Hepatocellular carcinoma (HCC) exhibits marked sexual dimorphism, with females demonstrating superior survival, yet the underlying molecular mechanisms remain unclear. We integrated bulk transcriptomics (GSE39791, TCGA-LIHC, GSE14520) and single-cell RNA sequencing (five datasets, n = 58 patients, 238,982 cells) with machine learning (LASSO, SVM, random forest) to identify female-protective genes driving HCC disparities. Activating transcription factor 5 (ATF5) emerged as a female-protective gene with higher expression in females versus males across cohorts. Single-cell analyses revealed ATF5 defines a female-enriched, low-grade malignant subcluster with elevated apoptotic programs and reduced proliferative signaling, and pseudotime analysis showed coordinated ATF5-GPER1 downregulation during malignant progression (Spearman ρ = -0.52 and -0.48; both p < 0.001). In the immune compartment, ATF5 marked a female-enriched IFN-γ⁺ macrophage state with enhanced immunostimulatory programs and preferential CXCL9/10-CXCR3-mediated communication with CD8/NK cells. Mechanistically, ATF5 transcriptionally activates G protein-coupled estrogen receptor 1 (GPER1), forming an estrogen-responsive regulatory module that functionally suppresses proliferation, induces apoptosis (HepG2: 26.45% vs. 11.88%, p < 0.0001), and inhibits migration in a GPER1-dependent manner as demonstrated by rescue experiments. Tissue microarray validation (n = 167) confirmed high ATF5 expression predicts improved recurrence-free survival specifically in female patients (HR = 0.34, p = 0.040) but not males (p = 0.080). The ATF5-GPER1 axis represents a female-protective circuit operating through tumor-intrinsic suppression and immune remodeling, offering mechanistic insight into HCC sexual dimorphism and identifying ATF5 as a sex-specific prognostic biomarker with potential therapeutic implications.

Successful surgical resection of solid tumours requires highly reliable real-time intraoperative tools to accurately delineate tumour boundaries, which remains challenging in routine clinical standards. Here, we identify endogenous substances with intense autofluorescence in the second near-infrared window (NIR-II, 1,000–1,700 nm) that are abundant in human liver tissues but negligible in cancerous tissues. Inspired by this discovery, we develop a label-free and wide-field imaging approach, named tissue autofluorescence NIR-II imaging (TANI) for visualizing human liver malignancies. TANI demonstrates exceptional contrast (7.69 ± 0.52), sensitivity (97.8

Oncolytic adenovirus H101 has shown antitumor activity in hepatocellular carcinoma (HCC), but the molecular determinants of treatment response remain unclear. In this study, a Hepa1-6 subcutaneous tumor model was established in C57BL/6 mice and treated with intratumoral H101, followed by integrated transcriptomic and proteomic analyses to identify candidate genes associated with H101 response. PRXL2B was selected for further investigation using public multi-omics datasets, tissue microarray-based immunohistochemistry, in vitro functional assays, mechanistic analyses, and in vivo validation experiments. Integrated multi-omics analyses identified PRXL2B as a candidate gene downregulated after H101 treatment. Public datasets and tissue-based validation further showed that PRXL2B was upregulated in HCC tissues. In MHCC97H and HCCLM3 cells, PRXL2B knockdown inhibited proliferation, migration, and invasion, promoted apoptosis and cell-cycle arrest, and enhanced the antitumor effect of H101. Mechanistically, PRXL2B silencing reduced AKT phosphorylation and PD-L1 expression. In vivo, PRXL2B knockdown suppressed tumor growth, and the combination of PRXL2B knockdown and H101 produced the strongest antitumor effect. These findings indicate that PRXL2B promotes malignant phenotypes in HCC and may modulate H101 efficacy through the PI3K/AKT/PD-L1 axis. Targeting PRXL2B may therefore represent a potential strategy to enhance the therapeutic efficacy of oncolytic virus therapy in HCC.

