bims-glucam 2021-01-24 papers

bims-glucam

Biomed News

on Glutamine cancer metabolism

Issue of 2021–01–24
sixteen papers selected by
Sreeparna Banerjee, Middle East Technical University

Glioma cells require one-carbon metabolism to survive glutamine starvation.
Glutaminolysis is a metabolic route essential for survival and growth of prostate cancer cells and a target of 5α-dihydrotestosterone regulation.
The role of the glutamine transporter ASCT2 in antineoplastic therapy.
More than just protein building blocks: How amino acids and related metabolic pathways fuel macrophage polarization.
Host cell glutamine metabolism as a potential antiviral target.
Targeting the Proline-Glutamine-Asparagine-Arginine Metabolic Axis in Amino Acid Starvation Cancer Therapy.
Personalized Genome-Scale Metabolic Models Identify Targets of Redox Metabolism in Radiation-Resistant Tumors.
The metabolism of cancer cells during metastasis.
Molecular Mechanisms Linking Nutrition to Metabolic Homeostasis: An Overview Picture of Current Understanding.
The Source of Glycolytic Intermediates in Mammalian Tissues.
Human pancreatic cancer cells under nutrient deprivation are vulnerable to redox system inhibition.
Aglycemic growth enhances carbohydrate metabolism and induces sensitivity to menadione in cultured tumor-derived cells.
Clinically-targetable vulnerabilities in cancer metabolism: A systematic review and meta-analysis.
The pleiotropic functions of autophagy in metastasis.
Crystal structures of glutathione- and inhibitor-bound human GGT1: critical interactions within the cysteinylglycine binding site.
MYCN-amplified neuroblastoma is addicted to iron and vulnerable to inhibition of the system Xc-/glutathione axis.

Acta Neuropathol Commun. 2021 Jan 19. 9(1): 16

Glioma cells require one-carbon metabolism to survive glutamine starvation.

Kazuhiro Tanaka, Takashi Sasayama, Hiroaki Nagashima, Yasuhiro Irino, Masatomo Takahashi, Yoshihiro Izumi, Takiko Uno, Naoko Satoh, Akane Kitta, Katsusuke Kyotani, Yuichi Fujita, Mitsuru Hashiguchi, Tomoaki Nakai, Masaaki Kohta, Yoichi Uozumi, Masakazu Shinohara, Kohkichi Hosoda, Takeshi Bamba, Eiji Kohmura.

  Cancer cells optimize nutrient utilization to supply energetic and biosynthetic pathways. This metabolic process also includes redox maintenance and epigenetic regulation through nucleic acid and protein methylation, which enhance tumorigenicity and clinical resistance. However, less is known about how cancer cells exhibit metabolic flexibility to sustain cell growth and survival from nutrient starvation. Here, we find that serine and glycine levels were higher in low-nutrient regions of tumors in glioblastoma multiforme (GBM) patients than they were in other regions. Metabolic and functional studies in GBM cells demonstrated that serine availability and one-carbon metabolism support glioma cell survival following glutamine deprivation. Serine synthesis was mediated through autophagy rather than glycolysis. Gene expression analysis identified upregulation of methylenetetrahydrofolate dehydrogenase 2 (MTHFD2) to regulate one-carbon metabolism. In clinical samples, MTHFD2 expression was highest in the nutrient-poor areas around "pseudopalisading necrosis." Genetic suppression of MTHFD2 and autophagy inhibition caused tumor cell death and growth inhibition of glioma cells upon glutamine deprivation. These results highlight a critical role for serine-dependent one-carbon metabolism in surviving glutamine starvation and suggest new therapeutic targets for glioma cells adapting to a low-nutrient microenvironment.

Keywords:  Glioblastoma multiforme; Glutamine starvation; One-carbon metabolism; Serine synthesis

DOI:  https://doi.org/10.1186/s40478-020-01114-1
Cell Oncol (Dordr). 2021 Jan 19.

Glutaminolysis is a metabolic route essential for survival and growth of prostate cancer cells and a target of 5α-dihydrotestosterone regulation.

Henrique J Cardoso, Marília I Figueira, Cátia V Vaz, Tiago M A Carvalho, Luís A Brás, Patrícia A Madureira, Paulo J Oliveira, Vilma A Sardão, Sílvia Socorro.

