bims-mascan Biomed News
on Mass spectrometry in cancer research
Issue of 2019–11–10
twenty-two papers selected by
Giovanny Rodríguez Blanco, The Beatson Institute for Cancer Research
  1. Lipid signalling drives proteolytic rewiring of mitochondria by YME1L.
  2. The PI3K-AKT network at the interface of oncogenic signalling and cancer metabolism.
  3. Targeting Esterified Oxylipins by LC-MS - Effect of Sample Preparation on Oxylipin Pattern.
  4. EGFL9 promotes breast cancer metastasis by inducing cMET activation and metabolic reprogramming.
  5. Plasma High-Resolution Metabolomics Differentiates Adults with Normal Weight Obesity from Lean Individuals.
  6. Plasma proteomic approach in patients with heart failure: insights into pathogenesis of disease progression and potential novel treatment targets.
  7. Lactate production is a prioritised feature of adipocyte metabolism.
  8. Using PCI-IS Assisted Quantitative Metabolomics for Establishing Prediction Models for Breast Cancer Detection.
  9. Metabolic Profiling of Human Plasma and Urine, Targeting Tryptophan, Tyrosine and Branched Chain Amino Acid Pathways.
  10. Proteomics analysis of Formalin Fixed Paraffin Embedded tissues in the investigation of prostate cancer.
  11. Racial differences in distribution of fatty acids in prostate cancer and benign prostatic tissues.
  12. FUNCTIONAL METABOLOMICS DECIPHER BIOCHEMICAL FUNCTIONS AND ASSOCIATED MECHANISMS UNDERLIE SMALL-MOLECULE METABOLISM.
  13. Reproducibility, specificity and accuracy of relative quantification using spectral library-based data-independent acquisition.
  14. Isolating Urinary Extracellular Vesicles as Biomarkers for Diabetic Disease.
  15. CD9 identifies pancreatic cancer stem cells and modulates glutamine metabolism to fuel tumour growth.
  16. Hepatic stearoyl CoA desaturase 1 deficiency increases glucose uptake in adipose tissue partially through the PGC-1α-FGF21 axis in mice.
  17. Inter-Laboratory Comparison of Metabolite Measurements for Metabolomics Data Integration.
  18. Lipid Profile Characterization and Lipoprotein Comparison of Extracellular Vesicles from Human Plasma and Serum.
  19. Role of Sphingosylphosphorylcholine in Tumor and Tumor Microenvironment.
  20. Glutamine blockade induces divergent metabolic programs to overcome tumor immune evasion.
  21. MetaboLights: a resource evolving in response to the needs of its scientific community.
  22. Novel MYC-Targeting Drug Is Effective in Mouse Prostate Cancer Models.
  1. Nature. 2019 Nov 06.
    Lipid signalling drives proteolytic rewiring of mitochondria by YME1L.
    Thomas MacVicar, Yohsuke Ohba, Hendrik Nolte, Fiona Carola Mayer, Takashi Tatsuta, Hans-Georg Sprenger, Barbara Lindner, Yue Zhao, Jiahui Li, Christiane Bruns, Marcus Krüger, Markus Habich, Jan Riemer, Robin Schwarzer, Manolis Pasparakis, Sinika Henschke, Jens C Brüning, Nicola Zamboni, Thomas Langer.
      Reprogramming of mitochondria provides cells with the metabolic flexibility required to adapt to various developmental transitions such as stem cell activation or immune cell reprogramming, and to respond to environmental challenges such as those encountered under hypoxic conditions or during tumorigenesis1-3. Here we show that the i-AAA protease YME1L rewires the proteome of pre-existing mitochondria in response to hypoxia or nutrient starvation. Inhibition of mTORC1 induces a lipid signalling cascade via the phosphatidic acid phosphatase LIPIN1, which decreases phosphatidylethanolamine levels in mitochondrial membranes and promotes proteolysis. YME1L degrades mitochondrial protein translocases, lipid transfer proteins and metabolic enzymes to acutely limit mitochondrial biogenesis and support cell growth. YME1L-mediated mitochondrial reshaping supports the growth of pancreatic ductal adenocarcinoma (PDAC) cells as spheroids or xenografts. Similar changes to the mitochondrial proteome occur in the tumour tissues of patients with PDAC, suggesting that YME1L is relevant to the pathophysiology of these tumours. Our results identify the mTORC1-LIPIN1-YME1L axis as a post-translational regulator of mitochondrial proteostasis at the interface between metabolism and mitochondrial dynamics.
    DOI:  https://doi.org/10.1038/s41586-019-1738-6
  2. Nat Rev Cancer. 2019 Nov 04.
    The PI3K-AKT network at the interface of oncogenic signalling and cancer metabolism.
    Gerta Hoxhaj, Brendan D Manning.