Background Phase III trials establish the core safety profiles of tyrosine kinase inhibitors (TKIs) for hepatocellular carcinoma (HCC), but real-world comparative data remain limited. This study aimed to compare the real-world adverse event (AE) profiles of four HCC TKIs, cross-reference findings with pivotal trials, and identify underrecognized toxicities. Methods AE reports for sorafenib, lenvatinib, regorafenib, and cabozantinib were retrieved from the FDA Adverse Event Reporting System (FAERS) from approval through Q1 2025. Disproportionality and time-to-onset analyses were performed to characterize AE distributions and temporal patterns. Findings were cross-referenced with phase III trial data. Results While gastrointestinal and cutaneous AEs were shared, distinct drug-specific profiles emerged. Sorafenib and regorafenib exhibited strong dermatologic signals, with sorafenib notably associated with pancreatic enzyme elevation and regorafenib with peripheral neuropathy. Lenvatinib demonstrated the broadest multisystem toxicity, including prominent signals for hypertension, proteinuria, hypothyroidism, and severe underemphasized events like posterior reversible encephalopathy syndrome and interstitial lung disease. Cabozantinib was characterized by marked oral mucosal toxicity. Although FAERS data were broadly consistent with phase III trials, several clinically relevant AEs appeared underrepresented in trial settings. Time-to-onset analyses revealed distinct temporal risk windows. Conclusions This real-world analysis reveals meaningful differences in the AE profiles and onset timelines of four HCC TKIs, complementing phase III trial evidence. Identifying these underrecognized, drug-specific toxicities provides actionable insights for individualized surveillance and early intervention in routine clinical practice.

Lenvatinib, a first-line tyrosine kinase inhibitor for advanced hepatocellular carcinoma (HCC), faces clinical challenges due to acquired drug resistance. While metabolic reprogramming has been implicated in therapeutic resistance, the precise mechanistic links remain elusive. Here, we identified ACSS2-mediated metabolic-epigenetic crosstalk as a critical driver of Lenvatinib resistance. Transcriptomic and metabolomic profiling identified enhanced pyruvate metabolism in resistant HCC cells, with ACSS2 expression showing the strongest association with Lenvatinib resistance. Genetic manipulation experiments demonstrated that ACSS2 dictates therapeutic sensitivity, with knockdown restoring drug response and overexpression conferring resistance. Mechanistically, ACSS2-driven palmitate biosynthesis facilitates EGFR palmitoylation, which shields the receptor from ubiquitin-dependent degradation. This stabilization sustains oncogenic EGFR signaling, ultimately mediating therapeutic escape. Crucially, pharmacological inhibition of ACSS2 synergized with Lenvatinib to overcome resistance in both subcutaneous and hydrodynamic transfection HCC models. Our findings not only delineate the ACSS2/EGFR axis as a metabolic vulnerability in resistant HCC but also propose ACSS2-targeted therapy as a promising strategy to reverse Lenvatinib resistance, providing a novel therapeutic approach for advanced HCC management.

Background: Microvascular invasion (MVI) is a major determinant of recurrence and poor prognosis in hepatocellular carcinoma (HCC), yet accurate preoperative assessment remains challenging. We investigated the incremental value of multi-regional spatial interaction (MSI) features derived from supervoxel-based habitat analysis on Gd-EOB-DTPA-enhanced MRI for preoperative MVI prediction. Methods: In this retrospective dual-centre study, patients with HCC from two institutions who underwent preoperative Gd-EOB-DTPA-enhanced MRI were included. Tumour habitats were generated from T1-weighted, arterial phase, portal venous phase, and hepatobiliary phase images. Habitat radiomic and MSI features were extracted to build single-sequence models and a four-sequence fusion model. Centre 1 served as the development cohort and Centre 2 as the external validation cohort. Parallel subgroup analyses were performed for tumours smaller than 5 cm, 3 cm, and 2 cm. Findings: Four habitat subregions were consistently identified across sequences and showed differences in relative proportion, entropy, and radiomic feature distribution. Among single-sequence models, the hepatobiliary phase model performed best and retained the most MSI features. The MSI-inclusive fusion model achieved AUCs of 0.85 in the internal test cohort and 0.89 in the external validation cohort, with favourable performance maintained across all small-tumour subgroups. Interpretation: MSI features provide complementary information beyond conventional habitat radiomics for characterising spatial heterogeneity in HCC. Multi-sequence integration of MSI features improves preoperative MVI prediction and may support risk stratification.