   PURPOSE: Resistance to androgen-deprivation therapies and progression to so-called castrate-resistant prostate cancer (CRPC) remain challenges in prostate cancer (PCa) management and treatment. Among other alterations, CRPC has been associated with metabolic reprogramming driven by androgens. Here, we investigated the role of androgens in regulating glutaminolysis in PCa cells and determined the relevance of this metabolic route in controlling the survival and growth of androgen-sensitive (LNCaP) and CRPC (DU145 and PC3) cells.
METHODS: PCa cells (LNCaP, DU145 and PC3) and 3-month old rats were treated with 5α-dihydrotestosterone (DHT). Alternatively, LNCaP cells were exposed to the glutaminase inhibitor BPTES, alone or in combination with the anti-androgen bicalutamide. Biochemical, Western blot and extracellular flux assays were used to evaluate the viability, proliferation, migration and metabolism of PCa cells in response to DHT treatment or glutaminase inhibition.
RESULTS: We found that DHT up-regulated the expression of the glutamine transporter ASCT2 and glutaminase, both in vitro in LNCaP cells and in vivo in rat prostate cells. BPTES diminished the viability and migration of PCa cells, while increasing caspase-3 activity. CRPC cells were found to be more dependent on glutamine and more sensitive to glutaminase inhibition. BPTES and bicalutamide co-treatment had an additive effect on suppressing LNCaP cell viability. Finally, we found that inhibition of glutaminolysis differentially affected glycolysis and lipid metabolism in both androgen-sensitive and CRPC cells.
CONCLUSION: Our data reveal glutaminolysis as a central metabolic route controlling PCa cell fate and highlight the relevance of targeting glutaminase for CRPC treatment.

Keywords:  5α-dihydrotestosterone; ASCT2; BPTES; Bicalutamide; Castrate resistance; Glutamine; Glutaminolysis; Prostate cancer

DOI:  https://doi.org/10.1007/s13402-020-00575-9
Cancer Chemother Pharmacol. 2021 Jan 19.

The role of the glutamine transporter ASCT2 in antineoplastic therapy.

Estefânia Teixeira, Cláudia Silva, Fátima Martel.

  Cancer cells are metabolically reprogrammed to support their high rates of proliferation, continuous growth, survival, invasion, metastasis, and resistance to cancer treatments. Among changes in cancer cell bioenergetics, the role of glutamine metabolism has been receiving increasing attention. Increased glutaminolysis in cancer cells is associated with increased expression of membrane transporters that mediate the cellular uptake of glutamine. ASCT2 (Alanine, Serine, Cysteine Transporter 2) is a Na+-dependent transmembrane transporter overexpressed in cancer cells and considered to be the primary transporter for glutamine in these cells. The possibility of inhibiting ASCT2 for antineoplastic therapy is currently under investigation. In this article, we will present the pharmacological agents currently known to act on ASCT2, which have been attracting attention in antineoplastic therapy research. We will also address the impact of ASCT2 inhibition on the prognosis of some cancers. We conclude that ASCT2 inhibition and combination of ASCT2 inhibitors with other anti-tumor therapies may be a promising antineoplastic strategy. However, more research is needed in this area.

Keywords:  ASCT2; Antineoplastic therapy; Glutamine uptake; Metabolic reprogramming

DOI:  https://doi.org/10.1007/s00280-020-04218-6
FEBS J. 2021 Jan 18.

More than just protein building blocks: How amino acids and related metabolic pathways fuel macrophage polarization.

Markus Kieler, Melanie Hofmann, Gernot Schabbauer.

  Macrophages represent the first line of defence in innate immune responses and additionally serve important functions for the regulation of host inflammation and tissue homeostasis. The M1/M2 model describes the two extremes of macrophage polarization states, which can be induced by multiple stimuli, most notably by LPS/IFN-γ and IL-4/IL-13. Historically, the expression of two genes encoding for enzymes, which use the same amino acid as their substrate, iNOS and ARG1, has been used to define classically activated M1 (iNOS) and alternatively activated M2 (ARG1) macrophages. This "arginine dichotomy" has recently become a matter of debate; however, in parallel with the emerging field of immunometabolism it is becoming more and more evident that these two enzymes and their related metabolites are fundamentally involved in the intrinsic regulation of macrophage polarization and function. The aim of this review is to highlight recent advances in macrophage biology and immunometabolism with a specific focus on amino acid metabolism and their related metabolic pathways: urea cycle (arginine), TCA cycle and OXPHOS (glutamine) as well as the one carbon metabolism (serine, glycine).

