bims-mascan 2025-03-23 papers

bims-mascan

Biomed News

on Mass spectrometry in cancer research

Issue of 2025–03–23
29 papers selected by
Giovanny Rodríguez Blanco, Uniklinikum Graz

Clinical Lipidomics Reveals High Individuality and Sex Specificity of Circulatory Lipid Signatures: A Prospective Healthy Population Study.
Benchmarking of quantitative proteomics workflows for Limited proteolysis mass spectrometry.
Quantitative Lipid Analysis of Extracellular Vesicle Preparations: A Perspective.
Beyond Targeted Newborn Screening: A Nontargeted Metabolomics Workflow to Investigate Birthweight-Metabolome Correlations.
Advances in total glycomic analysis including sialylated sub-glycan isomers by SALSA method.
Frontiers in Mass Spectrometry-Based Spatial Metabolomics: Current Applications and Challenges in the Context of Biomedical Research.
Determination of Free Amino Acid Concentrations in Bovine Plasma Using High-Pressure Liquid Chromatography With Electrospray Ionization Mass Spectrometry Detection.
Quetzal: Comprehensive Peptide Fragmentation Annotation and Visualization.
Toward Real-Time Proteomics: Blood to Biomarker Quantitation in under One Hour.
A novel, high throughput, and low-cost method for the detection of 40 amines relevant to inborn errors of metabolism, in under 60 min, using reverse phase high performance liquid chromatography.
Trends in Mass Spectrometry-Based Single-Cell Proteomics.
Light-Reactive Norharmane Derivatization of Lipid Isomers by Matrix-Assisted Laser Desorption/Ionization Mass Spectrometry.
Multidimensional Separations for Characterization of Isomeric PFAS Using SLIM High-Resolution Ion Mobility and Tandem Mass Spectrometry.
The metabolomic profile features of some bio logical fluids in serous ovarian adenocarcinoma patients.
Application of rapid evaporative ionization mass spectrometry in preclinical and clinical analyses of steatotic liver tissues and cells.
Untargeted pixel-by-pixel metabolite ratio imaging as a novel tool for biomedical discovery in mass spectrometry imaging.
Monitoring Functional Posttranslational Modifications Using a Data-Driven Proteome Informatic Pipeline.
MGVB: a New Proteomics Toolset for Fast and Efficient Data Analysis.
High resolution profiling of cell cycle-dependent protein and phosphorylation abundance changes in non-transformed cells.
A Single-Step Protein Extraction for Lung Extracellular Matrix Proteomics Enabled by the Photocleavable Surfactant Azo and timsTOF Pro.
The time is now: accounting for time-of-day effects to improve reproducibility and translation of metabolism research.
Metabolic Reprogramming into a Glycolysis Phenotype Induced by Extracellular Vesicles Derived from Prostate Cancer Cells.
Plasma-based untargeted metabolomics reveals potential biomarkers for screening and distinguishing of ovarian tumors.
Lactate dehydrogenase A-coupled NAD+ regeneration is critical for acute myeloid leukemia cell survival.
Genetically encoding ε-N-methacryllysine into proteins in live cells.
Addressing NHS chemistry: Efficient quenching of excess TMT reagent and reversing TMT-over labelling in proteomic samples by methylamine.
Targeted proteomics of serum IGF-I, -II, IGFBP-2, -3, -4, -5, -6 and ALS.
LipidSigR: a R-based solution for integrated lipidomics data analysis and visualization.
Circulating metabolite signatures indicate differential gut-liver crosstalk in lean and obese MASLD.

J Lipid Res. 2025 Mar 18. pii: S0022-2275(25)00040-9. [Epub ahead of print] 100780

Clinical Lipidomics Reveals High Individuality and Sex Specificity of Circulatory Lipid Signatures: A Prospective Healthy Population Study.

Jessica Medina, Nicolas Goss, Gonçalo Dos Santos Correia, Rebecca Borreggine, Tony Teav, Zoltan Kutalik, Pedro Marques Vidal, Hector Gallart-Ayala, Julijana Ivanisevic.

  Lipid metabolism and circulatory lipid levels are tightly associated with the (cardio)metabolic health. Consequently, mass spectrometry (MS)-based lipidomics has emerged as a powerful phenotyping tool in epidemiological, human population, and in clinical intervention studies. However, ensuring high throughput and reproducible measurement of a wide panel of circulatory lipid species in large-scale studies poses a significant challenge. Here, we applied a recently developed quantitative LC-MS/MS lipidomics approach to a subset of 1086 fasted plasma samples belonging to apparently healthy participants from prospective Lausanne population study. This high-coverage and high-throughput HILIC-based methodology allowed for the robust measurement of 782 circulatory lipid species spanning 22 lipid classes and six orders of magnitude-wide concentration range. This was achieved by combining semi-automated sample preparation using a stable isotope dilution approach and the alternate analysis of NIST plasma reference material, as a quality control. Based on NIST QC analysis, median between-batch reproducibility was 8.5%, over the course of analysis of 13 independent batches comprising 1086 samples collected from 364 individuals at three time points. Importantly, the biological variability, per lipid species, was significantly higher than the batch-to-batch analytical variability. Furthermore, the significantly lower between-subject (compared to within-subject) variability, and unsupervised sample clustering demonstrated the high individuality and sex-specificity of circulatory lipidome. The most prominent sex differences were reported for sphingomyelins and ether-linked phospholipids present in significantly higher concentrations in female plasma. The high individuality and sex specificity of circulatory lipidome constitute important advantages for the application of lipidomics in next-generation metabolic health monitoring.

Keywords:  Circulatory lipids; Clinical lipidomics; HILIC-MS/MS; Personalized signatures; Prospective healthy population; Sex differences

DOI:  https://doi.org/10.1016/j.jlr.2025.100780
Mol Cell Proteomics. 2025 Mar 13. pii: S1535-9476(25)00043-X. [Epub ahead of print] 100945

Benchmarking of quantitative proteomics workflows for Limited proteolysis mass spectrometry.

Tomas Koudelka, Claudio Bassot, Ilaria Piazza.

  Limited proteolysis coupled with mass spectrometry (LiP-MS) has emerged as a powerful technique for detecting protein structural changes and drug-protein interactions on a proteome-wide scale. However, there is no consensus on the best quantitative proteomics workflow for analyzing LiP-MS data. In this study, we comprehensively benchmarked two major quantification approaches-data-independent acquisition (DIA) and tandem mass tag (TMT) isobaric labeling-in combination with LiP-MS, using a drug-target deconvolution assay as a model system. Our results show that while TMT labeling enabled the quantification of more peptides and proteins with lower coefficients of variation (CVs), DIA-MS exhibited greater accuracy in identifying true drug targets and stronger dose-response correlation in protein targets peptides. Additionally, we evaluated the performance of freely available (FragPipe) versus commercial (Spectronaut) software tools for DIA-MS analysis, revealing that the choice between precision (FragPipe) and sensitivity (Spectronaut) largely depends on the specific experimental context. Our findings underscore the importance of selecting the appropriate LiP-MS quantification strategy based on the study objectives. This work provides valuable guidelines for researchers in structural proteomics and drug discovery, and highlights how advancements in mass spectrometry instrumentation, such as the Astral mass spectrometer, may further improve sensitivity and protein sequence coverage, potentially reducing the need for TMT labeling.

