Molecular & Cellular Proteomics最新文献

{"title":"ERAP1 activity modulates the immunopeptidome but also affects the proteome, metabolism and stress responses in cancer cells.","authors":"Martha Nikopaschou, Martina Samiotaki, Elli-Anna Stylianaki, Kamila Król, Paula Gragera, Aroosha Raja, Vassilis Aidinis, Angeliki Chroni, Doriana Fruci, George Panayotou, Efstratios Stratikos","doi":"10.1016/j.mcpro.2025.100964","DOIUrl":"https://doi.org/10.1016/j.mcpro.2025.100964","url":null,"abstract":"<p><p>Endoplasmic reticulum (ER) aminopeptidase 1 (ERAP1) metabolizes peptides inside the ER and shapes the peptide repertoire available for binding to Major Histocompatibility Complex Class I molecules (MHC-I). However, it may have additional effects on cellular homeostasis, which have not been explored. To address these questions, we used both genetic silencing of ERAP1 expression as well as treatment with a selective allosteric ERAP1 inhibitor to probe changes in the immunopeptidome and proteome of the A375 melanoma cancer cell line. We observed significant immunopeptidome shifts with both methods of functional ERAP1 disruption, which were distinct for each method. Both methods of inhibition led to an enhancement, albeit slight, in tumor cell killing by stimulated human PBMCs and in significant proteomic alterations in pathways related to metabolism and cellular stress. Similar proteomic changes were also observed in the leukemia cell line THP-1. Biochemical analyses suggested that ERAP1 inhibition affected sensitivity to ER stress, reactive oxygen species production and mitochondrial metabolism. Although the proteomics shifts were significant, their potential in shaping immunopeptidome shifts was limited since only 9.6% of differentially presented peptides belonged to proteins with altered expression and only 4.0% of proteins with altered expression were represented in the immunopeptidome shifts. Taken together, our findings suggest that modulation of ERAP1 activity can generate unique immunopeptidomes, mainly due to altered peptide processing in the ER, but also induce changes in the cellular proteome and metabolic state which may have further effects on tumor cells.</p>","PeriodicalId":18712,"journal":{"name":"Molecular & Cellular Proteomics","volume":" ","pages":"100964"},"PeriodicalIF":6.1,"publicationDate":"2025-04-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143795850","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"CellEKT: a robust chemical proteomics workflow to profile cellular target engagement of kinase inhibitors.","authors":"Joel Rüegger, Berend Gagestein, Antonius P A Janssen, Alexandra Valeanu, Alger Lazo Mori, Marielle van der Peet, Michael S Boutkan, Bogdan I Florea, Alex A Henneman, Remo Hochstrasser, Haiyan Wang, Paul Westwood, Andreas Topp, Patricia M Gomez Barila, Jan Paul Medema, Connie R Jimenez, Bigna Woersdoerfer, Stephan Kirchner, Jitao David Zhang, Uwe Grether, Arne C Rufer, Mario van der Stelt","doi":"10.1016/j.mcpro.2025.100961","DOIUrl":"https://doi.org/10.1016/j.mcpro.2025.100961","url":null,"abstract":"<p><p>The human genome encodes 518 protein kinases that are pivotal for drug discovery in various therapeutic areas such as cancer and autoimmune disorders. The majority of kinase inhibitors target the conserved ATP-binding pocket, making it difficult to develop selective inhibitors. To characterize and prioritize kinase-inhibiting drug candidates, efficient methods are desired to determine target engagement across the cellular kinome. In this study, we present CellEKT (Cellular Endogenous Kinase Targeting), an optimized and robust chemical proteomics platform for investigating cellular target engagement of endogenously expressed kinases using the sulfonyl fluoride-based probe XO44 and two new probes ALX005 and ALX011. The optimized workflow enabled the determination of the kinome interaction landscape of covalent and non-covalent drugs across over 300 kinases, expressed as half maximum inhibitory concentration (IC₅₀), which were validated using distinct platforms like phosphoproteomics and NanoBRET. With CellEKT, target engagement profiles were linked to their substrate space. CellEKT has the ability to decrypt drug actions and to guide the discovery and development of drugs.