BACKGROUND:Most patients with biliary tract cancer (BTC) do not derive durable clinical benefit (DCB) from immune checkpoint inhibitors (ICIs), underscoring the urgent need for predictive biomarkers. While urinary proteomics represents a non-invasive approach for biomarker discovery and mechanism exploration, its utility in ICI-treated patients with cancer remains unexplored. OBJECTIVE:We aimed to establish urinary proteomics as a predictive tool for ICI responsiveness and to elucidate its relationship with tumour dynamics and tumour microenvironment (TME) remodelling in BTC. DESIGN:We performed a staged mass spectrometry (MS)-based discovery-validation proteomics workflow in 211 urine samples from 97 treatment-naïve patients with BTC undergoing ICI-based therapy. A machine learning model was developed based on baseline proteomic features for ICI response prediction. Single-cell transcriptomics of 11 pretreatment tumour biopsies and spatial transcriptomics were integrated to explore the link between urinary proteomics and TME. RESULTS:Patients achieving DCB exhibited enrichment of immune activation and systemic inflammatory pathways, whereas non-durable benefit was correlated with protumourigenic processes. Longitudinal urinary proteomic dynamics could mirror TME remodelling and tumour evolution. A machine learning-derived 4-urinary protein panel (protein tyrosine phosphatase non-receptor 13 (PTPN13), SUB1, MICAL-L1, VARS1) robustly predicted DCB and early responses. Subsequent external validation in an independent cohort (n=24) using parallel reaction monitoring-MS further confirms its generalisability. PTPN13+ malignant cells were identified as key regulators of proapoptotic TME states, contributing to sustained ICI responsiveness. CONCLUSIONS:This study pioneers the application of urinary proteomics in immuno-oncology, providing a non-invasive approach to predict and monitor ICI responsiveness, while offering mechanistic insights into TME dynamics in BTC.

Background and Aims: The immunosuppressive tumor microenvironment (TME) limits immunotherapy efficacy in intrahepatic cholangiocarcinoma (ICC). Understanding the molecular drivers of this TME is essential for developing new therapies. This study aimed to identify novel oncogenes that modulate the immune landscape of ICC using a multi-omics approach. Methods: We integrated transcriptomic and proteomic data from our ICC cohorts with public datasets (TCGA-CHOL, GSE107943, OEP002768) to identify genes coupregulated with PD-L1 (CD274). Single-cell RNA sequencing (scRNA-seq) was used to analyze cell-type-specific expression and intercellular communication. Clinical significance was validated through tissue microarrays and multiplex immunofluorescence in an independent ICC cohort. Results: Multi-omics screening identified TACC3 as a key candidate in ICC. Elevated TACC3 expression in ICC tissues correlated with poor prognosis and promoted tumor cell proliferation and migration. TACC3 activated the STAT3 pathway, increasing PD-L1 transcription. scRNA-seq showed TACC3/PD-L1 interaction in malignant epithelial cells, with PD-L1 co-expressed with FOXP3 in regulatory T cells (Tregs). Cell-cell communication analysis predicted strong interactions between malignant cells and Tregs. TACC3 knockdown reduced PD-L1 expression and inhibited STAT3 and AKT phosphorylation. Clinical validation confirmed co-expression of TACC3, PD-L1, and FOXP3, with high TACC3 levels linked to worse clinicopathological features and shorter progression-free survival. Conclusions: Our study defines a TACC3-STAT3-PD-L1 axis driving immunosuppression in ICC. TACC3 fosters an immunosuppressive TME by upregulating PD-L1 and is associated with a Treg-rich contexture, suggesting that TACC3 may serve as a potential therapeutic target to overcome ICC immunosuppression.

