Raman-based machine learning platform reveals unique metabolic differences between IDHmut and IDHwt glioma.

BACKGROUND: Formalin-fixed, paraffin-embedded (FFPE) tissue slides are routinely used in cancer diagnosis, clinical decision-making, and stored in biobanks, but their utilization in Raman spectroscopy-based studies has been limited due to the background coming from embedding media. METHODS: Spontaneous Raman spectroscopy was used for molecular fingerprinting of FFPE tissue from 46 patient samples with known methylation subtypes. Spectra were used to construct tumor/non-tumor, IDH1WT/IDH1mut, and methylation-subtype classifiers. Support vector machine and random forest were used to identify the most discriminatory Raman frequencies. Stimulated Raman spectroscopy was used to validate the frequencies identified. Mass spectrometry of glioma cell lines and TCGA were used to validate the biological findings. RESULTS: Here we develop APOLLO (rAman-based PathOLogy of maLignant glioma) - a computational workflow that predicts different subtypes of glioma from spontaneous Raman spectra of FFPE tissue slides. Our novel APOLLO platform distinguishes tumors from nontumor tissue and identifies novel Raman peaks corresponding to DNA and proteins that are more intense in the tumor. APOLLO differentiates isocitrate dehydrogenase 1 mutant (IDH1mut) from wildtype (IDH1WT) tumors and identifies cholesterol ester levels to be highly abundant in IDHmut glioma. Moreover, APOLLO achieves high discriminative power between finer, clinically relevant glioma methylation subtypes, distinguishing between the CpG island hypermethylated phenotype (G-CIMP)-high and G-CIMP-low molecular phenotypes within the IDH1mut types. CONCLUSIONS: Our results demonstrate the potential of label-free Raman spectroscopy to classify glioma subtypes from FFPE slides and to extract meaningful biological information thus opening the door for future applications on these archived tissues in other cancers.

DMT1 contributes to MF- 438 - mediated inhibition of glioma cells.

Elevated SCD1 expression has been associated with enhanced cancer cell survival, proliferation, and resistance to therapy in many cancer types including gliomas. Hereby, we investigate the impact of MF-438 on SCD1-mediated lipid metabolism and its consequences on glioma growth and survival. Our data reveals an IDH mut -specific inhibitory effect of MF438 on gliomas. Also, we delineate a dual mechanism of action: while SCD1-mediated lipid metabolism is hindered by MF-438 treatment, MF-438 also exerts an SCD1-independent inhibition on DMT1 expression. Supporting data from the DMT1 blocker underscores its significance in MF-438's anti-glioma efficacy.

Targeting the sphingolipid rheostat in IDH1 mut glioma alters cholesterol homeostasis and triggers apoptosis via membrane degradation.

The cross-regulation of metabolism and trafficking is not well understood for the vital sphingolipids and cholesterol constituents of cellular compartments. While reports are starting to surface on how sphingolipids like sphingomyelin (SM) dysregulate cholesterol levels in different cellular compartments (Jiang et al., 2022), limited research is available on the mechanisms driving the relationship between sphingolipids and cholesterol homeostasis, or its biological implications. Previously, we have identified sphingolipid metabolism as a unique vulnerability for IDH1 mut gliomas via a rational drug design. Herein, we show how modulating sphingolipid levels affects cholesterol homeostasis in brain tumors. However, we unexpectedly discovered for the first time that C17 sphingosine and NDMS addition to cancer cells alters cholesterol homeostasis by impacting its cellular synthesis, uptake, and efflux leading to a net decrease in cholesterol levels and inducing apoptosis. Our results reflect a reverse correlation between the levels of sphingosines, NDMS, and unesterified, free cholesterol in the cells. We show that increasing sphingosine and NDMS (a sphingosine analog) levels alter not only the trafficking of cholesterol between membranes but also the efflux and synthesis of cholesterol. We also demonstrate that despite the effort to remove free cholesterol by ABCA1-mediated efflux or by suppressing machinery for the influx (LDLR) and biosynthetic pathway (HMGCR), apoptosis is inevitable for IDH1 mut glioma cells. This is the first study that shows how altering sphingosine levels directly affects cholesterol homeostasis in cancer cells and can be used to manipulate this relationship to induce apoptosis in IDH1 mut gliomas.

Targeting IDH1-Mutated Oligodendroglioma with Acid Ceramidase Inhibitors.

