bims-glucam 2023-07-30 papers

bims-glucam

Biomed News

on Glutamine cancer metabolism

Issue of 2023–07–30
eightteen papers selected by
Sreeparna Banerjee, Middle East Technical University

Targeting Glutamine Metabolism through Glutaminase Inhibition Suppresses Cell Proliferation and Progression in Nasopharyngeal Carcinoma.
Monitoring and modelling the glutamine metabolic pathway: a review and future perspectives.
Metabolic Profiles Point Out Metabolic Pathways Pivotal in Two Glioblastoma (GBM) Cell Lines, U251 and U-87MG.
PI3K signaling-regulated metabolic reprogramming: From mechanism to application.
Loss of the mitochondrial citrate carrier, Slc25a1/CIC disrupts embryogenesis via 2-Hydroxyglutarate.
3-hydroxykynurenine is a ROS-inducing cytotoxic tryptophan metabolite that disrupts the TCA cycle.
Surviving the hunger games: Metabolic reprogramming in medulloblastoma.
Metabolic stress-induced reciprocal loop of long noncoding RNA ZFAS1 and ZEB1 promotes epithelial-mesenchymal transition and metastasis of pancreatic cancer cells.
Atractylenolide III Ameliorates Bile Duct Ligation-Induced Liver Fibrosis by Inhibiting the PI3K/AKT Pathway and Regulating Glutamine Metabolism.
Metabolism, metabolites, and macrophages in cancer.
A Prognostic Model of Head and Neck Cancer Based on Amino Acid Metabolism-Related Signature and Its Implication for Immunosuppressive Microenvironment.
Differential Inhibition of Anaplerotic Pyruvate Carboxylation and Glutaminolysis-Fueled Anabolism Underlies Distinct Toxicity of Selenium Agents in Human Lung Cancer.
The Fe-S cluster assembly protein IscU2 increases α-ketoglutarate catabolism and DNA 5mC to promote tumor growth.
Remodelling of the Mitochondrial Bioenergetic Pathways in Human Cultured Fibroblasts with Carbohydrates.
VHL-deficiency leads to reductive stress in renal cells.
Quantification of glutathione with high throughput live-cell imaging.
Engineering of a GSH activatable photosensitizer for enhanced photodynamic therapy through disrupting redox homeostasis.
Therapeutic Efficacy, Radiotoxicity and Abscopal Effect of BNCT at the RA-3 Nuclear Reactor Employing Oligo-Fucoidan and Glutamine as Adjuvants in an Ectopic Colon Cancer Model in Rats.

Anticancer Agents Med Chem. 2023 Jul 27.

Targeting Glutamine Metabolism through Glutaminase Inhibition Suppresses Cell Proliferation and Progression in Nasopharyngeal Carcinoma.

Chang Su, Minghan Li, Yuxin Yang, Ziying Wang, Qianru Wang, Weijia Wang, Xuemin Ma, Rongrong Jie, Huaihong Chen, Xiangping Li, Juan Lu.

   BACKGROUND: Glutaminase (GLS), the key enzyme involved in glutamine metabolism, has been identified as a critical player in tumor growth and progression. The GLS inhibitor CB-839 has entered several clinical trials against a variety of tumors.
OBJECTIVE: Our study aimed to investigate the role and underlying mechanism of GLS and its inhibitor CB-839 in nasopharyngeal carcinoma (NPC).
METHODS: The expression, downstream genes, and signaling pathways of GLS in NPC were determined by real-time polymerase chain reaction (RT-PCR), PCR array, western blotting (WB), and immunohistochemical staining (IHC), and the phenotype of GLS was confirmed by in vivo experiments of subcutaneous tumor formation in mice and in vitro experiments of functional biology, including Cell Counting Kit-8 (CCK-8), colony formation, flow cytometry, transwell migration, and Boyden invasion assay. Finally, it was also verified whether the treatment of NPC cells by GLS inhibitor CB-839 can change various biological functions and protein expression to achieve the purpose of blocking tumor progression.
RESULTS: GLS was remarkably overexpressed in NPC cells and tissues, predicting a poor overall survival of NPC patients. GLS promoted cell cycle, proliferation, colony formation, migratory, and invasive capacities by regulating Cyclin D2 (CCND2) via PI3K/AKT/mTOR pathway in NPC in vitro and in vivo. Notably, CB-839 showed an effective anti-NPC tumor effect by blocking the biological functions of the tumor.
CONCLUSION: The first innovative proof is that GLS promotes cell proliferation by regulating CCND2 via PI3K/AKT/mTOR pathway in NPC, and GLS inhibitor CB-839 may serve as a new potential therapeutic target for NPC treatment.