Keywords:  DIA software benchmarking; DIA-MS; FAIMS; FragPipe; LiP-MS; TMT; structural proteomics

DOI:  https://doi.org/10.1016/j.mcpro.2025.100945
J Extracell Vesicles. 2025 Mar;14(3): e70049

Quantitative Lipid Analysis of Extracellular Vesicle Preparations: A Perspective.

Tore Skotland, Kim Ekroos, Alicia Llorente, Kirsten Sandvig.

  Quantitative lipidomic analysis performed by mass spectrometry is required for determination of the lipid content of extracellular vesicles (EVs). Such methods can provide information about the total amount of lipids, the lipid species composition, the purity of EV samples as well as the cellular origin of the EVs. There are, however, many pitfalls when performing lipid analyses. Thus, any non-specialist should collaborate with experts in lipidomics. In addition to many good review articles giving advice about lipid analyses, we recommend the information and guidelines published by the Lipidomic Standard Initiative, an interest group affiliated with the International Lipidomics Society.

Keywords:  exosomes; extracellular vesicles; lipid analyses; mass spectrometry; membranes

DOI:  https://doi.org/10.1002/jev2.70049
Anal Chem. 2025 Mar 18.

Beyond Targeted Newborn Screening: A Nontargeted Metabolomics Workflow to Investigate Birthweight-Metabolome Correlations.

Carter K Asef, Samuel G Moore, Charles Austin Pickens, Carlos A Saavedra-Matiz, Joseph J Orsini, Konstantinos Petritis, David A Gaul, Facundo M Fernández.

Newborn screening (NBS) is one of the United States' largest, most successful preventative public health initiatives, improving outcomes for newborns with inborn errors of metabolism. Most disorders on the Recommended Uniform Screening Panel are screened using triple-quadrupole mass spectrometry and flow injection analysis. While these methods are sensitive and well suited for high-throughput quantitative applications, the breadth of measured analytes is limited to a relatively small number of biomarkers, which often have considerable overlaps between healthy and diseased populations. High-resolution liquid chromatography-mass spectrometry (LC-MS)-based metabolomics is now capable of profiling thousands of metabolites, making it well suited for exploratory and biomarker discovery studies. To this end, we developed a robust workflow for performing nontargeted LC-MS analysis on dried bloodspot (DBS) specimens with coverage across many metabolic pathways relevant to NBS. HILIC chromatography enabled quantitation of amino acid and acylcarnitine species while also retaining lipid species, such as lyso-phosphatidylcholines. We analyzed 810 newborn-derived DBS samples across a wide range of newborn birthweights, identifying correlations with metabolites that help to better account for the lower accuracy observed for some NBS markers (e.g., isovalerylcarnitine). Additionally, we leveraged this nontargeted workflow to capture new biomarkers and metabolic phenotypes in newborns associated with parenteral nutrition administration and maternal nicotine exposure. Two critical biomarkers were identified as useful additions to targeted screening panels: N-acetyltyrosine as a qualitative marker for parenteral nutrition administration and N-acetylputrescine as a quantitative marker for controlling birthweight variability.

DOI: https://doi.org/10.1021/acs.analchem.4c06061
BBA Adv. 2025 ;7 100144

Advances in total glycomic analysis including sialylated sub-glycan isomers by SALSA method.

Masaki Kurogochi, Chiharu Suzuki, Hisatoshi Hanamatsu, Jun-Ichi Furukawa.

  All eukaryotic cell surfaces are coated with various types of glycans, which are essential molecules in biological events. In this review, we summarize recent integrated glycomics studies using various biological samples. We introduce an improved sialic acid linkage-specific alkylamidation (SALSA) method for sialylated glycan analysis and an automated glycosphingolipid-glycan preparation system for large-scale glycomic analysis of human plasma/serum. Finally, we explain the importance of integrated glycomics of glycoconjugates through total glycomic analysis of human serum and mouse brain tissue, and discuss prospects for exploring glycans as effective biomarkers of biological phenomena.

Keywords:  Automation; Glycoconjugate; Glycomics; Mass spectrometry; Sialic acid

DOI:  https://doi.org/10.1016/j.bbadva.2025.100144
Trends Analyt Chem. 2024 Jun;pii: 117713. [Epub ahead of print]175

Frontiers in Mass Spectrometry-Based Spatial Metabolomics: Current Applications and Challenges in the Context of Biomedical Research.

Kate Wheeler, Camil Gosmanov, Michael Jimenez Sandoval, Zhibo Yang, Laura-Isobel McCall.

  Metabolites are critical products and mediators of cellular and tissue function, and key signals in cell-to-cell, organ-to-organ and cross-organism communication. Many of these interactions are spatially segregated. Thus, spatial metabolomics can provide valuable insight into healthy tissue function and disease pathogenesis. Here, we review major mass spectrometry-based spatial metabolomics techniques and the biological insights they have enabled, with a focus on brain and microbiota function and on cancer, neurological diseases and infectious diseases. These techniques also present significant translational utility, for example in cancer diagnosis, and for drug development. However, spatial mass spectrometry techniques still encounter significant challenges, including artifactual features, metabolite annotation, open data, and ethical considerations. Addressing these issues represent the future challenges in this field.

Keywords:  DESI mass spectrometry imaging; MALDI mass spectrometry imaging; chemical cartography; mass spectrometry; metabolomics; secondary ion mass spectrometry; spatial analysis

DOI:  https://doi.org/10.1016/j.trac.2024.117713
Rapid Commun Mass Spectrom. 2025 Apr 15. 39(11): e10027

Determination of Free Amino Acid Concentrations in Bovine Plasma Using High-Pressure Liquid Chromatography With Electrospray Ionization Mass Spectrometry Detection.

Laurie A Reinhardt, Levi Svaren, Ritvik Marathe, Geoffrey I Zanton, Michael L Sullivan.