      The altered metabolic programme of cancer cells facilitates their cell-autonomous proliferation and survival. In normal cells, signal transduction pathways control core cellular functions, including metabolism, to couple the signals from exogenous growth factors, cytokines or hormones to adaptive changes in cell physiology. The ubiquitous, growth factor-regulated phosphoinositide 3-kinase (PI3K)-AKT signalling network has diverse downstream effects on cellular metabolism, through either direct regulation of nutrient transporters and metabolic enzymes or the control of transcription factors that regulate the expression of key components of metabolic pathways. Aberrant activation of this signalling network is one of the most frequent events in human cancer and serves to disconnect the control of cell growth, survival and metabolism from exogenous growth stimuli. Here we discuss our current understanding of the molecular events controlling cellular metabolism downstream of PI3K and AKT and of how these events couple two major hallmarks of cancer: growth factor independence through oncogenic signalling and metabolic reprogramming to support cell survival and proliferation.
    DOI:  https://doi.org/10.1038/s41568-019-0216-7
  3. Prostaglandins Other Lipid Mediat. 2019 Nov 04. pii: S1098-8823(19)30135-2. [Epub ahead of print] 106384
    Targeting Esterified Oxylipins by LC-MS - Effect of Sample Preparation on Oxylipin Pattern.
    Annika I Ostermann, Elisabeth Koch, Katharina M Rund, Laura Kutzner, Malwina Mainka, Nils Helge Schebb.
      A major part of oxygenated metabolites of polyunsaturated fatty acids - i.e. eicosanoids and other oxylipins - in biological samples occurs in the esterified form. Yet, their biological role is only poorly understood. For quantification of esterified oxylipins in biological samples current protocols mostly apply alkaline hydrolysis with or without prior lipid extraction to release oxylipins in their free form which are subsequently quantified via liquid chromatography mass spectrometry. Herein, a detailed protocol for precise and reproducible quantification of esterified oxylipins in plasma is presented comprising i) extraction of lipids and removal of proteins with iso-propanol, ii) base hydrolysis with potassium hydroxide to saponify lipids and iii) solid phase extraction of the liberated oxylipins on a C8/anion exchange mixed mode cartridge. Method development revealed, that unequal extraction of internal standards and lipid classes led to distorted concentrations, emphasizing that the choice of extraction solvent is important to minimize discrimination. Regarding the hydrolysis conditions sufficient hydrolysis time is needed and at least 30 min incubation at 60 °C is required with 0.1 M KOH. Drying of the SPE cartridges is a critical parameter due to autoxidation processes of PUFA, which are present in high concentrations after cleavage leading to artificial formation of epoxy fatty acids. With the developed protocol, inter-day, intra-day and inter-operator variance was <25% for most oxylipins including hydroxy-, dihydroxy-, and epoxy-PUFA. The applicability of the developed methodology is demonstrated by the investigation of the changes in oxylipin pattern following omega-3 fatty acid feeding to rats.
    Keywords:  Esterified oxylipins; alkaline hydrolysis; n3-PUFA feeding; protein precipitation; solid phase extraction; targeted LC-MS/MS
    DOI:  https://doi.org/10.1016/j.prostaglandins.2019.106384
  4. Nat Commun. 2019 Nov 06. 10(1): 5033
    EGFL9 promotes breast cancer metastasis by inducing cMET activation and metabolic reprogramming.
    Fanyan Meng, Ling Wu, Lun Dong, Allison V Mitchell, C James Block, Jenney Liu, Haijun Zhang, Qing Lu, Won-Min Song, Bin Zhang, Wei Chen, Jiani Hu, Jian Wang, Qifeng Yang, Maik Hüttemann, Guojun Wu.
      The molecular mechanisms driving metastatic progression in triple-negative breast cancer (TNBC) patients are poorly understood. In this study, we demonstrate that epidermal growth factor-like 9 (EGFL9) is significantly upregulated in basal-like breast cancer cells and associated with metastatic progression in breast tumor samples. Functionally, EGFL9 is both necessary and sufficient to enhance cancer cell migration and invasion, as well as distant metastasis. Mechanistically, we demonstrate that EGFL9 binds cMET, activating cMET-mediated downstream signaling. EGFL9 and cMET co-localize at both the cell membrane and within the mitochondria. We further identify an interaction between EGFL9 and the cytochrome c oxidase (COX) assembly factor COA3. Consequently, EGFL9 regulates COX activity and modulates cell metabolism, promoting a Warburg-like metabolic phenotype. Finally, we show that combined pharmacological inhibition of cMET and glycolysis reverses EGFL9-driven stemness. Our results identify EGFL9 as a therapeutic target for combating metastatic progression in TNBC.
    DOI:  https://doi.org/10.1038/s41467-019-13034-3
  5. Obesity (Silver Spring). 2019 Nov;27(11): 1729-1737
    Plasma High-Resolution Metabolomics Differentiates Adults with Normal Weight Obesity from Lean Individuals.
    Moriah P Bellissimo, Qingpo Cai, Thomas R Ziegler, Ken H Liu, Phong H Tran, Miriam B Vos, Greg S Martin, Dean P Jones, Tianwei Yu, Jessica A Alvarez.
       OBJECTIVE: This study explored underlying metabolism-related dysfunction by examining metabolomic profiles in adults categorized as lean, as having normal weight obesity (NWO), or as having overweight/obesity.