Cholangiocarcinoma (CCA) is an aggressive malignancy with dismal prognosis; PD-1/PD-L1 blockade benefits few patients, as the tumor microenvironment (TME) is immunologically “cold”. A key CCA-TME feature is massive tumor-associated neutrophil (TAN) infiltration releasing neutrophil extracellular traps (NETs). Conventionally deemed pro-tumorigenic, NETs’ anti-tumor potential is overlooked, leaving their dual roles and regulatory circuits in CCA undefined. Our study finds prominent NETs enrichment in CCA clinical specimens and preclinical models, correlating with poor outcomes. Functional studies show NETs exert dual effects: a dominant pro-tumor arm accelerating growth/metastasis, and a latent immunostimulatory arm rendering the TME “hot”. Integrated multi-omics and bioinformatics analyses dissect these functions into distinct molecular clusters with divergent prognostic value. Pro-tumor clusters are selectively activated by MAPK signaling, present in ~25% of CCA with KRAS mutations. Mechanistically, uPA-loaded NETs engage uPAR on CCA cells; TLR co-reception licenses downstream MAPK activation, tipping toward tumor promotion. We devised a cluster-directed combination: DNase I dismantling NETs scaffolds plus uPAR blockade neutralizing residual pro-tumor fragments. This strategy abolishes oncogenic signaling while sparing—even boosting—STING-dependent anti-tumor immunity, sensitizing KRAS-mutant and wild-type CCA to anti-PD-L1 therapy. Human transcriptomic datasets link low pro-tumor/high immunostimulatory NETs signatures with durable immunotherapy responses. We establish a functional-cluster framework for CCA NETs biology and provide a precision co-targeting regimen turning pro-tumor function into immunotherapeutic opportunity.

Dietary intervention provides a novel approach for cancer therapy. Elaidic acid (EA), which accounts for 80-90% of total trans fatty acids in foods, has recently been found to exert anti-tumor effects. However, the biological functions and underlying mechanisms of EA remain elusive in hepatocellular carcinoma (HCC). In this study, targeted fatty acid metabolomics demonstrated that among 44 types of fatty acids, the concentration of EA decreased most significantly when comparing plasma from HCC patients with plasma from healthy people. Through in vivo assays using HCC orthotopic and xenograft mouse models, we further revealed that dietary EA attenuates HCC growth. Notably, when gut microbiota was depleted using a cocktail of antibiotics, the anti-tumor effect of EA was diminished, confirming that EA suppresses HCC tumor growth by modulating gut microbiota. Mechanistically, analysis of 16S ribosomal RNA sequencing showed that dietary EA markedly increases the abundance of intestinal Ligilactobacillus murinus (L. murinus). Subsequent untargeted metabolomic sequencing analysis further demonstrated that dietary EA drives the production of L. murinus-derived spermidine (SPD), which attenuates HCC growth in vitro as well as in vivo. The observed impact correlated with the phosphorylation of p38 MAPK and the upregulation of biomarkers pertinent to apoptosis and proliferation, including tumor protein 53, bcl-2-associated X protein, and cysteine-requiring aspartate protease 3. Taken together, our findings highlight the important role of intestinal L. murinus-derived SPD in EA-mediated HCC suppression, thereby offering a promising dietary strategy for HCC treatment.

