bims-cesirm 2026-03-01 papers

bims-cesirm

Biomed News

on Cell Signaling mediated regulation of metabolism

Issue of 2026–03–01
34 papers selected by
Tigist Tamir, University of North Carolina

Moonlighting Functions of Mammalian Peroxiredoxins in Cellular Signaling.
Targeting de novo pyrimidine synthesis confers vulnerability to copper-mediated ATR inactivation in PARP inhibitor-resistant ovarian cancer.
Acylcarnitines in Cancer Metabolism: Mechanistic Insights and Stratification Potential.
Metabolic Vulnerabilities as a Therapeutic Target in Breast Cancer.
Loss of HuD Sensitizes Neuroblastoma Cells to Palmitate-Driven Stress-Induced Premature Senescence via PPARα Downregulation and FAO Impairment.
Discoidin Domain Receptor 1 Translocation to the Mitochondria Promotes Oxidative Stress and Apoptosis in Acute Kidney Injury.
Receptor-Mitochondria Crosstalk in the Kynurenine Metabolic Pathway: Integrating Metabolomics and Clinical Mass Spectrometry.
Insights into the Function of a Conserved Cys120 in Human Neuroglobin in Oxidative Stress Regulation of Breast Cancer Cells.
Antitumor Activity of the ACC Inhibitor Firsocostat in Breast Cancer Cell Lines: A Proof-of-Concept In Vitro Study.
A Large-Scale Method to Measure the Stoichiometries of Protein Poly-ADP-Ribosylation.
Cooperative Architecture of Mitochondrial Proteome Homeostasis.
Integrative Analysis of 4-Hydroxynonenal-Modified Proteins and Plasma Metabolome in Breast Cancer Patients.
Lipoprotein Metabolism in Hematological Malignancies: A Role in Shaping the Tumor Cell Microenvironment?
Aspartate treatment ameliorates metabolic dysfunction-associated fatty liver disease through upregulation of CBS and suppression of ferroptosis.
Ferroptosis Suppressor Protein 1 (FSP1)-CoQ10-NADPH-Axis Is Responsible for Erastin Resistance in MCF-7 Breast Cancer Cells.
Pho-Tip: One-Pot Dephosphorylation for Rapid and Sensitive Analysis of DIA Phosphoproteomics Data.
Thioredoxin 1 Suppresses TXNIP-Driven Control of Glucose Metabolism in Human Cells.
A Scalable Design for Proximity-Inducing Molecules.
Alpha-ketoglutarate dehydrogenase: more than just a TCA cycle enzyme.
Vitamin B3 derivatives support pancreatic cancer cell survival and chemotherapy resistance.
High-throughput chemical proteomics workflow for profiling protein citrullination dynamics.
Proteomic landscape of crotonylation and acetylation in pig testes across male reproduction.
ARID1A Deficiency in Diffuse-Type Gastric Cancer Promotes a Pyrimidine Metabolic Vulnerability.
Evaluation and application of chemical decrosslinking in the context of histopathological spatial proteomics.
Decoding the substrate specificity landscape of a promiscuous enzyme through multi-substrate mutational scanning.
Targeting PFKFB3 to enhance CDK4/6 inhibitor response in ER+ breast cancer.
Best practices for cysteine analysis.
Tumor cell villages define the co-dependency of tumor and microenvironment in liver cancer.
A Computational Model of Tumor Interactions with Bone-Resident Cells Predicts Tumor-Type-Specific Responses to Perturbations.
Aromatase Inhibitor Therapy Is Associated with Distinct Plasma Lipidomic Profiles in Postmenopausal Breast Cancer Patients.
Virtual Cells Need Context, Not Just Scale.
A Gly-to-Ala Substitution Confers Choline Phosphotransferase Activity on the CDP-Ethanolamine-Specific Mammalian EPT1.
High-fat diet and a high amyloid load interact to induce PKC-α dependent synaptic insulin resistance.
MiProChip: A Scalable Microfluidic Platform for Multiplexed Single-Cell Proteomics via Isobaric Labeling.

Antioxidants (Basel). 2026 Feb 10. pii: 231. [Epub ahead of print]15(2):

Moonlighting Functions of Mammalian Peroxiredoxins in Cellular Signaling.

Yosup Kim, Eun-Kyung Kim, Ho Hee Jang.

  Peroxiredoxins (Prdxs) are a family of thiol-specific peroxidases that play a central role in maintaining intracellular redox homeostasis. In addition to their classical antioxidant activities, Prdxs function as peroxide sensors, modulators of redox signaling, and molecular chaperones. In this review, we summarize the peroxide-reducing activity, their redox-switch mechanism driven by reversible hyperoxidation, and the chaperone function that arises through oligomerization and accompanying structural changes. We also highlight that the Prdx1-Prdx6 isoforms exhibit distinct subcellular localizations and perform isoform-specific functions, thereby contributing to a wide range of physiological and pathological processes. Furthermore, we compile recent findings showing that diverse post-translational modifications (PTMs), including phosphorylation, acetylation, ubiquitination, glutathionylation, sumoylation, and S-nitrosylation, not only regulate Prdx activity but also contribute to cellular signaling processes. Overall, this review emphasizes that Prdxs are more than simple antioxidant enzymes: they serve as guardians of cellular redox balance and dynamic regulators of signaling networks, underscoring their potential as disease biomarkers and therapeutic targets.

Keywords:  antioxidant enzymes; chaperone; peroxidase activity; peroxiredoxin; post-translational modifications; protein interaction; reactive oxygen species

DOI:  https://doi.org/10.3390/antiox15020231
Nat Commun. 2026 Feb 25.

Targeting de novo pyrimidine synthesis confers vulnerability to copper-mediated ATR inactivation in PARP inhibitor-resistant ovarian cancer.

Yabing Nan, Kunyu Wang, Menghan Hu, Qingyu Luo, Xiaowei Wu, Xiao Yu, Xuantong Zhou, Li Chen, Bin Li, Zhumei Cui, Zhihua Liu.

Although poly(ADP-ribose) polymerase (PARP) inhibitors (PARPi) as monotherapy or in combination with other DNA-damaging agents exhibit promising clinical efficacy, the therapeutic responses are usually transient, with subsequent development of acquired resistance posing a significant challenge. Here, through a small-molecule compound screening, we identify elesclomol, a potent copper ionophore, which sensitizes BRCA-proficient ovarian cancer cells to PARPi by inhibiting activation of the ATR-CHK1 pathway. Mechanistically, we demonstrate that copper directly binds to ATRIP, a critical cofactor of ATR activation, disrupting the ATR-ATRIP interaction, further impairing ATR-mediated DNA damage repair signaling and potentiating PARPi sensitivity. Importantly, we reveal a secondary metabolic vulnerability in PARPi-resistant ovarian cancer associated with de novo pyrimidine synthesis, suggesting that targeting this pathway as an effective strategy to eradicate drug-adaptive residual tumors and resistant patient-derived xenograft models following ATR and PARP co-inhibition. These findings propose de novo pyrimidine synthesis as an adaptive metabolic vulnerability that can be therapeutically targeted to overcome PARPi resistance in BRCA-proficient ovarian cancer.

DOI: https://doi.org/10.1038/s41467-026-70001-5
Cancers (Basel). 2026 Feb 23. pii: 713. [Epub ahead of print]18(4):

Acylcarnitines in Cancer Metabolism: Mechanistic Insights and Stratification Potential.

Hwa Pyoung Lee, Jieun Oh, Nury Lee, Yujin Jung, Jisu Yum, Minsu Kim, Maro Yoo, Jae Gwang Park, Jae Youl Cho.

  Metabolic reprogramming constitutes a fundamental hallmark of malignancy, enabling cancer cells to sustain proliferation and survival under physiological stress. While aerobic glycolysis is well characterized, fatty acid oxidation (FAO) has emerged as a decisive driver of oncogenic progression and therapeutic resistance. Acylcarnitines (ACs), obligatory intermediates for the mitochondrial transport of long-chain fatty acids, have transcended their traditional categorization as passive metabolic byproducts to function as potent signaling entities and functional readouts of mitochondrial oxidative throughput. This review delineates the AC metabolic axis in oncology, examining the coordinated biochemical machinery, including the carnitine palmitoyltransferase (CPT) system, carnitine-acylcarnitine translocase (CACT; SLC25A20), and organic cation/carnitine transporter 2 (OCTN2), that governs cellular AC homeostasis. We further evaluate the clinical utility of altered AC profiles as non-invasive biomarkers for early diagnosis and risk stratification across diverse malignancies, highlighting their capacity to reflect metabolic bottlenecks and flux dynamics. Additionally, we scrutinize therapeutic strategies targeting the AC-FAO axis, demonstrating how the inhibition of key transporters and enzymes sensitizes tumors to conventional chemotherapy and immunotherapy. Ultimately, deciphering the systemic and spatial dynamics of ACs remains essential for advancing precision metabolic oncology and developing personalized therapeutic strategies based on metabolic profiling.

