Multiomic characterization of pancreatic cancer-associated macrophage polarization reveals deregulated metabolic programs driven by the GM-CSF-PI3K pathway.

The pancreatic ductal adenocarcinoma microenvironment is composed of a variety of cell types and marked by extensive fibrosis and inflammation. Tumor-associated macrophages (TAMs) are abundant, and they are important mediators of disease progression and invasion. TAMs are polarized in situ to a tumor promoting and immunosuppressive phenotype via cytokine signaling and metabolic crosstalk from malignant epithelial cells and other components of the tumor microenvironment. However, the specific distinguishing features and functions of TAMs remain poorly defined. Here, we generated tumor-educated macrophages (TEMs) in vitro and performed detailed, multiomic characterization (i.e., transcriptomics, proteomics, metabolomics). Our results reveal unique genetic and metabolic signatures of TEMs, the veracity of which were queried against our in-house single-cell RNA sequencing dataset of human pancreatic tumors. This analysis identified expression of novel, metabolic TEM markers in human pancreatic TAMs, including ARG1, ACLY, and TXNIP. We then utilized our TEM model system to study the role of mutant Kras signaling in cancer cells on TEM polarization. This revealed an important role for granulocyte-macrophage colony-stimulating factor (GM-CSF) and lactate on TEM polarization, molecules released from cancer cells in a mutant Kras-dependent manner. Lastly, we demonstrate that GM-CSF dysregulates TEM gene expression and metabolism through PI3K-AKT pathway signaling. Collectively, our results define new markers and programs to classify pancreatic TAMs, how these are engaged by cancer cells, and the precise signaling pathways mediating polarization.

Chemical augmentation of mitochondrial electron transport chains tunes T cell activation threshold in tumors.

BACKGROUND: Cancer immunotherapy shows insufficient efficacy for low immunogenic tumors. Furthermore, tumors often downregulate antigen and major histocompatibility complex expression to escape recognition by T cells, resulting in insufficient T cell receptor (TCR) stimulation in the tumor microenvironment. Thus, augmenting TCR-mediated recognition of tumor antigens is a useful strategy to improve the efficacy of cancer immunotherapy. METHODS: We screened 310 small molecules from our library and identified PQDN, a small molecule that activates CD8 T cells after TCR engagement, even when antigen stimulation is too weak for their activation. We used inhibitors of mitochondrial functions and Seahorse Flux Analyzer to investigate the mechanism underlying the effect of PQDN on T cells. Effect of PQDN on tumor-infiltrating CD8 T cells was examined using flow cytometry and TCR repertoire analysis. RESULTS: PQDN increased mitochondrial reciprocal capacity through enhancement of electron transport chains (ETCs) and facilitated glycolysis via mTOR/AKT signaling, resulting in augmented CD8 T cell activation, even when antigen stimulation is extremely weak. Intratumoral administration of this compound into tumor-bearing mice tunes inactivated T cell with tumor antigen recognition potent and expanded functional T cell receptor diversity of tumor-infiltrating T cells, augmenting antitumor immune responses and retarding tumor growth. Furthermore, PQDN has a synergistic potent with T cell dependent immunotherapy, such as checkpoint inhibitory therapy or adoptive cell therapy, even in a low immunogenic tumor. We also demonstrated that this compound enhances the activation of human CD8 T cells. CONCLUSIONS: These data suggest that tuning the T cell activation threshold by chemical activation of mitochondrial ETC is a new strategy for improving therapeutic efficacy through the activation of low-avidity tumor-specific T cells.

G6PD functions as a metabolic checkpoint to regulate granzyme B expression in tumor-specific cytotoxic T lymphocytes.