Keywords:  CCND2; GLS; Nasopharyngeal carcinoma; PI3K/AKT/mTOR pathway; Proliferation

DOI:  https://doi.org/10.2174/1871520623666230727104825
Metabolomics. 2023 Jul 23. 19(8): 67

Monitoring and modelling the glutamine metabolic pathway: a review and future perspectives.

Zohreh Mirveis, Orla Howe, Paul Cahill, Nitin Patil, Hugh J Byrne.

   BACKGROUND: Analysis of the glutamine metabolic pathway has taken a special place in metabolomics research in recent years, given its important role in cell biosynthesis and bioenergetics across several disorders, especially in cancer cell survival. The science of metabolomics addresses the intricate intracellular metabolic network by exploring and understanding how cells function and respond to external or internal perturbations to identify potential therapeutic targets. However, despite recent advances in metabolomics, monitoring the kinetics of a metabolic pathway in a living cell in situ, real-time and holistically remains a significant challenge.
AIM: This review paper explores the range of analytical approaches for monitoring metabolic pathways, as well as physicochemical modeling techniques, with a focus on glutamine metabolism. We discuss the advantages and disadvantages of each method and explore the potential of label-free Raman microspectroscopy, in conjunction with kinetic modeling, to enable real-time and in situ monitoring of the cellular kinetics of the glutamine metabolic pathway.
KEY SCIENTIFIC CONCEPTS: Given its important role in cell metabolism, the ability to monitor and model the glutamine metabolic pathways are highlighted. Novel, label free approaches have the potential to revolutionise metabolic biosensing, laying the foundation for a new paradigm in metabolomics research and addressing the challenges in monitoring metabolic pathways in living cells.

Keywords:  Chemometrics; Computational modelling; Flux analysis; Glutaminolysis; Metabolic pathways analysis; Vibrational spectroscopy

DOI:  https://doi.org/10.1007/s11306-023-02031-9
Biomedicines. 2023 Jul 20. pii: 2041. [Epub ahead of print]11(7):

Metabolic Profiles Point Out Metabolic Pathways Pivotal in Two Glioblastoma (GBM) Cell Lines, U251 and U-87MG.

Filipa Martins, David van der Kellen, Luís G Gonçalves, Jacinta Serpa.

  Glioblastoma (GBM) is the most lethal central nervous system (CNS) tumor, mainly due to its high heterogeneity, invasiveness, and proliferation rate. These tumors remain a therapeutic challenge, and there are still some gaps in the GBM biology literature. Despite the significant amount of knowledge produced by research on cancer metabolism, its implementation in cancer treatment has been limited. In this study, we explored transcriptomics data from the TCGA database to provide new insights for future definition of metabolism-related patterns useful for clinical applications. Moreover, we investigated the impact of key metabolites (glucose, lactate, glutamine, and glutamate) in the gene expression and metabolic profile of two GBM cell lines, U251 and U-87MG, together with the impact of these organic compounds on malignancy cell features. GBM cell lines were able to adapt to the exposure to each tested organic compound. Both cell lines fulfilled glycolysis in the presence of glucose and were able to produce and consume lactate. Glutamine dependency was also highlighted, and glutamine and glutamate availability favored biosynthesis observed by the increase in the expression of genes involved in fatty acid (FA) synthesis. These findings are relevant and point out metabolic pathways to be targeted in GBM and also reinforce that patients' metabolic profiling can be useful in terms of personalized medicine.

Keywords:  cancer metabolism; gene expression profiles; glioblastoma; metabolic remodeling; metabolomics

DOI:  https://doi.org/10.3390/biomedicines11072041
Biochim Biophys Acta Rev Cancer. 2023 Jul 24. pii: S0304-419X(23)00101-4. [Epub ahead of print] 188952

PI3K signaling-regulated metabolic reprogramming: From mechanism to application.

Wen Ya Su, Lu Yao Tian, Lan Pin Guo, Lu Qi Huang, Wen Yuan Gao.

  Oncogenic signaling involved in tumor metabolic reprogramming. Tumorigenesis was not only determined by the mutations or deletion of oncogenes but also accompanied by the reprogramming of cellular metabolism. Metabolic alterations play a crucial regulatory role in the development and progression of tumors. Oncogenic PI3K/AKT signaling mediates the metabolic switch in cancer cells and immune cells in the tumor microenvironment. PI3K/AKT and its downstream effector branch off and connect to multiple steps of metabolism, such as glucose, lipids, and amino acids. Thus, PI3K inhibitor could effectively regulate metabolic pathway and impede the oncogenic process and some key metabolic proteins or critical enzymes also constitute biomarkers for tumor diagnosis and treatment. In the current review, we summarize the significant effect of PI3K/AKT signaling toward tumor metabolism, it enables us to obtain the better understanding for this interaction and develop more effective therapeutic strategies targeting cancer cell metabolism.