Keywords:  TCA cycle; arginase/iNOS; glutamine; immunometabolism; macrophage polarization; nitric oxide; oxidative phosphorylation; polyamines; serine; α-ketoglutarate

DOI:  https://doi.org/10.1111/febs.15715
Clin Sci (Lond). 2021 Jan 29. 135(2): 305-325

Host cell glutamine metabolism as a potential antiviral target.

Sandro Massao Hirabara, Renata Gorjao, Adriana Cristina Levada-Pires, Laureane Nunes Masi, Elaine Hatanaka, Maria Fernanda Cury-Boaventura, Eliane Borges da Silva, Laiane Cristina Dos Santos-Oliveira, Vinicius Leonardo Sousa Diniz, Tamires Afonso Duarte Serdan, Vivian Araujo Barbosa de Oliveira, Diego Ribeiro de Souza, Raquel Bragante Gritte, Talita Souza-Siqueira, Raquel Freitas Zambonatto, Tania Cristina Pithon-Curi, Roberto Barbosa Bazotte, Philip Newsholme, Rui Curi.

  A virus minimally contains a nucleic acid genome packaged by a protein coat. The genome and capsid together are known as the nucleocapsid, which has an envelope containing a lipid bilayer (mainly phospholipids) originating from host cell membranes. The viral envelope has transmembrane proteins that are usually glycoproteins. The proteins in the envelope bind to host cell receptors, promoting membrane fusion and viral entry into the cell. Virus-infected host cells exhibit marked increases in glutamine utilization and metabolism. Glutamine metabolism generates ATP and precursors for the synthesis of macromolecules to assemble progeny viruses. Some compounds derived from glutamine are used in the synthesis of purines and pyrimidines. These latter compounds are precursors for the synthesis of nucleotides. Inhibitors of glutamine transport and metabolism are potential candidate antiviral drugs. Glutamine is also an essential nutrient for the functions of leukocytes (lymphocyte, macrophage, and neutrophil), including those in virus-infected patients. The increased glutamine requirement for immune cell functions occurs concomitantly with the high glutamine utilization by host cells in virus-infected patients. The development of antiviral drugs that target glutamine metabolism must then be specifically directed at virus-infected host cells to avoid negative effects on immune functions. Therefore, the aim of this review was to describe the landscape of cellular glutamine metabolism to search for potential candidates to inhibit glutamine transport or glutamine metabolism.

Keywords:  Coronavirus; Glutamine Inhibitors; Glutaminolysis; Metabolic Reprogramming; glutamate

DOI:  https://doi.org/10.1042/CS20201042
Pharmaceuticals (Basel). 2021 Jan 18. pii: E72. [Epub ahead of print]14(1):

Targeting the Proline-Glutamine-Asparagine-Arginine Metabolic Axis in Amino Acid Starvation Cancer Therapy.

Macus Tien Kuo, Helen H W Chen, Lynn G Feun, Niramol Savaraj.

  Proline, glutamine, asparagine, and arginine are conditionally non-essential amino acids that can be produced in our body. However, they are essential for the growth of highly proliferative cells such as cancers. Many cancers express reduced levels of these amino acids and thus require import from the environment. Meanwhile, the biosynthesis of these amino acids is inter-connected but can be intervened individually through the inhibition of key enzymes of the biosynthesis of these amino acids, resulting in amino acid starvation and cell death. Amino acid starvation strategies have been in various stages of clinical applications. Targeting asparagine using asparaginase has been approved for treating acute lymphoblastic leukemia. Targeting glutamine and arginine starvations are in various stages of clinical trials, and targeting proline starvation is in preclinical development. The most important obstacle of these therapies is drug resistance, which is mostly due to reactivation of the key enzymes involved in biosynthesis of the targeted amino acids and reprogramming of compensatory survival pathways via transcriptional, epigenetic, and post-translational mechanisms. Here, we review the interactive regulatory mechanisms that control cellular levels of these amino acids for amino acid starvation therapy and how drug resistance is evolved underlying treatment failure.