   RATIONALE: Quantification of free amino acid concentrations in plasma has become an important tool in monitoring the health of dairy cows and health of their offspring under various management regimes, especially diet. Consequently, it was desirable to develop a robust, accurate, medium-throughput method to quantitate free amino acid concentrations in bovine plasma.
METHODS: Bovine plasma was deproteinated with methanol and amino acids partially purified using cation exchange resin. Samples were then subjected to precolumn derivatization with phenyl isothiocyanate, followed by high-pressure liquid chromatography with positive electrospray ionization single quadrupole mass spectrometry detection for analysis. The corresponding 13C and 15N labeled amino acids (mass unit difference > 3) were used as internal standards, while deuterium labeled standards were used for other metabolites.
RESULTS: All 20 amino acids showed linear fits to their individual calibration curves (correlation coefficients > 0.99) with concentration range of amino acids measured from 5 to 600 μM. Coefficient of variation (CV) values for the concentrations measured for all amino acids ranged from 2.0 to 6.7 for intraday aliquots and from 1.0 to 4.6 for interday aliquots with the exception of aspartic acid (11.1 and 12.6 for intraday and interday, respectively).
CONCLUSIONS: The use of a stable isotope labeled version of each amino acid analyte as internal standard added to plasma samples at the beginning of the procedure corrected for any losses, instrument variability, and chemistry of derivatization. Use of this method to quantify bovine plasma amino acids will allow better understanding of physiological processes underlying nutritional interventions in dairy production systems and may be more broadly applicable to ruminant and other animal production systems.

Keywords:  amino acid analysis; dairy cow; plasma

DOI:  https://doi.org/10.1002/rcm.10027
J Proteome Res. 2025 Mar 20.

Quetzal: Comprehensive Peptide Fragmentation Annotation and Visualization.

Eric W Deutsch, Luis Mendoza, Robert L Moritz.

  Proteomics data-dependent acquisition data sets collected with high-resolution mass-spectrometry (MS) can achieve very high-quality results, but nearly every analysis yields results that are thresholded at some accepted false discovery rate, meaning that a substantial number of results are incorrect. For study conclusions that rely on a small number of peptide-spectrum matches being correct, it is thus important to examine at least some crucial spectra to ensure that they are not one of the incorrect identifications. We present Quetzal, a peptide fragment ion spectrum annotation tool to assist researchers in annotating and examining such spectra to ensure that they correctly support study conclusions. We describe how Quetzal annotates spectra using the new Human Proteome Organization (HUPO) Proteomics Standards Initiative (PSI) mzPAF standard for fragment ion peak annotation, including the Python-based code, a web-service end point that provides annotation services, and a web-based application for annotating spectra and producing publication-quality figures. We illustrate its functionality with several annotated spectra of varying complexity. Quetzal provides easily accessible functionality that can assist in the effort to ensure and demonstrate that crucial spectra support study conclusions. Quetzal is publicly available at https://proteomecentral.proteomexchange.org/quetzal/.

Keywords:  Quetzal; Universal Spectrum Identifier; mass spectrometry; mzPAF; proteomics; spectrum annotation

DOI:  https://doi.org/10.1021/acs.jproteome.5c00092
Anal Chem. 2025 Mar 20.

Toward Real-Time Proteomics: Blood to Biomarker Quantitation in under One Hour.

Steven M Yannone, Vikas Tuteja, Olena Goleva, Donald Y M Leung, Aleksandr Stotland, Angel J Keoseyan, Nathan G Hendricks, Sarah Parker, Jennifer E Van Eyk, Simion Kreimer.

Multistep multihour tryptic proteolysis has limited the utility of bottom-up proteomics for cases that require immediate quantitative information. The power of proteomics to quantify biomarkers of health status cannot practically assist in clinical care if the dynamics of disease outpaces the turnaround of analysis. The recently available hyperthermoacidic archaeal (HTA) protease "Krakatoa" digests samples in a single 5 to 30 min step at pH 3 and >80 °C in conditions that disrupt most cells and tissues, denature proteins, and block disulfide reformation thereby dramatically expediting and simplifying sample preparation. The combination of quick single-step proteolysis with high-throughput dual-trapping single analytical column (DTSC) liquid chromatography-mass spectrometry (LC-MS) returns actionable data in less than 1 h from collection of unprocessed biofluid. The systematic evaluation of this methodology finds that over 160 proteins are quantified in less than 1 h from 1 μL of whole blood. Furthermore, labile Angiotensin I and II bioactive peptides along with a panel of protein species can be measured at 8 min intervals with a 20 min initial lag using targeted MS. With these methods, we analyzed serum and plasma from 53 individuals and quantified Angiotensin I and II and over 150 proteins including at least 46 that were not detected with trypsin. We discuss some of the implications of real-time proteomics including the immediate potential to advance several clinical and research applications.

DOI: https://doi.org/10.1021/acs.analchem.4c05172
Mol Genet Metab Rep. 2025 Jun;43 101202

A novel, high throughput, and low-cost method for the detection of 40 amines relevant to inborn errors of metabolism, in under 60 min, using reverse phase high performance liquid chromatography.

Kirkland A Wilson, Yun Zhou, Gary Cunningham, Kimberly Chapman, Marshall Summar, Debra Regier.

   Objectives: An assessment of amino acid and amine concentrations is important for the diagnosis and management of inherited metabolic disorders (IMDs). Methods exist that measure these biologically important metabolites but are cost-prohibitive and/or time consuming. We therefore sought to develop a novel methodology, applicable to IMDs, that is both high-throughput and low cost.
Methods: Previously, we developed a methodology for rapid, repeatable, and cost-efficient separation of approximately 20 amines as a proof of concept and now expand it to amines relevant to IMDs. We describe our separation methodology using reverse phase high performance liquid chromatography with ultraviolet-visible spectrum absorbance paired with pre-column derivatization with o-pthalaldehyde.
Results: We show reproducibility via concentration assessments, in triplicate, for each amine. We assess amines in prepared standard solutions and in biologic samples from patients with IMDs. We also detected and assessed the amino group containing compounds glutathione (oxidized and reduced forms) and ammonia. Validation was established using absolute area under the curve (AUC) and via comparison using a single internal standard.
Conclusions: We report good separation of 40 primary amino group containing metabolites, in a single, 53 min run. This rapid, low cost, and accurate methodology only requires a small volume of sample and can greatly increase availability and access. Finally, the numerous amines and unique compounds detected in our single run has large utility and can potentially increase clinical efficiency and broaden access to research, both important as the need for analysis of amines grows globally.

Keywords:  Amine; Amino acid; Chromatography; Inborn error of metabolism; Metabolomic; RP-HPLC

DOI:  https://doi.org/10.1016/j.ymgmr.2025.101202
Anal Chem. 2025 Mar 16.

Trends in Mass Spectrometry-Based Single-Cell Proteomics.

Ximena Sanchez-Avila, Raphaela M de Oliveira, Siqi Huang, Chao Wang, Ryan T Kelly.

DOI: https://doi.org/10.1021/acs.analchem.5c00661
Anal Chem. 2025 Mar 15.

Light-Reactive Norharmane Derivatization of Lipid Isomers by Matrix-Assisted Laser Desorption/Ionization Mass Spectrometry.

Thomas R Fredriksen, Joseph H Holbrook, Lorena Rosas, Quetzalli D Angeles-Lopez, Ana L Mora, Mauricio Rojas, Amanda B Hummon.