Keywords:  Amino acid metabolism; Fatty acid metabolism; Glycolysis; Metabolic reprogramming; PI3K/AKT

DOI:  https://doi.org/10.1016/j.bbcan.2023.188952
bioRxiv. 2023 Jul 23. pii: 2023.07.18.549409. [Epub ahead of print]

Loss of the mitochondrial citrate carrier, Slc25a1/CIC disrupts embryogenesis via 2-Hydroxyglutarate.

Anna Kasprzyk-Pawelec, Mingjun Tan, Yu Leng Phua, Raneen Rahhal, Alec McIntosh, Harvey Fernandez, Rami Mosaoa, Michael Girgis, Amrita Cheema, Lei Jiang, Lawrence F Kroemer, Anastas Popratiloff, Cheryl Clarkson, Brian M Kirmsa, Gray W Pearson, Eric Glasgow, Christopher Albanese, Jerry Vockley, Maria Laura Avantaggiati.

Biallelic germline mutations in the SLC25A1 gene lead to combined D/L-2-hydroxyglutaric aciduria (D/L-2HGA), a fatal systemic disease uniquely characterized by the accumulation of both enantiomers of 2-hydroxyglutaric acid (2HG). How SLC25A1 deficiency contributes to D/L-2HGA and the role played by 2HG is unclear and no therapy exists. Both enantiomers act as oncometabolites, but their activities in normal tissues remain understudied. Here we show that mice lacking both SLC25A1 alleles exhibit developmental abnormalities that mirror human D/L-2HGA. SLC25A1 deficient cells undergo premature mitochondrial dysfunction associated senescence, suggesting that loss of proliferative capacity underlies the pathogenesis of D/L-2HGA. Remarkably, D- and L-2HG directly induce mitochondrial respiratory deficit and treatment of zebrafish embryos with the combination of D- and L-2HG phenocopies SLC25A1 loss, leading to developmental abnormalities in an additive fashion relative to either enantiomer alone. Metabolic analyses demonstrated that loss of SLC25A1 leads to global remodeling towards glutamine metabolism, with glutamine serving as a source for 2HG synthesis. Therefore, we explored the pre-clinical relevance of phenylbutyrate, an FDA-approved drug that reduces the blood glutamine levels, and found that it reduces 2HG accumulation reversing metabolic abnormalities in patients affected by D/L-2HGA. These results reveal pathogenic and growth suppressive activities of 2HG in the context of SLC25A1 deficiency and expose metabolic vulnerabilities for the clinical management of this disease.

DOI: https://doi.org/10.1101/2023.07.18.549409
bioRxiv. 2023 Jul 10. pii: 2023.07.10.548411. [Epub ahead of print]

3-hydroxykynurenine is a ROS-inducing cytotoxic tryptophan metabolite that disrupts the TCA cycle.

Jane L Buchanan, Adam J Rauckhorst, Eric B Taylor.

Tryptophan is an essential amino acid that is extensively characterized as a regulator of cellular function through its metabolism by indoleamine 2,3-deoxygenase (IDO) into the kynurenine pathway. However, despite decades of research on tryptophan metabolism, the metabolic regulatory roles of it and its metabolites are not well understood. To address this, we performed an activity metabolomics screen of tryptophan and most of its known metabolites in cell culture. We discovered that treatment of human colon cancer cells (HCT116) with 3-hydroxykynurenine (3-HK), a metabolite of kynurenine, potently disrupted TCA cycle function. Citrate and aconitate levels were increased, while isocitrate and all downstream TCA metabolites were decreased, suggesting decreased aconitase function. We hypothesized that 3HK or one of its metabolites increased reactive oxygen species (ROS) and inhibited aconitase activity. Accordingly, we observed almost complete depletion of reduced glutathione and a decrease in total glutathione levels. We observed a dose-dependent decrease in cell viability after 48 hours of 3HK treatment. These data suggest that raising the intracellular levels of 3HK could be sufficient to induce ROS-mediated apoptosis. We modulated the intracellular levels of 3HK by combined induction of IDO and knockdown of kynureninase (KYNU) in HCT116 cells. Cell viability decreased significantly after 48 hours of KYNU knockdown compared to controls, which was accompanied by increased ROS production and Annexin V staining revealing apoptosis. Finally, we identify xanthommatin production from 3-HK as a candidate radical-producing, cytotoxic mechanism. Our work indicates that KYNU may be a target for disrupting tryptophan metabolism. Interestingly, many cancers exhibit overexpression of IDO, providing a cancer-specific metabolic vulnerability that could be exploited by KYNU inhibition.