Keywords:  acylcarnitine; cancer metabolism; fatty acid oxidation; metabolic reprogramming; non-invasive biomarkers; therapeutic targets

DOI:  https://doi.org/10.3390/cancers18040713
Curr Oncol. 2026 Feb 23. pii: 129. [Epub ahead of print]33(2):

Metabolic Vulnerabilities as a Therapeutic Target in Breast Cancer.

Sabrina Guo, Christina L Addison.

  Metabolic reprogramming is a defining feature of breast cancer, enabling tumor cells to sustain rapid proliferation, survive under stress, and resist therapy. Key pathways including glycolysis, glutaminolysis, lipid metabolism, and one-carbon metabolism, play central roles in meeting the energetic and biosynthetic demands of malignant cells. Enhanced glycolytic flux supports ATP generation and lactate production, while glutamine metabolism fuels the tricarboxylic acid cycle and provides nitrogen for nucleotide synthesis. Lipid metabolic pathways, particularly fatty acid synthesis, contribute to membrane biogenesis and signaling, and one-carbon metabolism driven by serine and glycine supplies methyl groups for epigenetic regulation and nucleotide production. These metabolic adaptations not only promote tumor growth but also create vulnerabilities that can be exploited therapeutically. Inhibiting these pathways has shown promise in preclinical models; however, challenges such as metabolic plasticity, tumor heterogeneity, and potential toxicity in normal tissues underscore the need for biomarker-driven strategies and rational combination therapies. Herein, we describe current knowledge of the role of these pathways in breast cancer progression, highlighting the role of key enzymes in promoting breast cancer tumor cell growth and in breast cancer prognoses.

Keywords:  TCA cycle; breast cancer; fatty acids; glutaminolysis; glycolysis; metabolism; pentose phosphate pathway; prognosis; serine biosynthesis; tumor growth

DOI:  https://doi.org/10.3390/curroncol33020129
Cells. 2026 Feb 07. pii: 316. [Epub ahead of print]15(4):

Loss of HuD Sensitizes Neuroblastoma Cells to Palmitate-Driven Stress-Induced Premature Senescence via PPARα Downregulation and FAO Impairment.

Seungyeon Ryu, Jiyoon Seo, Ye Eun Sim, Se Hoon Jung, Wei Zhang, Seung Min Jeong, Eun Kyung Lee.

  Metabolic stress caused by lipid overload is a key driver of cellular dysfunction in aging and disease. Excess saturated fatty acids such as palmitate impair fatty acid oxidation (FAO), promote lipid accumulation, and increase reactive oxygen species (ROS), ultimately triggering premature senescence-like states. Senescence further amplifies vulnerability by worsening mitochondrial dysfunction, enhancing lipid imbalance, and sustaining pro-inflammatory signaling. Here, we investigated the role of the neuron-enriched RNA-binding protein HuD (ELAVL4) in protecting cells against lipotoxic stress. Using Neuro2a neuroblastoma cells, we found that HuD knockdown suppressed FAO, leading to increased lipid accumulation and elevated ROS following palmitate exposure. HuD-deficient cells also exhibited cytosolic mitochondrial DNA release, IRF phosphorylation, and upregulation of senescence markers. Mechanistically, RNA immunoprecipitation revealed that HuD binds directly to PPARα mRNA, sustaining its expression by competing with the PPARα-targeting microRNAs miR-9-5p and miR-22-3p. Loss of HuD reduced PPARα levels, thereby weakening the FAO capacity and sensitizing cells to palmitate-induced lipotoxic stress. These findings identify a previously unrecognized HuD-PPARα-FAO axis that restrains metabolic stress and senescence. By linking post-transcriptional regulation to lipid metabolism and inflammatory signaling, this work highlights stress-induced premature senescence as both an outcome and a propagator of metabolic dysfunction, providing insight into mechanisms of aging-related vulnerability.

Keywords:  HuD; PPARα; fatty acid oxidation; mitochondria; senescence

DOI:  https://doi.org/10.3390/cells15040316
J Am Soc Nephrol. 2026 Feb 24.

Discoidin Domain Receptor 1 Translocation to the Mitochondria Promotes Oxidative Stress and Apoptosis in Acute Kidney Injury.

Gema Bolas, Corina M Borza, Fabian Bock, Xinyu Dong, Oscar Hanson, Manuel Chiusa, Shirong Cao, Ming-Tsun Tsai, Agnes B Fogo, Ming-Zhi Zhang, Raymond C Harris, Craig R Brooks, Roy Zent, Ambra Pozzi.

BACKGROUND: Mitochondrial damage with overproduction of mitochondrial reactive oxygen species (mtROS) and apoptosis is a hallmark of acute kidney injury (AKI). Discoidin Domain Receptor 1 (DDR1) is a collagen receptor tyrosine kinase that contributes to AKI. Mass spectrometry analysis of DDR1 interacting proteins identified several mitochondrial proteins, suggesting that DDR1 associated with mitochondria. Thus, we analyzed whether DDR1 translocated to mitochondria and promoted mitochondrial dysfunction following AKI.
METHODS: We analyzed DDR1 localization in kidneys of patients with AKI and mice following ischemia reperfusion-induced AKI. To determine whether mitochondrial DDR1 (mitDDR1) regulated mitochondrial functions, we generated kidney cells expressing wild-type or a kinase dead DDR1. Then, we investigated the location of wild-type or mutated DDR1 upon collagen stimulation; the steps involved in DDR1 mitochondrial translocation; and the contribution of mitDDR1 in regulating mtROS production and apoptosis.
RESULTS: mitDDR1 was detected in injured human and mice kidneys and collagen-activated DDR1 translocated to the mitochondria where it increased mtROS production and tubule cell apoptosis. Collagen-activated DDR1 translocated to the outer membrane of mitochondria through its association with the chaperone mtHsp60 and induced oxidative stress and apoptosis by promoting tyrosine phosphorylation of p66Shc, a regulator of the cellular redox state and apoptosis. Moreover, cells expressing a kinase dead DDR1, treated with a DDR1 inhibitor, or expressing p66Shc mutated in the DDR1-targeted phosphorylation sites had reduced mtROS and apoptosis.
CONCLUSIONS: We describe a novel non-canonical pathway whereby activated DDR1 translocates to the mitochondria to promote oxidative stress and cell apoptosis.

DOI: https://doi.org/10.1681/ASN.0000001045
Antioxidants (Basel). 2026 Feb 19. pii: 261. [Epub ahead of print]15(2):

Receptor-Mitochondria Crosstalk in the Kynurenine Metabolic Pathway: Integrating Metabolomics and Clinical Mass Spectrometry.

László Juhász, Zsolt Galla, Masaru Tanaka, László Vécsei.

  Mitochondria govern energy transfer, redox balance, and cell fate. Tryptophan catabolism generates kynurenines (KYNs) that can tune mitochondrial function, with growing evidence that G protein-coupled receptor 35 (GPR35), aryl hydrocarbon receptor (AhR), and N-methyl-D-aspartate receptors (NMDA receptors) link extracellular cues to adenosine 5 prime triphosphate (ATP) maintenance, calcium (Ca2+) handling, mitophagy, and inflammasome control. In parallel, quinolinic acid (QA)-driven de novo nicotinamide adenine dinucleotide (NAD+) synthesis connects KYN flux to tricarboxylic acid (TCA) cycle activity and sirtuin programs across tissues. Key gaps remain: receptor pharmacology is rarely integrated with NAD+ economics and respiration, and clinical workflows still lack single-run assays that quantify both kynurenine and TCA nodes. We therefore integrate receptor proximal signaling, QA-driven NAD+ supply, and unified liquid chromatography-mass spectrometry (LC-MS) measurement into one translational framework spanning kynurenic acid (KYNA), KYN, 3-hydroxykynurenine (3-HK), and QA, using mitochondrial endpoints as the common readout. We synthesize evidence for mitochondrial GPR35 signaling that preserves ATP, AhR programs that tune oxidative defenses and mitophagy, and NMDA receptor antagonism that limits excitotoxic stress. These mechanisms are linked to QA-dependent NAD+ biogenesis and alpha ketoglutarate control points, then aligned with chromatography and ionization choices suited to routine LC-MS workflows. This receptor to organelle framework couples KYN flux to respiratory control and provides a practical roadmap for standardized single-run LC-MS panels. It can strengthen target validation in ischemia, neurodegeneration, psychiatry, and oncology while improving biomarker qualification through harmonized analytics and decision-grade readouts.