Keywords:  amino acid starvation therapy; arginine; asparagine; drug resistance; glutamine; proline

DOI:  https://doi.org/10.3390/ph14010072
Cell Syst. 2021 Jan 20. pii: S2405-4712(20)30502-0. [Epub ahead of print]12(1): 68-81.e11

Personalized Genome-Scale Metabolic Models Identify Targets of Redox Metabolism in Radiation-Resistant Tumors.

Joshua E Lewis, Tom E Forshaw, David A Boothman, Cristina M Furdui, Melissa L Kemp.

  Redox cofactor production is integral toward antioxidant generation, clearance of reactive oxygen species, and overall tumor response to ionizing radiation treatment. To identify systems-level alterations in redox metabolism that confer resistance to radiation therapy, we developed a bioinformatics pipeline for integrating multi-omics data into personalized genome-scale flux balance analysis models of 716 radiation-sensitive and 199 radiation-resistant tumors. These models collectively predicted that radiation-resistant tumors reroute metabolic flux to increase mitochondrial NADPH stores and reactive oxygen species (ROS) scavenging. Simulated genome-wide knockout screens agreed with experimental siRNA gene knockdowns in matched radiation-sensitive and radiation-resistant cancer cell lines, revealing gene targets involved in mitochondrial NADPH production, central carbon metabolism, and folate metabolism that allow for selective inhibition of glutathione production and H2O2 clearance in radiation-resistant cancers. This systems approach represents a significant advancement in developing quantitative genome-scale models of redox metabolism and identifying personalized metabolic targets for improving radiation sensitivity in individual cancer patients.

Keywords:  NADPH; The Cancer Genome Atlas; flux balance analysis; genome-scale; glutathione; hydrogen peroxide; personalized models; radiation resistance; reactive oxygen species; redox metabolism

DOI:  https://doi.org/10.1016/j.cels.2020.12.001
Nat Rev Cancer. 2021 Jan 18.

The metabolism of cancer cells during metastasis.

Gabriele Bergers, Sarah-Maria Fendt.

Metastasis formation is the major cause of death in most patients with cancer. Despite extensive research, targeting metastatic seeding and colonization is still an unresolved challenge. Only recently, attention has been drawn to the fact that metastasizing cancer cells selectively and dynamically adapt their metabolism at every step during the metastatic cascade. Moreover, many metastases display different metabolic traits compared with the tumours from which they originate, enabling survival and growth in the new environment. Consequently, the stage-dependent metabolic traits may provide therapeutic windows for preventing or reducing metastasis, and targeting the new metabolic traits arising in established metastases may allow their eradication.

DOI: https://doi.org/10.1038/s41568-020-00320-2
Crit Rev Eukaryot Gene Expr. 2020 ;30(6): 543-564

Molecular Mechanisms Linking Nutrition to Metabolic Homeostasis: An Overview Picture of Current Understanding.

Carla Pignatti, Stefania D'Adamo, Flavio Flamigni, Silvia Cetrulllo.

Increasing evidence supports the notion that in humans many pathological conditions including obesity, metabolic syndrome, and type 2 diabetes are closely related to the amount and quality of each nutritional component and to an impairment of the metabolic homeostatic mechanisms of their utilization. Cell signaling pathways that sense the availability of nutrients and the energy status of the cells communicate with signaling pahways triggered by hormones and growth factors to coordinately regulate whole-body metabolic homeostasis. The aim of this review is to provide an overview picture of current knowledge about the main molecular mechanisms that connect nutritional status, hormones, and nutrient levels with gene expression, metabolic homeostasis, and nutrient sensing. We recapitulate molecular mechanisms governing fuel selection between glucose and fatty acids in different nutritional conditions, highlighting metabolic flexibility as mechanism to ensure metabolic health. Disrupted metabolic flexibility, or metabolic inflexibility, is associated with many pathological conditions including metabolic syndrome, type 2 diabetes mellitus, and cancer. We also describe how macronutrients that can be used as energy sources may reciprocally modulate their own metabolism as well as directly interact with transcriptional factors, nutrient sensors and nutrient sensing pathways in order to achieve metabolic homeostasis.

DOI: https://doi.org/10.1615/CritRevEukaryotGeneExpr.2020037120
Cell Metab. 2021 Jan 18. pii: S1550-4131(20)30728-2. [Epub ahead of print]

The Source of Glycolytic Intermediates in Mammalian Tissues.