DOI: https://doi.org/10.1101/2023.07.10.548411
Biochem Pharmacol. 2023 Jul 20. pii: S0006-2952(23)00288-5. [Epub ahead of print]215 115697

Surviving the hunger games: Metabolic reprogramming in medulloblastoma.

Lorenzo Manfreda, Elena Rampazzo, Luca Persano, Giampietro Viola, Roberta Bortolozzi.

  Medulloblastoma is a highly malignant pediatric brain tumor characterized by its aggressive nature and limited treatment options. Metabolic changes have recently emerged as key factors in the development, progression, and response to therapy in various types of cancer. Cancer cells exhibit remarkable adaptability by modulating glucose, lipids, amino acids, and nucleotide metabolism to survive in nutrient- and oxygen-deprived environments. Although medulloblastoma has been extensively studied from a genomic perspective, leading to the identification of four subgroups and their respective subcategories, the investigation of its metabolic phenotype has remained relatively understudied. This review focus on the available literature, aiming to summarize the current knowledge about the main metabolic pathways that are deregulated in medulloblastoma tumors, while emphasizing the controversial aspects and the progress that is yet to be made. Furthermore, we underscored the insights gained so far regarding the impact of metabolism on the development of drug resistance in medulloblastoma and the therapeutic strategies employed to target specific metabolic pathways.

Keywords:  Central-carbon metabolism; Drugs targeting metabolism; Medulloblastoma; Medulloblastoma subgroups; Metabolic reprogramming; Metabolic vulnerability

DOI:  https://doi.org/10.1016/j.bcp.2023.115697
Cancer Sci. 2023 Jul 24.

Metabolic stress-induced reciprocal loop of long noncoding RNA ZFAS1 and ZEB1 promotes epithelial-mesenchymal transition and metastasis of pancreatic cancer cells.

Wenfeng Zhuo, Zhu Zeng, Yuhang Hu, Ping Hu, Shengbo Han, Decai Wang, Fan Wang, Yong Zhao, Yan Huang, Jie Wang, Guozheng Lv, Hongda Wang, Yang Li, Eryang Zhao, Kailin Cai, Gang Zhao.

  Pancreatic cancer (PC) development faces significant metabolic stress due to metabolic reprogramming and a distinct hypovascular nature, often leading to glucose and glutamine depletion. However, the adaption mechanisms by which PC adapts to these metabolic challenges have not yet been completely explored. Here, we found that metabolic stress induced by glucose and glutamine deprivation led to an overexpression of ZNFX1 antisense RNA 1 (ZFAS1). This overexpression played a significant role in instigating PC cell epithelial-mesenchymal transition (EMT) and metastasis. Mechanistically, ZFAS1 enhanced the interaction between AMPK, a key kinase, and ZEB1, the primary regulator of EMT. This interaction resulted in the phosphorylation and subsequent stabilization of ZEB1. Interestingly, ZEB1 also reciprocally influenced the transcription of ZFAS1 by binding to its promoter. Furthermore, when ZFAS1 was depleted, the nutrient deprivation-induced EMT of PC cells and lung metastasis in nude mice were significantly inhibited. Our investigations also revealed that ZFAS1-rich exosomes released from cells suffering glucose and glutamine deprivation promoted the EMT and metastasis of recipient PC cells. Corroborating these findings, a correlated upregulation of ZFAS1 and ZEB1 expression was observed in PC tissues and was associated with a poor overall survival rate for patients. Our findings highlight the involvement of a long noncoding RNA-driven metabolic adaptation in promoting EMT and metastasis of PC, suggesting ZFAS1 as a promising novel therapeutic target for PC metabolic treatment.

Keywords:  EMT; exosome; lncRNA; metabolic stress; metastasis; pancreatic cancer

DOI:  https://doi.org/10.1111/cas.15905
Molecules. 2023 Jul 19. pii: 5504. [Epub ahead of print]28(14):

Atractylenolide III Ameliorates Bile Duct Ligation-Induced Liver Fibrosis by Inhibiting the PI3K/AKT Pathway and Regulating Glutamine Metabolism.

Yan Wang, Kun Shi, Jiyuan Tu, Chang Ke, Niping Chen, Bo Wang, Yanju Liu, Zhongshi Zhou.