BACKGROUND: Granzyme B is a key effector of cytotoxic T lymphocytes (CTLs), and its expression level positively correlates with the response of patients with mesothelioma to immune checkpoint inhibitor immunotherapy. Whether metabolic pathways regulate Gzmb expression in CTLs is incompletely understood. METHODS: A tumor-specific CTL and tumor coculture model and a tumor-bearing mouse model were used to determine the role of glucose-6-phosphate dehydrogenase (G6PD) in CTL function and tumor immune evasion. A link between granzyme B expression and patient survival was analyzed in human patients with epithelioid mesothelioma. RESULTS: Mesothelioma cells alone are sufficient to activate tumor-specific CTLs and to enhance aerobic glycolysis to induce a PD-1hi Gzmblo CTL phenotype. However, inhibition of lactate dehydrogenase A, the key enzyme of the aerobic glycolysis pathway, has no significant effect on tumor-induced CTL activation. Tumor cells induce H3K9me3 deposition at the promoter of G6pd, the gene that encodes the rate-limiting enzyme G6PD in the pentose phosphate pathway, to downregulate G6pd expression in tumor-specific CTLs. G6PD activation increases acetyl-coenzyme A (CoA) production to increase H3K9ac deposition at the Gzmb promoter and to increase Gzmb expression in tumor-specific CTLs converting them from a Gzmblo to a Gzmbhi phenotype, thus increasing CTL tumor lytic activity. Activation of G6PD increases Gzmb+ tumor-specific CTLs and suppresses tumor growth in tumor-bearing mice. Consistent with these findings, GZMB expression level was found to correlate with increased survival in patients with epithelioid mesothelioma. CONCLUSION: G6PD is a metabolic checkpoint in tumor-activated CTLs. The H3K9me3/G6PD/acetyl-CoA/H3K9ac/Gzmb pathway is particularly important in CTL activation and immune evasion in epithelioid mesothelioma.

Metabolic reprograming shapes neutrophil functions in severe COVID-19.

To better understand the mechanisms at the basis of neutrophil functions during SARS-CoV-2, we studied patients with severe COVID-19 pneumonia. They had high blood proportion of degranulated neutrophils and elevated plasma levels of myeloperoxidase (MPO), elastase, and MPO-DNA complexes, which are typical markers of neutrophil extracellular traps (NET). Their neutrophils display dysfunctional mitochondria, defective oxidative burst, increased glycolysis, glycogen accumulation in the cytoplasm, and increase glycogenolysis. Hypoxia-inducible factor 1α (ΗΙF-1α) is stabilized in such cells, and it controls the level of glycogen phosphorylase L (PYGL), a key enzyme in glycogenolysis. Inhibiting PYGL abolishes the ability of neutrophils to produce NET. Patients displayed significant increases of plasma levels of molecules involved in the regulation of neutrophils' function including CCL2, CXCL10, CCL20, IL-18, IL-3, IL-6, G-CSF, GM-CSF, IFN-γ. Our data suggest that metabolic remodelling is vital for the formation of NET and for boosting neutrophil inflammatory response, thus, suggesting that modulating ΗΙF-1α or PYGL could represent a novel approach for innovative therapies.

COVID19 Disease Map, a computational knowledge repository of virus-host interaction mechanisms.

We need to effectively combine the knowledge from surging literature with complex datasets to propose mechanistic models of SARS-CoV-2 infection, improving data interpretation and predicting key targets of intervention. Here, we describe a large-scale community effort to build an open access, interoperable and computable repository of COVID-19 molecular mechanisms. The COVID-19 Disease Map (C19DMap) is a graphical, interactive representation of disease-relevant molecular mechanisms linking many knowledge sources. Notably, it is a computational resource for graph-based analyses and disease modelling. To this end, we established a framework of tools, platforms and guidelines necessary for a multifaceted community of biocurators, domain experts, bioinformaticians and computational biologists. The diagrams of the C19DMap, curated from the literature, are integrated with relevant interaction and text mining databases. We demonstrate the application of network analysis and modelling approaches by concrete examples to highlight new testable hypotheses. This framework helps to find signatures of SARS-CoV-2 predisposition, treatment response or prioritisation of drug candidates. Such an approach may help deal with new waves of COVID-19 or similar pandemics in the long-term perspective.

Metabolic Analysis of Potential Key Genes Associated with Systemic Lupus Erythematosus Using Liquid Chromatography-Mass Spectrometry.