The induction of osteogenic differentiation in preadipocytes may serve as a potential therapeutic approach for treating osteoporosis and osteoporotic fractures. All-trans retinoic acid (ATRA) promotes the bone morphogenetic protein 9 (BMP9)-induced osteogenic differentiation of preadipocytes. The present study further investigated whether vascular endothelial growth factor A (VEGFA) may play a role in this process and the effect of ATRA and BMP9 on osteoporotic fracture healing in rats. The results indicated that ATRA and BMP9 synergistically upregulated VEGFA expression in preadipocytes. Furthermore, knockdown of VEGFA expression abolished the stimulatory effect of ATRA on the BMP9-induced early and late osteogenic differentiation of preadipocytes in vitro, as evidenced by a decrease in alkaline phosphatase (ALP) activity, osteopontin and osteocalcin expression as well as mineralization. The in vivo cell implantation assay showed that ATRA failed to augment BMP9-induced ectopic bone formation in the absence of VEGFA. Subsequently, an osteoporotic femoral fracture rat model was established and micro-CT scans, alongside quantitative analysis, revealed that ATRA effectively promoted BMP9-stimulated callus formation during osteoporotic fracture healing. Moreover, ATRA and BMP9 acted together to significantly elevate VEGFA expression in bone calluses. Mechanistically, ATRA and BMP9 synergistically stimulated the osteogenic transcription factor, runt-related transcription factor 2 (Runx2). Transcriptomic and bioinformatic analyses further revealed that the phosphatidylinositol 3-kinase (PI3K)/protein kinase B (Akt) signaling pathway may be crucial for mediating the synergistic effects of ATRA and BMP9, as validated by detecting the phosphorylation of PI3K and Akt. However, the increases in Runx2 expression and Akt phosphorylation induced by the combination of ATRA and BMP9 were inhibited by VEGFA silencing. ATRA also failed to increase the BMP9-induced ALP activity in preadipocytes treated with an Akt inhibitor. These findings suggest that VEGFA may influence the potentiation effect of ATRA on the BMP9-mediated osteogenesis of preadipocytes and in osteoporotic fracture healing through Runx2 and the PI3K/Akt signaling pathway.

Immunotherapy has revolutionized cancer treatment, making it a challenge to noninvasively monitor immune infiltration. Metabolic reprogramming in cancers, including hepatocellular carcinoma (HCC), is closely linked to immune status. In this study, we aimed to evaluate the ability of carbon-11 acetate (11C-acetate) and fluorine-18 fluorodeoxyglucose (18F-FDG) PET/CT findings in predicting overall survival (OS) and immune infiltration in HCC patients. Totally 32 patients who underwent preoperative 18F-FDG and 11C-acetate PET/CT, followed by liver resection for HCC, were prospectively enrolled at authors' institute between January 2019 and October 2021. Tracer uptake was qualified. Densities of CD3+, CD8+, and granzyme B+ CD8+ immune cells were assessed and the Immunoscore was defined by combining the densities of CD3+ and CD8+ in tumor interior (TI) and invasion margin (IM). Patients with avid HCCs in 11C-acetate PET/CT demonstrated a longer OS. Those with only 11C-acetate-avid HCCs exhibited a longer OS compared to those with only 18F-FDG uptake. In contrast to 18F-FDG uptake, 11C-acetate uptake was positively associated with CD3+, CD8+, and granzyme B+ CD8+ cell infiltration. Patients with a higher Immunoscore exhibited a longer OS and an increased uptake of 11C-acetate rather than 18F-FDG. The sensitivity of 11C-acetate PET/CT in the detection of patients with immune infiltration was superior to that of 18F-FDG PET/CT (88% [21 of 24] vs. 58% [14 of 24]). These data show that preoperative 11C-acetate PET/CT may be a promising approach for the evaluation of immune status and postoperative outcome of HCCs.