  Liver fibrosis is one of the leading causes of hepatic sclerosis and hepatocellular carcinoma worldwide. However, the complex pathophysiological mechanisms of liver fibrosis are unknown, and no specific drugs are available to treat liver fibrosis. Atractylenolide III (ATL III) is a natural compound isolated from the plant Atractylodes lancea (Thunb.) DC. that possesses antioxidant properties and the ability to inhibit inflammatory responses. In this study, cholestatic hepatic fibrosis was induced in mice using a bile duct ligation (BDL) model and treated with 10 mg/kg and 50 mg/kg of ATL III via gavage for 14 days. ATL III significantly reduced the liver index, lowered serum ALT and AST levels, and reduced liver injury in bile-duct-ligated mice. In addition, ATL III significantly attenuated histopathological changes and reduced collagen deposition. ATL III reduced the expression of fibrosis-related genes α-smooth muscle actin (α-SMA), Collagen I (col1a1), Collagen IV (col4a2), and fibrosis-related proteins α-SMA and col1a1 in liver tissue. Using RNA sequencing (RNA-seq) to screen molecular targets and pathways, ATL III was found to affect the PI3K/AKT singling pathway by inhibiting the phosphorylation of PI3K and AKT, thereby ameliorating BDL-induced liver fibrosis. Gas chromatography-mass spectrometry (GC-MS) was used to evaluate the effect of ATL III on liver metabolites in BDL mice. ATL III further affected glutamine metabolism by down-regulating the activity of glutamine (GLS1) and glutamine metabolism. ATL III further affected glutamine metabolism by down-regulating the activity of glutaminase (GLS1), as well as glutamine metabolism. Therefore, we conclude that ATL III attenuates liver fibrosis by inhibiting the PI3K/AKT pathway and glutamine metabolism, suggesting that ATL III is a potential drug candidate for treating liver fibrosis.

Keywords:  Atractylenolide III; PI3K/AKT; bile duct ligation; glutaminase; liver fibrosis

DOI:  https://doi.org/10.3390/molecules28145504
J Hematol Oncol. 2023 07 25. 16(1): 80

Metabolism, metabolites, and macrophages in cancer.

Mengyuan Li, Yuhan Yang, Liting Xiong, Ping Jiang, Junjie Wang, Chunxiao Li.

  Tumour-associated macrophages (TAMs) are crucial components of the tumour microenvironment and play a significant role in tumour development and drug resistance by creating an immunosuppressive microenvironment. Macrophages are essential components of both the innate and adaptive immune systems and contribute to pathogen resistance and the regulation of organism homeostasis. Macrophage function and polarization are closely linked to altered metabolism. Generally, M1 macrophages rely primarily on aerobic glycolysis, whereas M2 macrophages depend on oxidative metabolism. Metabolic studies have revealed that the metabolic signature of TAMs and metabolites in the tumour microenvironment regulate the function and polarization of TAMs. However, the precise effects of metabolic reprogramming on tumours and TAMs remain incompletely understood. In this review, we discuss the impact of metabolic pathways on macrophage function and polarization as well as potential strategies for reprogramming macrophage metabolism in cancer treatment.

Keywords:  Cancer; Metabolism; Metabolism reprogramming; Tumour microenvironment; Tumour-associated macrophages

DOI:  https://doi.org/10.1186/s13045-023-01478-6
Int J Mol Sci. 2023 Jul 21. pii: 11753. [Epub ahead of print]24(14):

A Prognostic Model of Head and Neck Cancer Based on Amino Acid Metabolism-Related Signature and Its Implication for Immunosuppressive Microenvironment.

Xuran Li, Danni Li, Jiaojiao Li, Yiliang Chen, Zhenyu Cai, Fei Tan.

  Amino acid metabolism has been implicated in tumorigenesis and tumor progression. Alterations in intracellular and extracellular metabolites associated with metabolic reprogramming in cancer have profound effects on gene expression, cell differentiation, and tumor immune microenvironment. However, the prognostic significance of amino acid metabolism in head and neck cancer remains to be further investigated. In this study, we identified 98 differentially expressed genes related to amino acid metabolism in head and neck cancer in The Cancer Genome Atlas. Using batch univariate Cox regression and Lasso regression, we extracted nine amino acid metabolism-related genes. Based on that, we developed the amino acid metabolism index. The prognostic value of this index was validated in two Gene Expression Omnibus cohorts. The results show that this model can help predict tumor recurrence and prognosis. The infiltration of immune cells in the tumor microenvironment was analyzed, and it was discovered that the high index is associated with an immunosuppressive microenvironment. In addition, this study demonstrated the impact of the amino acid metabolism index on clinical indicators, survival of patients with head and neck cancer, and the prediction of treatment response to immune checkpoint inhibitors. We conducted several cell experiments and demonstrated that epigenetic drugs could affect the index and enhance tumor immunity. In conclusion, our study demonstrates that the index not only has important prognostic value in head and neck cancer patients but also facilitates patient stratification for immunotherapy.

