Faculty Bibliography

Gliomas are the most common primary brain tumors and a major source of mortality and morbidity in adults and children. Recent genomic studies have identified multiple molecular subtypes; however metabolic characterization of these tumors has thus far been limited. We performed metabolic profiling of 114 adult and pediatric primary gliomas and integrated metabolomic data with transcriptomics and DNA methylation classes. We identified that pediatric tumors have higher levels of glucose and reduced lactate compared to adult tumors regardless of underlying genetics or grade, suggesting differences in availability of glucose and/or utilization of glucose for downstream pathways. Differences in glucose utilization in pediatric gliomas may be facilitated through overexpression of SLC2A4, which encodes the insulin-stimulated glucose transporter GLUT4. Transcriptomic comparison of adult and pediatric tumors suggests that adult tumors may have limited access to glucose and experience more hypoxia, which is supported by enrichment of lactate, 2-hydroxyglutarate (2-HG), even in isocitrate dehydrogenase (IDH) wild-type tumors, and 3-hydroxybutyrate, a ketone body that is produced by oxidation of fatty acids and ketogenic amino acids during periods of glucose scarcity. Our data support adult tumors relying more on fatty acid oxidation, as they have an abundance of acyl carnitines compared to pediatric tumors and have significant enrichment of transcripts needed for oxidative phosphorylation. Our findings suggest striking differences exist in the metabolism of pediatric and adult gliomas, which can provide new insight into metabolic vulnerabilities for therapy.

Cancer cells often experience nutrient-limiting conditions because of their robust proliferation and inadequate tumour vasculature, which results in metabolic adaptation to sustain proliferation. Most cancer cells rapidly consume glucose, which is severely reduced in the nutrient-scarce tumour microenvironment. In CRISPR-based genetic screens to identify metabolic pathways influenced by glucose restriction, we find that tumour-relevant glucose concentrations (low glucose) protect cancer cells from inhibition of de novo pyrimidine biosynthesis, a pathway that is frequently targeted by chemotherapy. We identify two mechanisms to explain this result, which is observed broadly across cancer types. First, low glucose limits uridine-5-diphosphate-glucose synthesis, preserving pyrimidine nucleotide availability and thereby prolonging the time to replication fork stalling. Second, low glucose directly modulates apoptosis downstream of replication fork stalling by suppressing BAK activation and subsequent cytochrome c release, key events that activate caspase-9-dependent mitochondrial apoptosis. These results indicate that the low glucose levels frequently observed in tumours may limit the efficacy of specific chemotherapeutic agents, highlighting the importance of considering the effects of the tumour nutrient environment on cancer therapy.

UNLABELLED:Iron-sulfur clusters (ISCs) are cell-essential cofactors present in ∼60 proteins including subunits of OXPHOS complexes I-III, DNA polymerases, and iron-sensing proteins. Dysfunctions in ISC biosynthesis are associated with anemias, neurodegenerative disorders, and metabolic diseases. To assess consequences of acute ISC inhibition in a whole body setting, we developed a mouse model in which key ISC biosynthetic enzyme NFS1 can be acutely and reversibly suppressed. Contrary to in vitro ISC inhibition and pharmacological OXPHOS suppression, global NFS1 inhibition rapidly enhances lipid utilization and decreases adiposity without affecting caloric intake and physical activity. ISC proteins decrease, including key proteins involved in OXPHOS (SDHB), lipoic acid synthesis (LIAS), and insulin mRNA processing (CDKAL1), causing acute metabolic inflexibility. Age-related metabolic changes decelerate loss of adiposity substantially prolonged survival of mice with NFS1 inhibition. Thus, the observation that ISC metabolism impacts organismal fuel choice will aid in understanding the mechanisms underlying ISC diseases with increased risk for diabetes. HIGHLIGHTS/UNASSIGNED:Acute ISC inhibition leads to rapid loss of adiposity in miceMulti-metabolic pathway disruption upon ISC deficiency blocks energy storageNfs1 inhibition induces glucose dyshomeostasis due to ISC deficiency in β-cellsEnergy distress caused by inhibition of ISC synthesis is attenuated in aged mice.

Cells require micronutrients for numerous basic functions. Among these, iron, copper, and selenium are particularly critical for redox metabolism, and their importance is heightened during oncogene-driven perturbations in cancer. In this review, which particularly focuses on iron, we describe how these micronutrients are carefully chaperoned about the body and made available to tissues, a process that is designed to limit the toxicity of free iron and copper or by-products of selenium metabolism. We delineate perturbations in iron metabolism and iron-dependent proteins that are observed in cancer, and describe the current approaches being used to target iron metabolism and iron-dependent processes.