The biological mechanism underlying the pathogenesis of systemic lupus erythematosus (SLE) remains unclear. In this study, we found 21 proteins upregulated and 38 proteins downregulated by SLE relative to normal protein metabolism in our samples using liquid chromatography-mass spectrometry. By PPI network analysis, we identified 9 key proteins of SLE, including AHSG, VWF, IGF1, ORM2, ORM1, SERPINA1, IGF2, IGFBP3, and LEP. In addition, we identified 4569 differentially expressed metabolites in SLE sera, including 1145 reduced metabolites and 3424 induced metabolites. Bioinformatics analysis showed that protein alterations in SLE were associated with modulation of multiple immune pathways, TP53 signaling, and AMPK signaling. In addition, we found altered metabolites associated with valine, leucine, and isoleucine biosynthesis; one carbon pool by folate; tyrosine metabolism; arginine and proline metabolism; glycine, serine, and threonine metabolism; limonene and pinene degradation; tryptophan metabolism; caffeine metabolism; vitamin B6 metabolism. We also constructed differently expressed protein-metabolite network to reveal the interaction among differently expressed proteins and metabolites in SLE. A total of 481 proteins and 327 metabolites were included in this network. Although the role of altered metabolites and proteins in the diagnosis and therapy of SLE needs to be further investigated, the present study may provide new insights into the role of metabolites in SLE.

Metabolomics analysis of grains of wheat infected and noninfected with Tilletia controversa Kühn.

Dwarf bunt caused by the pathogen Tilletia controversa Kühn is one of the most serious quarantine diseases of winter wheat. Metabolomics studies provide detailed information about the biochemical changes at the cell and tissue levels of plants. In the present study, a liquid chromatography/mass spectrometry (LC/MS) metabolomics approach was used to investigate the changes in the grain metabolomics of infected and noninfected with T. controversa samples. PCA suggested that T. controversa-infected and noninfected samples were separated during the interaction. LC/MS analysis showed that 62 different metabolites were recorded in the grains, among which a total of 34 metabolites were upregulated and 28 metabolites were downregulated. Prostaglandins (PGs) and 9-hydroxyoctadecadienoic acids (9-HODEs) are fungal toxin-related substances, and their expression significantly increased in T. controversa-infected grains. Additionally, the concentrations of cucurbic acid and octadecatrienoic acid changed significantly after pathogen infection, which play a large role in plant defense. The eight different metabolic pathways activated during T. controversa and wheat plant interactions included phenylalanine metabolism, isoquinoline alkaloid biosynthesis, starch and sucrose metabolism, tyrosine metabolism, sphingolipid metabolism, arginine and proline metabolism, alanine, aspartate, and glutamate metabolism, and tryptophan metabolism. In conclusion, we found differences in the metabolic profiles of wheat grains after T. controversa infection. To our knowledge, this is the first study to evaluate the metabolites in wheat grains after T. controversa infection.

Altered kynurenine pathway metabolism in patients with ankylosing spondylitis.

BACKGROUND: Various studies reported that increased proinflammatory cytokines in patients with ankylosing spondylitis (AS). Proinflammatory cytokines can affect the expression of various kynurenine pathway enzymes and therefore lead to metabolic changes that can affect the inflammatory response and immunity. Our aim was to measure serum levels of kynurenine pathway metabolites in patients with AS. METHODS: The study included 85 patients with AS and 50 healthy volunteers. Serum tryptophan, kynurenine, kynurenic acid, 3-hydroxyanthranilic acid, 3-hydroxykynurenine, quinolinic acid concentrations were measured with tandem mass spectrometry. In addition, participants were divided into four groups according to the treatment regimen: TNF-α inhibitor group, conventional therapy group, control group and newly diagnosed AS group. These groups were compared in terms of kynurenine pathways metabolites, interleukin 6 (IL-6), erythrocyte sedimentation rate (ESR) and C-reactive protein (CRP) levels. RESULTS: Serum tryptophan, kynurenic acid, 3-hydroxykynurenine levels were significantly decreased (p < 0.05) in both AS groups compared to the control group, while the levels of kynurenine, quinolinic acid, CRP, ESR, and IL-6 were higher (p < 0.05). The Kynurenine/Tryptophan ratio and CRP levels of the conventional therapy and anti-TNF therapy group were significantly lower than the newly diagnosed AS patients (p < 0.05). CONCLUSION: As a result of our study, we found that altered kynurenine pathway metabolism in patients with AS. Conventional therapy and anti-TNF-α therapy are effective in reducing the Kynurenine/Tryptophan ratio and CRP levels, although the effect of both treatments on other metabolites appears to be limited.