Keywords:  amino acid metabolism; epigenetic drugs; head and neck cancer; immune checkpoint therapy; tumor microenvironment

DOI:  https://doi.org/10.3390/ijms241411753
Metabolites. 2023 Jun 21. pii: 774. [Epub ahead of print]13(7):

Differential Inhibition of Anaplerotic Pyruvate Carboxylation and Glutaminolysis-Fueled Anabolism Underlies Distinct Toxicity of Selenium Agents in Human Lung Cancer.

Teresa W-M Fan, Jason Winnike, Ahmad Al-Attar, Alexander C Belshoff, Pawel K Lorkiewicz, Jin Lian Tan, Min Wu, Richard M Higashi, Andrew N Lane.

  Past chemopreventive human trials on dietary selenium supplements produced controversial outcomes. They largely employed selenomethionine (SeM)-based diets. SeM was less toxic than selenite or methylseleninic acid (MSeA) to lung cancer cells. We thus investigated the toxic action of these Se agents in two non-small cell lung cancer (NSCLC) cell lines and ex vivo organotypic cultures (OTC) of NSCLC patient lung tissues. Stable isotope-resolved metabolomics (SIRM) using 13C6-glucose and 13C5,15N2-glutamine tracers with gene knockdowns were employed to examine metabolic dysregulations associated with cell type- and treatment-dependent phenotypic changes. Inhibition of key anaplerotic processes, pyruvate carboxylation (PyC) and glutaminolysis were elicited by exposure to MSeA and selenite but not by SeM. They were accompanied by distinct anabolic dysregulation and reflected cell type-dependent changes in proliferation/death/cell cycle arrest. NSCLC OTC showed similar responses of PyC and/or glutaminolysis to the three agents, which correlated with tissue damages. Altogether, we found differential perturbations in anaplerosis-fueled anabolic pathways to underlie the distinct anti-cancer actions of the three Se agents, which could also explain the failure of SeM-based chemoprevention trials.

Keywords:  chemoprevention; glutaminolysis; methylseleninic acid; non-small cell lung cancer; pyruvate carboxylation; selenite; selenomethionine; stable isotope resolved metabolomics

DOI:  https://doi.org/10.3390/metabo13070774
Cell Discov. 2023 Jul 25. 9(1): 76

The Fe-S cluster assembly protein IscU2 increases α-ketoglutarate catabolism and DNA 5mC to promote tumor growth.

Xiaojun Ren, Jimei Yan, Qiongya Zhao, Xinzhu Bao, Xinyu Han, Chen Zheng, Yan Zhou, Lifang Chen, Bo Wang, Lina Yang, Xi Lin, Dandan Liu, Yuyan Lin, Min Li, Hezhi Fang, Zhimin Lu, Jianxin Lyu.

IscU2 is a scaffold protein that is critical for the assembly of iron-sulfur (Fe-S) clusters and the functions of Fe-S-containing mitochondrial proteins. However, the role of IscU2 in tumor development remains unclear. Here, we demonstrated that IscU2 expression is much higher in human pancreatic ductal adenocarcinoma (PDAC) tissues than in adjacent normal pancreatic tissues. In PDAC cells, activated KRAS enhances the c-Myc-mediated IscU2 transcription. The upregulated IscU2 stabilizes Fe-S cluster and regulates the activity of tricarboxylic acid (TCA) cycle enzymes α-ketoglutarate (α-KG) dehydrogenase and aconitase 2, which promote α-KG catabolism through oxidative and reductive TCA cycling, respectively. In addition to promoting mitochondrial functions, activated KRAS-induced and IscU2-dependent acceleration of α-KG catabolism results in reduced α-KG levels in the cytosol and nucleus, leading to an increase in DNA 5mC due to Tet methylcytosine dioxygenase 3 (TET3) inhibition and subsequent expression of genes including DNA polymerase alpha 1 catalytic subunit for PDAC cell proliferation and tumor growth in mice. These findings underscore a critical role of IscU2 in KRAS-promoted α-KG catabolism, 5mC-dependent gene expression, and PDAC growth and highlight the instrumental and integrated regulation of mitochondrial functions and gene expression by IscU2 in PDAC cells.

DOI: https://doi.org/10.1038/s41421-023-00558-8
Biology (Basel). 2023 Jul 14. pii: 1002. [Epub ahead of print]12(7):

Remodelling of the Mitochondrial Bioenergetic Pathways in Human Cultured Fibroblasts with Carbohydrates.

Margherita Protasoni, Jan-Willem Taanman.