The lipidome, encompassing the comprehensive lipid fingerprint of a biological system, includes thousands of unique isomeric and isobaric lipid species. Mass spectrometry (MS) is an effective technique for characterizing the lipidome, although the resolution of isomeric lipid species through MS typically requires specialized or modified equipment. In this study, we introduce a novel matrix derivatization technique that leverages the unique photoreactive properties of unsaturated lipids to reveal the double-bond location in conventional matrix-assisted laser desorption-ionization mass spectrometry (MALDI-MS) experiments. The principle mechanistic framework of this technique is type II photosensitization, where the MALDI matrix norharmane acts as an organic photosensitizer to generate singlet oxygen upon light exposure. The singlet oxygen then reacts with unsaturated lipid species, forming hydroperoxide derivatives at acyl group carbon double bonds, facilitating their identification. The labile nature of these hydroperoxide-functionalized lipids allows for further decomposition under normal MALDI laser exposure, enhancing the analytical resolution of isomeric lipids without additional experiments. With this approach, we were able to distinguish the 18:1 (Δ6-cis) and 18:1 (Δ9-cis) PC lipid isomers. We also demonstrated that the approach works in an imaging context, mapping lipid species in both mouse tissue and 3D cell cultures.

DOI: https://doi.org/10.1021/acs.analchem.4c05629
Anal Chem. 2025 Mar 17.

Multidimensional Separations for Characterization of Isomeric PFAS Using SLIM High-Resolution Ion Mobility and Tandem Mass Spectrometry.

Heidi M Sabatini, Terra Pettit-Bacovin, Ralph Aderorho, Christopher D Chouinard.

Per- and polyfluoroalkyl substances (PFAS) are synthetic organofluorine compounds that accumulate in the environment due to significant industrial use and resistance to degradation. PFAS are of global interest because of their environmental and health concerns. They exist in a variety of linear and nonlinear forms containing a variety of isomers, as well as differing functional headgroups for each class. That structural complexity requires advanced analytical techniques, beyond current high-resolution mass spectrometry (HRMS) methods, for their accurate identification and quantification in a wide range of samples. Herein, we demonstrate the power of Structures for Lossless Ion Manipulations (SLIM)-based high-resolution ion mobility (HRIM) for separation of complex PFAS branched isomers. SLIM is integrated into a multidimensional LC-SLIM IM-MS/MS workflow, developed for the extensive characterization of a wide range of PFAS compounds. As we surveyed sulfonate and carboxylic acid classes of PFAS, we observed unique arrival time vs m/z trend lines that were representative of each class; these trend lines are important for allowing identification of emerging species based on their placement in that two-dimensional space. Next, we used complementary tandem mass spectrometry (MS/MS) approaches with all ion fragmentation (AIF), as well as energy-resolved MS/MS, to further investigate the structure of mobility-separated species. This allowed both investigation of fragmentation mechanism and identification of unique fragment ions that could allow differentiation of isomers when ion mobility was insufficient. Overall, the combination of chromatography, high-resolution SLIM, and MS/MS provided a comprehensive workflow capable of identifying unknown emerging PFAS compounds in complex environmental samples.

DOI: https://doi.org/10.1021/acs.analchem.4c06985
Klin Onkol. 2025 ;38(1): 38-44

The metabolomic profile features of some bio logical fluids in serous ovarian adenocarcinoma patients.

D S Kutilin, F E Filippov, O N Guskova, I A Alliluev, Y S Enin, A Y Maksimov.

   BACKGROUND: The search for effective biomarkers for ovarian cancer (OC) early diagnosis is an urgent task of modern oncogynecology. Metabolic profiling by ultra-high performance liquid chromatography and mass spectrometry (UHPLC-MS) provides information on the totality of all low molecular weight metabolites of patient's biological fluids sample, reflecting the processes occurring in the body. The aim of the study was to research blood plasma and urine metabolomic profile of patients with serous ovarian adenocarcinoma by UHPLC-MS.
MATERIAL AND METHODS: To perform metabolomic analysis, 60 blood plasma samples and 60 urine samples of patients diagnosed with serous ovarian carcinoma and 20 samples of apparently healthy volunteers were taken. Chromatographic separation was performed on a Vanquish Flex UHPLC System chromatograph (Thermo Scientific, Germany). Mass spectrometric analysis was performed on an Orbitrap Exploris 480 (Thermo Scientific, Germany) equipped with an electrospray ionization source. Bioinformatic analysis was performed using Compound Discoverer Software (Thermo Fisher Scientific, USA), statistical data analysis was performed in the Python programming language using the SciPy library.
RESULTS: Using UHPLC-MS, 1,049 metabolites of various classes were identified in blood plasma. In patients with OC, 8 metabolites had a significantly lower concentration (P < 0.01) compared with conditionally healthy donors, while the content of 19 compounds, on the contrary, increased (P < 0.01). During the metabolomic profiling of urine samples, 417 metabolites were identified: 12 compounds had a significantly lower concentration compared to apparently healthy individuals, the content of 14 compounds increased (P < 0.01). In patients with ovary serous adenocarcinoma, a significant change in the metabolome of blood plasma and urine was found, expressed in abnormal concentrations of lipids and their derivatives, fatty acids and their derivatives, acylcarnitines, phospholipids, amino acids and their derivatives, derivatives of nitrogenous bases and steroids. At the same time, kynurenine, myristic acid, lysophosphatidylcholine and L-octanoylcarnitine are the most promising markers of this disease.
CONCLUSION: The revealed changes in the metabolome can become the basis for improving approaches to the diagnosis of serous ovarian adenocarcinoma.

Keywords:  Urine; blood plasma; metabolomic profile; serous ovarian adenocarcinoma; ultra-high performance liquid chromatography and mass spectrometry; urine

DOI:  https://doi.org/10.48095/ccko202538
Sci Rep. 2025 Mar 18. 15(1): 9226

Application of rapid evaporative ionization mass spectrometry in preclinical and clinical analyses of steatotic liver tissues and cells.

Julian Connor Eckel, Lena Seidemann, Mohamed Albadry, Gerda Schicht, Marija Skvoznikova, Sandra Nickel, René Hänsel, Daniel Seehofer, Grit Gesine Ruth Hiller, Hans-Michael Tautenhahn, Uta Dahmen, Georg Damm.