    METHODS: Participants (N = 179) had fasting plasma analyzed by liquid chromatography and high-resolution mass spectrometry for high-resolution metabolomics. Body composition was assessed by dual-energy x-ray absorptiometry. NWO was defined as BMI < 25 and body fat > 30% for women and > 23% for men. Differentiating metabolomic features were determined by using linear regression models and likelihood ratio tests with false discovery rate correction. Mummichog was used for pathway and network analyses.
    RESULTS: A total of 222 metabolites significantly differed between the groups at a false discovery rate of q = 0.2. Linoleic acid, β-alanine, histidine, and aspartate/asparagine metabolism pathways were significantly enriched (all P < 0.01) by metabolites that were similarly upregulated in the NWO and overweight/obesity groups compared with the lean group. A module analysis linked branched-chain amino acids and amino acid metabolites as elevated in the NWO and overweight/obesity groups compared with the lean group (all P < 0.05).
    CONCLUSIONS: Metabolomic profiles of individuals with NWO reflected similar metabolic disruption as those of individuals with overweight/obesity. High-resolution metabolomics may help identify people at risk for developing obesity-related disease, despite normal BMI.
    DOI:  https://doi.org/10.1002/oby.22654
  6. Eur J Heart Fail. 2019 Nov 06.
    Plasma proteomic approach in patients with heart failure: insights into pathogenesis of disease progression and potential novel treatment targets.
    Thong H Cao, Donald J L Jones, Adriaan A Voors, Paulene A Quinn, Jatinderpal K Sandhu, Daniel C S Chan, Helen M Parry, Mohapradeep Mohan, Ify R Mordi, Iziah E Sama, Stefan D Anker, John G Cleland, Kenneth Dickstein, Gerasimos Filippatos, Hans L Hillege, Marco Metra, Piotr Ponikowski, Nilesh J Samani, Dirk J Van Veldhuisen, Faiez Zannad, Chim C Lang, Leong L Ng.
       AIMS: To provide insights into pathogenesis of disease progression and potential novel treatment targets for patients with heart failure by investigation of the plasma proteome using network analysis.
    METHODS AND RESULTS: The plasma proteome of 50 patients with heart failure who died or were rehospitalised were compared with 50 patients with heart failure, matched for age and sex, who did not have an event. Peptides were analysed on two-dimensional liquid chromatography coupled to tandem mass spectrometry (2D LC ESI-MS/MS) in high definition mode (HDMSE). We identified and quantified 3001 proteins, of which 51 were significantly up-regulated and 46 down-regulated with more than two-fold expression changes in those who experienced death or rehospitalisation. Gene ontology enrichment analysis and protein-protein interaction networks of significant differentially expressed proteins discovered the central role of metabolic processes in clinical outcomes of patients with heart failure. The findings revealed that a cluster of proteins related to glutathione metabolism, arginine and proline metabolism, and pyruvate metabolism in the pathogenesis of poor outcome in patients with heart failure who died or were rehospitalised.
    CONCLUSIONS: Our findings show that in patients with heart failure who died or were rehospitalised, the glutathione, arginine and proline, and pyruvate pathways were activated. These pathways might be potential targets for therapies to improve poor outcomes in patients with heart failure.
    Keywords:  Heart failure; Mass spectrometry; Metabolism; Pathogenesis; Proteomics; Treatment target
    DOI:  https://doi.org/10.1002/ejhf.1608
  7. J Biol Chem. 2019 Nov 05. pii: jbc.RA119.011178. [Epub ahead of print]
    Lactate production is a prioritised feature of adipocyte metabolism.
    James R Krycer, Lake-Ee Quek, Deanne Francis, Daniel J Fazakerley, Sarah D Elkington, Alexis Diaz-Vegas, Kristen C Cooke, Fiona C Weiss, Xiaowen Duan, Sergey Kurdyukov, Ping-Xin Zhou, Uttam K Tambar, Akiyoshi Hirayama, Satsuki Ikeda, Yushi Kamei, Tomoyoshi Soga, Gregory J Cooney, David E James.
      Adipose tissue is essential for whole-body glucose homeostasis, with a primary role in lipid storage. It has been previously observed that lactate production is also an important metabolic feature of adipocytes, but its relationship to adipose and whole-body glucose disposal remains unclear. Therefore, using a combination of metabolic labeling techniques, here we closely examined lactate production of cultured and primary mammalian adipocytes. Insulin treatment increased glucose uptake and conversion to lactate, with the latter responding more to insulin than did other metabolic fates of glucose. However, lactate production did not just serve as a mechanism to dispose of excess glucose, since we also observed that lactate production in adipocytes did not solely depend on glucose availability and even occurred independently of glucose metabolism. This suggests that lactate production is prioritized in adipocytes. Furthermore, knocking down lactate dehydrogenase specifically in the fat body of Drosophila flies lowered circulating lactate and improved whole-body glucose disposal. These results emphasize lactate production is an additional metabolic role of adipose tissue beyond lipid storage and release.
    Keywords:  Drosophila; adipocyte; cell metabolism; fat tissue; glucose disposal; insulin; insulin resistance; lactate; metabolic regulation; whole-body glucose homeostasis
    DOI:  https://doi.org/10.1074/jbc.RA119.011178
  8. Rapid Commun Mass Spectrom. 2019 Nov 06.
    Using PCI-IS Assisted Quantitative Metabolomics for Establishing Prediction Models for Breast Cancer Detection.