Oligodendroglioma is genetically defined as a tumor harboring isocitrate dehydrogenase 1 or 2 mutations (IDH1 mut /IDH2 mut ) and 1p/19q co-deletions. Previously, we reported that in IDH1 mut gliomas, D-2HG, the product of IDH1 mutant enzyme produces an increase in monounsaturated fatty acid levels that are incorporated into ceramides, tilting the S1P-to-ceramide rheostat toward apoptosis. Herein, we exploited this imbalance to further induce and IDH mut -specific glioma cell death. We report for the first time that the inhibition of acid ceramidase (AC) induces apoptosis and provides a benefit in mice survival in IDH1 mut oligodendroglioma. We demonstrated an IDH1 mut -specific cytotoxicity of SABRAC, an irreversible inhibitor of AC, in patient-derived oligodendroglioma cells. Exploring the mechanism of action of this drug, we found that SABRAC activates both extrinsic and intrinsic apoptosis in an ER stress-independent manner, pointing to a direct action of AC-related ceramides in mitochondria permeability. The activation of apoptosis detected under SABRAC treatment was associated with up to 30-fold increase in some ceramide levels and its derivatives from the salvage pathway. We propose that this novel enzyme, AC, has the potential to increase survival in oligodendroglioma with IDH1 mut and should be considered in the future.

Resolving Challenges in Detection and Quantification of D-2-hydroxyglutarate and L-2-hydroxyglutarate via LC/MS.

D-2-Hydroxyglutarate and L-2-Hydroxyglutarate (D-2HG/L-2HG) are typically metabolites of non-specific enzymatic reactions that are kept in check by the housekeeping enzymes, D-2HG /L-2HG dehydrogenase (D-2HGDH/L-2HGDH). In certain disease states, such as D-2HG or L-2HG aciduria and cancers, accumulation of these biomarkers interferes with oxoglutarate-dependent enzymes that regulate bioenergetic metabolism, histone methylation, post-translational modification, protein expression and others. D-2HG has a complex role in tumorigenesis that drives metabolomics investigations. Meanwhile, L-2HG is produced by non-specific action of malate dehydrogenase and lactate dehydrogenase under acidic or hypoxic environments. Characterization of divergent effects of D-2HG/L-2HG on the activity of specific enzymes in diseased metabolism depends on their accurate quantification via mass spectrometry. Despite advancements in high-resolution quadrupole time-of-flight mass spectrometry (HR-QTOF-MS), challenges are typically encountered when attempting to resolve of isobaric and isomeric metabolites such as D-2HG/L-2HG for quantitative analysis. Herein, available D-2HG/L-2HG derivatization and liquid chromatography (LC) MS quantification methods were examined. The outcome led to the development of a robust, high-throughput HR-QTOF-LC/MS approach that permits concomitant quantification of the D-2HG and L-2HG enantiomers with the benefit to quantify the dysregulation of other intermediates within interconnecting pathways. Calibration curve was obtained over the linear range of 0.8-104 nmol/mL with r 2 ≥ 0.995 for each enantiomer. The LC/MS-based assay had an overall precision with intra-day CV % ≤ 8.0 and inter-day CV % ≤ 6.3 across the quality control level for commercial standard and pooled biological samples; relative error % ≤ 2.7 for accuracy; and resolution, R s = 1.6 between 2HG enantiomers (m/z 147.030), D-2HG and L-2HG (at retention time of 5.82 min and 4.75 min, respectively) following chiral derivatization with diacetyl-L-tartaric anhydride (DATAN). Our methodology was applied to disease relevant samples to illustrate the implications of proper enantioselective quantification of both D-2HG and L-2HG. The stability of the method allows scaling to large cohorts of clinical samples in the future.

The patatin-like protein PlpD forms structurally dynamic homodimers in the Pseudomonas aeruginosa outer membrane.

Members of the Omp85 superfamily of outer membrane proteins (OMPs) found in Gram-negative bacteria, mitochondria and chloroplasts are characterized by a distinctive 16-stranded ß-barrel transmembrane domain and at least one periplasmic POTRA domain. All previously studied Omp85 proteins promote critical OMP assembly and/or protein translocation reactions. Pseudomonas aeruginosa PlpD is the prototype of an Omp85 protein family that contains an N-terminal patatin-like (PL) domain that is thought to be translocated across the OM by a C-terminal ß-barrel domain. Challenging the current dogma, we find that the PlpD PL-domain resides exclusively in the periplasm and, unlike previously studied Omp85 proteins, PlpD forms a homodimer. Remarkably, the PL-domain contains a segment that exhibits unprecedented dynamicity by undergoing transient strand-swapping with the neighboring ß-barrel domain. Our results show that the Omp85 superfamily is more structurally diverse than currently believed and suggest that the Omp85 scaffold was utilized during evolution to generate novel functions.

