade. Finally, we demonstrate that Ngb increases Akt phosphorylation and prevents H2 O2 -provoked inhibition of ERK and Akt phosphorylation. Taken together, these data demonstrate for the first time that Ngb is a glioprotective agent that prevents H2 O2 -induced oxidative stress and apoptotic astroglial cell death. Protection of astrocytes from oxidative insult may thus contribute to the neuroprotective effect of Ngb.

Oxidative stress induced by inactivation of TP53INP1 cooperates with KrasG12D to initiate and promote pancreatic carcinogenesis in the murine pancreas.

Tumor protein p53-induced nuclear protein 1 (TP53INP1) is involved in cell stress response. Its expression is lost at the pancreatic intraepithelial neoplasia 1b (PanIN1b)/PanIN2 stage of pancreatic carcinogenesis. Our objective was to determine whether TP53INP1 loss of expression contributes to pancreatic cancer formation in a conditional KrasG12D mouse model. We generated Kras-INP1KO mice using LSL-Kras(G12D/+);Pdx1-Cre(+/-) mice (Kras mice) and TP53INP1(-/-) mice. Analysis of pancreases during ageing shows that in the presence of activated Kras, TP53INP1 loss of expression accelerated PanIN formation and increased pancreatic injury and the number of high-grade lesions as compared with what occurs in Kras mice. Moreover, cystic lesions resembling intraductal papillary mucinous neoplasm (IPMN) were observed as early as 2 months of age. Remarkably, TP53INP1 is down-regulated in human IPMN. Activation of the small GTPase Rac1 shows that more oxidative stress is generated in Kras-INP1KO than in Kras mice pancreas despite elevated levels of the Nrf2 antioxidant regulator. We firmly establish the link between Kras-INP1KO pancreatic phenotype and oxidative stress with rescue of the phenotype by the antioxidant action of N-acetylcysteine. Our data provide in vivo functional demonstration that TP53INP1 deficiency accelerates progression of pancreatic cancer, underlining its role in the occurrence of IPMN and highlighting the importance of TP53INP1 in the control of oxidative status during development of pancreatic cancer.

Combining the antianginal drug perhexiline with chemotherapy induces complete pancreatic cancer regression in vivo.

Pancreatic ductal adenocarcinoma (PDAC) remains one of the human cancers with the poorest prognosis. Interestingly, we found that mitochondrial respiration in primary human PDAC cells depends mainly on the fatty acid oxidation (FAO) to meet basic energy requirements. Therefore, we treated PDAC cells with perhexiline, a well-recognized FAO inhibitor used in cardiac diseases. Some PDAC cells respond efficiently to perhexiline, which acts synergistically with chemotherapy (gemcitabine) in vitro and in two xenografts in vivo. Importantly, perhexiline in combination with gemcitabine induces complete tumor regression in one PDAC xenograft. Mechanistically, this co-treatment causes energy and oxidative stress promoting apoptosis but does not exert inhibition of FAO. Yet, our molecular analysis indicates that the carnitine palmitoyltransferase 1C (CPT1C) isoform is a key player in the response to perhexiline and that patients with high CPT1C expression have better prognosis. Our study reveals that repurposing perhexiline in combination with chemotherapy is a promising approach to treat PDAC.

Transcription profiling of Prss16 (Tssp) can be used to find additional peptidase genes that are candidates for self-peptide generation in the thymus.

Positive selection (PS) in the thymus involves the presentation of self-peptides that are bound to MHC class II on the surface of cortical thymus epithelial cells (cTECs). Prss16 gene corresponds to one important element regulating the PS of CD4(+) T lymphocytes, which encodes Thymus-specific serine protease (Tssp), a cTEC serine-type peptidase involved in the proteolytic generation of self-peptides. Nevertheless, additional peptidase genes participating in the generation of self-peptides need to be found. Because of its role in the mechanism of PS and its expression in cTECs, the Prss16 gene might be used as a transcriptional marker to identify new genes that share the same expression profile and that encode peptidases in the thymus. To test this hypothesis, we compared the differential thymic expression of 4,500 mRNAs of wild-type (WT) C57BL/6 mice with their respective Prss16-knockout (KO) mutants by using microarrays. From these, 223 genes were differentially expressed, of which 115 had known molecular/biological functions. Four endopeptidase genes (Casp1, Casp2, Psmb3 and Tpp2) share the same expression profile as the Prss16 gene; i.e., induced in WT and repressed in KO while one endopeptidase gene, Capns1, features opposite expression profile. The Tpp2 gene is highlighted because it encodes a serine-type endopeptidase functionally similar to the Tssp enzyme. Profiling of the KO mice featured down-regulation of Prss16, as expected, along with the genes mentioned above. Considering that the Prss16-KO mice featured impaired PS, the shared regulation of the four endopeptidase genes suggested their participation in the mechanism of self-peptide generation and PS.