  Mitochondrial oxidative phosphorylation defects underlie many neurological and neuromuscular diseases. Patients' primary dermal fibroblasts are one of the most commonly used in vitro models to study mitochondrial pathologies. However, fibroblasts tend to rely more on glycolysis than oxidative phosphorylation for their energy when cultivated in standard high-glucose medium, rendering it difficult to expose mitochondrial dysfunctions. This study aimed to systematically investigate to which extent the use of galactose- or fructose-based medium switches the fibroblasts' energy metabolism to a more oxidative state. Highly proliferative cells depend more on glycolysis than less proliferative cells. Therefore, we investigated two primary dermal fibroblast cultures from healthy subjects: a highly proliferative neonatal culture and a slower-growing adult culture. Cells were cultured with 25 mM glucose, galactose or fructose, and 4 mM glutamine as carbon sources. Compared to glucose, both galactose and fructose reduce the cellular proliferation rate, but the galactose-induced drop in proliferation is much more profound than the one observed in cells cultivated in fructose. Both galactose and fructose result in a modest increase in mitochondrial content, including mitochondrial DNA, and a disproportionate increase in protein levels, assembly, and activity of the oxidative phosphorylation enzyme complexes. Galactose- and fructose-based media induce a switch of the prevalent biochemical pathway in cultured fibroblasts, enhancing aerobic metabolism when compared to glucose-based medium. While both galactose and fructose stimulate oxidative phosphorylation to a comparable degree, galactose decreases the cellular proliferation rate more than fructose, suggesting that a fructose-based medium is a better choice when studying partial oxidative phosphorylation defects in patients' fibroblasts.

Keywords:  fructose; galactose; glucose; mitochondria; mitochondrial DNA; mitochondrial respiration; oxidative phosphorylation

DOI:  https://doi.org/10.3390/biology12071002
Free Radic Biol Med. 2023 Jul 26. pii: S0891-5849(23)00558-0. [Epub ahead of print]

VHL-deficiency leads to reductive stress in renal cells.

Hans Vellama, Kattri-Liis Eskla, Hillar Eichelmann, Andria Hüva, Daniel A Tennant, Alpesh Thakker, Jennie Roberts, Toomas Jagomäe, Rando Porosk, Agu Laisk, Vello Oja, Heikko Rämma, Vallo Volke, Eero Vasar, Hendrik Luuk.

  Heritable renal cancer syndromes (RCS) are associated with numerous chromosomal alterations including inactivating mutations in von Hippel-Lindau (VHL) gene. Here we identify a novel aspect of the phenotype in VHL-deficient human renal cells. We call it reductive stress as it is characterised by increased NADH/NAD+ ratio that is associated with impaired cellular respiration, impaired CAC activity, upregulation of reductive carboxylation of glutamine and accumulation of lipid droplets in VHL-deficient cells. Reductive stress was mitigated by glucose depletion and supplementation with pyruvate or resazurin, a redox-reactive agent. This study demonstrates for the first time that reductive stress is a part of the phenotype associated with VHL-deficiency in renal cells and indicates that the reversal of reductive stress can augment respiratory activity and CAC activity, suggesting a strategy for altering the metabolic profile of VHL-deficient tumours.

Keywords:  HIF; Reductive stress; Renal cell carcinoma; Respiration; VHL; VHL syndrome

DOI:  https://doi.org/10.1016/j.freeradbiomed.2023.07.029
bioRxiv. 2023 Jul 12. pii: 2023.07.11.548586. [Epub ahead of print]

Quantification of glutathione with high throughput live-cell imaging.

Xiaoli Qi, Jianwei Chen, Xiqian Jiang, Dong Lu, Xin Yu, Hanfeng Lin, Erika Y Monroy, Meng C Wang, Jin Wang.

Reduction oxidation (redox) reactions are central in life and altered redox state is associated with a spectrum of human diseases. Glutathione (GSH) is the most abundant antioxidant in eukaryotic cells and plays critical roles in maintaining redox homeostasis. Thus, measuring intracellular GSH level is an important method to assess the redox state of organism. The currently available GSH probes are based on irreversible chemical reactions with glutathione and can't monitor the real-time glutathione dynamics. Our group developed the first reversible reaction based fluorescent probe for glutathione, which can measure glutathione levels at high resolution using a confocal microscope and in the bulk scale with a flow cytometry. Most importantly it can quantitatively monitor the real-time GSH dynamics in living cells. Using the 2 nd generation of GSH probe, RealThiol (RT), this study measured the GSH level in living Hela cells after treatment with varying concentrations of DL-Buthionine sulfoximine (BSO) which inhibits GSH synthesis, using a high throughput imaging system, Cytation™ 5 cell imaging reader. The results revealed that GSH probe RT at the concentration of 2.0 µM accurately monitored the BSO treatment effect on GSH level in the Hela cells. The present results demonstrated that the GSH probe RT is sensitive and precise in GSH measurement in living cells at a high throughput imaging platform and has the potential to be applied to any cell lines.