    Marisa Huang, Hung-Yuan Li, Hsiao-Wei Liao, Ching-Hung Lin, Chin-Yi Wang, Wen-Hung Kuo, Ching-Hua Kuo.
       RATIONALE: Breast cancer is one of the most common cancers among women and its associated mortality has been on the rise. Metabolomics is a potential strategy for breast cancer detection. The post column infused internal standard (PCI-IS) assisted liquid chromatography-tandem mass spectrometry (LC-MS/MS) method has been demonstrated as an effective strategy for quantitative metabolomics. In this study, we evaluated the performance of targeted metabolomics with the PCI-IS quantification method to identify women with breast cancer.
    METHODS: We used metabolite profiling to identify 17 dysregulated metabolites in breast cancer patients. Two LC-MS/MS methods in combination with the PCI-IS strategy were developed to quantify these metabolites in plasma samples. Detection models were built through the analysis of plasma samples from 176 subjects consisting of healthy volunteers and breast cancer patients.
    RESULTS: Three isotope standards were selected as the PCI-IS for the metabolites. The accuracy was within 82.8-114.16 %, except for citric acid and lactic acid at high concentration levels. The repeatability and intermediate precision were all lower than 15% relative standard deviation. We have identified several metabolites that indicate the presence of breast cancer. The area under the receiver operating characteristics (AUROC) curve, sensitivity and specificity of the linear combinations of metabolite concentrations and age with the highest AUROC were 0.940 (0.889-0.992), 88.4% and 94.2% for pre-menopausal woman, respectively, and 0.828 (0.734-0.922), 73.5% and 85.1% for post-menopausal women, respectively.
    CONCLUSIONS: The targeted metabolomics with PCI-IS quantification method successfully established prediction models for breast cancer detection. Further study is essential to validate these proposed markers.
    DOI:  https://doi.org/10.1002/rcm.8581
  9. Metabolites. 2019 Nov 01. pii: E261. [Epub ahead of print]9(11):
    Metabolic Profiling of Human Plasma and Urine, Targeting Tryptophan, Tyrosine and Branched Chain Amino Acid Pathways.
    Andrea Anesi, Josep Rubert, Kolade Oluwagbemigun, Ximena Orozco-Ruiz, Ute Nöthlings, Monique M B Breteler, Fulvio Mattivi.
      Tryptophan and tyrosine metabolism has a major effect on human health, and disorders have been associated with the development of several pathologies. Recently, gut microbial metabolism was found to be important for maintaining correct physiology. Here, we describe the development and validation of a UHPLC-ESI-MS/MS method for targeted quantification of 39 metabolites related to tryptophan and tyrosine metabolism, branched chain amino acids and gut-derived metabolites in human plasma and urine. Extraction from plasma was optimised using 96-well plates, shown to be effective in removing phospholipids. Urine was filtered and diluted ten-fold. Metabolites were separated with reverse phase chromatography and detected using triple quadrupole MS. Linear ranges (from ppb to ppm) and correlation coefficients (r2 > 0.990) were established for both matrices independently and the method was shown to be linear for all tested metabolites. At medium spiked concentration, recovery was over 80% in both matrices, while analytical precision was excellent (CV < 15%). Matrix effects were minimal and retention time stability was excellent. The applicability of the methods was tested on biological samples, and metabolite concentrations were found to be in agreement with available data. The method allows the analysis of up to 96 samples per day and was demonstrated to be stable for up to three weeks from acquisition.
    Keywords:  LC-MS/MS; branched chain amino acids; clinical studies; gut microbiota metabolites; human plasma; targeted metabolomics; tryptophan metabolism; tyrosine metabolism; urine
    DOI:  https://doi.org/10.3390/metabo9110261
  10. J Proteome Res. 2019 Nov 04.
    Proteomics analysis of Formalin Fixed Paraffin Embedded tissues in the investigation of prostate cancer.
    Anna Mantsiou, Manousos Makridakis, Konstantinos Fasoulakis, Ioannis Katafigiotis, Constantinos A Constantinides, Jerome Zoidakis, Maria G Roubelakis, Antonia Vlahou, Vasiliki Lygirou.
      Prostate cancer (PCa) is one of the leading causes of death in men worldwide. The molecular features, associated with the onset and progression of the disease, are under vigorous investigation. Formalin-fixed paraffin-embedded (FFPE) tissues are valuable resources for large-scale studies, however, their application in proteomics is limited due to protein cross-linking. In this study, the adjustment of a protocol for the proteomic analysis of FFPE tissues was performed which was followed by a pilot application on FFPE PCa clinical samples to investigate whether the optimized protocol can provide biologically relevant data for the investigation of PCa. For the optimization, FFPE mouse tissues were processed using seven protein extraction protocols including combinations of homogenization methods (beads, sonication, boiling) and buffers (SDS based and Urea-Thiourea based). The proteome extraction efficacy was then evaluated based on protein identifications and reproducibility using SDS electrophoresis and high resolution LC-MS/MS analysis. Comparison between the FFPE and matched fresh frozen (FF) tissues, using an optimized protocol involving protein extraction with an SDS-based buffer following beads homogenization and boiling, showed a substantial overlap in protein identifications with a strong correlation in relative abundances (rs=0.819, p <0.001). Next, FFPE tissues (3 sections, 15μm each per sample) from 10 patients with PCa corresponding to tumor (GS=6 or GS≥8) and adjacent benign regions were processed with the optimized protocol. Extracted proteins were analyzed by GeLC-MS/MS followed by statistical and bioinformatics analysis. Proteins significantly deregulated between PCa GS≥8 and PCa GS=6 represented extracellular matrix organisation, gluconeogenesis and phosphorylation pathways. Proteins deregulated between cancerous and adjacent benign tissues, reflected increased translation, peptide synthesis and protein metabolism in the former, which is consistent with the literature. In conclusion, the results support the relevance of the proteomic findings in the context of PCa and the reliability of the optimized protocol for proteomics analysis of FFPE material.