Tara TeSlaa, Caroline R Bartman, Connor S R Jankowski, Zhaoyue Zhang, Xincheng Xu, Xi Xing, Lin Wang, Wenyun Lu, Sheng Hui, Joshua D Rabinowitz.

  Glycolysis plays a central role in organismal metabolism, but its quantitative inputs across mammalian tissues remain unclear. Here we use 13C-tracing in mice to quantify glycolytic intermediate sources: circulating glucose, intra-tissue glycogen, and circulating gluconeogenic precursors. Circulating glucose is the main source of circulating lactate, the primary end product of tissue glycolysis. Yet circulating glucose highly labels glycolytic intermediates in only a few tissues: blood, spleen, diaphragm, and soleus muscle. Most glycolytic intermediates in the bulk of body tissue, including liver and quadriceps muscle, come instead from glycogen. Gluconeogenesis contributes less but also broadly to glycolytic intermediates, and its flux persists with physiologic feeding (but not hyperinsulinemic clamp). Instead of suppressing gluconeogenesis, feeding activates oxidation of circulating glucose and lactate to maintain glucose homeostasis. Thus, the bulk of the body slowly breaks down internally stored glycogen while select tissues rapidly catabolize circulating glucose to lactate for oxidation throughout the body.

Keywords:  compartmentalized metabolism; glucose homeostasis; glycogen; glycolysis; glycolytic intermediates; glycolytic specialist; isotope tracing; metabolic heterogeneity; red muscle

DOI:  https://doi.org/10.1016/j.cmet.2020.12.020
J Biol Chem. 2020 Dec 04. pii: S0021-9258(17)50484-6. [Epub ahead of print]295(49): 16678-16690

Human pancreatic cancer cells under nutrient deprivation are vulnerable to redox system inhibition.

Takefumi Onodera, Isao Momose, Hayamitsu Adachi, Yohko Yamazaki, Ryuichi Sawa, Shun-Ichi Ohba, Manabu Kawada.

  Large regions in tumor tissues, particularly pancreatic cancer, are hypoxic and nutrient-deprived because of unregulated cell growth and insufficient vascular supply. Certain cancer cells, such as those inside a tumor, can tolerate these severe conditions and survive for prolonged periods. We hypothesized that small molecular agents, which can preferentially reduce cancer cell survival under nutrient-deprived conditions, could function as anticancer drugs. In this study, we constructed a high-throughput screening system to identify such small molecules and screened chemical libraries and microbial culture extracts. We were able to determine that some small molecular compounds, such as penicillic acid, papyracillic acid, and auranofin, exhibit preferential cytotoxicity to human pancreatic cancer cells under nutrient-deprived compared with nutrient-sufficient conditions. Further analysis revealed that these compounds target to redox systems such as GSH and thioredoxin and induce accumulation of reactive oxygen species in nutrient-deprived cancer cells, potentially contributing to apoptosis under nutrient-deprived conditions. Nutrient-deficient cancer cells are often deficient in GSH; thus, they are susceptible to redox system inhibitors. Targeting redox systems might be an attractive therapeutic strategy under nutrient-deprived conditions of the tumor microenvironment.

Keywords:  auranofin; cancer therapy; chemical biology; drug discovery; drug screening; glutathione; metabolism; oxidation reduction (redox); oxidative stress; papyracillic acid; penicillic acid; redox regulation; thioredoxin

DOI:  https://doi.org/10.1074/jbc.RA120.013893
Cancer Metab. 2021 Jan 19. 9(1): 3

Aglycemic growth enhances carbohydrate metabolism and induces sensitivity to menadione in cultured tumor-derived cells.

Cameron A Schmidt, Kelsey L McLaughlin, Ilya N Boykov, Rafiq Mojalagbe, Arthi Ranganathan, Katherine A Buddo, Chien-Te Lin, Kelsey H Fisher-Wellman, P Darrell Neufer.