Integration of metabolomics, genomics, and immune phenotypes reveals the causal roles of metabolites in disease.

BACKGROUND: Recent studies highlight the role of metabolites in immune diseases, but it remains unknown how much of this effect is driven by genetic and non-genetic host factors. RESULT: We systematically investigate circulating metabolites in a cohort of 500 healthy subjects (500FG) in whom immune function and activity are deeply measured and whose genetics are profiled. Our data reveal that several major metabolic pathways, including the alanine/glutamate pathway and the arachidonic acid pathway, have a strong impact on cytokine production in response to ex vivo stimulation. We also examine the genetic regulation of metabolites associated with immune phenotypes through genome-wide association analysis and identify 29 significant loci, including eight novel independent loci. Of these, one locus (rs174584-FADS2) associated with arachidonic acid metabolism is causally associated with Crohn's disease, suggesting it is a potential therapeutic target. CONCLUSION: This study provides a comprehensive map of the integration between the blood metabolome and immune phenotypes, reveals novel genetic factors that regulate blood metabolite concentrations, and proposes an integrative approach for identifying new disease treatment targets.

Kynurenic acid may underlie sex-specific immune responses to COVID-19.

Coronavirus disease 2019 (COVID-19) has poorer clinical outcomes in males than in females, and immune responses underlie these sex-related differences. Because immune responses are, in part, regulated by metabolites, we examined the serum metabolomes of COVID-19 patients. In male patients, kynurenic acid (KA) and a high KA-to-kynurenine (K) ratio (KA:K) positively correlated with age and with inflammatory cytokines and chemokines and negatively correlated with T cell responses. Males that clinically deteriorated had a higher KA:K than those that stabilized. KA inhibits glutamate release, and glutamate abundance was lower in patients that clinically deteriorated and correlated with immune responses. Analysis of data from the Genotype-Tissue Expression (GTEx) project revealed that the expression of the gene encoding the enzyme that produces KA, kynurenine aminotransferase, correlated with cytokine abundance and activation of immune responses in older males. This study reveals that KA has a sex-specific link to immune responses and clinical outcomes in COVID-19, suggesting a positive feedback between metabolites and immune responses in males.

Immunometabolism: A 'Hot' Switch for 'Cold' Pediatric Solid Tumors.

Despite the success of immunotherapies in adult solid cancers and pediatric hematological malignancies, limited progress has been made towards implementing these strategies in pediatric solid tumors. These tumors exhibit a high potential to escape antitumor immunity, making them difficult to target by current immunotherapies. This review highlights the altered metabolic pathways in pediatric solid tumors that promote immune escape, and discusses current novel strategies targeting these pathways. We further explore how these strategies could be applied to potentiate immunotherapies for pediatric solid cancers and pose key questions yet to be addressed. Translational challenges to facilitate clinical application of antimetabolic strategies through personalized medicine are identified. We propose preclinical testing of antimetabolic approaches in combination with immunotherapies for pediatric solid cancers.

Amino Acid Trp: The Far Out Impacts of Host and Commensal Tryptophan Metabolism.