    DOI:  https://doi.org/10.1021/acs.jproteome.9b00587
  11. Lipids Health Dis. 2019 Nov 03. 18(1): 189
    Racial differences in distribution of fatty acids in prostate cancer and benign prostatic tissues.
    Xinchun Zhou, Hao Mei, Joshua Agee, Timera Brown, Jinghe Mao.
       BACKGROUND: It remains controversial whether and which fatty acids are different between prostate cancer (PCa) and benign prostatic tissues (BPT) in association with occurrence, progression and racial disparity between African American (AA) and Caucasian American (CA) populations.
    METHODS: Total fatty acids (TFA) and free fatty acid (FFA) were determined on fresh frozen prostatic tissues including 26 PCa and 21 BPT from AA and CA patients by Gas chromatography with flame ionization detection (GC-FID) and Electrospray Ionization Mass Spectrometry (ESI-MS), respectively.
    RESULTS: In all studied population, TFA in 8 out of 16 individual species, in total and in groups of saturated total fatty acid (STFA), mono-unsaturated total fatty acid (MUTFA), poly-unsaturated total fatty acid (PUTFA) and n-6 TFA were significantly higher in PCa than in BPT; FFA in 4 out of 10 individual species, in total and in groups of MUFFA, PUFFA, n-6 FFA and n-3 FFA were significantly higher in PCa than in BPT. The concentrations of most fatty acid parameters correlated with Gleason's grade and clinical stage of PCa. As compared with CA men, AA men had higher concentrations of TFA, especially TFA with chains of 14-18 carbons than in BPT, and lower concentrations of TFA in PCa.
    CONCLUSIONS: Increasing in prostatic fatty acids in the form of TFA and FFA correlated to occurrence, progression and racial disparity of PCa.
    Keywords:  Free fatty acid (FFA); Lipidomics; Prostate cancer (PCa); Racial disparity; Total fatty acid (TFA)
    DOI:  https://doi.org/10.1186/s12944-019-1130-4
  12. Mass Spectrom Rev. 2019 Nov 04.
    FUNCTIONAL METABOLOMICS DECIPHER BIOCHEMICAL FUNCTIONS AND ASSOCIATED MECHANISMS UNDERLIE SMALL-MOLECULE METABOLISM.
    Longlong Hu, Jingjing Liu, Wenhua Zhang, Tianyu Wang, Ning Zhang, Yie Hou Lee, Haitao Lu.
      Metabolism is the collection of biochemical reactions enabled by chemically diverse metabolites, which facilitate different physiological processes to exchange substances and synthesize energy in diverse living organisms. Metabolomics has emerged as a cutting-edge method to qualify and quantify the metabolites in different biological matrixes, and it has the extraordinary capacity to interrogate the biological significance that underlies metabolic modification and modulation. Liquid chromatography combined with mass spectrometry (LC/MS), as a robust platform for metabolomics analysis, has increased in popularity over the past 10 years due to its excellent sensitivity, throughput, and versatility. However, metabolomics investigation currently provides us with only phenotype data without revealing the biochemical functions and associated mechanisms. This limitation indeed weakens the core value of metabolomics data in a broad spectrum of the life sciences. In recent years, the scientific community has actively explored the functional features of metabolomics and translated this cutting-edge approach to be used to solve key multifaceted questions, such as disease pathogenesis, the therapeutic discovery of drugs, nutritional issues, agricultural problems, environmental toxicology, and microbial evolution. Here, we are the first to briefly review the history and applicable progression of LC/MS-based metabolomics, with an emphasis on the applications of metabolic phenotyping. Furthermore, we specifically highlight the next era of LC/MS-based metabolomics to target functional metabolomes, through which we can answer phenotype-related questions to elucidate biochemical functions and associated mechanisms implicated in dysregulated metabolism. Finally, we propose many strategies to enhance the research capacity of functional metabolomics by enabling the combination of contemporary omics technologies and cutting-edge biochemical techniques. The main purpose of this review is to improve the understanding of LC/MS-based metabolomics, extending beyond the conventional metabolic phenotype toward biochemical functions and associated mechanisms, to enhance research capability and to enlarge the applicable scope of functional metabolomics in small-molecule metabolism in different living organisms.