   BACKGROUND: Hepatocellular carcinoma (HCC) is the most prevalent form of liver malignancy and carries poor prognoses due to late presentation of symptoms. Treatment of late-stage HCC relies heavily on chemotherapeutics, many of which target cellular energy metabolism. A key platform for testing candidate chemotherapeutic compounds is the intrahepatic orthotopic xenograft (IOX) model in rodents. Translational efficacy from the IOX model to clinical use is limited (in part) by variation in the metabolic phenotypes of the tumor-derived cells that can be induced by selective adaptation to subculture conditions.
METHODS: In this study, a detailed multilevel systems approach combining microscopy, respirometry, potentiometry, and extracellular flux analysis (EFA) was utilized to examine metabolic adaptations that occur under aglycemic growth media conditions in HCC-derived (HEPG2) cells. We hypothesized that aglycemic growth would result in adaptive "aerobic poise" characterized by enhanced capacity for oxidative phosphorylation over a range of physiological energetic demand states.
RESULTS: Aglycemic growth did not invoke adaptive changes in mitochondrial content, network complexity, or intrinsic functional capacity/efficiency. In intact cells, aglycemic growth markedly enhanced fermentative glycolytic substrate-level phosphorylation during glucose refeeding and enhanced responsiveness of both fermentation and oxidative phosphorylation to stimulated energy demand. Additionally, aglycemic growth induced sensitivity of HEPG2 cells to the provitamin menadione at a 25-fold lower dose compared to control cells.
CONCLUSIONS: These findings indicate that growth media conditions have substantial effects on the energy metabolism of subcultured tumor-derived cells, which may have significant implications for chemotherapeutic sensitivity during incorporation in IOX testing panels. Additionally, the metabolic phenotyping approach used in this study provides a practical workflow that can be incorporated with IOX screening practices to aid in deciphering the metabolic underpinnings of chemotherapeutic drug sensitivity.

Keywords:  Cancer; Confocal microscopy; Galactose; Glycolysis; HEPG2; Mitochondria; Oroboros; Oxidative phosphorylation; Seahorse xf24

DOI:  https://doi.org/10.1186/s40170-021-00241-0
J Pak Med Assoc. 2020 Dec;70(12(A)): 2226-2238

Clinically-targetable vulnerabilities in cancer metabolism: A systematic review and meta-analysis.

Arslaan Javaeed, Sanniya Khan Ghauri.

   OBJECTIVE: To investigate the efficacy and safety of targeting cancer metabolic vulnerabilities with specific anticancer agents.
METHODS: The systematic review and meta-analysis entailed search on PubMed, Embase and Google Scholar databases for cohort-based studies or clinical trials which reported hazard ratio for overall survival and/or median overall survival of patients treated with metabolicallyactive anticancer drugs. Data was analysed using the Number Cruncher Statistical System version 11.
RESULTS: There were 16 studies published between 1989 and 2018 that reported improvement in the overall survival (p=0.05) despite the reported significant heterogeneity across the studies (I2=70%). Exploiting amino acid metabolic vulnerabilities was associated with a favourable prognostic outcome (p=0.05), while targeting glycolysis and nucleic acid synthesis had no significant clinical importance (p>0.05).
CONCLUSIONS: There is an urgent need to develop future therapies relying on the synergistic actions of nucleotide biosynthesis, glycolysis and amino acid metabolism.

Keywords:   Metabolic vulnerabilities, Cancer, Chemotherapy, Cell metabolism, Metabolic enzymes.

DOI:  https://doi.org/10.47391/JPMA.079
J Cell Sci. 2021 Jan 22. pii: jcs247056. [Epub ahead of print]134(2):

The pleiotropic functions of autophagy in metastasis.

Timothy Marsh, Bhairavi Tolani, Jayanta Debnath.

  Autophagy is deregulated in many cancers and represents an attractive target for therapeutic intervention. However, the precise contributions of autophagy to metastatic progression, the principle cause of cancer-related mortality, is only now being uncovered. While autophagy promotes primary tumor growth, metabolic adaptation and resistance to therapy, recent studies have unexpectedly revealed that autophagy suppresses the proliferative outgrowth of disseminated tumor cells into overt and lethal macrometastases. These studies suggest autophagy plays unexpected and complex roles in the initiation and progression of metastases, which will undoubtedly impact therapeutic approaches for cancer treatment. Here, we discuss the intricacies of autophagy in metastatic progression, highlighting and integrating the pleiotropic roles of autophagy on diverse cell biological processes involved in metastasis.

Keywords:  Autophagy; Cancer; Metastasis; Selective Autophagy

DOI:  https://doi.org/10.1242/jcs.247056
J Biol Chem. 2020 Nov 22. pii: S0021-9258(20)00052-6. [Epub ahead of print]296 100066

Crystal structures of glutathione- and inhibitor-bound human GGT1: critical interactions within the cysteinylglycine binding site.