Tryptophan (Trp) is an essential amino acid primarily derived from the diet for use by the host for protein synthesis. The intestinal tract is lined with cells, both host and microbial, that uptake and metabolize Trp to also generate important signaling molecules. Serotonin (5-HT), kynurenine and its downstream metabolites, and to a lesser extent other neurotransmitters are generated by the host to signal onto host receptors and elicit physiological effects. 5-HT production by neurons in the CNS regulates sleep, mood, and appetite; 5-HT production in the intestinal tract by enterochromaffin cells regulates gastric motility and inflammation in the periphery. Kynurenine can signal onto the aryl hydrocarbon receptor (AHR) to elicit pleiotropic responses from several cell types including epithelial and immune cells, or can be further metabolized into bioactive molecules to influence neurodegenerative disease. There is a remarkable amount of cross-talk with the microbiome with regard to tryptophan metabolites as well. The gut microbiome can regulate the production of host tryptophan metabolites and can use dietary or recycled trp to generate bioactive metabolites themselves. Trp derivatives like indole are able to signal onto xenobiotic receptors, including AHR, to elicit tolerogenic effects. Here, we review studies that demonstrate that tryptophan represents a key intra-kingdom signaling molecule.

Physiological and Pathological Inflammation Induced by Antibodies and Pentraxins.

Macrophages play a key role in induction of inflammatory responses. These inflammatory responses are mostly considered to be instigated by activation of pattern recognition receptors (PRRs) or cytokine receptors. However, recently it has become clear that also antibodies and pentraxins, which can both activate Fc receptors (FcRs), induce very powerful inflammatory responses by macrophages that can even be an order of magnitude greater than PRRs. While the physiological function of this antibody-dependent inflammation (ADI) is to counteract infections, undesired activation or over-activation of this mechanism will lead to pathology, as observed in a variety of disorders, including viral infections such as COVID-19, chronic inflammatory disorders such as Crohn's disease, and autoimmune diseases such as rheumatoid arthritis. In this review we discuss how physiological ADI provides host defense by inducing pathogen-specific immunity, and how erroneous activation of this mechanism leads to pathology. Moreover, we will provide an overview of the currently known signaling and metabolic pathways that underlie ADI, and how these can be targeted to counteract pathological inflammation.

Targeting immunometabolism of neoplasms by interleukins: A promising immunotherapeutic strategy for cancer treatment.

In recent years, tumor metabolism has become a prevalent research topic for scientists and pharmaceutical companies. As research in the field has progressed, the metabolism-based therapy of tumors has ushered in new opportunities. Most tumors emerge and evolve under selective pressure from their microenvironment, which promotes the diversification of both neoplastic and non-neoplastic compartments of the tumor microenvironment (TME), and finally reaches a certain degree of intratumoral heterogeneity. As a result of the tumor intratumoral heterogeneity, tumor cells often possess a complex energy metabolism phenotype. During tumor progression, the metabolism for both tumor parenchyma and stroma is reprogrammed. The tumor stroma mainly consists of the extracellular matrix, fibroblasts, and immune cells. Interestingly, tumor-infiltrating immune cells utilize different metabolites based on their subtype and function, and these immunometabolic pathways can be modified in the TME. In particular, interleukins play a vital role in the activation and differentiation of immune cells and have exhibited multiple effects on tumor cell neoplasia, invasion, and metastasis. In this review, we summarize the common mechanisms of interleukins affecting the tumor and tumor-infiltrating immune cells metabolically and discuss how these mechanisms may lead to novel therapeutic opportunities. This review might contribute to the novel development of cancer immunotherapy.

Autophagy Regulates Stress Responses, Metabolism, and Anticancer Immunity.

Autophagy is a catabolic intracellular nutrient-scavenging pathway triggered by nutrient deprivation and stress that captures and degrades intracellular proteins and organelles in lysosomes. The breakdown products are then recycled into metabolic pathways to sustain survival. Organelle turnover by autophagy contributes to quality control and suppresses inflammation. Autophagy is upregulated in many cancers and supports their growth, survival, and malignancy in a tumor cell-autonomous fashion. Host autophagy also promotes tumor growth by maintaining a supply of essential nutrients and suppressing innate and adaptive antitumor immune responses. Autophagy is also upregulated in response to cancer therapy and confers treatment resistance. Thus, autophagy is a cancer vulnerability and its inhibition is under investigation as a novel therapeutic approach.