</p>","PeriodicalId":18712,"journal":{"name":"Molecular & Cellular Proteomics","volume":" ","pages":"100961"},"PeriodicalIF":6.1,"publicationDate":"2025-04-03","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143788541","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Cold Atmospheric Plasma Jet Promotes Wound Healing through CK2-coordinated PI3K/AKT and MAPK Signaling Pathways.","authors":"Pei-Shan Wu, Tzu-Hsuan Wong, Chun-Wei Hou, Teng-Ping Chu, Jyh-Wei Lee, Bih-Show Lou, Miao-Hsia Lin","doi":"10.1016/j.mcpro.2025.100962","DOIUrl":"https://doi.org/10.1016/j.mcpro.2025.100962","url":null,"abstract":"<p><p>The promising role of cold atmospheric plasma jet (CAPJ) treatment in promoting wound healing has been widely documented in therapeutic implications. However, the fact that not all subjects respond equally to CAPJ necessitates the investigation of the underlying cellular mechanisms, which have been rarely understood so far. Given that wound healing is a complex and prolonged process, post-plasma-activated medium (PAM) treated keratinocytes were collected at two time points, 2 hours (receiving) and 24 hours (recovery), for (phospho)proteomic analysis to systematically dissect the molecular basis of CAPJ-promoted wound healing. The receiving (phospho)proteomics datasets, referred to the time point of 2 hours, revealed an apparent increase in the phosphorylation of CK2 and its-mediated PI3K/AKT and MAPK signaling pathways, accompanied by a prompted downstream physiological response of cell migration. Additionally, incorporating the network analysis of predicted kinases and their direct interactors, we reiterated that CAPJ influenced cell growth and migration, thereby paving the way for its role in subsequent wound healing processes. Further determining the proteome profiles at recovery phase, which is the time point of 24 hours, displayed a totally different view from the receiving proteome which had almost no change. The up-regulation of ROBOs/SLITs expression and vesicle trafficking and fusion-related proteins, along with the abundant presence of 14-3-3 family proteins, indicated that the persistent effect of PAM on the wound healing process could potentially promote keratinocyte-fibroblast crosstalk and stimulate extracellular matrix (ECM) synthesis upon epithelialization. Consistent with proteome patterns, CAPJ-treated wound tissues indeed showed a denser and well-organized ECM architecture, implying hastened epithelialization during wound healing. Collectively, we delineated the molecular basis of CAPJ-accelerated wound healing at early and late responses, providing valuable insights for treatment selection and the development of therapeutic strategies to achieve better outcomes.</p>","PeriodicalId":18712,"journal":{"name":"Molecular & Cellular Proteomics","volume":" ","pages":"100962"},"PeriodicalIF":6.1,"publicationDate":"2025-04-03","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143788544","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"A probe-based target engagement assay for kinases in live cells.","authors":"Ursula M Glocker, Florian Braun, H Christian Eberl, Marcus Bantscheff","doi":"10.1016/j.mcpro.2025.100963","DOIUrl":"https://doi.org/10.1016/j.mcpro.2025.100963","url":null,"abstract":"<p><p>The efficacy and safety of kinase inhibitor drugs are largely influenced by their selectivity. Available profiling technologies are primarily based on over-expressed or endogenously expressed kinases in cell extracts. We compared kinase capture with the cell penetrant covalent probe XO44 to three derivatives and found that replacing the alkyne handle with a trans-cyclooctene group allowed the development of a more robust kinase capture and enrichment protocol. An intracellular chemoproteomics target profiling and engagement assay was devised by optimizing probe concentration and incubation time and using an isobaric mass tag-based strategy for relative quantification. Comparing intracellular kinase profiles of the marketed drug Dasatinib and the tool compound Dinaciclib with the lysate-based kinobeads assay revealed excellent agreement in rank-order of binding. Dinaciclib showed a systematic shift to higher IC₅₀s suggesting that intracellular co-substrate concentrations, cell penetration of the compound as well as kinase localization and complexes in live cells influence target profiles. Further, we show that sepiapterin reductase SPR and Multidrug Resistance Protein 1 ABCC1 are off-targets of kinase inhibitor scaffolds with potential implications on efficacy and safety.