Keywords:  G protein-coupled receptors; N-methyl-D-aspartate (NMDA); aryl hydrocarbon receptor (AhR); kynurenic acid (KYNA); liquid chromatography–mass spectrometry (LC-MS); metabolomics; mitochondria; mitophagy; nicotinamide adenine dinucleotide (NAD+); receptors; tricarboxylic acid (TCA) cycle

DOI:  https://doi.org/10.3390/antiox15020261
Biomolecules. 2026 Jan 31. pii: 215. [Epub ahead of print]16(2):

Insights into the Function of a Conserved Cys120 in Human Neuroglobin in Oxidative Stress Regulation of Breast Cancer Cells.

Shu-Qin Gao, Wen Shi, Si-Qi Xia, Zi-Lei He, Ying-Wu Lin.

  Human neuroglobin (Ngb) is a globin featuring a disulfide bond (Cys46-Cys55) and a redox-active cysteine residue (Cys120) and plays a dual role in cellular stress responses. In this study, we investigated how wild-type (WT) Ngb and its two mutants, C120S Ngb, in which Cys120 is replaced by serine, and A15C Ngb, which contains an engineered Cys15-Cys120 disulfide bridge, modulate oxidative stress in triple-negative breast cancer (MDAMB231) and hormone receptor-positive breast cancer (MCF-7) cells. In both cell lines, WT Ngb enhanced cell survival under H2O2-induced oxidative stress by scavenging reactive oxygen species (ROS) through oxidation of Cys120. In contrast, the C120S and A15C mutants lost this protective capacity and instead promoted apoptosis. Mass spectrometry analysis confirmed the oxidation of Cys120 to sulfenic acid in WT Ngb, whereas both mutants exhibited impaired redox activity, leading to elevated ROS levels, lipid peroxidation, and activation of caspase-9/3. AO/EB staining further revealed that WT Ngb attenuated DNA damage, while the mutants exacerbated apoptosis in both MDAMB231 and MCF-7 cells. These results demonstrate that Cys120 acts as a critical redox switch, dictating whether Ngb exerts cytoprotective or pro-apoptotic effects across different breast cancer cell types. Our findings suggest that WT Ngb may help protect normal tissues during cancer therapy, whereas engineered Ngb mutants could be used to selectively sensitize both triple-negative and hormone receptor-positive breast cancer cells to oxidative damage, offering a novel redox-targeted therapeutic strategy.

Keywords:  apoptosis; breast cancer cells; neuroglobin; oxidative stress; redox regulation

DOI:  https://doi.org/10.3390/biom16020215
Pharmaceuticals (Basel). 2026 Jan 24. pii: 201. [Epub ahead of print]19(2):

Antitumor Activity of the ACC Inhibitor Firsocostat in Breast Cancer Cell Lines: A Proof-of-Concept In Vitro Study.

Simona Picerno, Eugenia Giglio, Martina Giuseffi, Marcello Radino, Marzia Sichetti, Marisabel Mecca.

  Background/Objectives: Breast cancer is the most frequently diagnosed malignancy among women and is characterized by marked heterogeneity in treatment response. Metabolic reprogramming, particularly enhanced de novo lipogenesis, represents a hallmark of cancer progression and a promising therapeutic target. Firsocostat, a selective allosteric inhibitor of acetyl-CoA carboxylase (ACC), has previously been investigated in metabolic diseases but has never been evaluated in breast cancer models. This study aimed to assess the antitumor effects of firsocostat on breast cancer cell lines. Methods: We investigated the cytotoxic and metabolic effects of firsocostat in four breast cancer cell lines-MCF7 (luminal A HR+), SK-BR-3 (HER2-positive), MDA-MB-231 (triple-negative), and HCC1937 (triple-negative, BRCA1-mutated)-together with the non-tumorigenic MCF-10A line. Dose- and time-dependent responses were evaluated using phase-contrast microscopy for morphological evaluation, Trypan Blue exclusion assays, and MTS-based viability assays. Results: Firsocostat significantly reduced cell viability across all breast cancer subtypes in a concentration- and time-dependent manner, with IC50 values ranging from 80 to 93 µM. In contrast, non-tumorigenic MCF-10A cells were less affected, indicating a selective cytotoxic effect toward malignant cells. Conclusions: Firsocostat exerts robust cytotoxic effects in breast cancer models, identifying it as a promising metabolism-targeting therapeutic candidate capable of selectively impairing breast cancer cell survival by disrupting fatty acid biosynthesis. These results indicate that firsocostat could represent a viable candidate as a metabolic-based therapeutic approach for breast cancer. Given its established clinical safety profile in metabolic diseases, firsocostat warrants further preclinical investigation and supports further mechanistic and preclinical evaluation.

Keywords:  ACC inhibition; breast cancer; drug repurposing; firsocostat; lipid metabolism; metabolic reprogramming; tumor metabolism

DOI:  https://doi.org/10.3390/ph19020201
ACS Chem Biol. 2026 Feb 26.

A Large-Scale Method to Measure the Stoichiometries of Protein Poly-ADP-Ribosylation.

Peng Li, Yajie Zhang, Chiho Kim, Yonghao Yu.

Poly-ADP-ribosylation (PARylation) is a reversible post-translational modification that occurs in higher eukaryotes. While thousands of PARylated substrates have been identified, the specific biological functions of most PARylated proteins remain elusive. PARylation stoichiometry is a critical parameter to assess the potential functions of a PARylated protein. Here, we developed a large-scale strategy to measure the stoichiometries of protein PARylation. By integrating chemically mild cell lysis conditions, boronate enrichment, and carefully designed titration experiments, we were able to determine the PARylation stoichiometries for a total of 235 proteins. Importantly, this approach enables the capture of all PARylation events, regardless of their amino acid acceptor linkages. We revealed that PARylation occupancy spans over 3 orders of magnitude. However, most PARylation events occur at low stoichiometric values (median 0.58%). Notably, we observed that high-stoichiometry PARylation (>1%) predominantly targets proteins involved in transcription regulation and chromatin remodeling. Thus, our study provides a system-scale, quantitative view of PARylation stoichiometries under genotoxic conditions, which serves as an invaluable resource for future functional studies of this important protein post-translational modification.

DOI: https://doi.org/10.1021/acschembio.5c00817
medRxiv. 2026 Feb 09. pii: 2026.02.06.26345691. [Epub ahead of print]

Cooperative Architecture of Mitochondrial Proteome Homeostasis.

Patrick Forny, Merima Forny, Andrew J Smith, Andrew Y Sung, Kaixian Liu, David J Pagliarini.

Mitochondria are semi-autonomous organelles whose generation and maintenance demand precise expression, processing, and assembly of >1,000 proteins encoded across two genomes. To explore this cooperativity, we performed multiomic analyses on >200 cell lines harboring mitochondrial gene perturbations, generating >26M molecular measurements. Our data reveal that mitochondrial proteome homeostasis is heavily influenced by post-transcriptional processes. Through nearest neighbor analyses, we reveal diverse protein activities undergirding this regulation, including MDH2's regulation of MT-ND3 transcription via FASTKD1 binding and CLPP's processing of the mitoribosomal assembly factor MALSU1, which we establish as a disease gene. Through entropy analysis, we reveal unexpectedly heterogeneous protein-level variability across complexes and use complexome profiling to identify new complex-specific membership, including C15orf61's association with complex V. We further observe substantial mtDNA copy number variation, notably upon disruption of the disease-related cobalamin biosynthesis protein MMADHC. Together, we establish new protein functions and provide a multilayered view into mitochondrial proteome regulation.
Highlights: Multiomic signatures across perturbations reveal extensive post-transcriptional regulationThe TCA cycle enzyme MDH2 binds FASTKD1 to modulate MT-ND3 transcript levelsMALSU1 is a CLPP protease substrate whose deficiency causes a mitochondrial diseaseC15orf61 binds ATP synthase and negatively regulates its higher order assemblyMMADHC inversely affects mtDNA levels potentially mediated through LONP1.

DOI: https://doi.org/10.64898/2026.02.06.26345691
Antioxidants (Basel). 2026 Feb 21. pii: 265. [Epub ahead of print]15(2):

Integrative Analysis of 4-Hydroxynonenal-Modified Proteins and Plasma Metabolome in Breast Cancer Patients.

Morana Jaganjac, Matea Nikolac Perkovic, Tea Horvat, David Rojo, Marija Krizic, Natalija Dedic Plavetic, Damir Vrbanec, Biserka Orehovec, Kamelija Zarkovic, Neven Zarkovic.