TGF-ß uncouples glycolysis and inflammation in macrophages and controls survival during sepsis.

Changes in metabolism of macrophages are required to sustain macrophage activation in response to different stimuli. We showed that the cytokine TGF-ß (transforming growth factor-ß) regulates glycolysis in macrophages independently of inflammatory cytokine production and affects survival in mouse models of sepsis. During macrophage activation, TGF-ß increased the expression and activity of the glycolytic enzyme PFKL (phosphofructokinase-1 liver type) and promoted glycolysis but suppressed the production of proinflammatory cytokines. The increase in glycolysis was mediated by an mTOR-c-MYC-dependent pathway, whereas the inhibition of cytokine production was due to activation of the transcriptional coactivator SMAD3 and suppression of the activity of the proinflammatory transcription factors AP-1, NF-κB, and STAT1. In mice with LPS-induced endotoxemia and experimentally induced sepsis, the TGF-ß-induced enhancement in macrophage glycolysis led to decreased survival, which was associated with increased blood coagulation. Analysis of septic patient cohorts revealed that the expression of PFKL, TGFBRI (which encodes a TGF-ß receptor), and F13A1 (which encodes a coagulation factor) in myeloid cells positively correlated with COVID-19 disease. Thus, these results suggest that TGF-ß is a critical regulator of macrophage metabolism and could be a therapeutic target in patients with sepsis.

T cell exhaustion in malignant gliomas.

Despite advances in understanding tumor biology, malignant gliomas remain incurable. While immunotherapy has improved outcomes in other cancer types, comparable efficacy has not yet been demonstrated for primary cancers of the central nervous system (CNS). T cell exhaustion, defined as a progressive decrease in effector function, sustained expression of inhibitory receptors, metabolic dysfunction, and distinct epigenetic and transcriptional alterations, contributes to the failure of immunotherapy in the CNS. Herein, we describe recent advances in understanding the drivers of T cell exhaustion in the glioma microenvironment. We discuss the extrinsic and intrinsic factors that contribute to exhaustion and highlight potential avenues for reversing this phenotype. Our ability to directly target specific immunosuppressive drivers in brain cancers would be a major advance in immunotherapy.

Metabolic biomarkers of radiotherapy response in plasma and tissue of an IDH1 mutant astrocytoma mouse model.

Astrocytomas are the most common subtype of brain tumors and no curative treatment exist. Longitudinal assessment of patients, usually via Magnetic Resonance Imaging (MRI), is crucial since tumor progression may occur earlier than clinical progression. MRI usually provides a means for monitoring the disease, but it only informs about the structural changes of the tumor, while molecular changes can occur as a treatment response without any MRI-visible change. Radiotherapy (RT) is routinely performed following surgery as part of the standard of care in astrocytomas, that can also include chemotherapy involving temozolomide. Monitoring the response to RT is a key factor for the management of patients. Herein, we provide plasma and tissue metabolic biomarkers of treatment response in a mouse model of astrocytoma that was subjected to radiotherapy. Plasma metabolic profiles acquired over time by Liquid Chromatography Mass Spectrometry (LC/MS) were subjected to multivariate empirical Bayes time-series analysis (MEBA) and Receiver Operating Characteristic (ROC) assessment including Random Forest as the classification strategy. These analyses revealed a variation of the plasma metabolome in those mice that underwent radiotherapy compared to controls; specifically, fumarate was the best discriminatory feature. Additionally, Nuclear Magnetic Resonance (NMR)-based 13C-tracing experiments were performed at end-point utilizing [U-13C]-Glutamine to investigate its fate in the tumor and contralateral tissues. Irradiated mice displayed lower levels of glycolytic metabolites (e.g. phosphoenolpyruvate) in tumor tissue, and a higher flux of glutamine towards succinate was observed in the radiation cohort. The plasma biomarkers provided herein could be validated in the clinic, thereby improving the assessment of brain tumor patients throughout radiotherapy. Moreover, the metabolic rewiring associated to radiotherapy in tumor tissue could lead to potential metabolic imaging approaches for monitoring treatment using blood draws.

Probabilistic model checking of cancer metabolism.

