Aug 2026· Clinical and Translational Medicine· Vol 16· 0 citations· 40 references
Medicine
TL;DR
CD155 is identified as a key driver of anti‐PD‐1 resistance in NSCLC and targeting CD155 combined with anti‐PD‐1 overcomes resistance, supporting a dual‐target therapeutic strategy.
Abstract
Abstract Objective Anti‐PD‐1 therapy resistance remains a critical barrier in non‐small cell lung cancer (NSCLC) management, and the underlying mechanisms are incompletely defined. Methods We generated CD155‑knockout (KO) NSCLC cell lines using the CRISPR‑Cas9 system and performed systematic multi‑omics analyses, including single‑cell RNA‑seq, bulk RNA‑seq, proteomics, and metabolomics. The key molecular mechanisms were further validated by immunohistochemistry (IHC), western blotting, and chromatin immunoprecipitation (ChIP). Functional assays assessed cell proliferation, migration, and metabolic phenotypes, while the therapeutic efficacy was assessed in vivo using AAV9_shCD155. Results Single‐cell sequencing revealed aberrantly high CD155 expression in NSCLC patients with poor response to anti‐PD‐1 therapy. High CD155 expression in NSCLC tissues correlated with unfavourable prognosis. ETS1 was identified as a direct transcriptional driver of CD155. Multi‐omics analysis and functional assays demonstrated that CD155 upregulates the expression of key glycolytic proteins (GLUT1, GLUT3, LDHB) by activating the PI3K/AKT/HIF‐1α signalling axis, thereby driving glycolytic metabolism, proliferation, and migration of tumour cells. CD155 knockout significantly suppressed these malignant phenotypes. In xenograft mouse models, monotherapy with AAV9_shCD155 effectively inhibited tumour growth and postoperative recurrence. More importantly, in humanised mouse models, combining AAV9_shCD155 with pembrolizumab produced synergistic anti‐tumour effects, more significantly suppressing tumour growth and promoting immune cell infiltration into the tumour microenvironment. Conclusion CD155 mediates anti‐PD‐1 resistance by activating PI3K/AKT/HIF‐1α‐driven glycolytic reprogramming. Targeting CD155 combined with anti‐PD‐1 overcomes resistance, supporting a dual‐target therapeutic strategy. Key points CD155 is identified as a key driver of anti‐PD‐1 resistance in NSCLC. CD155 promotes tumour glycolysis and malignant progression via the PI3K/AKT/HIF‐1α signalling axis. AAV9_shCD155 combined with anti‐PD‐1 markedly inhibits tumour growth and promotes immune infiltration.
Background/Objectives: Immune checkpoint blockade targeting the PD-1/PD-L1 axis is a standard treatment for urothelial carcinoma (UC), but 20–30% of patients develop resistance. The underlying mechanisms, particularly regarding tumor microenvironment metabolic reprogramming, remain unclear. Methods: We performed single-cell RNA sequencing on pre-treatment tumors from eight pembrolizumab-treated UC patients (three responders, five non-responders). Bioinformatic analyses included cell clustering, pathway enrichment, and cell–cell communication, validated using public datasets. Functional experiments involved SGPL1 knockdown in MB49 cells and a syngeneic PD-1-resistant murine model (MB49-R5). Results: Responders exhibited a higher proportion of infiltrating T-cells. Unexpectedly, CD4+ subsets, rather than CD8+ cells, were numerically enriched in responders. However, all T-cell subsets in responders demonstrated enhanced oxidative phosphorylation-dominant metabolic activity. Enhanced macrophage–T-cell crosstalk was observed, with macrophages exhibiting M1-like polarization driven by CD4+ T-cell-derived IFN-γ and CD40L signaling. In contrast, non-responders displayed dominant tumor-derived MIF–CD74 signaling and M2 polarization. Integration of public datasets identified SGPL1 as a key metabolic regulator associated with poor prognosis and reduced immune activation. Functional experiments demonstrated that SGPL1 knockdown inhibited tumor proliferation and restored anti-PD-1 sensitivity, accompanied by increased T-cell infiltration. Conclusions: T-cell metabolic activation drives anti-PD-1 responses in UC, and CD4+ T-cell-mediated M1 polarization and tumor-intrinsic SGPL1 crucially shape therapeutic outcomes, highlighting SGPL1 as a candidate metabolic target to overcome PD-1 resistance.