  Breast cancer is a highly heterogeneous malignancy, characterized by diverse genetic, epigenetic, and phenotypic variations, as well as by metabolic reprogramming and oxidative stress. Lipid peroxidation bioactive product 4-hydroxynonenal (4-HNE) plays a significant role in the development and progression of cancer. In this study, we quantified circulating 4-HNE-modified proteins and performed comprehensive untargeted metabolomic profiling of the patients' plasma using LC-ESI-QTOF-MS and GC-EI-QMS, aiming to investigate systemic metabolic pathways associated with oxidative damage in breast cancer. Significantly elevated levels of 4-HNE-modified proteins were detected in breast cancer patients compared to healthy controls, accompanied by distinct metabolomic signatures enriched in lipid metabolism. Several metabolites, including specific long-chain fatty acids, exhibited significant correlations with circulating 4-HNE-modified proteins, suggesting an interaction between lipid peroxidation-driven protein modification and breast cancer-associated metabolic reprogramming. Overall, this study provides evidence of associations between systemic 4-HNE-mediated protein modification and altered metabolic profiles in breast cancer, highlighting oxidative stress-related metabolites as potential biomarkers and pointing to redox-metabolic crosstalk in breast cancer patients.

Keywords:  4-hydroynonenal; breast cancer; lipid peroxidation; metabolome remodeling

DOI:  https://doi.org/10.3390/antiox15020265
Metabolites. 2026 Feb 20. pii: 145. [Epub ahead of print]16(2):

Lipoprotein Metabolism in Hematological Malignancies: A Role in Shaping the Tumor Cell Microenvironment?

Manal Sellam, Mélanie Lambert, Nadine Varin-Blank, Kevin Saitoski.

  The tumor microenvironment (TME) plays a key role in driving tumor progression, metastasis, and resistance to therapy. The TME is a highly variable ecosystem composed of both cancer and surrounding normal cells, immune survey cells and the extracellular matrix, also composed of signaling molecules that mediate interactions between them. Blood cancer cells pose a unique challenge because of their circulation and widespread distribution along with their capacity to invade various niches, interacting with a wide range of host cells such as fibroblasts, immune cells, endothelial cells, and adipocytes. Metabolism reprogramming in this tumor context, notably referring to elevated cholesterol and fatty acid metabolism, emerges as a crucial event in shaping an immune-suppressive microenvironment that promotes tumor progression. Cholesterol and fatty acids are supplied by both de novo biosynthesis and exogenous uptake from lipoproteins. Lipoproteins are pseudo-micellar structures, designed to transport essential water-insoluble metabolites, including triacylglycerols and cholesterol, in the plasma, lymph, and interstitial fluids. A number of studies have reported abnormal circulating lipoprotein levels in leukemic patients and have suggested that lipoproteins are key for cancer cells to thrive. However, the role of lipoprotein metabolism in cancer cells in the context of the TME is still incompletely discussed so far. The aim of this review is to consider the importance of lipoprotein metabolism in shaping the tumor microenvironment in the context of hematological malignancies.

Keywords:  leukemia; lipoprotein metabolism; lymphoma; tumor progression

DOI:  https://doi.org/10.3390/metabo16020145
J Biol Chem. 2026 Feb 25. pii: S0021-9258(26)00188-2. [Epub ahead of print] 111318

Aspartate treatment ameliorates metabolic dysfunction-associated fatty liver disease through upregulation of CBS and suppression of ferroptosis.

Jie Zhang, Shihua Lin, Tiantian Zhang, Xing Son, Chaohui Yu.

  Dysregulation of non-essential amino acids (NEAAs) contributes to metabolic dysfunction-associated fatty liver disease (MAFLD) progression. Aspartate (ASP), an NEAA serving as a critical precursor for purine, asparagine, and arginine biosynthesis, exhibits unexplored pathophysiological significance in MAFLD. Mass spectrometry profiling identified hepatic ASP as the sole NEAA consistently depleted in both human MAFLD patients and animal models. Dietary or orally administered ASP supplementation in mice significantly improved insulin sensitivity, suppressed de novo lipogenesis, and enhanced mitochondrial respiration, collectively ameliorating diet-induced MAFLD. Integrative multi-omics analyses revealed that ASP activates the trans-sulfuration pathway through cystathionine β-synthase (CBS) upregulation in hepatocytes, thereby attenuating ferroptosis - evidenced by elevated glutathione and reduced lipid peroxidation. Crucially, pharmacological ferroptosis inhibition (liproxstatin-1) abolished ASP-mediated hepatic improvements, while CBS blockade completely abrogated ASP's therapeutic effects. These findings establish ASP as a novel ferroptosis inhibitor with therapeutic promise for MAFLD.

Keywords:  Aspartate; CBS; ferroptosis; hepatic steatosis

DOI:  https://doi.org/10.1016/j.jbc.2026.111318
Antioxidants (Basel). 2026 Feb 11. pii: 239. [Epub ahead of print]15(2):

Ferroptosis Suppressor Protein 1 (FSP1)-CoQ10-NADPH-Axis Is Responsible for Erastin Resistance in MCF-7 Breast Cancer Cells.

Brian B Silver, Carri Murphy, Erik J Tokar, Birandra K Sinha.

  Ferroptosis has emerged as a promising therapeutic strategy for drug-resistant cancers; however, the molecular mechanisms governing ferroptosis susceptibility in breast cancer remain poorly defined. Here, we have investigated distinct ferroptosis resistance mechanisms in parental MCF-7 breast cancer cells and their BCRP-overexpressing derivative, MCF-7/MXR. MCF-7/MXR cells displayed robust erastin (ER)-induced ferroptosis characterized by extensive lipid peroxidation, ROS accumulation, and suppression of the xCT-GSH-GPX4 axis. In contrast, MCF-7 cells were resistant to ER, exhibiting minimal lipid damage despite measurable ER-induced oxidative stress. We found that this resistance is mediated not by the canonical GPX4 pathway, but by a potent compensatory antioxidant system centered on the FSP1-CoQ10-NADPH axis. Pharmacological inhibition of FSP1 strongly sensitized MCF-7 cells to ER, with minimal effects in MXR cells. ER differentially regulated ferroptosis-associated genes, downregulating GPX4, and SLC7A11 in MXR cells but upregulating the GPX4 pathway in MCF-7 cells. Additionally, ER downregulated FSP1 and NQO1 in MCF-7 cells without affecting their expressions in MXR cells. This mechanistic divergence highlights that ferroptosis resistance in breast cancer is context-dependent and mediated by pathway-specific antioxidant programs. Co-targeting FSP1 and GPX4 therefore represents a rational strategy to overcome ferroptosis resistance in MCF-7-like breast cancers.

Keywords:  CoQ10; FSP1; NQO1; breast cancer; erastin; ferroptosis; resistance

DOI:  https://doi.org/10.3390/antiox15020239
Anal Chem. 2026 Feb 23.

Pho-Tip: One-Pot Dephosphorylation for Rapid and Sensitive Analysis of DIA Phosphoproteomics Data.

Katharina D Faisst, Kate Lau, Ludwig R Sinn, Lukasz Szyrwiel, Vadim Demichev.

Recent advances in instrumentation and data processing have transformed data-independent acquisition (DIA) proteomics into a reliable technology for quantitative profiling of post-translational modifications. However, analysis of DIA phosphoproteomics data is challenging due to the large search space, wherein all combinations of phosphosites on a peptide need to be considered. Current approaches therefore face significant hurdles in detecting low-abundant phosphorylated peptides, in particular when working with low sample amounts. Here we introduce Pho-Tip, a lossless one-pot dephosphorylation strategy. We show that Pho-Tip enables comprehensive mapping of phosphorylated peptide sequences, facilitating streamlined creation of experiment-focused in silico predicted spectral libraries and thus rapid and sensitive analysis of DIA phosphoproteomics experiments.

DOI: https://doi.org/10.1021/acs.analchem.5c07139
Antioxid Redox Signal. 2026 Feb 24. 15230864261421616

Thioredoxin 1 Suppresses TXNIP-Driven Control of Glucose Metabolism in Human Cells.

Shayida Maimaiti, Markus Dagnell, Lucia Coppo, Wenchao Zhao, Peter Geserick, Kai Kappert, Elias S J Arnér.