    Keywords:  LC/MS; biochemical mechanisms; functional metabolomics; phenotypic metabolomics; small-molecule metabolism
    DOI:  https://doi.org/10.1002/mas.21611
  13. Mol Cell Proteomics. 2019 Nov 07. pii: mcp.RA119.001714. [Epub ahead of print]
    Reproducibility, specificity and accuracy of relative quantification using spectral library-based data-independent acquisition.
    Katalin Barkovits, Sandra Pacharra, Kathy Pfeiffer, Simone Steinbach, Martin Eisenacher, Katrin Marcus, Julian Uszkoreit.
      Currently data-dependent acquisition (DDA) is the method of choice for mass spectrometry-based proteomics discovery experiments, but data-independent acquisition (DIA) is steadily becoming more important. One of the most important requirements to perform a DIA analysis is the availability of suitable spectral libraries for peptide identification and quantification. Several studies were performed addressing the evaluation of spectral library performance for protein identification in DIA measurements. But so far only few experiments estimate the effect of these libraries on the quantitative level.In this work we created a gold standard spike-in sample set with known contents and ratios of proteins in a complex protein matrix that allowed a detailed comparison of DIA quantification data obtained with different spectral library approaches. We utilized in-house generated sample-specific spectral libraries created using varying sample preparation approaches and repeated DDA measurement. In addition, two different search engines were tested for protein identification from DDA data and subsequent library generation. In total, eight different spectral libraries were generated and the quantification results compared to a library free method, as well as a default DDA analysis. Not only the number of identifications on peptide and protein level in the spectral libraries and the corresponding DIA analysis results was inspected, but also the number of expected and identified differentially abundant protein groups and their ratios.We found, that while libraries of prefractionated samples were generally larger, there was no significant increase in DIA identifications compared to repetitive non-fractionated measurements. Furthermore, we show that the accuracy of the quantification is strongly dependent on the applied spectral library and whether the quantification is based on peptide or protein level. Overall, the reproducibility and accuracy of DIA quantification is superior to DDA in all applied approaches.Data has been deposited to the ProteomeXchange repository with identifiers PXD012986, PXD012987, PXD012988 and PXD014956.
    Keywords:  Bioinformatics software; Label-free quantification; Mass Spectrometry; Quantification; Target identification; data-independent acquisition (DIA); peptide identification; proteomics; spectral library
    DOI:  https://doi.org/10.1074/mcp.RA119.001714
  14. Methods Mol Biol. 2020 ;2067 175-188
    Isolating Urinary Extracellular Vesicles as Biomarkers for Diabetic Disease.
    Karina Barreiro, Tobias B Huber, Harry Holthofer.
      Extracellular vesicles are lipid bilayer enclosed structures secreted by all cell types. Their cargo includes proteins, lipids, RNAs, and DNA, which reflect the physiological state of their cells of origin. Recently, urinary extracellular vesicles have emerged as a valuable source of biomarkers for kidney and systemic disease.Unfortunately, all existing methods for extracellular vesicle isolation from urine are time consuming and/or expensive. Thus, they are not adaptable to large-scale studies and unsuitable for clinical use without special equipment in the laboratory. Recently, our group has devised a set of new, quick, simple, and inexpensive techniques, based on hydrostatic filtration dialysis (HFD) of urine extremely suitable for diagnostic purposes. This novel approach represents a great potential for new diagnostics and understanding disease biology in general and brings the biomarker detection to the scope of all laboratories.
    Keywords:  Electron microscopy; Hydrostatic filtration dialysis; Proteomics; Transcriptomic; Urinary extracellular vesicle isolation
    DOI:  https://doi.org/10.1007/978-1-4939-9841-8_13
  15. Nat Cell Biol. 2019 Nov 04.
    CD9 identifies pancreatic cancer stem cells and modulates glutamine metabolism to fuel tumour growth.
    Victoria M-Y Wang, Rute M M Ferreira, Jorge Almagro, Theodore Evan, Nathalie Legrave, May Zaw Thin, David Frith, Joana Carvalho, David J Barry, Ambrosius P Snijders, Eleanor Herbert, Emma L Nye, James I MacRae, Axel Behrens.
      Pancreatic ductal adenocarcinoma (PDAC) shows great cellular heterogeneity, with pronounced epithelial and mesenchymal cancer cell populations. However, the cellular hierarchy underlying PDAC cell diversity is unknown. Here we identify the tetraspanin CD9 as a marker of PDAC tumour-initiating cells. CD9high cells had increased organoid formation capability, and generated tumour grafts in vivo at limiting dilutions. Tumours initiated from CD9high cells recapitulated the cellular heterogeneity of primary PDAC, whereas CD9low cells produced only duct-like epithelial progeny. CD9 knockdown decreased the growth of PDAC organoids, and heterozygous CD9 deletion in Pdx1-Cre; LSL-KRasG12D; p53F/F mice prolonged overall survival. Mechanistically, CD9 promoted the plasma membrane localization of the glutamine transporter ASCT2, enhancing glutamine uptake in PDAC cells. Thus, our study identifies a PDAC subpopulation capable of initiating PDAC and giving rise to PDAC heterogeneity, suggesting that the cellular diversity of PDAC is generated by PDAC stem cell differentiation.