Grape seed and skin extract mitigates garlic-induced oxidative stress in rat liver.

Garlic is a commonly used spice in folk medicine that can exert adverse health effects when given at a high dose. Grape seed and skin extract (GSSE) exhibits a variety of beneficial effects even at a high dose. In the present study we evaluated the toxicity of high-dose garlic treatment on liver and the protective effect of GSSE. Rats were intraperitoneally administered either with garlic extract (5 g·(kg body weight)(-1)) or GSSE (500 mg·(kg body weight)(-1)) or a combination of garlic and GSSE at the same doses daily for 1 month. Plasma and hepatic levels of cholesterol, triacylglycerol, and transaminases and liver antioxidant status were evaluated. Data showed that a high garlic dose induced liver toxicity and a pro-oxidative status characterized by increased malondialdehyde and decreased antioxidant enzyme activities as catalase, peroxidase, and superoxide dismutase. Garlic increased intracellular H(2)O(2) but decreased free iron and Ca(2+). GSSE alone or in co-treatment with garlic had the reverse effect and counteracted almost all garlic-induced deleterious impacts to near control levels. In conclusion, a high garlic dose induced a pro-oxidative state characterized by the Fenton reaction between H(2)O(2) and free iron, inducing Ca(2+) depletion, while GSSE exerted antioxidant properties and Ca(2+) repletion.

A novel anticancer pharmacological agent targeting mitochondrial complex I.

Targeting metabolic reprogramming has proven successful in oncology, but this field requires better identification of drugs that inhibit mitochondrial metabolism in cancer cells. Recent work from Dr Wolf's group reveals that the primary target of the antitumor compound SMIP004-7 is mitochondrial complex I (NDUFS2 subunit), inhibition of which promotes anticancer immune surveillance.

Autophagy Contributes to Metabolic Reprogramming and Therapeutic Resistance in Pancreatic Tumors.

Metabolic reprogramming is a feature of cancers for which recent research has been particularly active, providing numerous insights into the mechanisms involved. It occurs across the entire cancer process, from development to resistance to therapies. Established tumors exhibit dependencies for metabolic pathways, constituting vulnerabilities that can be targeted in the clinic. This knowledge is of particular importance for cancers that are refractory to any therapeutic approach, such as Pancreatic Ductal Adenocarcinoma (PDAC). One of the metabolic pathways dysregulated in PDAC is autophagy, a survival process that feeds the tumor with recycled intracellular components, through both cell-autonomous (in tumor cells) and nonautonomous (from the local and distant environment) mechanisms. Autophagy is elevated in established PDAC tumors, contributing to aberrant proliferation and growth even in a nutrient-poor context. Critical elements link autophagy to PDAC including genetic alterations, mitochondrial metabolism, the tumor microenvironment (TME), and the immune system. Moreover, high autophagic activity in PDAC is markedly related to resistance to current therapies. In this context, combining autophagy inhibition with standard chemotherapy, and/or drugs targeting other vulnerabilities such as metabolic pathways or the immune response, is an ongoing clinical strategy for which there is still much to do through translational and multidisciplinary research.

A pharmacological strategy to recapitulate exercise-induced antitumoral immunity.

The antitumor activity of exercise by means of enhanced immune activation is documented, but better identification of the underlying mechanisms is required to develop new therapeutic strategies. Recent work from the Dr Bar-Sagi group reveals that exercise engages IL-15 signaling and pharmacological activation of the IL-15/IL-15R axis mimics the exercise-driven immune cell-mediated cytotoxicity in pancreatic cancer.

Pesticides and pancreatic adenocarcinoma: A transversal epidemiological, environmental and mechanistic narrative review.

Pancreatic adenocarcinoma (PA) incidence is rising worldwide, especially in France. The evolution of known risk factors such as tobacco smoking, obesity, type 2 diabetes, chronic pancreatitis, or constitutional mutations is not sufficient to explain this trend. Pesticides are known risk factors in other malignancies. Previous studies have outlined pesticides' influence in PA, such as dichlorodiphenyltrichloroethane as plausible risk factors. The general population is directly or indirectly exposed to pesticides through air, food or water. Some of these chemicals may accumulate in the body all along lifetime and may harm carriers. The toxic mixing effects of these chemicals are not well documented. Several hypotheses have been put forward to explain how pesticides can induce indirect (fatty pancreas, induced diabetes) or direct (oxidative stress, cell damage) carcinogenesis in pancreatic cells through inflammation. A strong corpus exists acknowledging pesticides as a PA risk factor. However, published studies do not provide a sufficient level of evidence to prove causality and current prospective case-control studies are still ongoing.