OBJECTIVE:To study the effects of Short Chain Fatty Acids (SCFAs) on arthritic bone remodeling. METHODS:CD4Cre mice, with SCFA supplemented water. We also performed in vitro osteoclast differentiation assays in the presence of serum-level SCFAs to evaluate the direct impact of these microbial metabolites on maturation and function of osteoclasts. We further characterized the molecular mechanism of SCFAs by transcriptional analysis. RESULTS:CD4Cre mice. Further interrogation revealed that bone marrow derived OCPs from diseased mice expressed a higher level of SCFA receptors than that of control mice and that the progenitor cells in the bone marrow of SCFA-treated mice presented a modified transcriptomic landscape, suggesting a direct impact of SCFAs on bone marrow progenitors in the context of osteoporosis. CONCLUSION/CONCLUSIONS:We demonstrated how gut microbiota-derived SCFAs can regulate distal pathology, i.e., osteoporosis, and identified a potential therapeutic option for restoring bone density in rheumatic disease, further highlighting the critical role of the gut-bone axis in these disorders.

Glioblastoma (GBM) is the most common and aggressive primary brain malignancy. Adhesion G protein-coupled receptors (aGPCRs) have attracted interest for their potential as treatment targets. Here, we show that CD97 (ADGRE5) is the most promising aGPCR target in GBM, by virtue of its de novo expression compared to healthy brain tissue. CD97 knockdown or knockout significantly reduces the tumor initiation capacity of patient-derived GBM cultures (PDGCs) in vitro and in vivo. We find that CD97 promotes glycolytic metabolism via the mitogen-activated protein kinase (MAPK) pathway, which depends on phosphorylation of its C terminus and recruitment of β-arrestin. We also demonstrate that THY1/CD90 is a likely CD97 ligand in GBM. Lastly, we show that an anti-CD97 antibody-drug conjugate selectively kills tumor cells in vitro. Our studies identify CD97 as a regulator of tumor metabolism, elucidate mechanisms of receptor activation and signaling, and provide strong scientific rationale for developing biologics to target it therapeutically in GBM.

Mitochondria must maintain adequate amounts of metabolites for protective and biosynthetic functions. However, how mitochondria sense the abundance of metabolites and regulate metabolic homeostasis is not well understood. In this work, we focused on glutathione (GSH), a critical redox metabolite in mitochondria, and identified a feedback mechanism that controls its abundance through the mitochondrial GSH transporter, SLC25A39. Under physiological conditions, SLC25A39 is rapidly degraded by mitochondrial protease AFG3L2. Depletion of GSH dissociates AFG3L2 from SLC25A39, causing a compensatory increase in mitochondrial GSH uptake. Genetic and proteomic analyses identified a putative iron-sulfur cluster in the matrix-facing loop of SLC25A39 as essential for this regulation, coupling mitochondrial iron homeostasis to GSH import. Altogether, our work revealed a paradigm for the autoregulatory control of metabolic homeostasis in organelles.

Kinases are key players in cancer-relevant pathways and are the targets of many successful precision cancer therapies (1, 2). Phosphoproteomics is a powerful approach to study kinase activity and has been used increasingly for the characterization of tumor samples leading to the identification of novel chemotherapeutic targets and biomarkers (3-10). Finding co-regulated phosphorylation sites which represent potential kinase-substrate sets or members of the same signaling pathway allows us to harness this data to identify clinically relevant and targetable alterations in signaling cascades. Unfortunately, studies have found that databases of co-regulated phosphorylation sites (11, 12) are only experimentally supported in a small number of substrate sets (13, 14). To address the inherent challenge of defining co-regulated phosphorylation modules relevant to a given dataset, we developed PhosphoDisco, a toolkit for determining co-regulated phosphorylation modules. We applied this approach to tandem mass spectrometry based phosphoproteomic data for breast and non-small cell lung cancer and identified canonical as well as putative new phosphorylation site modules. Our analysis identified several interesting modules in each cohort. Among these was a new cell cycle checkpoint module enriched in basal breast cancer samples and a module of PRKC isozymes putatively co-regulated by CDK12 in lung cancer. We demonstrate that modules defined by PhosphoDisco can be used to further personalized cancer treatment strategies by establishing active signaling pathways in a given patient tumor or set of tumors, and in providing new ways to classify tumors based on signaling activity.

Metabolism is a fundamental cellular process that is coordinated with cell cycle progression. Despite this association, a mechanistic understanding of cell cycle phase-dependent metabolic pathway regulation remains elusive. Here we report the mechanism by which human de novo pyrimidine biosynthesis is allosterically regulated during the cell cycle. Combining traditional synchronization methods and metabolomics, we characterize metabolites by their accumulation pattern during cell cycle phases and identify cell cycle phase-dependent regulation of carbamoyl-phosphate synthetase 2, aspartate transcarbamylase and dihydroorotase (CAD), the first, rate-limiting enzyme in de novo pyrimidine biosynthesis. Through systematic mutational scanning and structural modelling, we find allostery as a major regulatory mechanism that controls the activity change of CAD during the cell cycle. Specifically, we report evidence of two Animalia-specific loops in the CAD allosteric domain that involve sensing and binding of uridine 5'-triphosphate, a CAD allosteric inhibitor. Based on homology with a mitochondrial carbamoyl-phosphate synthetase homologue, we identify a critical role for a signal transmission loop in regulating the formation of a substrate channel, thereby controlling CAD activity.