    DOI:  https://doi.org/10.1038/s41556-019-0407-1
  16. J Biol Chem. 2019 Nov 05. pii: jbc.RA119.009868. [Epub ahead of print]
    Hepatic stearoyl CoA desaturase 1 deficiency increases glucose uptake in adipose tissue partially through the PGC-1α-FGF21 axis in mice.
    Ahmed AlJohani, Mohammad Imran Khan, Abram Bonneville, Changan Guo, Justin Jeffery, Lucas O'Neill, Deeba Nadeem Syed, Sarah A Lewis, Maggie Burhans, Hasan Mukhtar, James M Ntambi.
      Increased carbohydrate consumption increases hepatic de novo lipogenesis, which has been linked to the development of chronic metabolic diseases, including obesity, hepatic steatosis, and insulin resistance. Stearoyl CoA desaturase 1 (SCD1) is a critical lipogenic enzyme that catalyzes the synthesis of two monounsaturated fatty acids (MUFA), oleate and palmitoleate, from the saturated fatty acids, stearate and palmitate, respectively. SCD1-deficient mouse models are protected against diet-induced adiposity, hepatic steatosis, and hyperglycemia. However, the mechanism of this protection by SCD1 deficiency is unclear. Using a liver-specific SCD1 knockout (LKO) mouse model fed a high-carbohydrate, low-fat diet (HCD), we show that hepatic SCD1 deficiency increases systemic glucose uptake. Hepatic SCD1 deficiency enhanced glucose transporter type 1 (GLUT1) expression in the liver and also up-regulated GLUT4 and adiponectin expression in the adipose tissue. The enhanced glucose uptake correlated with increased liver expression and elevated plasma levels of fibroblast growth factor 21 (FGF21), a hepatokine known to increase systemic insulin sensitivity and to regulate whole-body lipid metabolism. Feeding LKO mice with triolein-supplemented, but not tristearin-supplemented, HCD reduced FGF21 expression and plasma levels. Consistently, SCD1 inhibition in primary hepatocytes induced FGF21 expression, which was repressed by treatment with oleate but not palmitoleate. Moreover, deletion of the transcriptional coactivator PPARg coactivator 1α (PGC-1α) reduced hepatic and plasma FGF21 and white adipocyte tissue-specific GLUT4 expression and raised plasma glucose levels in the LKO mice. These results suggest that hepatic oleate regulates glucose uptake in adipose tissue either directly or partially by modulating the hepatic PGC-1α-FGF21 axis.
    Keywords:  Glucose transporters-Insulin sensitivity; Heptokine-FGF21; Liver-adipocyte-crosstalk; Monounsaturated fatty acids; Stearoyl-CoA Desaturase-1; adiponectin; carbohydrate metabolism; fibroblast growth factor (FGF); glucose metabolism; glucose transport; lipid metabolism; lipid signaling; liver; liver metabolism; obesity
    DOI:  https://doi.org/10.1074/jbc.RA119.009868
  17. Metabolites. 2019 Oct 31. pii: E257. [Epub ahead of print]9(11):
    Inter-Laboratory Comparison of Metabolite Measurements for Metabolomics Data Integration.
    Yoshihiro Izumi, Fumio Matsuda, Akiyoshi Hirayama, Kazutaka Ikeda, Yoshihiro Kita, Kanta Horie, Daisuke Saigusa, Kosuke Saito, Yuji Sawada, Hiroki Nakanishi, Nobuyuki Okahashi, Masatomo Takahashi, Motonao Nakao, Kosuke Hata, Yutaro Hoshi, Motohiko Morihara, Kazuhiro Tanabe, Takeshi Bamba, Yoshiya Oda.
       BACKGROUND: One of the current problems in the field of metabolomics is the difficulty in integrating data collected using different equipment at different facilities, because many metabolomic methods have been developed independently and are unique to each laboratory.
    METHODS: In this study, we examined whether different analytical methods among 12 different laboratories provided comparable relative quantification data for certain metabolites. Identical samples extracted from two cell lines (HT-29 and AsPc-1) were distributed to each facility, and hydrophilic and hydrophobic metabolite analyses were performed using the daily routine protocols of each laboratory.
    RESULTS: The results indicate that there was no difference in the relative quantitative data (HT-29/AsPc-1) for about half of the measured metabolites among the laboratories and assay methods. Data review also revealed that errors in relative quantification were derived from issues such as erroneous peak identification, insufficient peak separation, a difference in detection sensitivity, derivatization reactions, and extraction solvent interference.
    CONCLUSION: The results indicated that relative quantification data obtained at different facilities and at different times would be integrated and compared by using a reference materials shared for data normalization.
    Keywords:  data integration; inter-laboratory comparison; metabolomics; method validation; quality control sample; relative quantification
    DOI:  https://doi.org/10.3390/metabo9110257
  18. Metabolites. 2019 Nov 01. pii: E259. [Epub ahead of print]9(11):
    Lipid Profile Characterization and Lipoprotein Comparison of Extracellular Vesicles from Human Plasma and Serum.
    Yuchen Sun, Kosuke Saito, Yoshiro Saito.