   AIMS: Cytosolic thioredoxin 1 (Trx1, TXN, TRX) is a central player in redox control. Thioredoxin interacting protein (TXNIP), an α-arrestin regulating glucose metabolism and inflammation, is widely regarded to inhibit TRX activity. However, the interactions between the two proteins across various cellular contexts remain poorly understood; in addition, only a limited number of studies have yet been conducted in human primary cells. We thus aimed here to investigate the functional relationship between TRX and TXNIP in human primary cells. We studied whether TXNIP inhibits TRX cellular activity in these primary cells and how this interaction influences cellular redox biology or glucose metabolism.
RESULTS: In primary cells, TXNIP deficiency did not increase cellular TRX activity. Instead, TXNIP deficiency elevated PGC-1α and PDK4 transcripts, increased PDHA1 Ser293 phosphorylation, and raised basal GLUT4, consistent with enhanced glucose uptake and restrained flux through the pyruvate dehydrogenase complex. Conversely, lowering TRX expression levels triggered higher TXNIP levels. This in turn correlated with suppressed transcripts for PGC-1α and PDK4, a lower extent of PDHA1 phosphorylation at Ser293, and decreased glucose uptake.
INNOVATION: Our findings suggest that TXNIP, against common belief, may not necessarily be an endogenous inhibitor of TRX but, rather, that TRX can be an inhibitor of TXNIP.
CONCLUSION: This study reveals that the key intracellular redox protein TRX inversely regulates TXNIP, suggesting that modulation of the TRX system may provide a previously unrecognized therapeutic avenue for modulation of glucose metabolism. Antioxid. Redox Signal. 00, 000-000.

Keywords:  glucose transporter; glucose uptake; peroxiredoxins; pyruvate dehydrogenase; thioredoxin; thioredoxin interacting protein

DOI:  https://doi.org/10.1177/15230864261421616
bioRxiv. 2026 Feb 21. pii: 2026.02.20.706349. [Epub ahead of print]

A Scalable Design for Proximity-Inducing Molecules.

Endri Karaj, Varsha Venkatarangan, Shaimaa H Sindi, Surached Siriwongsup, Chaiheon Lee, Rajaiah Pergu, Vedagopuram Sreekanth, Karishma Kailass, Kien Tran, Prashant Singh, Sameek Singh, Junya Kawai, Jeffrey E Fung, Mahilet Tefera, Rohil Dhaliwal, Santosh K Chaudhary, April Keyes, Ananthan Sadagopan, Lisa Boatner, Neel H Shah, Charlie Fehl, Keriann M Backus, Amit Choudhary.

Chimeric molecules, which bring together an effector enzyme and a protein-of-interest (POI) to add/remove post-translational modifications (PTMs), are furnishing transformative modalities (e.g., PROTACs). However, these chimeras' scalability is limited as they employ rare, non-inhibitory binders of effectors. We report GRoup-transfer chimeras for Inducing Proximity (GRIPs) that employ abundantly available effectors' inhibitors to append POI binder on the effector using group-transfer handles. To demonstrate scalability, we develop 6 GRIPs classes for 3 PTMs utilizing diverse inhibitor, spanning 16 effector‒POI pairs. Furthermore, we report a toolbox of 42 tunable group-transfer handles for Cys/Lys residues and ∼5000 inhibitor‒residue pairs for diverse effectors. Using global proteomics, we confirm the specificity for group transfer and PTM editing. GRIPs endowed new functionalities to POI drugs, including preventing rebound signaling upon drug withdrawal, a more potent/persistent inhibition, and inhibitor-induced pathway activation in 4 fully-endogenous systems. In diverse hemi -endogenous systems (tagged POI), GRIPs induced condensate formation with reduced off-targets, cleared pathogenic PTMs, and initiated PTM crosstalk. Overall, GRIPs provide a scalable and versatile platform for PTM editing.
Graphical abstract:

DOI: https://doi.org/10.64898/2026.02.20.706349
Free Radic Biol Med. 2026 Feb 19. pii: S0891-5849(26)00147-4. [Epub ahead of print]248 210-221

Alpha-ketoglutarate dehydrogenase: more than just a TCA cycle enzyme.

Ryan J Mailloux.

Alpha-ketoglutarate dehydrogenase (KGDH; EC 1.2.4.2) catalyzes the fourth step of the tricarboxylic acid (TCA) cycle and links carbohydrate, fatty acid and amino acid metabolism to the aerobic production of ATP. KGDH is classically viewed as indispensable to energy metabolism and strictly located to mitochondria. Therefore, it is generally thought that the loss of its activity has catastrophic consequences for mammalian cells. However, recent advances in molecular biology and redox biology tools coupled with the implementation of new genetically modified mouse lines and cultured cells knocked down for components of KGDH have revealed it is a multifunctional cellular enzyme that localizes to the mitochondria and nucleus where it uses superoxide (O2•-)/hydrogen peroxide (H2O2) and metabolites related to its catalysis (e.g., alpha-ketoglutarate (KG), succinyl-CoA, succinate) to control cell fate decisions. In addition, it has been revealed that over-stimulation of KGDH causes severe oxidative stress through the hyper-production of O2•-/H2O2 and disturbs cell signals and epigenome regulation, which has been linked to cancer cell transformation, metabolic diseases like metabolic dysfunction-associated steatotic liver disease (MASLD), and inflammation. Furthermore, inhibition of KGDH with competitive inhibitors, redox modifications, or shRNAs has shown that the targeted disruption of the enzyme can alleviate these diseases. The aim of this review is to update the literature on KGDH. It is not just a TCA cycle enzyme anymore.

DOI: https://doi.org/10.1016/j.freeradbiomed.2026.02.050
Cancer Lett. 2026 Feb 20. pii: S0304-3835(26)00097-2. [Epub ahead of print] 218334

Vitamin B3 derivatives support pancreatic cancer cell survival and chemotherapy resistance.

Faith Nakazzi, Mehrdad Zarei, Mariana Lopes, Hallie J Graor, William C Beegan, Eric Gu, Sakineh Rezaei, Peder J Lund, Jordan M Winter.

  Nicotinamide adenine dinucleotide (NAD+) is a central metabolic cofactor essential for cell survival and stress response in normal tissues. Its dietary precursors, commonly referred to as vitamin B3 derivatives, including nicotinamide (NAM), nicotinamide riboside (NR), and nicotinamide mononucleotide (NMN), are marketed as nutraceuticals with potential energy-boosting, cardioprotective, and neuroprotective benefits. Consequently, many cancer patients utilize NAD+ precursors to alleviate chemotherapy-induced toxicity and promote health. However, the impact of NAD+ supplements on intrinsic tumor biology and progression remains controversial and incompletely characterized. In this study, we assessed the impact of common vitamin B3 derivatives, NAM, NR, and NMN, on chemotherapy efficacy in pancreatic ductal adenocarcinoma (PDAC) using both in vitro and in vivo models. Among the compounds tested, NMN exhibited the strongest protective effect on cancer cells, enhancing resistance to oxaliplatin, 5-fluorouracil, and gemcitabine in vitro. Mechanistically, NAD+ precursors promoted mitochondrial function, reduced oxidative stress, and suppressed DNA damage and apoptosis in treated cancer cells, all contributing to chemotherapy resistance. In murine models, both immunocompetent and immunodeficient, supplementation with NAM and NMN similarly conferred resistance to standard chemotherapy and supported cancer growth. Our findings highlight a potentially concerning role for NAD+-boosting supplements in the context of an active cancer, especially when used in conjunction with chemotherapy. These data underscore the need for careful evaluation of nutraceutical use in cancer patients, particularly those with PDAC, as vitamin B3 derivatives may inadvertently promote tumor cell survival and compromise treatment efficacy.

Keywords:  DNA repair; NAD(+); NAD(+) precursors; Pancreatic cancer; chemotherapy resistance; mitochondria; oxidative stress; vitamin B3 derivatives

DOI:  https://doi.org/10.1016/j.canlet.2026.218334
Nat Commun. 2026 Feb 21. pii: 1982. [Epub ahead of print]17(1):

High-throughput chemical proteomics workflow for profiling protein citrullination dynamics.

Rebecca Meelker González, Sophia Laposchan, Erik Riedel, Anna Fürst, Naomi O'Sullivan, Wassim Gabriel, Mathias Wilhelm, Percy A Knolle, Guillaume Médard, Bernhard Kuster, Chien-Yun Lee.

Citrullination is a post-translational modification implicated in autoimmune and inflammatory diseases, yet its low abundance and lack of effective enrichment tools have limited proteome-wide analysis. Here, we develop a robust chemical proteomics workflow with improved specificity and throughput. This method builds upon glyoxal-based derivatization and incorporates a cleavable biotin linker for efficient peptide enrichment, release, and identification via mass spectrometry. Benchmarking demonstrates a > 10-fold increase in the detection of citrullinated peptides at sub-0.1% abundance. Applying this workflow to primary human neutrophils, we successfully monitor dynamic regulation, quantifying dose-dependent activation and inhibition by the PAD4 inhibitor GSK484. Furthermore, stimulation with the fungal pathogen Candida albicans reveals a "core citrullinome" conserved across distinct stimuli. Notably, extensive citrullination of linker histone H1 and structural proteins like lamin B1 suggests broad remodeling of cell architecture during NET formation. This workflow enables proteome-wide mapping of citrullination sites and facilitates its study across diverse biological contexts.