  Rapid evaporative ionization mass spectrometry (REIMS) shows promise as a preparation-free tissue analysis tool with the prospect for real-time diagnostics. Given that hepatic steatosis is characterized by shifts in lipid species and abundance, we selected it as basis for method development, as REIMS specifically measures lipidomic profiles. However, further validation and protocol refinement are necessary to establish its clinical utility. In this study, we applied REIMS to steatotic human liver tissues, focusing on its ability to differentiate varying degrees of steatosis. We established standardized protocols for tissue handling and lipid analysis, which were essential for reliable data interpretation. Notably, our findings revealed that tissue size impacts REIMS sensitivity, with smaller samples yielding lower total ion counts and altered lipid profiles. Through principal component analysis, we identified key lipid classes, namely triacylglycerides, fatty acids, and glycerophospholipids. Despite a missing link between triacylglyceride abundance and degree of steatosis, we successfully identified condition-specific lipid patterns, with ceramides emerging as markers of advanced steatosis. Our study provides a protocol for the measurements of lipid standards showing the detailed degradation of specific lipids using iKnife-coupled REIMS. It highlights the pitfalls and limitations and provides critical recommendations for REIMS use. It also emphasizes the need for standardized biobanking and tissue preparation to ensure accurate lipid profiling, laying the groundwork for future protocol adjustments required for clinical application.

Keywords:  Lipidomics; Lipids; MASLD; Rapid evaporative ionization mass spectrometry; Steatosis

DOI:  https://doi.org/10.1038/s41598-025-93305-w
Elife. 2025 Mar 18. pii: RP96892. [Epub ahead of print]13

Untargeted pixel-by-pixel metabolite ratio imaging as a novel tool for biomedical discovery in mass spectrometry imaging.

Huiyong Cheng, Dawson Miller, Nneka Southwell, Paola Porcari, Joshua L Fischer, Isobel Taylor, J Michael Salbaum, Claudia Kappen, Fenghua Hu, Cha Yang, Kayvan R Keshari, Steven S Gross, Marilena D'Aurelio, Qiuying Chen.

  Mass spectrometry imaging (MSI) is a powerful technology used to define the spatial distribution and relative abundance of metabolites across tissue cryosections. While software packages exist for pixel-by-pixel individual metabolite and limited target pairs of ratio imaging, the research community lacks an easy computing and application tool that images any metabolite abundance ratio pairs. Importantly, recognition of correlated metabolite pairs may contribute to the discovery of unanticipated molecules in shared metabolic pathways. Here, we describe the development and implementation of an untargeted R package workflow for pixel-by-pixel ratio imaging of all metabolites detected in an MSI experiment. Considering untargeted MSI studies of murine brain and embryogenesis, we demonstrate that ratio imaging minimizes systematic data variation introduced by sample handling, markedly enhances spatial image contrast, and reveals previously unrecognized metabotype-distinct tissue regions. Furthermore, ratio imaging facilitates identification of novel regional biomarkers and provides anatomical information regarding spatial distribution of metabolite-linked biochemical pathways. The algorithm described herein is applicable to any MSI dataset containing spatial information for metabolites, peptides or proteins, offering a potent hypothesis generation tool to enhance knowledge obtained from current spatial metabolite profiling technologies.

Keywords:  adipose; brain; computational biology; embryo; mouse; systems biology

DOI:  https://doi.org/10.7554/eLife.96892
Proteomics. 2025 Mar 18. e202400238

Monitoring Functional Posttranslational Modifications Using a Data-Driven Proteome Informatic Pipeline.

Payman Nickchi, Uladzislau Vadadokhau, Mehdi Mirzaie, Marc Baumann, Amir A Saei, Mohieddin Jafari.

Posttranslational modifications (PTMs) are of significant interest in molecular biomedicine due to their crucial role in signal transduction across various cellular and organismal processes. Characterizing PTMs, distinguishing between functional and inert modifications, quantifying their occupancies, and understanding PTM crosstalk are challenging tasks in any biosystem. Studying each PTM often requires a specific, labor-intensive experimental design. Here, we present a PTM-centric proteome informatic pipeline for predicting relevant PTMs in mass spectrometry-based proteomics data without prior information. Once predicted, these in silico identified PTMs can be incorporated into a refined database search and compared to measured data. As a practical application, we demonstrate how this pipeline can be used to study glycoproteomics in oral squamous cell carcinoma based on the proteome profile of primary tumors. Subsequently, we experimentally identified cellular proteins that are differentially expressed in cells treated with multikinase inhibitors dasatinib and staurosporine using mass spectrometry-based proteomics. Computational enrichment analysis was then employed to determine the potential PTMs of differentially expressed proteins induced by both drugs. Finally, we conducted an additional round of database search with the predicted PTMs. Our pipeline successfully analyzed the enriched PTMs, and detected proteins not identified in the initial search. Our findings support the effectiveness of PTM-centric searching of MS data in proteomics based on computational enrichment analysis, and we propose integrating this approach into future proteomics search engines.

DOI: https://doi.org/10.1002/pmic.202400238
J Proteome Res. 2025 Mar 16.

MGVB: a New Proteomics Toolset for Fast and Efficient Data Analysis.

Metodi V Metodiev.

  MGVB is a collection of tools for proteomics data analysis. It covers data processing from in silico digestion of protein sequences to comprehensive identification of post-translational modifications and solving the protein inference problem. The toolset is developed with efficiency in mind. It enables analysis at a fraction of the resources cost typically required by existing commercial and free tools. MGVB, as it is a native application, is faster than existing proteomics tools such as MaxQuant and, at the same time, finds very similar, in some cases even larger, numbers of peptides at a chosen level of statistical significance. It implements a probabilistic scoring function to match spectra to sequences, a novel combinatorial search strategy for finding post-translational modifications, and a Bayesian approach to locate modification sites. This report describes the algorithms behind the tools, presents benchmarking data sets analysis comparing MGVB performance to MaxQuant/Andromeda, and provides step by step instructions for using it in typical analytical scenarios.

Keywords:  MS/MS search engine; mass spectrometry; post-translational modifications; shotgun analysis; software for computational proteomics

DOI:  https://doi.org/10.1021/acs.jproteome.4c00994
Nat Commun. 2025 Mar 16. 16(1): 2579

High resolution profiling of cell cycle-dependent protein and phosphorylation abundance changes in non-transformed cells.

Camilla Rega, Ifigenia Tsitsa, Theodoros I Roumeliotis, Izabella Krystkowiak, Maria Portillo, Lu Yu, Julia Vorhauser, Jonathon Pines, Jörg Mansfeld, Jyoti Choudhary, Norman E Davey.

The cell cycle governs a precise series of molecular events, regulated by coordinated changes in protein and phosphorylation abundance, that culminates in the generation of two daughter cells. Here, we present a proteomic and phosphoproteomic analysis of the human cell cycle in hTERT-RPE-1 cells using deep quantitative mass spectrometry by isobaric labelling. By analysing non-transformed cells and improving the temporal resolution and coverage of key cell cycle regulators, we present a dataset of cell cycle-dependent protein and phosphorylation site oscillation that offers a foundational reference for investigating cell cycle regulation. These data reveal regulatory intricacies including proteins and phosphorylation sites exhibiting cell cycle-dependent oscillation, and proteins targeted for degradation during mitotic exit. Integrated with complementary resources, our data link cycle-dependent abundance dynamics to functional changes and are accessible through the Cell Cycle database (CCdb), an interactive web-based resource for the cell cycle community.