Cancer cell metabolism is often deregulated as a result of adaption to meeting energy and biosynthesis demands of rapid growth or direct mutation of key metabolic enzymes. Better understanding of such deregulation can provide new insights on targetable vulnerabilities, but is complicated by the difficulty in probing cell metabolism at different levels of resolution and under different experimental conditions. We construct computational models of glucose and glutamine metabolism with focus on the effect of IDH1/2-mutations in cancer using a combination of experimental metabolic flux data and patient-derived gene expression data. Our models demonstrate the potential of computational exploration to reveal biologic behavior: they show that an exogenously-mutated IDH1 experimental model utilizes glutamine as an alternative carbon source for lactate production under hypoxia, but does not fully-recapitulate the patient phenotype under normoxia. We also demonstrate the utility of using gene expression data as a proxy for relative differences in metabolic activity. We use the approach of probabilistic model checking and the freely-available Probabilistic Symbolic Model Checker to construct and reason about model behavior.

Single cell metabolism: current and future trends.

Single cell metabolomics is an emerging and rapidly developing field that complements developments in single cell analysis by genomics and proteomics. Major goals include mapping and quantifying the metabolome in sufficient detail to provide useful information about cellular function in highly heterogeneous systems such as tissue, ultimately with spatial resolution at the individual cell level. The chemical diversity and dynamic range of metabolites poses particular challenges for detection, identification and quantification. In this review we discuss both significant technical issues of measurement and interpretation, and progress toward addressing them, with recent examples from diverse biological systems. We provide a framework for further directions aimed at improving workflow and robustness so that such analyses may become commonly applied, especially in combination with metabolic imaging and single cell transcriptomics and proteomics.

Advances in measuring cancer cell metabolism with subcellular resolution.

Characterizing metabolism in cancer is crucial for understanding tumor biology and for developing potential therapies. Although most metabolic investigations analyze averaged metabolite levels from all cell compartments, subcellular metabolomics can provide more detailed insight into the biochemical processes associated with the disease. Methodological limitations have historically prevented the wider application of subcellular metabolomics in cancer research. Recently, however, ways to distinguish and identify metabolic pathways within organelles have been developed, including state-of-the-art methods to monitor metabolism in situ (such as mass spectrometry-based imaging, Raman spectroscopy and fluorescence microscopy), to isolate key organelles via new approaches and to use tailored isotope-tracing strategies. Herein, we examine the advantages and limitations of these developments and look to the future of this field of research.

Targeting the Sphingolipid Rheostat in Gliomas.

Gliomas are highly aggressive cancer types that are in urgent need of novel drugs and targeted therapies. Treatment protocols have not improved in over a decade, and glioma patient survival remains among the worst of all cancer types. As a result, cancer metabolism research has served as an innovative approach to identifying novel glioma targets and improving our understanding of brain tumors. Recent research has uncovered a unique metabolic vulnerability in the sphingolipid pathways of gliomas that possess the IDH1 mutation. Sphingolipids are a family of lipid signaling molecules that play a variety of second messenger functions in cellular regulation. The two primary metabolites, sphingosine-1-phosphate (S1P) and ceramide, maintain a rheostat balance and play opposing roles in cell survival and proliferation. Altering the rheostat such that the pro-apoptotic signaling of the ceramides outweighs the pro-survival S1P signaling in glioma cells diminishes the hallmarks of cancer and enhances tumor cell death. Throughout this review, we discuss the sphingolipid pathway and identify the enzymes that can be most effectively targeted to alter the sphingolipid rheostat and enhance apoptosis in gliomas. We discuss each pathway's steps based on their site of occurrence in the organelles and postulate novel targets that can effectively exploit this vulnerability.

Cryo-EM structures reveal multiple stages of bacterial outer membrane protein folding.

Transmembrane ß barrel proteins are folded into the outer membrane (OM) of Gram-negative bacteria by the ß barrel assembly machinery (BAM) via a poorly understood process that occurs without known external energy sources. Here, we used single-particle cryo-EM to visualize the folding dynamics of a model ß barrel protein (EspP) by BAM. We found that BAM binds the highly conserved "ß signal" motif of EspP to correctly orient ß strands in the OM during folding. We also found that the folding of EspP proceeds via "hybrid-barrel" intermediates in which membrane integrated ß sheets are attached to the essential BAM subunit, BamA. The structures show an unprecedented deflection of the membrane surrounding the EspP intermediates and suggest that ß sheets progressively fold toward BamA to form a ß barrel. Along with in vivo experiments that tracked ß barrel folding while the OM tension was modified, our results support a model in which BAM harnesses OM elasticity to accelerate ß barrel folding.