DOI: https://doi.org/10.1038/s41467-026-69490-1
J Proteomics. 2026 Feb 20. pii: S1874-3919(26)00031-X. [Epub ahead of print]327 105628

Proteomic landscape of crotonylation and acetylation in pig testes across male reproduction.

Lulu Wang, Hong Ao, Xinran Niu, Liwei Zhai, Jianfeng Liu, Chuduan Wang, Kai Xing.

  Lysine crotonylation (Kcr) and acetylation (Kac) are evolutionarily conserved post-translational modifications that alter protein's charge, polarity, and structure. To explore the potential roles and critical modified proteins in male reproduction, we employed the affinity enrichment of pan-anti-Kcr and pan-anti-Kac antibodies with high-resolution LC-MS/MS from testes tissues of six boars (three at 5 months and three at 24 months of age), respectively. As a result, a large number of Kcr and Kac proteins were identified in testicular tissue between two groups. Differential analysis further revealed that Kcr proteins were significantly enriched to Gene Ontology biological process terms concerning reproductive system development, response to steroid hormone and response to hydroxy isoflavone, whereas Kac proteins primarily regulated mitochondria-localized proteins involved in testicular metabolism. RPS5, RPS6, CYP17A1, CYP11A1, and CYP19A1, which may act as important regulators of testicular development through crotonylation modification. In addition, we observed indications of crosstalk between the two modifications, involving shared enzymes (including HDAC1-3), overlapping substrates, and CoA donor metabolism. This study performed a large-scale analysis of Kcr and Kac proteins in porcine testis, providing a post-translational modification perspective to testis development. SIGNIFICANCE: The testis is a key organ determining male reproductive capacity, and pigs of different ages exhibit distinct reproductive performance. Most previous studies have focused on transcriptomic and proteomic differences in testes across developmental stages, while post-translational modifications (PTMs) remain poorly explored. Here, using affinity enrichment with pan-anti-Kcr and pan-anti-Kac antibodies combined with high-resolution LC-MS/MS, we profiled lysine crotonylation (Kcr) and acetylation (Kac) in pig testes. Bioinformatic analyses identified CYP17A1, CYP19A1, CYP11A1, RPS5, and RPS6 as potential regulators of testicular maturation via crotonylation, while acetylation was found to regulate metabolic activity by modifying mitochondrial proteins. Furthermore, crosstalk between crotonylation and acetylation at the metabolic level was revealed. These findings advance our understanding of PTMs in porcine testis development and provide novel data resources for future studies.

Keywords:  Acetylation; Crotonylation; Pig; Post-translational modifications; Testes

DOI:  https://doi.org/10.1016/j.jprot.2026.105628
Mol Cancer Res. 2026 Feb 27.

ARID1A Deficiency in Diffuse-Type Gastric Cancer Promotes a Pyrimidine Metabolic Vulnerability.

Harumi Hirano, Hideki Makinoshima, Hideaki Ogiwara.

Loss-of-function mutations in ARID1A define an aggressive subtype of diffuse gastric cancer (DGC) that is often resistant to standard chemotherapy. Here, we uncover a precise metabolic vulnerability in ARID1A-deficient DGC driven by a specific transporter defect. Through integrated metabolomic and transcriptomic analyses, we demonstrate that ARID1A loss transcriptionally represses the high-affinity nucleoside transporter SLC28A3. Our profiling revealed a critical lack of redundancy in the concentrative transporter family (CNT) in DGC, establishing a strict reliance on SLC28A3 for maintaining intracellular deoxycytidine (dC) pools. Consequently, ARID1A deficiency creates a severe "low-dCTP" metabolic bottleneck. We show that the dC analog Gemcitabine exploits this state through a distinct functional dichotomy: it enters via intact equilibrative transporters (ENTs) to target cells that have lost their competitive dC barrier. Mechanistically, Gemcitabine exerts a "dual-hit" effect by outcompeting the scarce dC pool for DNA incorporation while simultaneously inhibiting ribonucleotide reductase, thereby blocking de novo nucleotide synthesis. This synergistic collapse of pyrimidine metabolism was validated in patient-derived ex vivo cultures and in vivo peritoneal dissemination models. Our findings provide a robust mechanistic basis for repurposing Gemcitabine as a precision therapy for ARID1A-deficient DGC, offering a potent strategy for this intractable malignancy. Implications: This work uncovers a metabolic vulnerability in ARID1A-deficient gastric cancer caused by SLC28A3 loss. It provides a compelling rationale for repurposing Gemcitabine as a targeted therapy for this intractable malignancy.

DOI: https://doi.org/10.1158/1541-7786.MCR-25-1069
bioRxiv. 2026 Feb 09. pii: 2026.02.06.704439. [Epub ahead of print]

Evaluation and application of chemical decrosslinking in the context of histopathological spatial proteomics.

Andikan J Nwosu, Liang Chen, Rashmi Kumar, Yumi Kwon, Shaun M Goodyear, Adel Kardosh, James M Fulcher, Ljiljana Paša-Tolić.

Laser capture microdissection (LCM) - based spatial mass spectrometry proteomics is a rapidly emerging technique with strong potential for use in formalin-fixed, paraffin-embedded (FFPE) tissues. Several sample-preparation methods have been developed to decrosslink FFPE proteins for spatial proteomics; however, residual crosslinks often remain, and depth can remain impaired relative to fresh frozen tissue samples. To increase proteome coverage in spatially resolved LCM-FFPE samples, we investigated a panel of chemical compounds with the potential to catalyze the decrosslinking of nucleophilic functional groups on proteins. Systematic screening and optimization of temperature, incubation time, and reagent concentration led to the identification of 3,4-diaminobenzoic acid as an effective agent for improving proteome coverage in FFPE pancreatic tissue. This compound could boost precursor identifications by more than 10% at both reduced (70 °C) and high (90 °C) temperatures. Application of this chemical-decrosslinking strategy to a pancreatic ductal adenocarcinoma tissue section enabled the identification of numerous cell-type-enriched proteins with clinical and therapeutic relevance. Taken together, our findings show that chemical decrosslinking can increase proteome coverage in FFPE tissues, thereby advancing our understanding of tissue microenvironments in physiological and pathological contexts.

DOI: https://doi.org/10.64898/2026.02.06.704439
Nat Commun. 2026 Feb 26.

Decoding the substrate specificity landscape of a promiscuous enzyme through multi-substrate mutational scanning.

Rosario Vanella, Sean Boult, Christoph Küng, Michael A Nash.

Substrate specificity is a defining feature of enzyme function, but its molecular underpinnings remain difficult to decode and engineer. Here, we leverage enzyme proximity sequencing (EP-Seq) to systematically map how single-point and combinatorial mutations reshape the substrate preferences of D-amino acid oxidase (DAOx) from Rhodotorula gracilis, a model promiscuous enzyme. We generate ~40,000 sequence-phenotype pairs, enabling us to profile the activities of ~6,500 unique DAOx variants against five D-amino acid substrates with distinct physicochemical properties. Our analysis reveals that substrate-specific mutations are distributed throughout the enzyme structure. Mutations near the active site drive strong specificity shifts but also incur catalytic penalties, while distal mutations subtly rewire intramolecular contacts in order to modulate specificity with minimal loss of activity. We identify and validate positional hotspots that act allosterically to influence specificity, and characterize key variants that acquire exclusive substrate specificity or exhibit up to 230-fold changes in substrate preference. Combining mutations with complementary effects further sharpens substrate discrimination, enabling rational design of highly selective biocatalysts. This work establishes a powerful framework for decoding enzyme specificity and provides foundational datasets to advance AI-guided enzyme engineering.

DOI: https://doi.org/10.1038/s41467-026-69913-z
Res Sq. 2026 Feb 10. pii: rs.3.rs-8412774. [Epub ahead of print]

Targeting PFKFB3 to enhance CDK4/6 inhibitor response in ER+ breast cancer.

Sucheta Telang, Brian F Clem, Ariamna A Herrera Miret, Leanne Price, Susan M Dougherty, Anna Schmitz, Xinmin Yin, Xipeng Ma, Xiang Zhang, Jason Chesney, Yoannis Imbert-Fernandez.