DOI: https://doi.org/10.1038/s41467-025-57537-8
Mol Cell Proteomics. 2025 Mar 17. pii: S1535-9476(25)00048-9. [Epub ahead of print] 100950

A Single-Step Protein Extraction for Lung Extracellular Matrix Proteomics Enabled by the Photocleavable Surfactant Azo and timsTOF Pro.

Anna G Towler, Andrew J Perciaccante, Timothy J Aballo, Yanlong Zhu, Fei Wang, Sarah Lloyd, Kuniko Kadoya, Yupeng He, Yu Tian, Ying Ge.

The extracellular matrix (ECM) is a dynamic, complex network of proteins, collectively known as the 'matrisome', which not only provides essential structural support to cells and tissues but also regulates critical cellular processes. Dysregulation of the ECM is implicated in many diseases, underscoring the need to characterize the matrisome to better understand disease mechanisms. We have previously developed a dual-step protocol enabled by the photocleavable surfactant Azo for the extraction of ECM proteins from tissue using pH-neutral decellularization followed by solubilization by Azo. While effective for characterization of the ECM proteins, such a dual-step protocol requires two extracts per sample, limiting the throughput and complicating the comparison of protein quantitation across different extraction conditions. Here, we develop a single-step Azo-enabled protein extraction for the solubilization of ECM proteins from lung tissue to improve the throughput for studies with large sample sizes. Using this method, we identified 324 ECM proteins, including 137 core ECM and 187 ECM associated proteins. Core ECM proteins including elastin, fibronectin, and fibrillar collagens were reproducibly identified and quantified. We observed a 94.6% overlap in the ECM proteins identified between the single-step and dual-step Azo extracts, indicating the single-step Azo extraction achieves ECM protein coverage comparable to the dual-step extraction. Overall, we have demonstrated that this single-step Azo extraction is not only highly efficient but also comprehensive for ECM protein identification and quantification, making it a powerful method for ECM proteomics, especially for studies with large sample size.

DOI: https://doi.org/10.1016/j.mcpro.2025.100950
Nat Metab. 2025 Mar 17.

The time is now: accounting for time-of-day effects to improve reproducibility and translation of metabolism research.

Shaunak Deota, Julie S Pendergast, Ullas Kolthur-Seetharam, Karyn A Esser, Frédéric Gachon, Gad Asher, Charna Dibner, Salvador Aznar Benitah, Carolina Escobar, Deborah M Muoio, Eric Erquan Zhang, Gökhan S Hotamışlıgil, Joseph Bass, Joseph S Takahashi, Joshua D Rabinowitz, Katja A Lamia, Rafael de Cabo, Shingo Kajimura, Valter D Longo, Ying Xu, Mitchell A Lazar, Eric Verdin, Juleen R Zierath, Johan Auwerx, Daniel J Drucker, Satchidananda Panda.

The constant expansion of the field of metabolic research has led to more nuanced and sophisticated understanding of the complex mechanisms that underlie metabolic functions and diseases. Collaborations with scientists of various fields such as neuroscience, immunology and drug discovery have further enhanced the ability to probe the role of metabolism in physiological processes. However, many behaviours, endocrine and biochemical processes, and the expression of genes, proteins and metabolites have daily ~24-h biological rhythms and thus peak only at specific times of the day. This daily variation can lead to incorrect interpretations, lack of reproducibility across laboratories and challenges in translating preclinical studies to humans. In this Review, we discuss the biological, environmental and experimental factors affecting circadian rhythms in rodents, which can in turn alter their metabolic pathways and the outcomes of experiments. We recommend that these variables be duly considered and suggest best practices for designing, analysing and reporting metabolic experiments in a circadian context.

DOI: https://doi.org/10.1038/s42255-025-01237-6
Mol Cell Proteomics. 2025 Mar 13. pii: S1535-9476(25)00042-8. [Epub ahead of print] 100944

Metabolic Reprogramming into a Glycolysis Phenotype Induced by Extracellular Vesicles Derived from Prostate Cancer Cells.

Yoon-Jin Lee, Chul Won Seo, Shinwon Chae, Chang Yeol Lee, Sang Soo Kim, Yoon-Hee Shin, Hyun-Mee Park, Yong Song Gho, Seongho Ryu, Sang-Han Lee, Dongsic Choi.

Most cancer cells adopt a less efficient metabolic process of aerobic glycolysis with high level of glucose uptake followed by lactic acid production, known as the Warburg effect. This phenotypic transition enables cancer cells to achieve increased cellular survival and proliferation in a harsh low-oxygen tumor microenvironment. Also, the resulting acidic microenvironment causes inactivation of the immune system such as T-cell impairment that favors escape by immune surveillance. While lots of studies have revealed that tumor-derived EVs can deliver parental materials to adjacent cells and contribute to oncogenic reprogramming, their functionality in energy metabolism is not well addressed. In this study, we established prostate cancer cells PC3-AcT resistant to cellular death in an acidic culture medium driven by lactic acid. Quantitative proteomics between EVs derived from PC-3 and PC-3AcT cells identified 935 confident EV proteins. According to cellular adaptation to lactic acidosis, we revealed 159 regulated EV proteins related to energy metabolism, cellular shape, and extracellular matrix. These EVs contained a high abundance of glycolytic enzymes. In particular, PC-3AcT EVs were enriched with apolipoproteins including apolipoprotein B100 (APOB). APOB on PC-3AcT EVs could facilitate their endocytic uptake depending on low density lipoprotein receptor of recipient PC-3 cells, encouraging increases of cellular proliferation and survival in acidic culture media via increased activity and expression of hexokinases and phosphofructokinase. The activation of recipient PC-3 cells can increase glucose consumption and ATP generation, representing an acquired metabolic reprogramming into the Warburg phenotype. Our study first revealed that EVs derived from prostate cancer cells could contribute to energy metabolic reprogramming and that the acquired metabolic phenotypic transition of recipient cells could favor cellular survival in tumor microenvironment.

DOI: https://doi.org/10.1016/j.mcpro.2025.100944
Clin Chim Acta. 2025 Mar 17. pii: S0009-8981(25)00125-1. [Epub ahead of print] 120246

Plasma-based untargeted metabolomics reveals potential biomarkers for screening and distinguishing of ovarian tumors.

Shen Peng, Yiming Zhu, Jing Zhu, Zhongjian Chen, Yi Tao.