IDH-mutated gliomas promote epileptogenesis through d-2-hydroxyglutarate-dependent mTOR hyperactivation.

BACKGROUND: Uncontrolled seizures in patients with gliomas have a significant impact on quality of life and morbidity, yet the mechanisms through which these tumors cause seizures remain unknown. Here, we hypothesize that the active metabolite d-2-hydroxyglutarate (d-2-HG) produced by the IDH-mutant enzyme leads to metabolic disruptions in surrounding cortical neurons that consequently promote seizures. METHODS: We use a complementary study of in vitro neuron-glial cultures and electrographically sorted human cortical tissue from patients with IDH-mutant gliomas to test this hypothesis. We utilize micro-electrode arrays for in vitro electrophysiological studies in combination with pharmacological manipulations and biochemical studies to better elucidate the impact of d-2-HG on cortical metabolism and neuronal spiking activity. RESULTS: We demonstrate that d-2-HG leads to increased neuronal spiking activity and promotes a distinct metabolic profile in surrounding neurons, evidenced by distinct metabolomic shifts and increased LDHA expression, as well as upregulation of mTOR signaling. The increases in neuronal activity are induced by mTOR activation and reversed with mTOR inhibition. CONCLUSION: Together, our data suggest that metabolic disruptions in the surrounding cortex due to d-2-HG may be a driving event for epileptogenesis in patients with IDH-mutant gliomas.

Cysteine is a limiting factor for glioma proliferation and survival.

Nutritional intervention is becoming more prevalent as adjuvant therapy for many cancers in view of the tumor dependence on external sources for some nutrients. However, little is known about the mechanisms that make cancer cells require certain nutrients from the microenvironment. Herein, we report the dependence of glioma cells on exogenous cysteine/cystine, despite this amino acid being nonessential. Using several 13 C-tracers and analysis of cystathionine synthase and cystathioninase levels, we revealed that glioma cells were not able to support glutathione synthesis through the transsulfuration pathway, which allows methionine to be converted to cysteine in cysteine/cystine-deprived conditions. Therefore, we explored the nutritional deprivation in a mouse model of glioma. Animals subjected to a cysteine/cystine-free diet survived longer, although this increase did not attain statistical significance, with concomitant reductions in plasma glutathione and cysteine levels. At the end point, however, tumors displayed the ability to synthesize glutathione, even though higher levels of oxidative stress were detected. We observed a compensation from the nutritional intervention revealed as the recovery of cysteine-related metabolite levels in plasma. Our study highlights a time window where cysteine deprivation can be exploited for additional therapeutic strategies.

Reversing Epigenetic Gene Silencing to Overcome Immune Evasion in CNS Malignancies.

Glioblastoma (GBM) is an aggressive brain malignancy with a dismal prognosis. With emerging evidence to disprove brain-immune privilege, there has been much interest in examining immunotherapy strategies to treat central nervous system (CNS) cancers. Unfortunately, the limited success of clinical studies investigating immunotherapy regimens, has led to questions about the suitability of immunotherapy for these cancers. Inadequate inherent populations of tumor infiltrating lymphocytes (TILs) and limited trafficking of systemic, circulating T cells into the CNS likely contribute to the poor response to immunotherapy. This paucity of TILs is in concert with the finding of epigenetic silencing of genes that promote immune cell movement (chemotaxis) to the tumor. In this study we evaluated the ability of GSK126, a blood-brain barrier (BBB) permeable small molecule inhibitor of EZH2, to reverse GBM immune evasion by epigenetic suppression of T cell chemotaxis. We also evaluated the in vivo efficacy of this drug in combination with anti-PD-1 treatment on tumor growth, survival and T cell infiltration in syngeneic mouse models. GSK126 reversed H3K27me3 in murine and human GBM cell lines. When combined with anti-PD-1 treatment, a significant increase in activated T cell infiltration into the tumor was observed. This resulted in decreased tumor growth and enhanced survival both in sub-cutaneous and intracranial tumors of immunocompetent, syngeneic murine models of GBM. Additionally, a significant increase in CXCR3+ T cells was also seen in the draining lymph nodes, suggesting their readiness to migrate to the tumor. Closer examination of the mechanism of action of GSK126 revealed its ability to promote the expression of IFN-γ driven chemokines CXCL9 and CXCL10 from the tumor cells, that work to traffic T cells without directly affecting T maturation and/or proliferation. The loss of survival benefit either with single agent or combination in immunocompromised SCID mice, suggest that the therapeutic efficacy of GSK126 in GBM is primarily driven by lymphocytes. Taken together, our data suggests that in glioblastoma, epigenetic modulation using GSK126 could improve current immunotherapy strategies by reversing the epigenetic changes that enable immune cell evasion leading to enhanced immune cell trafficking to the tumor.