DOI: https://doi.org/10.1101/2023.07.11.548586
RSC Adv. 2023 Jul 19. 13(32): 22367-22374

Engineering of a GSH activatable photosensitizer for enhanced photodynamic therapy through disrupting redox homeostasis.

Datian Fu, Yan Wang, Kaiwen Lin, Liangjiu Huang, Jin Xu, Haimei Wu.

Although disrupted redox homeostasis has emerged as a promising approach for tumor therapy, most existing photosensitizers are not able to simultaneously improve the reactive oxygen species level and reduce the glutathione (GSH) level. Therefore, designing photosensitizers that can achieve these two aspects of this goal is still urgent and challenging. In this work, an organic activatable near-infrared (NIR) photosensitizer, CyI-S-diCF3, is developed for GSH depletion-assisted enhanced photodynamic therapy. CyI-S-diCF3, composed of an iodinated heptamethine cyanine skeleton linked with a recognition unit of 3,5-bis(trifluoromethyl)benzenethiol, can specifically react with GSH by nucleophilic substitution, resulting in intracellular GSH depletion and redox imbalance. Moreover, the activated photosensitizer can produce abundant singlet oxygen (1O2) under NIR light irradiation, further heightening the cellular oxidative stress. By this unique nature, CyI-S-diCF3 exhibits excellent toxicity to cancer cells, followed by inducing earlier apoptosis. Thus, our study may propose a new strategy to design an activatable photosensitizer for breaking the redox homeostasis in tumor cells.

DOI: https://doi.org/10.1039/d3ra04074g
Life (Basel). 2023 Jul 11. pii: 1538. [Epub ahead of print]13(7):

Therapeutic Efficacy, Radiotoxicity and Abscopal Effect of BNCT at the RA-3 Nuclear Reactor Employing Oligo-Fucoidan and Glutamine as Adjuvants in an Ectopic Colon Cancer Model in Rats.

Debora N Frydryk Benitez, Mónica A Palmieri, Yanina V Langle, Andrea Monti Hughes, Emiliano C C Pozzi, Silvia I Thorp, Marcela A Garabalino, Paula Curotto, Paula S Ramos, María L Paparella, Lucas Polti, Ana Eiján, Amanda E Schwint, Verónica A Trivillin.

  Boron neutron capture therapy (BNCT) is based on the preferential uptake of 10B compounds by tumors, followed by neutron irradiation. The aim of this study was to assess, in an ectopic colon cancer model, the therapeutic efficacy, radiotoxicity, abscopal effect and systemic immune response associated with (BPA/Borophenylalanine+GB-10/Decahydrodecaborate)-BNCT (Comb-BNCT) alone or in combination with Oligo-Fucoidan (O-Fuco) or Glutamine (GLN), compared to the "standard" BPA-BNCT protocol usually employed in clinical trials. All treatments were carried out at the RA-3 nuclear reactor. Boron biodistribution studies showed therapeutic values above 20 ppm 10B in tumors. At 7 weeks post-treatment, the ratio of tumor volume post-/pre-BNCT was significantly smaller for all BNCT groups vs. SHAM (p < 0.05). The parameter "incidence of tumors that underwent a reduction to ≤50% of initial tumor volume" exhibited values of 62% for Comb-BNCT alone, 82% for Comb-BNCT+GLN, 73% for Comb-BNCT+O-Fuco and only 30% for BPA-BNCT. For BPA-BNCT, the incidence of severe dermatitis was 100%, whereas it was significantly below 70% (p ≤ 0.05) for Comb-BNCT, Comb-BNCT+O-Fuco and Comb-BNCT+GLN. Considering tumors outside the treatment area, 77% of Comb-BNCT animals had a tumor volume lower than 50 mm3 vs. 30% for SHAM (p ≤ 0.005), suggesting an abscopal effect of Comb-BNCT. Inhibition of metastatic spread to lymph nodes was observed in all Comb-BNCT groups. Considering systemic aspects, CD8+ was elevated for Comb-BNCT+GLN vs. SHAM (p ≤ 0.01), and NK was elevated for Comb-BNCT vs. SHAM (p ≤ 0.05). Comb-BNCT improved therapeutic efficacy and reduced radiotoxicity compared to BPA-BNCT and induced an immune response and an abscopal effect.

Keywords:  BNCT; BPA and GB-10; Glutamine; Oligo-Fucoidan; RA-3 nuclear reactor; abscopal effect; biodistribution; lymph nodes; radiotoxicity; systemic immune response

DOI:  https://doi.org/10.3390/life13071538