Background Cyclin-dependent kinase 4 and 6 (CDK4/6) inhibitors are widely used in the treatment of estrogen receptor-positive (ER⁺) breast cancer; however, the metabolic adaptations induced by CDK4/6 inhibition remain incompletely defined. In ER⁺ breast cancer, estrogen signaling plays a central role in coordinating cell cycle progression and metabolic programs that support tumor growth. Glycolytic flux is regulated at the level of phosphofructokinase-1 (PFK1) through the inducible enzyme 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase 3 (PFKFB3), which is transcriptionally regulated by estrogen receptor signaling and has been shown to promote glycolysis and proliferation in ER⁺ breast cancer cells. Yet, how CDK4/6 inhibition intersects with estrogen-regulated glycolytic control to rewire glucose utilization in ER⁺ breast cancer has not been explored. Methods Glucose metabolism was assessed using extracellular flux analysis, untargeted metabolomics, and stable isotope tracing with uniformly labeled 13 C-glucose in ER + breast cancer cell lines. In vivo metabolic tracing was performed following bolus administration of [U- 13 C]-glucose. The effects of pharmacologic PFKFB3 inhibition, alone and in combination with CDK4/6 inhibitors, were evaluated in vitro and in patient-derived xenograft (PDX) models. Statistical analyses were performed using appropriate tests with correction for multiple comparisons where applicable. Results CDK4/6 inhibition increased glycolytic flux, as evidenced by elevated basal and compensatory glycolysis, accumulation of early glycolytic intermediates, and increased 13 C labeling of fructose 1,6-bisphosphate. PFKFB3 silencing abrogated the CDK4/6 inhibitor-induced increase in glycolytic flux. Despite increased glycolysis, stable isotope tracing revealed markedly reduced incorporation of glucose-derived carbon into nucleotide biosynthesis and lipid-associated metabolites, consistent with reduced anabolic demand during G1 cell cycle arrest. In vivo glucose tracing demonstrated a dissociation between increased glycolytic flux and downstream biosynthetic utilization. Pharmacologic inhibition of PFKFB3 imposed additional constrains on glucose utilization and significantly enhanced the antitumor efficacy of CDK4/6 inhibition in PDX models. Conclusions CDK4/6 inhibition rewires glucose metabolism in ER + breast cancer by increasing glycolytic flux while limiting downstream glucose utilization, resulting in heightened reliance on regulated glycolytic control to maintain metabolic homeostasis during cell cycle arrest. Disruption of this adaptive metabolic state through PFKFB3 inhibition enhances the antitumor effects of CDK4/6 inhibition and supports the therapeutic potential of targeting glycolytic regulation in combination with CDK4/6 inhibitor-directed therapies.

DOI: https://doi.org/10.21203/rs.3.rs-8412774/v1
Ferroptosis Oxid Stress. 2026 ;pii: 202508. [Epub ahead of print]2

Best practices for cysteine analysis.

Feroza K Choudhury, Gina M DeNicola.

  Accurate measurement of cysteine and related thiol-containing metabolites is essential for understanding cellular redox regulation. However, the intrinsic reactivity and instability of cysteine present substantial analytical challenges. This review summarizes the biochemical context of cysteine and glutathione metabolism, emphasizing their dynamic redox equilibria and physiological relevance. We critically examine existing analytical approaches, including mass spectrometry-based, enzyme-coupled, and colorimetric methods, and discuss their respective strengths and limitations. Particular attention is given to sample preparation, derivatization strategies, and reagent selection, as these steps are crucial for preserving native thiol-disulfide status. Among various alkylating agents, N-ethylmaleimide is identified as the most reliable for thiol stabilization in liquid chromatography-mass spectrometry (LC-MS) workflows, while specific reagents such as monobromobimane or β-(4-hydroxyphenyl)ethyl iodoacetamide (HPE-IAM) are required for persulfide and polysulfide detection. The review also highlights the pitfalls of using indirect surrogates-such as glutathione or cystathionine levels-to infer cysteine availability, which can lead to significant misinterpretation of metabolic states. We conclude that direct LC-MS-based quantification of cysteine and glutathione, combined with careful derivatization and sample handling, remains the most reliable and accurate approach currently available for the assessment of thiol metabolism and redox homeostasis.

Keywords:  Cysteine; LC-MS; N-ethylmaleimide; derivatization; glutathione; redox homeostasis; thiol analysis

DOI:  https://doi.org/10.70401/fos.2025.0010
Nat Commun. 2026 Feb 21. pii: 1986. [Epub ahead of print]17(1):

Tumor cell villages define the co-dependency of tumor and microenvironment in liver cancer.

Meng Liu, Maria O Hernandez, Darko Castven, Hsin-Pei Lee, Wenqi Wu, Limin Wang, Marshonna Forgues, Jonathan M Hernandez, Jens U Marquardt, Lichun Ma.

Spatial cellular context is crucial in shaping intratumor heterogeneity. However, understanding how each tumor establishes its unique spatial landscape and what factors drive the landscape for tumor fitness remains significantly challenging. Here, we analyze over 2 million cells from 50 tumor biospecimens using spatial single-cell imaging and single-cell RNA sequencing. We develop a deep learning-based strategy to spatially map tumor cell states and their surrounding environmental architecture, and find that different tumor cell states can be organized into distinct clusters, or "villages," each supported by unique microenvironments. Notably, tumor cell villages exhibit village-specific molecular co-dependencies between tumor cells and their microenvironment and are associated with patient outcomes. Perturbation of molecular co-dependencies via random spatial shuffling of the microenvironment results in destabilization of the corresponding villages. We validate our findings using single-cell, spatial, and bulk transcriptome data from 740 liver cancer patients. This study provides insights into understanding tumor spatial landscape and its impact on tumor aggressiveness.

DOI: https://doi.org/10.1038/s41467-026-69797-z
bioRxiv. 2026 Feb 18. pii: 2026.02.16.706164. [Epub ahead of print]

A Computational Model of Tumor Interactions with Bone-Resident Cells Predicts Tumor-Type-Specific Responses to Perturbations.

Alexandra Gutierrez Vega, Natalie E Bennett, Erik P Beadle, Saja Alshafeay, Ramcharan Chitturi, Anishka Nagarimadugu, Hrishikesh Villur, Arsh Jaiswal, Julie A Rhoades, Leonard A Harris.

Tumor-induced bone disease (TIBD) arises from a complex interplay between metastatic cancer cells and the bone microenvironment, creating a self-reinforcing "vicious cycle" of bone destruction and tumor growth. Experimental evidence from our group (Buenrostro et al., Bone 113:77-88, 2018) suggests that tumor cells in the bone microenvironment early in disease rely more heavily on bone-derived growth factors, such as transforming growth factor-β (TGF-β), to sustain proliferation than tumor cells late in disease, which may grow independently of these factors. Here, we integrate a mechanistic, population-dynamics model of tumor-bone interactions with in vivo data to test the hypothesis that inhibiting bone resorption suppresses growth of non-adapted but not bone-adapted tumors. The model includes key regulators of TIBD, including TGF-β-driven tumor proliferation, parathyroid hormone-related protein (PTHrP) secretion, and osteoblast (OB)-osteoclast (OC) coupling. Parameter calibration using data from mice injected intratibially with parental (non-adapted) and bone-adapted breast cancer cells reveals distinct parameter values for each tumor type. Bone-adapted cells exhibit a higher basal division rate and reduced sensitivity to TGF-β-mediated stimulation, whereas parental-derived tumor cells depend more strongly on TGF-β and secrete PTHrP at higher rates to compensate for their slower growth. Model simulations reproduce the greater bone loss observed experimentally for bone-adapted tumors and predict that, for non-adapted tumors, bone destruction results from a slower but meaningful rise in OC activity and a possible moderate decline in OBs. Simulated treatment of bone-adapted tumors with the bisphosphonate zoledronic acid stabilizes bone density but has limited or highly variable effects on tumor growth. These results suggest that OC inhibition alone may be insufficient to restrain tumor expansion once tumors have adapted to the bone microenvironment. Together, these findings support the hypothesis that tumor adaptation to the bone microenvironment governs dependence on bone-derived growth factors and response to OC-targeted therapy, underscoring the value of mechanistic modeling for elucidating tumor-bone interactions and guiding tumor-type-specific treatment strategies for TIBD.

DOI: https://doi.org/10.64898/2026.02.16.706164
Int J Mol Sci. 2026 Feb 17. pii: 1926. [Epub ahead of print]27(4):

Aromatase Inhibitor Therapy Is Associated with Distinct Plasma Lipidomic Profiles in Postmenopausal Breast Cancer Patients.

Aleksandra Arsic, Ales Kvasnicka, David Friedecky, Nebojsa Ivanovic, Maja Milosevic, Vesna Vucic.