</p>","PeriodicalId":18712,"journal":{"name":"Molecular & Cellular Proteomics","volume":" ","pages":"100963"},"PeriodicalIF":6.1,"publicationDate":"2025-04-03","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143788433","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Multi-Omics Analysis Unveils Nsun5-Mediated Molecular Alterations in the Somatosensory Cortex and its Impact on Pain Sensation.","authors":"Peipei Chen, Heyu Zhao, Xia Gao, Junchao Xu, Zhili Huang, Huali Shen","doi":"10.1016/j.mcpro.2025.100960","DOIUrl":"https://doi.org/10.1016/j.mcpro.2025.100960","url":null,"abstract":"<p><p>Nsun5 assumes a pivotal role in the regulation of RNA methylation, and its deficiency has been linked to the advancement of hepatocellular carcinoma, gliomas, tetralogy of Fallot, cognitive deficits in Williams-Beuren syndrome (WBS), and brain development. This underscores Nsun5's significant involvement in the nervous system. In this study, we present evidence of Nsun5's influence on the structure of the primary somatosensory cortex. Through comprehensive multi-omics analyses, we unveil a spectrum of systematically altered genes and proteins, collectively engaged in the orchestration of translation, neurotransmitter metabolism, nerve conduction, synaptic transmission, and other functions. Notably, there are discernible changes in molecules associated with pain sensation, strongly indicating that Nsun5 deficiency undermines pain-related behavior. This study establishes a clear link between Nsun5 deficiency and transcriptional and proteomic changes, as well as neurotransmitter expression within the primary somatosensory cortex, and uncovers its novel role in impaired pain perception.</p>","PeriodicalId":18712,"journal":{"name":"Molecular & Cellular Proteomics","volume":" ","pages":"100960"},"PeriodicalIF":6.1,"publicationDate":"2025-04-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143780782","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Offset mass carrier proteome improves quantification of multiplexed single cell proteomics.","authors":"Tommy K Cheung, Ying Zhu, Christopher M Rose","doi":"10.1016/j.mcpro.2025.100959","DOIUrl":"https://doi.org/10.1016/j.mcpro.2025.100959","url":null,"abstract":"<p><p>Multiplexed single cell proteomics by mass spectrometry (scpMS) approaches currently offer the highest throughput as measured by cells analyzed per day. These methods employ isobaric labels and typically a carrier proteome - a sample added at 20-500x the single cell level that improves peptide sampling and identification. Peptides from the carrier and single cell proteomes exist within the same precursor isotopic cluster and are co-isolated for identification and quantification. This represents a challenge as high levels of carrier proteome limit the sampling of peptide ions from single cell samples and can potentially lead to decreased accuracy of quantitative measurements. Here, we address this limitation by introducing a triggered by offset mass acquisition method for scpMS (toma-scpMS) that utilizes a carrier proteome labeled with non-isobaric tags that have the same chemical composition but different mass as the labels used for quantitative multiplexing. Within toma-scpMS the carrier proteome and single cell proteome are separated at the precursor level, enabling separate isolation, fragmentation, and quantitation of the single cell samples. To enable this workflow we implemented a custom data acquisition scheme within inSeqAPI, an instrument application programming interface program that performed real-time identification of carrier proteome peptides and subsequent triggering of offset single cell quantification scans. We demonstrate that toma-scpMS is more robust to high-levels of carrier proteome and offers superior quantitative accuracy as compared to traditional multiplexed scpMS approaches when similar carrier proteome levels are employed.</p>","PeriodicalId":18712,"journal":{"name":"Molecular & Cellular Proteomics","volume":" ","pages":"100959"},"PeriodicalIF":6.1,"publicationDate":"2025-04-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143780704","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"An improved method for sampling and quantitative protein analytics of cerebrospinal fluid of individual mice.","authors":"Athanasios Lourbopoulos, Stephan A Müller, Georg Jocher, Manfred Wick, Nikolaus Plesnila, Stefan F Lichtenthaler","doi":"10.1016/j.mcpro.2025.100958","DOIUrl":"https://doi.org/10.1016/j.mcpro.2025.100958","url":null,"abstract":"<p><p>The mouse is the species most commonly used in preclinical research, but protein analytics of murine cerebrospinal fluid (CSF) remains challenging because of low sample volumes (often <10 μl) and frequent contaminations with blood. We developed an improved CSF sampling method that allows routine collection of larger volumes (20-30 μl) of pure CSF from individual mice, enabling multiple protein analytical assays from a single sample. Based on cell counts and hemoglobin ELISAs, we provide an easy quality control workflow for obtaining cell- and blood-free