Meeting report of the 4th biennial Metabolism and Cancer symposium.

The 4th International meeting Metabolism and Cancer initially programed to take place in Bordeaux (France) was held virtually on May 27-29, 2021. The three-day event was followed by around 600 participants daily from 47 countries around the world. The meeting hosted 21 speakers including selected talks and a keynote lecture from the Nobel Prize winner Sir Peter J. Ratcliffe (Oxford, UK). Presentations and discussions were divided in four scientific sessions: (a) Redox and energy metabolism (b) Redox and hypoxia (c) Metabolic profiling and epigenetic control and (d) Signalling, fuelling and metabolism in cancer and a general public session on cancer and nutrition. This report summarises the presentations and outcomes of the 4th annual Metabolism and Cancer symposium. We provide here a summary of the scientific highlights of this exciting meeting.

TP53INP1 exerts neuroprotection under ageing and Parkinson's disease-related stress condition.

TP53INP1 is a stress-induced protein, which acts as a dual positive regulator of transcription and of autophagy and whose deficiency has been linked with cancer and metabolic syndrome. Here, we addressed the unexplored role of TP53INP1 and of its Drosophila homolog dDOR in the maintenance of neuronal homeostasis under chronic stress, focusing on dopamine (DA) neurons under normal ageing- and Parkinson's disease (PD)-related context. Trp53inp1-/- mice displayed additional loss of DA neurons in the substantia nigra compared to wild-type (WT) mice, both with ageing and in a PD model based on targeted overexpression of α-synuclein. Nigral Trp53inp1 expression of WT mice was not significantly modified with ageing but was markedly increased in the PD model. Trp53inp2 expression showed similar evolution and did not differ between WT and Trp53inp1-/- mice. In Drosophila, pan-neuronal dDOR overexpression improved survival under paraquat exposure and mitigated the progressive locomotor decline and the loss of DA neurons caused by the human α-synuclein A30P variant. dDOR overexpression in DA neurons also rescued the locomotor deficit in flies with RNAi-induced downregulation of dPINK1 or dParkin. Live imaging, confocal and electron microscopy in fat bodies, neurons, and indirect flight muscles showed that dDOR acts as a positive regulator of basal autophagy and mitophagy independently of the PINK1-mediated pathway. Analyses in a mammalian cell model confirmed that modulating TP53INP1 levels does not impact mitochondrial stress-induced PINK1/Parkin-dependent mitophagy. These data provide the first evidence for a neuroprotective role of TP53INP1/dDOR and highlight its involvement in the regulation of autophagy and mitophagy in neurons.

Pancreatic cancer intrinsic PI3Kα activity accelerates metastasis and rewires macrophage component.

Pancreatic ductal adenocarcinoma (PDAC) patients frequently suffer from undetected micro-metastatic disease. This clinical situation would greatly benefit from additional investigation. Therefore, we set out to identify key signalling events that drive metastatic evolution from the pancreas. We searched for a gene signature that discriminate localised PDAC from confirmed metastatic PDAC and devised a preclinical protocol using circulating cell-free DNA (cfDNA) as an early biomarker of micro-metastatic disease to validate the identification of key signalling events. An unbiased approach identified, amongst actionable markers of disease progression, the PI3K pathway and a distinctive PI3Kα activation signature as predictive of PDAC aggressiveness and prognosis. Pharmacological or tumour-restricted genetic PI3Kα-selective inhibition prevented macro-metastatic evolution by hindering tumoural cell migratory behaviour independently of genetic alterations. We found that PI3Kα inhibition altered the quantity and the species composition of the produced lipid second messenger PIP3 , with a selective decrease of C36:2 PI-3,4,5-P3 . Tumoural PI3Kα inactivation prevented the accumulation of pro-tumoural CD206-positive macrophages in the tumour-adjacent tissue. Tumour cell-intrinsic PI3Kα promotes pro-metastatic features that could be pharmacologically targeted to delay macro-metastatic evolution.

Thymus-specific serine protease regulates positive selection of a subset of CD4+ thymocytes.

Thymus-specific serine protease (TSSP) was initially reported as a putative protease specifically expressed in the endosomal compartment of cortical thymic epithelial cells (cTEC). As such, TSSP is potentially involved in the presentation of the self-peptides that are bound to MHC class II molecules expressed at the cTEC surface and are involved in the positive selection of CD4(+) thymocytes. We tested this hypothesis by generating mutant mice deprived of Prss16, the gene encoding TSSP. TSSP-deficient mice produced normal numbers of T cells, despite a decrease in the percentage of cTEC expressing high surface levels of MHC class II. By using sensitive transgenic models expressing MHC class II-restricted TCR transgenes (Marilyn and OT-II), we showed that the absence of TSSP markedly impaired the selection of Marilyn and OT-II CD4(+) T cells. In contrast, selection of CD8(+) T cells expressing an MHC class I-restricted TCR transgene (OT-I) was unaffected. Therefore, TSSP is involved in the positive selection of some CD4(+) T lymphocytes and likely constitutes the first serine protease to play a function in the intrathymic presentation of self-peptides bound to MHC class II complexes.