The collagen β (1-O) glycosyltransferase 25 domain 1 (GLT25D1), a crucial collagen-modifying enzyme (CME), plays a pivotal role in multiple pathophysiological processes. However, its prognostic and biological roles in hepatocellular carcinoma (HCC) have not been reported. CME-related genes (CMEGs) were obtained from the Molecular Signatures Database (MSigDB), differentially expressed CMEGs (DECMEGs) and prognostic ones were identified. GLT25D1 expression was determined at the mRNA and protein levels in multiple datasets and in our HCC cohort. Its prognostic performance was evaluated and the immune microenvironment was investigated. The effects of GLT25D1 on tumorigenesis were further explored via in vitro and in vivo experiments. Four potential prognosis-associated DECMEGs, including GLT25D1, were identified. GLT25D1 was noticeably up-regulated in HCC tissues and significantly associated with advanced tumor grade and stage. Enrichment analysis revealed that GLT25D1 could participate in regulating immune responses and various carcinogenic processes. HCC patients with high GLT25D1 expression had decreased CD8+ T cells and increased M0 macrophages, leading to an immunosuppressive microenvironment. Our in vivo and in vitro experiments confirmed the increased GLT25D1 expression, and GLT25D1 knockdown impaired the HCC malignant phenotypes. Our results showed that GLT25D1 could be a carcinogenic indicator reflecting poor prognosis and might serve as a potential risk biomarker for HCC patients.

BackgroundThe hematopoietic ecosystem comprises both cellular components such as hematopoietic stem cells (HSCs) and immune cells as well as non-cellular components including iron. Systemic iron overload, which leads to serious complications and affects both patients’ quality of life and overall survival, is a common clinical challenge in patients with acute myeloid leukemia (AML). We previously elucidated the direct effects of iron overload on AML cells. It’s worth noting that iron overload remodels the hematopoietic ecosystem. However, whether and how remodeled leukemic microenvironment with overloaded iron regulates normal HSCs and immune cells, especially leukemia-associated macrophages (LAMs), in AML have not been elucidated.MethodsThe MLL-AF9-induced AML (MA9) cells were originated from c-kit+ BM cells enriched from C57BL/6J mice that infected with MSCV-MLL-AF9-GFP retrovirus. The MA9 AML mouse model was established by transplantation of MA9 cells into C57BL/6 mice. MA9 mice were i.p. administered with iron dextran every other day for a total of 6 times to established the iron overload MLL-AF9-induced AML mouse model (MA9/FE). HSC maintenance and differentiation was assessed by flow cytometry, cell proliferation, cell apoptosis, colony forming and competitive transplantation assays. LAM activation and function was analyzed by RNA-sequencing, flow cytometry and coculture assay. Intravenous clodronate liposome administration was employed to reduce LAMs in AML.ResultsIron overload skewed myeloid differentiation of normal HSCs. Furthermore, iron overload affected LAMs in the AML microenvironment by promoting LAM polarization toward an M2 phenotype. Functionally, iron overload decreased the phagocytic function of LAMs against leukemia cells and inhibited LAM-induced T cell activation by acquiring a tolerogenic phenotype with aberrant immune checkpoints. Moreover, depletion of LAMs attenuated iron overload caused acceleration of AML progression.ConclusionsCollectively, this study reveals the significance of iron overload in remodeling hematopoietic ecosystem and affecting HSC and LAM function in AML, providing new insights into the multifaceted role of iron overload in leukemia.

To assess the prognostic value of Fluorine 18-labeled fluorodeoxyglucose [18F]FDG, gallium 68-labeled fibroblast-activation protein inhibitor-04 [68Ga]Ga-FAPI-04, 11C-acetate in hepatocellular carcinoma (HCC) and evaluate the potential usefulness and advantages of different combinations for accurate diagnosis. Thirty-six patients with suspected hepatic masses were prospectively enrolled from May 2021 to September 2022 and underwent [18F]FDG, [68Ga]Ga-FAPI-04, and 11C-acetate PET/CT scans before surgery. PET/CT results and histopathologic examinations were independently interpreted by two radiologists and pathologists, respectively. Kaplan–Meier overall survival curves were calculated and the sensitivity among [18F]FDG, 11C-acetate, [68Ga]Ga-FAPI-04, and different combinations were compared. Of the 36 included patients (mean age, 59 years ± 10 (standard deviation)), 29 were diagnosed with HCC, four with non-HCC malignant tumors, and three with benign tumors. Patients with HCC lesions negative for 11C-acetate or [68Ga]Ga-FAPI-04 exhibited poorer overall survival. Out of 36 patients, 44 HCC lesions were detected. The dual-tracer [68Ga]Ga-FAPI-04/11C-acetate exhibited the highest sensitivity (39 of 44 lesions (88.6