Simon S Terzyan, Luong T Nguyen, Anthony W G Burgett, Annie Heroux, Clyde A Smith, Youngjae You, Marie H Hanigan.

  Overexpression of γ-glutamyl transpeptidase (GGT1) has been implicated in an array of human diseases including asthma, reperfusion injury, and cancer. Inhibitors are needed for therapy, but development of potent, specific inhibitors of GGT1 has been hampered by a lack of structural information regarding substrate binding and cleavage. To enhance our understanding of the molecular mechanism of substrate cleavage, we have solved the crystal structures of human GGT1 (hGGT1) with glutathione (a substrate) and a phosphate-glutathione analog (an irreversible inhibitor) bound in the active site. These are the first structures of any eukaryotic GGT with the cysteinylglycine region of the substrate-binding site occupied. These structures and the structure of apo-hGGT reveal movement of amino acid residues within the active site as the substrate binds. Asn-401 and Thr-381 each form hydrogen bonds with two atoms of GSH spanning the γ-glutamyl bond. Three different atoms of hGGT1 interact with the carboxyl oxygen of the cysteine of GSH. Interactions between the enzyme and substrate change as the substrate moves deeper into the active site cleft. The substrate reorients and a new hydrogen bond is formed between the substrate and the oxyanion hole. Thr-381 is locked into a single conformation as an acyl bond forms between the substrate and the enzyme. These data provide insight on a molecular level into the substrate specificity of hGGT1 and provide an explanation for seemingly disparate observations regarding the enzymatic activity of hGGT1 mutants. This knowledge will aid in the design of clinically useful hGGT1 inhibitors.

Keywords:  crystal structure; crystallography; enzyme; enzyme inhibitor; glutathione; structural biology; γ-glutamyl transferase; γ-glutamyl transpeptidase

DOI:  https://doi.org/10.1074/jbc.RA120.016265
Cancer Res. 2021 Jan 22. pii: canres.1641.2020. [Epub ahead of print]

MYCN-amplified neuroblastoma is addicted to iron and vulnerable to inhibition of the system Xc-/glutathione axis.

Konstantinos V Floros, JinYang Cai, Sheeba Jacob, Richard Kurupi, Carter K Fairchild, Mayuri Shende, Colin M Coon, Krista M Powell, Benjamin R Belvin, Bin Hu, Madhavi Puchalapalli, Sivapriya Ramamoorthy, Kimberly Swift, Janina P Lewis, Mikhail G Dozmorov, John Glod, Jennifer E Koblinski, Sosipatros A Boikos, Anthony C Faber.

MYCN is amplified in 20-25% of neuroblastoma, and MYCN-amplified neuroblastoma contributes to a large percent of pediatric cancer-related deaths. Therapy improvements for this subtype of cancer is a high priority. Here we uncover a MYCN-dependent therapeutic vulnerability in neuroblastoma. Namely, amplified MYCN rewired the cell through expression of key receptors, ultimately enhancing iron influx through increased expression of the iron import transferrin receptor 1 (TfR1). Accumulating iron caused reactive oxygen species (ROS) production, and MYCN-amplified neuroblastomas showed enhanced reliance on the system Xc- cystine/glutamate antiporter for ROS detoxification through increased transcription of this receptor. This dependence created a marked vulnerability to targeting the system Xc-/glutathione (GSH) pathway with ferroptosis inducers. This reliance can be exploited through therapy with FDA-approved rheumatoid arthritis (RA) drugs sulfasalazine (SAS) and auranofin: in MYCN-amplified, patient-derived xenograft models, both therapies blocked growth and induced ferroptosis. SAS and auranofin activity was largely mitigated by the ferroptosis inhibitor ferrostatin-1, antioxidants like NAC, or by the iron scavenger deferoxamine (DFO). DFO reduced auranofin-induced ROS, further linking increased iron capture in MYCN-amplified NB to a therapeutic vulnerability to ROS-inducing drugs. These data uncover an oncogene vulnerability to ferroptosis caused by increased iron accumulation and subsequent reliance on the system Xc-/GSH pathway.

DOI: https://doi.org/10.1158/0008-5472.CAN-20-1641