      Extracellular vesicles (EVs) consist of lipid bilayers, occur in various biofluids, and are invaluable in biomarker screening. Liquid chromatography coupled with high-resolution mass spectrometry (LC-MS) was recently used to study comprehensive EV lipid profiles in vitro. The aim of this study was to establish a lipidomics platform for human plasma and serum EVs for comprehensive characterization of their lipid profiles, and to compare them with those of other lipid-containing particles, such as high-density lipoproteins (HDL), and low/very low-density lipoproteins (LDL/VLDL). Isolation was validated by specific protein markers; CD9 and MHC class for EVs, apoA-I for HDL, and apoB-100 for LDL/VLDL. Lipidomics identified 264 lipids from isolated plasma EVs, HDL, and LDL/VLDL. The absolute lipid levels per unit protein content in the EVs were more than eight times lower than those of the lipoproteins. Moreover, the EVs had higher lysoglycerophospholipid levels than HDL or LDL/VLDL. Similar profiles were also determined for human serum. The present study found that the lipid profiles of EVs are unique and distinctly different from those of lipoproteins. The lipidomics platform applied to human plasma and serum EVs could generate important information for the exploration and qualification of biomarkers in disease diagnosis.
    Keywords:  extracellular vesicles; lipidomics; lipoproteins; mass spectrometry
    DOI:  https://doi.org/10.3390/metabo9110259
  19. Cancers (Basel). 2019 Oct 31. pii: E1696. [Epub ahead of print]11(11):
    Role of Sphingosylphosphorylcholine in Tumor and Tumor Microenvironment.
    Mi Kyung Park, Chang Hoon Lee.
      Sphingosylphosphorylcholine (SPC) is a unique type of lysosphingolipid found in some diseases, and has been studied in cardiovascular, neurological, and inflammatory phenomena. In particular, SPC's studies on cancer have been conducted mainly in terms of effects on cancer cells, and relatively little consideration has been given to aspects of tumor microenvironment. This review summarizes the effects of SPC on cancer and tumor microenvironment, and presents the results and prospects of modulators that regulate the various actions of SPC.
    Keywords:  inflammatory; lysosphingolipids; neurological; sphingosylphosphorylcholine; tumor microenvironment
    DOI:  https://doi.org/10.3390/cancers11111696
  20. Science. 2019 Nov 07. pii: eaav2588. [Epub ahead of print]
    Glutamine blockade induces divergent metabolic programs to overcome tumor immune evasion.
    Robert D Leone, Liang Zhao, Judson M Englert, Im-Meng Sun, Min-Hee Oh, Im-Hong Sun, Matthew L Arwood, Ian A Bettencourt, Chirag H Patel, Jiayu Wen, Ada Tam, Richard L Blosser, Eva Prchalova, Jesse Alt, Rana Rais, Barbara S Slusher, Jonathan D Powell.
      The metabolic characteristics of tumors present significant hurdles to immune cell function and cancer immunotherapy. Using a novel glutamine antagonist, we metabolically dismantled the immunosuppressive microenvironment of tumors. We demonstrate that glutamine blockade in tumor-bearing mice suppresses oxidative and glycolytic metabolism of cancer cells, leading to decreased hypoxia, acidosis, and nutrient depletion. In contrast, effector T cells responded to glutamine antagonism by markedly upregulating oxidative metabolism and adopting a long-lived, highly-activated phenotype. These divergent changes in cellular metabolism and programming form the basis for potent anti-tumor responses. Glutamine antagonism therefore exposes a previously undefined difference in metabolic plasticity between cancer cells and effector T cells that can be exploited as a "metabolic checkpoint" for tumor immunotherapy.
    DOI:  https://doi.org/10.1126/science.aav2588
  21. Nucleic Acids Res. 2019 Nov 06. pii: gkz1019. [Epub ahead of print]
    MetaboLights: a resource evolving in response to the needs of its scientific community.
    Kenneth Haug, Keeva Cochrane, Venkata Chandrasekhar Nainala, Mark Williams, Jiakang Chang, Kalai Vanii Jayaseelan, Claire O'Donovan.
      MetaboLights is a database for metabolomics studies, their raw experimental data and associated metadata. The database is cross-species and cross-technique and it covers metabolite structures and their reference spectra as well as their biological roles and locations. MetaboLights is the recommended metabolomics repository for a number of leading journals and ELIXIR, the European infrastructure for life science information. In this article, we describe the significant updates that we have made over the last two years to the resource to respond to the increasing amount and diversity of data being submitted by the metabolomics community. We refreshed the website and most importantly, our submission process was completely overhauled to enable us to deliver a far more user-friendly submission process and to facilitate the growing demand for reproducibility and integration with other 'omics. Metabolomics resources and data are available under the EMBL-EBI's Terms of Use via the web at https://www.ebi.ac.uk/metabolights and under Apache 2.0 at Github (https://github.com/EBI-Metabolights/).
    DOI:  https://doi.org/10.1093/nar/gkz1019
  22. Cancer Discov. 2019 Nov 08.
    Novel MYC-Targeting Drug Is Effective in Mouse Prostate Cancer Models.
      A new MYC inhibitor reduced tumor growth in mouse prostate cancer models.
    DOI:  https://doi.org/10.1158/2159-8290.CD-RW2019-167
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