Nuclear morphology plays a critical role in regulating gene expression and cell functions. While most research has focused on the direct effects of nuclear morphology on cell fate, its impact on the cell secretome and surrounding cells remains largely unexplored. In this study, we fabricate implants with a micropillar topography using methacrylated poly(octamethylene citrate)/hydroxyapatite (mPOC/HA) composites to investigate how micropillar-induced nuclear deformation influences cell secretome for osteogenesis and cranial bone regeneration. In vitro, cells with deformed nuclei show enhanced secretion of proteins that support extracellular matrix (ECM) organization, which promotes osteogenic differentiation in neighboring mesenchymal stromal cells (MSCs). In a female mouse model with critical-size cranial defects, nuclear-deformed MSCs on micropillar mPOC/HA implants elevate Col1a2 expression, contributing to bone matrix formation, and drive cell differentiation toward osteogenic progenitor cells. These findings indicate that micropillars modulate the secretome of hMSCs, thereby influencing the fate of surrounding cells through matricrine effects.

BACKGROUND: The precise role of Galectin-9, an immune checkpoint protein involved in immune responses, in hepatocellular carcinoma (HCC) remains elusive. Importantly, the prognostic value of serum Galectin-9 has not been clarified, and its association with infiltrating immune characteristics was unclear. METHODS: The association between serum Galectin-9 concentration and HCC recurrence was analyzed in two cohorts of HCC patients (training 133; validation 97) who received curative resection during 2018 and 2019. Bioinformatic analyses, including WGCNA, GSEA, GO, KEGG, Hallmark, CIBERSORT, QUANTISEQ, ssGSEA and TISIDB, were performed to systematically demonstrate the expression pattern, immunomodulation role, and prognostic value of Galectin-9 in HCC. These findings were further validated by immunohistochemistry staining. RESULTS: Patients with high serum Galectin-9 levels had significantly shorter time to tumor recurrence (TTR; P < 0.001) in both cohorts, and serum Galectin-9 was identified as an independent predictor of HCC recurrence, even in patients with low-AFP or early-stage. Bioinformatic analyzes revealed high Galectin-9 expression is involved in immune-evasive and inflammatory signaling pathways. It correlated with increased infiltration of exhausted CD8 + T cells, Tregs, TAMs and MDSCs. Interestingly, we found Galectin-9 was predominantly expressed on macrophages rather than malignant cells, and showed positively association with serum Galectin-9 concentration according to IHC results. Concordantly, high serum Galectin-9 levels also reflected an immune-evasive microenvironment composed by extensive CD163 + and FOXP3 + cell infiltrates. CONCLUSIONS: Elevated serum Galectin-9 was a novel indicator for worse prognosis in HCC. The high expression of Galectin-9 may reflect the immunosuppressive environment by increasing CD163 + and FOXP3 + cell infiltrates.

The lack of reliable non-invasive biomarkers for early colorectal cancer (CRC) diagnosis underscores the need for improved diagnostic tools. Extracellular vesicles (EVs) have emerged as promising candidates for liquid-biopsy-based cancer monitoring. Here, we propose a comprehensive workflow that integrates staged mass spectrometry (MS)-based discovery and verification with ELISA-based validation to identify EV protein biomarkers for CRC. Our approach, applied to 1,272 individuals, yields a machine learning model, ColonTrack, incorporating EV proteins HNRNPK, CTTN, and PSMC6. ColonTrack effectively distinguishes CRC from non-CRC cases and identifies early-stage CRC with high accuracy (combined area under the curve [AUC] >0.97, sensitivity ∼0.94, specificity ∼0.93). Our analysis of EV protein profiles from tissue and plasma demonstrates ColonTrack's potential as a robust non-invasive biomarker panel for CRC diagnosis and early detection.