murine CSF. Through mass spectrometry-based proteomics using an absolutely quantified external standard, we estimated concentrations for hundreds of mouse CSF proteins. While repeated CSF sampling from the same mouse was possible, it induced CSF proteome changes. Applying the improved method, we found that the mouse CSF proteome remains largely stable over time in wild-type mice, but that amyloid pathology in the 5xFAD mouse model of Alzheimer's disease massively changes the CSF proteome. Neurofilament light chain and TREM2, markers of neurodegeneration and activated microglia, respectively, were strongly upregulated and validated using immunoassays. In conclusion, our refined murine CSF collection method overcomes previous limitations, allowing multiple quantitative protein analyses for applications in biomedicine.</p>","PeriodicalId":18712,"journal":{"name":"Molecular & Cellular Proteomics","volume":" ","pages":"100958"},"PeriodicalIF":6.1,"publicationDate":"2025-03-27","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143743184","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Metal ion-enhanced ZIC-cHILIC StageTip for N-Glycoproteomic and Phosphoproteomic Profiling in EGFR-mutated Lung Cancer Cells.","authors":"Yi-Ju Chen, Yan-Lin Chen, Kun-Hao Chang, Hsiang-Chun Cheng, Chiao-Chun Chang, Yu-Ju Chen","doi":"10.1016/j.mcpro.2025.100957","DOIUrl":"https://doi.org/10.1016/j.mcpro.2025.100957","url":null,"abstract":"<p><p>Surface glycosylation and intracellular phosphorylation regulates the cell-cell communication and signaling cascades. Due to complex glycosylation and dynamic phosphorylation, exploring their interplay remains technically challenging. In this study, we reported a tandem ZIC-cHILIC StageTip strategy for streamlined and simultaneous (sialo)glycoproteomic and phosphoproteomic profiling. We first demonstrated that Fe ions expand the utility of ZIC-cHILIC strategy to phosphoproteomic analysis with greatly enhanced >4-fold coverage and high specificity for mono-phosphopeptides (95%). The Fe-ZIC-cHILIC tandem tips, leveraging stepwise fractionation, enable large-scale coverage of 10,536 glycopeptides, including highly confident 4,285 sialoglycopeptpides, and 11,329 phosphopeptides in a single cell type. To study the mechanism underlying the tyrosine kinase inhibitor (TKI) resistance in non-small cell lung cancer (NSCLC), application of the strategy to 4 NSCLC cells harboring different EGFR mutations reveals significantly differential 1,559 glycopeptides and 1,949 phosphopeptides either in EGFR mutation or TKI resistant cells. Without protein immunoprecipitation, the approach identified FDA-approved drug targets, such as EGFR, ERBB2, MET, and integrin family members. Most prominent alterations were observed in EGFR (auto-phosphorylation Y1197 and 10 bi- and triantennary fucosyl-sialo glycans at N603), downstream PI3K-Akt pathway (ERBB2-T1240, MET-S990/T992, AKT-S124/S126) and integrin family (sialo-fucosyl glycans), suggesting site-specific alteration between N-glycosylation and phosphorylation interplay in the TKI resistant L858R-T790M mutant NSCLC cells. The glycoproteomic and phosphoproteomic landscape may help to unravel the complex modification alterations underlying the resistant mechanism, offering insights for improving therapeutic strategies and patient outcomes.</p>","PeriodicalId":18712,"journal":{"name":"Molecular & Cellular Proteomics","volume":" ","pages":"100957"},"PeriodicalIF":6.1,"publicationDate":"2025-03-26","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143743185","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Bioinformatics-Guided Identification and Quantification of Biomarkers of Crotalus atrox Envenoming and its Neutralization by Antivenom.","authors":"Auwal A Bala, Anas Bedraoui, Salim El Mejjad, Nicholas K Willard, Joseph D Hatcher, Anton Iliuk, Joanne E Curran, Elda E Sanchez, Montamas Suntravat, Emelyn Salazar, Rachid El Fatimy, Tariq Daouda, Jacob A Galan","doi":"10.1016/j.mcpro.2025.100956","DOIUrl":"https://doi.org/10.1016/j.mcpro.2025.100956","url":null,"abstract":"<p><p>Quantitative mass spectrometry-based proteomics of extracellular vesicles (EVs) provides systems-level exploration for the analysis of snakebite envenoming (SBE) as the venom progresses, causing injuries such as hemorrhage, trauma, and death. Predicting EV biomarkers has become an essential aspect of this process, offering an avenue to explore the specific pathophysiological changes that occur after envenoming. As new omics approaches emerge to advance our understanding of SBE, further bioinformatics analyses are warranted to incorporate the use of