Jejunal mucosa proteomics unravel metabolic adaptive processes to mild chronic heat stress in dairy cows.

Climate change affects the duration and intensity of heat waves during summer months and jeopardizes animal health and welfare. High ambient temperatures cause heat stress in dairy cows resulting in a reduction of milk yield, feed intake, and alterations in gut barrier function. The objectives of this study were to investigate the mucosal amino acid, glucose and lactate metabolism, as well as the proteomic response of the small intestine in heat stressed (HS) Holstein dairy cows. Cows of the HS group (n = 5) were exposed for 4 days to 28 °C (THI = 76) in a climate chamber. Percentage decrease in daily ad libitum intake of HS cows was calculated to provide isocaloric energy intake to pair-fed control cows kept at 15 °C (THI = 60) for 4 days. The metabolite, mRNA and proteomic analyses revealed that HS induced incorrect protein folding, cellular destabilization, increased proteolytic degradation and protein kinase inhibitor activity, reduced glycolysis, and activation of NF-κB signaling, uronate cycling, pentose phosphate pathway, fatty acid and amino acid catabolism, mitochondrial respiration, ATPase activity and the antioxidative defence system. Our results highlight adaptive metabolic and immune mechanisms attempting to maintain the biological function in the small intestine of heat-stressed dairy cows.

Meet the author: Alice Clare Newman.

We talk to Alice Clare Newman, new mother, postdoc, and first author of "Immune-regulated IDO1-dependent tryptophan metabolism is source of one-carbon units for pancreatic cancer and stellate cells," about her paper, juggling family and career, and life-changing moments that led to the realization that life doesn't wait for science.

Ketogenic diet-mediated steroid metabolism reprogramming improves the immune microenvironment and myelin growth in spinal cord injury rats according to gene and co-expression network analyses.

The ketogenic diet has been widely used in the treatment of various nervous system and metabolic-related diseases. Our previous research found that a ketogenic diet exerts a protective effect and promotes functional recovery after spinal cord injury. However, the mechanism of action is still unclear. In this study, different dietary feeding methods were used, and myelin expression and gene level changes were detected among different groups. We established 15 RNA-seq cDNA libraries from among 4 different groups. First, KEGG pathway enrichment of upregulated differentially expressed genes and gene set enrichment analysis of the ketogenic diet and normal diet groups indicated that a ketogenic diet significantly improved the steroid anabolic pathway in rats with spinal cord injury. Through cluster analysis, protein-protein interaction analysis and visualization of iPath metabolic pathways, it was determined that Sqle, Sc5d, Cyp51, Dhcr24, Msmo1, Hsd17b7, and Fdft1 expression changed significantly. Second, through weighted gene co-expression network analysis showed that rats fed a ketogenic diet showed a significant reduction in the expression of genes involved in immune-related pathways, including those associated with immunity and infectious diseases. A ketogenic diet may improve the immune microenvironment and myelin growth in rats with spinal cord injury through reprogramming of steroid metabolism.

Conserved human effector Treg cell transcriptomic and epigenetic signature in arthritic joint inflammation.

Treg cells are critical regulators of immune homeostasis, and environment-driven Treg cell differentiation into effector (e)Treg cells is crucial for optimal functioning. However, human Treg cell programming in inflammation is unclear. Here, we combine transcriptional and epigenetic profiling to identify a human eTreg cell signature. Inflammation-derived functional Treg cells have a transcriptional profile characterized by upregulation of both a core Treg cell (FOXP3, CTLA4, TIGIT) and effector program (GITR, BLIMP-1, BATF). We identify a specific human eTreg cell signature that includes the vitamin D receptor (VDR) as a predicted regulator in eTreg cell differentiation. H3K27ac/H3K4me1 occupancy indicates an altered (super-)enhancer landscape, including enrichment of the VDR and BATF binding motifs. The Treg cell profile has striking overlap with tumor-infiltrating Treg cells. Our data demonstrate that human inflammation-derived Treg cells acquire a conserved and specific eTreg cell profile guided by epigenetic changes, and fine-tuned by environment-specific adaptations.