  Ovarian cancer (OC), a leading cause of gynecological cancer mortality, is frequently detected at advanced stages due to asymptomatic early progression. This study investigates plasma-based untargeted metabolomics for identifying biomarkers to screen and differentiate ovarian tumors (OT). Plasma samples from OC, benign ovarian tumors (BOT), and healthy controls (HC) were analyzed. Samples were randomized into train and test sets, with differential metabolites screened via two-tailed Student's t-test and partial least squares discriminant analysis. ROC models evaluated discriminatory capacity. Key metabolites demonstrated high predictive value: TMAO and hippuric acid distinguished OT from HC (AUC > 0.95), while linoleic acid, alpha-linolenic acid, and arachidonic acid (AUC > 0.9) further supported OT screening. Kynurenine differentiated OC from BOT (AUC = 0.808). Reduced levels of specific lysophosphatidylcholines (LPC (17:0/0:0), LPC (15:0/0:0)) also distinguished OT from HC (AUC = 0.771-0.89). These findings suggest plasma metabolomics holds promise for noninvasive biomarker discovery in OT screening and distinguishing between malignant and benign cases, though further validation of metabolite quantification is warranted prior to clinical application.

Keywords:  Kynurenine; Metabolomics; Ovarian tumor; Plasma, Metabolomics

DOI:  https://doi.org/10.1016/j.cca.2025.120246
bioRxiv. 2025 Mar 07. pii: 2025.03.03.641255. [Epub ahead of print]

Lactate dehydrogenase A-coupled NAD+ regeneration is critical for acute myeloid leukemia cell survival.

Ayşegül Erdem, Séléna Kaye, Francesco Caligiore, Manuel Johanns, Fleur Leguay, Jan Jacob Schuringa, Keisuke Ito, Guido Bommer, Nick van Gastel.

Enhanced glycolysis plays a pivotal role in fueling the aberrant proliferation, survival and therapy resistance of acute myeloid leukemia (AML) cells. Here, we aimed to elucidate the extent of glycolysis dependence in AML by focusing on the role of lactate dehydrogenase A (LDHA), a key glycolytic enzyme converting pyruvate to lactate coupled with the recycling of NAD+. We compared the glycolytic activity of primary AML patient samples to protein levels of metabolic enzymes involved in central carbon metabolism including glycolysis, glutaminolysis and the tricarboxylic acid cycle. To evaluate the therapeutic potential of targeting glycolysis in AML, we treated AML primary patient samples and cell lines with pharmacological inhibitors of LDHA and monitored cell viability. Glycolytic activity and mitochondrial oxygen consumption were analyzed in AML patient samples and cell lines post-LDHA inhibition. Perturbations in global metabolite levels and redox balance upon LDHA inhibition in AML cells were determined by mass spectrometry, and ROS levels were measured by flow cytometry. Among metabolic enzymes, we found that LDHA protein levels had the strongest positive correlation with glycolysis in AML patient cells. Blocking LDHA activity resulted in a strong growth inhibition and cell death induction in AML cell lines and primary patient samples, while healthy hematopoietic stem and progenitor cells remained unaffected. Investigation of the underlying mechanisms showed that LDHA inhibition reduces glycolytic activity, lowers levels of glycolytic intermediates, decreases the cellular NAD+ pool, boosts OXPHOS activity and increases ROS levels. This increase in ROS levels was however not linked to the observed AML cell death. Instead, we found that LDHA is essential to maintain a correct NAD+/NADH ratio in AML cells. Continuous intracellular NAD+ supplementation via overexpression of water-forming NADH oxidase from Lactobacillus brevis in AML cells effectively increased viable cell counts and prevented cell death upon LDHA inhibition. Collectively, our results demonstrate that AML cells critically depend on LDHA to maintain an adequate NAD+/NADH balance in support of their abnormal glycolytic activity and biosynthetic demands, which cannot be compensated for by other cellular NAD+ recycling systems. These findings also highlight LDHA inhibition as a promising metabolic strategy to eradicate leukemic cells.

DOI: https://doi.org/10.1101/2025.03.03.641255
Nat Commun. 2025 Mar 17. 16(1): 2623

Genetically encoding ε-N-methacryllysine into proteins in live cells.

Tian-Yi Zhu, Shi-Yi Chen, Mengdi Zhang, Heyu Li, Ting Wu, Emmanuel Ajiboye, Jia Wen Wang, Bi-Kun Jin, Dan-Dan Liu, Xintong Zhou, He Huang, Xiaobo Wan, Ke Sun, Peilong Lu, Yaxin Fu, Ying Yuan, Hai Song, Anna A Sablina, Chao Tong, Long Zhang, Ming Wu, Haifan Wu, Bing Yang.

Lysine acylation is a ubiquitous post-translational modification (PTM) that plays pivotal roles in various cellular processes, such as transcription, metabolism, protein localization and folding. Thousands of lysine acylation sites have been identified based on advances in antibody enrichment strategies, highly sensitive analysis by mass spectrometry (MS), and bioinformatics. However, only 27 lysine methacrylation (Kmea) sites have been identified exclusively in histone proteins. It is hard to separate, purify and differentiate the Kmea modification from its structural isomer lysine crotonylation (Kcr) using general biochemical approaches. Here, we identify Kmea sites on a non-histone protein, Cyclophillin A (CypA). To investigate the functions of Kmea in CypA, we develop a general genetic code expansion approach to incorporate a non-canonical amino acid (ncAA) ε-N-Methacryllysine (MeaK) into target proteins and identify interacting proteins of methacrylated CypA using affinity-purification MS. We find that Kmea at CypA site 125 regulates cellular redox homeostasis, and HDAC1 is the regulator of Kmea on CypA. Moreover, we discover that genetically encode Kmea can be further methylated to ε-N-methyl-ε-N-methacrylation (Kmemea) in live cells.

DOI: https://doi.org/10.1038/s41467-025-57969-2
Mol Cell Proteomics. 2025 Mar 13. pii: S1535-9476(25)00046-5. [Epub ahead of print] 100948

Addressing NHS chemistry: Efficient quenching of excess TMT reagent and reversing TMT-over labelling in proteomic samples by methylamine.

Yana Demyanenko, Xintong Sui, Andrew M Giltrap, Benjamin G Davis, Bernhard Küster, Shabaz Mohammed.

N-Hydroxysuccinimide (NHS) ester chemistry is used extensively across proteomics sample preparation. One of its increasingly prevalent applications is in isobaric reagent-based quantitation such as iTRAQ (isobaric tags for relative and absolute quantitation) and TMT (tandem mass tag) approaches. In these methods, labelling on the primary amines of lysine residues and N-termini of tryptic peptides via amide formation (N-derivatives) from corresponding NHS ester reagents is the intended reactive outcome. However, the role of NHS esters as activated carboxyls can also drive the formation of serine-, tyrosine-, and threonine- derived esters (O-derivatives). These O-derivative peptides are typically classed as over-labelled and are disregarded for quantitation, leading to loss of information and hence potential sensitivity. Their presence also unnecessarily increases sample complexity, which reduces the overall identification rates. One common approach for removing these unwanted labelling events has involved treatment with hydroxylamine. We show here that this approach is not efficient and can still leave substantial levels of unwanted over-labelled peptides. Through systematic study of nucleophilic aminolysis reagents and reaction conditions, we have now developed a robust method to efficiently remove over-labelled peptides. The new method reduces the proportion of over-labelled peptides in the sample to less than 1% without affecting the labelling rate or introducing other modifications, leading to superior identification rates and quantitation precision.