Mutant mouse models of oxidative stress.

Oxidative stress corresponds to an excess in reactive oxygen species (ROS) including free radicals which are highly reactive with cellular constituents. Thereby ROS induce damage to DNA, proteins and lipids, which are all involved in the etiology of numerous pathologies such as cancer. To prevent potential damage, a tight regulation of ROS level is achieved through numerous enzyme systems and small molecules such as glutathione and vitamin C. Mutant mouse models targeting antioxidant enzymes have confirmed their essential role in ROS level control, and have shown a limited redundancy of their activity. Additionally, a number of other mutant mouse models exhibit increased ROS levels, suggesting an antioxidant role for the corresponding targeted gene. This is the case for mice deficient for the transcription factors p53, JunD, FoxOs, and HIF-2alpha, which are involved in the modulation of antioxidant enzymes expression. Mice deficient either for the stress factor TP53INP1, which is a target of p53, or for ATM involved in DNA damage sensoring, also show a constitutive oxidative stress. Finally, the last reported case of mice with a permanent oxidative stress targets Bmi which is a transcriptional repressor of the polycomb family. Interestingly, most of these "oxidative stressed mice" either spontaneously develop cancers or are more susceptible than wild-type to tumor-induced protocols. Altogether, these models markedly reinforce the causal link between oxidative stress and cancer. In the future, they will be helpful tools for basic research aimed at unraveling the interplay between redox control actors as well as their relative importance. In addition, these oxidative stressed mouse models may be useful for applied research in particular in preclinical assays where redox status regulation is absolutely required.

Colitis and colitis-associated cancer are exacerbated in mice deficient for tumor protein 53-induced nuclear protein 1.

Tumor protein 53-induced nuclear protein 1 (TP53INP1) is an antiproliferative and proapoptotic protein involved in cell stress response. To address its physiological roles in colorectal cancer and colitis, we generated and tested the susceptibility of Trp53inp1-deficient mice to the development of colorectal tumors induced by injection of the carcinogen azoxymethane followed by dextran sulfate sodium (DSS)-induced chronic colitis. Trp53inp1-deficient mice showed an increased incidence and multiplicity of tumors compared to those of wild-type (WT) mice. Furthermore, acute colitis induced by DSS treatment was more severe in Trp53inp1-deficient mice than in WT mice. Treatment with the antioxidant N-acetylcysteine prevented colitis and colitis-associated tumorigenesis more efficiently in WT mice than in Trp53inp1-deficient mice, suggesting a higher oxidative load in the latter. Consistently, we demonstrated by electron spin resonance and spin trapping that colons derived from deficient mice produced more free radicals than those of the WT during colitis and that the basal blood level of the antioxidant ascorbate was decreased in Trp53inp1-deficient mice. Collectively, these results indicate that the oxidative load is higher in Trp53inp1-deficient mice than in WT mice, generating a more-severe DSS-induced colitis, which favors development of colorectal tumors in Trp53inp1-deficient mice. Therefore, TP53INP1 is a potential target for the prevention of colorectal cancer in patients with inflammatory bowel disease.

Mitochondrial Metabolism in PDAC: From Better Knowledge to New Targeting Strategies.

Cancer cells reprogram their metabolism to meet bioenergetics and biosynthetic demands. The first observation of metabolic reprogramming in cancer cells was made a century ago ("Warburg effect" or aerobic glycolysis), leading to the classical view that cancer metabolism relies on a glycolytic phenotype. There is now accumulating evidence that most cancers also rely on mitochondria to satisfy their metabolic needs. Indeed, the current view of cancer metabolism places mitochondria as key actors in all facets of cancer progression. Importantly, mitochondrial metabolism has become a very promising target in cancer therapy, including for refractory cancers such as Pancreatic Ductal AdenoCarcinoma (PDAC). In particular, mitochondrial oxidative phosphorylation (OXPHOS) is an important target in cancer therapy. Other therapeutic strategies include the targeting of glutamine and fatty acids metabolism, as well as the inhibition of the TriCarboxylic Acid (TCA) cycle intermediates. A better knowledge of how pancreatic cancer cells regulate mitochondrial metabolism will allow the identification of metabolic vulnerabilities and thus novel and more efficient therapeutic options for the benefit of each patient.