antivenom or other therapeutics to observe their global impact on various biological processes. Herein, we used an in vivo BALB/c mouse model and proteomics approach to analyze the physiological impacts of SBE and antivenom neutralization in intact animals; this was followed by bioinformatics methods to predict potential EV biomarkers. Groups of mice (n=5) were intramuscularly injected with Saline or Crotalus atrox venom. After 30 minutes, the mice received saline or antivenom (ANTIVIPMYN®) by intravenous injection. After 24 hours, blood was collected to extract the plasma to analyze the EV content and determine the exposome of C. atrox venom as well as the neutralizing capabilities of the antivenom. The predicted biomarkers consistently and significantly sensitive to antivenom treatment are Slc25a4, Rps8, Akr1c6, Naa10, Sult1d1, Hadha, Mbl2, Zc3hav, Tgfb1, Prxl2a, Coro1c, Tnni1, Ryr3, C8b, Mycbp, and Cfhr4. These biomarkers pointed towards specific physiological alterations, causing significant metabolic changes in mitochondrial homeostasis, lipid metabolism, immunity, and cytolysis, indicating hallmarks of traumatic injury. Here, we present a more comprehensive view of murine plasma EV proteome and further identify significant changes in abundance for potential biomarkers associated with antivenom treatment. The predicted biomarkers have the potential to enhance current diagnostic tools for snakebite management, thereby contributing significantly to the evolution of treatment strategies in the diagnosis and prognosis of SBE.</p>","PeriodicalId":18712,"journal":{"name":"Molecular & Cellular Proteomics","volume":" ","pages":"100956"},"PeriodicalIF":6.1,"publicationDate":"2025-03-25","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143730676","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Proteomic Analysis of Aqueous Humor Identified Clinically Relevant Molecular Targets for Neovascular Complications in Diabetic Retinopathy.","authors":"Jae Won Oh, Seong Joon Ahn, Jae Hun Jung, Tae Wan Kim, Kwang Pyo Kim","doi":"10.1016/j.mcpro.2025.100953","DOIUrl":"https://doi.org/10.1016/j.mcpro.2025.100953","url":null,"abstract":"<p><p>Diabetic retinopathy (DR) is a leading cause of blindness in adults under 40 in the developed world, with a significant proportion progressing to vision-threatening stages such as proliferative diabetic retinopathy (PDR) and neovascular glaucoma (NVG). This study aims to explore the molecular mechanisms underlying the progression from non-proliferative DR (NPDR) to PDR and NVG, focusing on identifying potential biomarkers and therapeutic targets. Utilizing discovery-based proteomics, specifically label-free quantification (LFQ) and Tandem mass tag (TMT), we analyzed aqueous humor (AH) proteins obtained during cataract surgery or anterior chamber paracentesis from patients with NPDR, PDR, and NVG. Validation of marker candidates for each disease state was conducted using triple quadrupole (QQQ)-MS for targeted protein quantification. Our proteomic analysis identified 2,255 proteins and gene ontology analysis and functional annotation highlighted key biological processes implicated in DR, such as lens development, immune responses and lipid metabolism. Validation of potential biomarkers identified 20 proteins with significant concentration changes, including several candidates with diagnostic utility based on ROC curve analysis. Further investigation into clinical relevance revealed that crystallin gamma-S (CRYGS) is strongly associated with cataract severity, highlighting its role as a potential marker for ocular complications in DR. Importantly, we identified that the pathological factors driving DR progression have a much greater impact than age, a previously known variable, in shaping the proteomic landscape of aqueous humor. Additionally, proteins associated with macular degeneration (CA1, CA2, and HBA1) were uncovered, providing new insights into overlapping mechanisms between DR and other retinal diseases. Finally, proteins linked to panretinal photocoagulation (PRP) treatment, including APOB and CST6, were identified, suggesting their involvement in the therapeutic response and post-treatment adaptation. These findings underscore the potential of AH proteomics in uncovering predictive biomarkers and elucidating the molecular pathogenesis of DR and its complications.</p>","PeriodicalId":18712,"journal":{"name":"Molecular & Cellular Proteomics","volume":" ","pages":"100953"},"PeriodicalIF":6.1,"publicationDate":"2025-03-19","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143674394","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}