DOI: https://doi.org/10.1016/j.mcpro.2025.100948
Clin Chem Lab Med. 2025 Mar 24.

Targeted proteomics of serum IGF-I, -II, IGFBP-2, -3, -4, -5, -6 and ALS.

Jakob Albrethsen, Lylia Drici, Lea Marie Slot Vilmann, Stine A Holmboe, Charlotte Ehlers Thomsen, Veronica Lykke Rogaczewska Groendahl, Maud Eline Ottenheijm, Annelaura Bach Nielsen, Christina Christoffersen, Lise Aksglaede, Casper P Hagen, Nicolai J Wewer Albrechtsen, Anders Juul.

   OBJECTIVES: The insulin-like growth factors (IGFs) regulate growth in humans. IGF-I and IGF binding protein (IGFBP)-3 are biomarkers in children with growth disorders. We investigate a targeted proteomics method for absolute quantitation of eight IGF protein family members in human serum, including the peptide hormones IGF-I and -II, and the six binding proteins IGFBP-2, -3, -4, -5, -6 and acid labile subunit (ALS).
METHODS: Serum preparation was optimized for targeted proteomics of IGF related proteins on a clinical LC-MS/MS platform (UHPLC coupled with Triple-Q MS). We created quality controls, standards and internal standards and 289 serum samples from healthy children and adolescents were measured in ten batches over two months. The method was compared to WHO reference standards, clinical and research immunoassays, and relative proteomics profiling.
RESULTS: The sensitivity and reproducibility were sufficient for most but not all IGF protein family members. Targeted proteomics correlated well with clinical immunoassays for IGF-I (R2=0.88) and for IGFBP-3 (R2=0.46), (p<0.001). The correlation between targeted proteomics and non-clinical immunoassays for IGF-II, IGFBP-2, -4, -5, -6 and ALS varied between proteins.
CONCLUSIONS: We present a method for parallel quantification of IGF-I, IGFBP-3, 5 and ALS for clinical verification studies, whereas targeted proteomics of the five remaining IGF related proteins (IGF-II, IGFBP-2, -4, and -6) require further examination. The sensitivity of our new IGF-I method suggests a possible diagnostic role for targeted proteomics of IGF-I in the management of children with extremely low levels of circulating IGF-I.

Keywords:  LC-MS/MS; insulin-like growth factor (IGF); proteomics; serum

DOI:  https://doi.org/10.1515/cclm-2024-1428
Bioinform Adv. 2025 ;5(1): vbaf047

LipidSigR: a R-based solution for integrated lipidomics data analysis and visualization.

Chia-Hsin Liu, Pei-Chun Shen, Meng-Hsin Tsai, Hsiu-Cheng Liu, Wen-Jen Lin, Yo-Liang Lai, Yu-De Wang, Mien-Chie Hung, Wei-Chung Cheng.

Motivation: Lipidomics is a rapidly expanding field focused on studying lipid species and classes within biological systems. As the field evolves, there is an increasing demand for user-friendly, open-source software tools capable of handling large and complex datasets while keeping pace with technological advancements. LipidSig, a widely used web-based platform, has been instrumental in data analysis and visualization of lipidomics. However, its limitations become evident when users want to build customized workflows. To address the limitation, we developed a companion R package, LipidSigR, based on the R code of the LipidSig web platform.
Results: LipidSigR offers greater flexibility, allowing researchers with basic R programming skills to modify and adapt workflows according to their needs. It has been rigorously tested following CRAN guidelines to ensure compatibility and reproducibility. In demonstrating its functionality, we analyze the case with commonly used experimental design, case versus control, in lipidomics studies. Researchers can follow the use case to explore the key capabilities and build customized lipidomics data analysis workflows using LipidSigR.
Availability and implementation: LipidSigR is freely available from https://lipidsig.bioinfomics.org/lipidsigr/index.html and https://github.com/BioinfOMICS/LipidSigR.

DOI: https://doi.org/10.1093/bioadv/vbaf047
JCI Insight. 2025 Mar 18. pii: e180943. [Epub ahead of print]

Circulating metabolite signatures indicate differential gut-liver crosstalk in lean and obese MASLD.

Mathias Haag, Stefan Winter, Aurino M Kemas, Julia Tevini, Alexandra Feldman, Sebastian K Eder, Thomas K Felder, Christian Datz, Bernhard Paulweber, Gerhard Liebisch, Oliver Burk, Volker M Lauschke, Elmar Aigner, Matthias Schwab.

   BACKGROUND: Alterations in circulating metabolites have been described in obese metabolic dysfunction-associated steatotic liver disease (MASLD), but data on lean MASLD are lacking. We investigated serum metabolites, including microbial bile acids (BAs) and short-chain fatty acids (SCFAs), and their association with lean and obese MASLD.
METHODS: Serum samples from 204 subjects of European descent were allocated to four groups: lean healthy (n=61), lean MASLD (n=49), obese healthy (n=47) and obese MASLD (n=47). LC/MS-based metabolomics was performed followed by linear model analysis. MASLD prediction was assessed based on LASSO regression. Functional effects of significantly altered molecules were confirmed in organotypic 3D primary human liver cultures.
RESULTS: Lean MASLD was characterized by elevated isobutyrate, along with higher methionine sulfoxide, propionate and phosphatidylcholines. Patients with obese MASLD had increased sarcosine and decreased lysine and asymmetric dimethylarginine. Using metabolites, sex and body mass index, MASLD vs. healthy could be predicted with a median AUC of 86.5% and 85.6% in the lean and obese subgroups, respectively. Functional experiments in organotypic 3D primary human liver cultures showed that propionate and isobutyrate induced lipid accumulation and altered expression of genes involved in lipid and glucose metabolism.
CONCLUSION: Our results indicate that lean MASLD is characterized by a distinct metabolite pattern related to amino acid metabolism, lipids and SCFAs, while metabolic pathways of lipid accumulation are differentially activated by microbial metabolites. Our findings highlight an important role of microbial metabolites in MASLD pathogenesis, with implications for the predictive and mechanistic assessment of liver disease across different weight categories.
FUNDING: The work received funding from the Robert Bosch Stiftung, Stuttgart, Germany, the Swedish Research Council [grant numbers 2021-02801, 2023-03015 and 2024-03401], the ERC Consolidator Grant 3DMASH [101170408], Ruth and Richard Julin Foundation for Gastroenterology [grant number 2021-00158], the SciLifeLab and Wallenberg National Program for Data-Driven Life Science [WASPDDLS22:006], and the Novo Nordisk Foundation [NNF23OC0085944 and NNF23OC0084420]. JT was supported by PMU-FFF [grant number E-18/28/148-FEL].

Keywords:  Hepatology; Metabolism; Obesity

DOI:  https://doi.org/10.1172/jci.insight.180943