A Single-Organelle Optical Omics Platform for Cell Science and Biomarker Discovery.

Research in fundamental cell biology and pathology could be revolutionized by developing the capacity for quantitative molecular analysis of subcellular structures. To that end, we introduce the Ramanomics platform, based on confocal Raman microspectrometry coupled to a biomolecular component analysis algorithm, which together enable us to molecularly profile single organelles in a live-cell environment. This emerging omics approach categorizes the entire molecular makeup of a sample into about a dozen of general classes and subclasses of biomolecules and quantifies their amounts in submicrometer volumes. A major contribution of our study is an attempt to bridge Raman spectrometry with big-data analysis in order to identify complex patterns of biomolecules in a single cellular organelle and leverage discovery of disease biomarkers. Our data reveal significant variations in organellar composition between different cell lines. We also demonstrate the merits of Ramanomics for identifying diseased cells by using prostate cancer as an example. We report large-scale molecular transformations in the mitochondria, Golgi apparatus, and endoplasmic reticulum that accompany the development of prostate cancer. Based on these findings, we propose that Ramanomics datasets in distinct organelles constitute signatures of cellular metabolism in healthy and diseased states.

Magnetic resonance spectroscopy for the study of cns malignancies.

Despite intensive research, brain tumors are amongst the malignancies with the worst prognosis; therefore, a prompt diagnosis and thoughtful assessment of the disease is required. The resistance of brain tumors to most forms of conventional therapy has led researchers to explore the underlying biology in search of new vulnerabilities and biomarkers. The unique metabolism of brain tumors represents one potential vulnerability and the basis for a system of classification. Profiling this aberrant metabolism requires a method to accurately measure and report differences in metabolite concentrations. Magnetic resonance-based techniques provide a framework for examining tumor tissue and the evolution of disease. Nuclear Magnetic Resonance (NMR) analysis of biofluids collected from patients suffering from brain cancer can provide biological information about disease status. In particular, urine and plasma can serve to monitor the evolution of disease through the changes observed in the metabolic profiles. Moreover, cerebrospinal fluid can be utilized as a direct reporter of cerebral activity since it carries the chemicals exchanged with the brain tissue and the tumor mass. Metabolic reprogramming has recently been included as one of the hallmarks of cancer. Accordingly, the metabolic rewiring experienced by these tumors to sustain rapid growth and proliferation can also serve as a potential therapeutic target. The combination of 13C tracing approaches with the utilization of different NMR spectral modalities has allowed investigations of the upregulation of glycolysis in the aggressive forms of brain tumors, including glioblastomas, and the discovery of the utilization of acetate as an alternative cellular fuel in brain metastasis and gliomas. One of the major contributions of magnetic resonance to the assessment of brain tumors has been the non-invasive determination of 2-hydroxyglutarate (2HG) in tumors harboring a mutation in isocitrate dehydrogenase 1 (IDH1). The mutational status of this enzyme already serves as a key feature in the clinical classification of brain neoplasia in routine clinical practice and pilot studies have established the use of in vivo magnetic resonance spectroscopy (MRS) for monitoring disease progression and treatment response in IDH mutant gliomas. However, the development of bespoke methods for 2HG detection by MRS has been required, and this has prevented the wider implementation of MRS methodology into the clinic. One of the main challenges for improving the management of the disease is to obtain an accurate insight into the response to treatment, so that the patient can be promptly diverted into a new therapy if resistant or maintained on the original therapy if responsive. The implementation of 13C hyperpolarized magnetic resonance spectroscopic imaging (MRSI) has allowed detection of changes in tumor metabolism associated with a treatment, and as such has been revealed as a remarkable tool for monitoring response to therapeutic strategies. In summary, the application of magnetic resonance-based methodologies to the diagnosis and management of brain tumor patients, in addition to its utilization in the investigation of its tumor-associated metabolic rewiring, is helping to unravel the biological basis of malignancies of the central nervous system.