  Aromatase inhibitors (AIs) are the standard adjuvant endocrine therapy for postmenopausal women with hormone receptor-positive breast cancer; however, their effects on lipid metabolism remain incompletely characterized. In this study, we investigated AI-associated alterations in the plasma lipidome using mass spectrometry-based lipidomics. Plasma samples were collected from 30 patients prior to AI initiation and 29 patients receiving non-steroidal AI therapy for at least 24 months. Ultra-high-performance liquid chromatography-tandem mass spectrometry identified and relatively quantified 649 lipid species across 23 lipid classes and subclasses. Lipidomic analysis revealed significant differences in specific lipid species. Several phosphatidylcholine, sphingomyelin, and lysophosphatidylethanolamine species were significantly more abundant in patient plasma prior to AI therapy, whereas higher levels of selected ceramides, hexosylceramides, phosphatidylinositol (PI 16:0_16:0), and a polyunsaturated diacylglycerol species were observed in patients receiving AI therapy. Multivariate analyses revealed patient group separation, and a Naive Bayes classification model based on lipid-class levels achieved an area under the curve of 0.79. Additionally, lipid network and hierarchical clustering analyses identified systematic lipid-class trends. Protein-protein interaction network analysis based on lipidomic profiles highlighted enzymes associated with sphingolipid metabolism pathways. These findings demonstrate that long-term AI therapy is associated with specific alterations in the plasma lipidome, consistent with estrogen-deprivation-related metabolic differences. Targeted lipidomic profiling may provide mechanistic insights into therapy-associated metabolic effects and support future efforts to optimize long-term management of breast cancer survivors.

Keywords:  aromatase inhibitor therapy; breast cancer; lipidomics; sphingolipid metabolism

DOI:  https://doi.org/10.3390/ijms27041926
bioRxiv. 2026 Feb 09. pii: 2026.02.04.703804. [Epub ahead of print]

Virtual Cells Need Context, Not Just Scale.

Payam Dibaeinia, Sudarshan Babu, Mei Knudson, Ali ElSheikh, Yibo Wen, Han Liu, Jason Perera, Aly A Khan.

The intersection of AI and biology has entered a phase of explosive growth, driven by the ambition to build "Virtual Cells" or computational models capable of predicting cellular responses to any perturbation. Following the success of structural biology (e.g., AlphaFold) and large language models, the field has converged on training massive, high-capacity models on large-scale single-cell data. This position paper argues that scaling model capacity is insufficient to solve the Virtual Cell problem because the primary failure mode is a lack of adequate coverage over diverse biological contexts , not insufficient model expressivity. We support this claim by reviewing recent studies showing that simple baselines perform on par with sophisticated architectures within a given biological context, and current models fail to consistently generalize across contexts. We connect this finding to the causal inference literature on transportability and contrast it with domains where scaling has succeeded. We substantiate our argument through analysis of a state-of-the-art model on a 22-million-cell immunology dataset. We conclude that the community faces a causal transport problem that cannot be solved by accumulating more data from the same distributions. Instead, we argue that contextual diversity and causal representation learning deserve increased emphasis, complementing ongoing scaling of model capacity and data volume.

DOI: https://doi.org/10.64898/2026.02.04.703804
Genes Cells. 2026 Mar;31(2): e70095

A Gly-to-Ala Substitution Confers Choline Phosphotransferase Activity on the CDP-Ethanolamine-Specific Mammalian EPT1.

Yasuhiro Horibata, Shoji Maeda, Akimitsu Konishi.

  Choline phosphotransferase (CPT) and ethanolamine phosphotransferase (EPT) catalyze the synthesis of choline and ethanolamine phospholipids using CDP-choline or CDP-ethanolamine as substrates, respectively. Although both enzymes share the CDP-alcohol phosphotransferase motif at their active sites, the structural determinants of their substrate specificities remain unclear. Previously, we resolved the structure of the CDP-choline-specific yeast CPT1 and identified a Trp residue that interacts with the choline moiety of CDP-choline. The spatial positioning of this Trp residue was influenced by a neighboring Ala residue. Replacing this Ala with Gly conserved in EPT caused the Trp to shift inward, thereby conferring specificity for CDP-ethanolamine. Here we explored the reciprocal scenario in the CDP-ethanolamine-specific mammalian EPT1. A Gly105-to-Ala mutant of mouse EPT1 was generated, and structural prediction revealed an outward shift of Trp37 residue. Enzymatic assays and metabolic labeling in HEK293 cells demonstrated that the G105A mutant acquired CDP-choline specificity, which is absent in the wild-type enzyme. These results demonstrate that the positioning of Trp37, influenced by the adjacent Ala/Gly residue, is one of the important determinants of substrate specificity, supporting a conserved structural mechanism across yeast and mammalian systems.

Keywords:  CDP‐alcohol phosphotransferase; choline phosphotransferase; ethanolamine phosphotransferase; lipid metabolism; phosphatidylcholine; phosphatidylethanolamine; phospholipid biosynthesis; selenoprotein I; substrate specificity; the Kennedy pathway

DOI:  https://doi.org/10.1111/gtc.70095
Mol Cell Proteomics. 2026 Feb 23. pii: S1535-9476(26)00033-2. [Epub ahead of print] 101537

High-fat diet and a high amyloid load interact to induce PKC-α dependent synaptic insulin resistance.

Alexander Wenger, Tingting Li, Chi Nguyen, Ali Celik, Eleonora Cuboni, Alexander Dityatev, Anna Karpova, Michael R Kreutz, Robert Ahrends.

  A plethora of studies suggest that a high-fat diet in combination with a high amyloid load causes synaptic insulin resistance and is a risk factor for Alzheimer's disease. Our understanding of the underlying mechanisms is still fragmented. To gain new insights, we conducted integrated proteomic and phosphoproteomic profiling of hippocampal synaptosomes from wild-type and a transgenic mouse line with a high amyloid load (heterozygous TBA2.1 mice) that show no overt signs of neurodegeneration and dementia. Mice were fed with a regular or high-fat diet. Data-independent acquisition quantified over 5,400 proteins, revealing a stable synaptic proteome across conditions. However, the combination of high amyloid load and high-fat diet triggered coordinated remodeling of lipid metabolism pathways, particularly mitochondrial and peroxisomal fatty acid catabolism. Phosphoproteomic analysis showed pronounced activation of lipid- and stress-responsive kinases, including PKC-α, along with increased inhibitory phosphorylation of insulin receptor substrates (IRS1/2). In vitro experiments indicate that blocking PKC-α indeed prevents synaptic insulin resistance in primary neurons. The findings suggest that this proteomic workflow, combined with kinase pathway analysis, can reveal nodal points for interventions in a complex disease state with a trajectory to Alzheimer's disease.

Keywords:  Alzheimer’s disease; TMT; enrichment analysis; high-fat diet; network analysis; phosphoproteomics; synaptic insulin resistance; synaptosomes

DOI:  https://doi.org/10.1016/j.mcpro.2026.101537
Anal Chem. 2026 Feb 25.

MiProChip: A Scalable Microfluidic Platform for Multiplexed Single-Cell Proteomics via Isobaric Labeling.

Tsai-Fang Chou, Huan-Chi Chiu, Sofani Tafesse Gebreyesus, Guan-Fu Chen, Yi-Ju Chen, Abigail Ruth F Velasquez, Kuo-I Lin, Yu-Ju Chen, Hsiung-Lin Tu.

Single-cell proteomics (SCP) platforms are widely sought-after biomedical tools to complement existing omics technologies. Here, we present MiProChip, a microfluidic platform that integrates cell capture, lysis, protein digestion, tandem mass tag (TMT) labeling, on-chip pooling, and desalting a streamlined workflow for multiplexed SCP profiling. We optimized a chip-compatible TMT multiplexing protocol with a carrier-boosting strategy, enabling high-throughput operation and deep proteome coverage. MiProChip was designed to effectively reduce the mass spectrometry (MS) operation time, minimize adsorptive losses, enhance mixing, and stabilize flow for on-chip pooling, leading to a superior performance in recovery. Using PC9 and H1975 cells with a 100-cell carrier, a total of 3362 protein groups with 2775 ± 36 proteins were confidently identified across TMT-10-plex single-cell channels. Demonstration on murine colon adenocarcinoma cells identified 3199 proteins with 1669 ± 261 proteins per single cell to characterize galectin-8- and TGF-β-specific responses. Single-cell principal component analysis (PCA) showed separation of the control from treated groups, partial overlap between galectin-8 and TGF-β, and close clustering of TGF-β with the combination treatment, supporting a dominant TGF-β effect. Pathway enrichment analysis reveals their responsive pathway and distinguishes galectin-8- and TGF-β-specific responses, revealing downregulation of metastasis-related markers to support antimetastasis potential of galectin-8, which was not detected by bulk proteomic analysis. Collectively, MiProChip captured subtle proteomic heterogeneity and treatment-dependent single-cell responses, establishing a sensitive and robust platform for high-throughput SCP analysis.

DOI: https://doi.org/10.1021/acs.analchem.5c07275