Background Drug therapy plays an essential role in the management of hepatocellular carcinoma (HCC). Recently, the use of natural products to suppress tumor cells has emerged as a promising direction for drug development. Juglone, a natural compound, exhibits anticancer activities across various cancer types. However, the precise mechanism underlying the anticancer effect of juglone, especially in HCC, remains elusive. Purpose This study aimed to investigate the potential inhibitory effects of juglone on HCC and pan-cancer, as well as elucidate the underlying mechanism. Methods Cell Counting Kit-8 and colony formation assays were used to examine cell proliferation. Transwell and wound healing assays were used to evaluate cell migration. Cell cycle distribution was assessed by flow cytometry. The in vivo effect of juglone on HCC was evaluated by establishing the HCC xenograft mice model. RNA sequencing and inhibitors targeting diverse modes of programmed cell death were applied to uncover the form of juglone-induced cell death. Integrated transcriptomic, and proteomic analyses unveiled the underlying mechanism. The dual-luciferase reporter assay was employed to verify the findings. The pan-cancer value of juglone was assessed using TCGA database analysis and cellular assays. Results Juglone suppressed HCC growth via ferroptosis in vitro and in vivo, which is evidenced by increased levels of iron, lipid peroxidation (LPO), reactive oxygen species (ROS), malondialdehyde (MDA), and decreased levels of glutathione (GSH). Omic analyses, gene silencing and functional analyses showed the upregulated HMOX1 and FOSL1 were the key effector molecule and transcriptional factor in juglone-induced ferroptosis, respectively. The binding site of FOSL1 at the promoter of HMOX1 was identified. Juglone could induce ferroptosis in pan-cancer by activating the FOSL1-HMOX1 axis. Conclusion Our findings, for the first time, demonstrate that juglone effectively inhibits tumor growth by inducing FOSL1-HMOX1-dependent ferroptosis, thereby offering a promising strategy for the development of anticancer drugs.

BRCA1/BRCA2-containing complex subunit 3 (BRCC3) serves as a deubiquitinating enzyme contributing to multiple inflammation-related disorders. However, the role of BRCC3 in modulating airway inflammation in asthma has not been investigated. This study aimed to examine the role of BRCC3 in airway inflammation using a mouse model of asthma induced by ovalbumin (OVA). BRCC3 levels were found to be elevated in mice with asthma. BRCC3 knockout (KO) mice demonstrated a notable improvement in pathological changes, accompanied by reduced levels of inflammatory cell infiltration and inflammatory cytokines, compared to wild-type (WT) mice following OVA challenge. The NLRP3 inflammasome was high activated in asthmatic mice, which was restrained by BRCC3 KO, as companied by a decrease in NLRP3, ASC, cleaved Caspase-1, cleaved Gasdermin D (GSDMD), IL-1β, and IL-18. In vitro studies demonstrated BRCC3 levels increased in airway epithelial cells in response to house dust mite (HDM) stimulation, depending on the dose and duration of exposure. Silencing BRCC3 in airway epithelial cells protected against HDM-induced cell injury and inflammation, along with inhibiting the NLRP3 inflammasome and pyroptosis. Conversely, the overexpression of BRCC3 in airway epithelial cells worsened DM-induced cell injury and inflammation while also enhancing the NLRP3 inflammasome and pyroptosis. Further investigations revealed that silencing BRCC3 increased the ubiquitination of NLRP3, whereas overexpressing BRCC3 decreased it. Pharmacological inhibition of the NLRP3 inflammasome diminished the effects of BRCC3 overexpression on the inflammation and pyroptosis induced by HDM in airway epithelial cells. Overall, these findings underscore the importance of BRCC3 in the pathogenesis of asthma. Deletion of BRCC3 alleviates airway inflammation in asthma by impeding the activation of the NLRP3 inflammasome, thus indicating that BRCC3 could serve as a potential target for asthma therapy.