Abstract Background Metabolic reprogramming within the tumor microenvironment is a pivotal barrier to effective immune checkpoint blockade (ICB). While programmed death ligand 1 (PD-L1) is well characterized as a ligand inhibiting T-cell function, its intrinsic ‘reverse signaling’ role in regulating tumor metabolism and shaping the immune landscape remains poorly understood. Here, we investigated the metabolic determinants of resistance to anti-programmed cell death protein 1 (anti-PD-1) therapy and the underlying molecular mechanisms. Methods Integrated metabolomics and transcriptomics were performed on tumor samples from patients with non-small cell lung cancer and cell lines. Mechanisms were delineated using RNA sequencing, cleavage under targets and tagmentation assays, metabolic flux analysis, and coculture systems. The therapeutic efficacy of targeting metabolic effectors was evaluated in syngeneic mouse models and correlated with immune profiling. Results We identified a distinct metabolic signature characterized by aberrant pyruvate accumulation in patients resistant to anti-PD-1 therapy. Mechanistically, we demonstrate that antibody-mediated ligation of PD-L1 triggers an intrinsic endoplasmic reticulum (ER) stress response via the PERK–ATF4–CHOP axis. ATF4 acts as a transcriptional activator that directly upregulates pyruvate dehydrogenase kinase 4 (PDK4) (blocking pyruvate oxidation) and glutaminase (GLS) (promoting glutaminolysis), creating a ‘dual-hit’ metabolic rewiring that drives intracellular pyruvate build-up. Subsequently, tumor-secreted pyruvate is taken up by tumor-associated macrophages (TAMs) via MCT1, inducing mitochondrial reactive oxygen species accumulation and driving them into a state of cellular senescence. These senescent TAMs upregulate PD-L1 via STAT3 signaling, thereby reinforcing an immunosuppressive feedback loop. Pharmacological inhibition of PDK4 and GLS effectively abolished pyruvate accumulation, prevented macrophage senescence, and restored CD8+ T-cell cytotoxicity. Conclusions Our study identifies a novel ‘PD-L1–ER stress–pyruvate–macrophage senescence’ axis as a key mechanism underlying primary resistance to ICB. These findings highlight the non-canonical reverse-signaling function of PD-L1 in metabolic remodeling and propose that targeting the PDK4/GLS-dependent pyruvate surge offers a promising therapeutic strategy to sensitize tumors to anti-PD-1 immunotherapy.
Wen-Xin Lian, Hui Yang, Yu-Qi Zhang et al.· Journal for ImmunoTherapy of...· 0 citations
Abstract Background As a pivotal metabolic enzyme, cysteine dioxygenase type 1 (CDO1) exerts tumour‐suppressive effects across diverse tumour types, and its expression is strongly correlated with clinical prognosis. However, the molecular mechanisms underlying CDO1‐mediated tumour suppression in gastric cancer (GC), its relationship with the tumour‐associated immune microenvironment, and pharmacological strategies to restore its expression remain poorly understood. Methods CDO1 expression and prognosis were evaluated by multi‐omics and tissue microarray analyses. Tumour microenvironment and immune infiltration were analyzed using ESTIMATE and ssGSEA. Downstream pathways and interacting proteins were identified by transcriptomics, co‐immunoprecipitation, and GST pull‐down. CDO1 function was assessed by proliferation, apoptosis, and migration assays in gain‐ and loss‐of‐function models. In vivo tumorigenesis and CDO1‐dependent decitabine efficacy were evaluated by subcutaneous xenografts. Patient‐derived organoids were used to assess decitabine sensitivity and 5‐FU synergy. Results Compared with normal controls, CDO1 expression was notably decreased in GC tissues, and its low expression was strongly linked to unfavourable prognosis, supporting its utility as a biomarker for prognosis. Elevated CDO1 levels correlated with an immune‐active tumour microenvironment and reduced metastatic signatures. Mechanistically, CDO1 directly bound to PI3K p85α, disrupting p85α‐p110α dimerization, thereby attenuating PI3K/AKT phosphorylation and downregulating THBS1 expression. CDO1 overexpression led to reduced proliferation, invasiveness, and EMT, accompanied by increased apoptosis. These effects were reversed by PI3K activation or THBS1 co‐overexpression. Decitabine was identified as an agent that epigenetically restores CDO1 expression. Critically, CDO1 knockdown significantly attenuated the anti‐tumour efficacy of decitabine in vivo, confirming that decitabine acts primarily through CDO1 reactivation. Decitabine synergized with 5‐FU in both organoids and xenografts. Conclusions Our data identify CDO1 as both a biomarker for prognosis and a tumour suppressor in gastric cancer. They reveal a CDO1–PI3K/AKT–THBS1 signalling axis and support the epigenetic reactivation of CDO1 by decitabine as a translatable therapeutic strategy. Key points CDO1 is frequently downregulated in gastric cancer and serves as an independent favourable prognostic biomarker. CDO1 directly binds PI3K p85α, disrupting p85α–p110α dimerization to suppress the PI3K/AKT–THBS1 signalling axis. Decitabine epigenetically restores CDO1 expression, and its anti‐tumour activity is critically CDO1‐dependent in vivo. Combining decitabine with 5‐FU synergistically overcomes gastric cancer growth in patient‐derived organoids and subcutaneous xenograft models.
Fazhi Wang, Run-Kai Zhou, Jinfeng Cai et al.· Clinical and Translational M...· 0 citations
ABSTRACT Background: Glioblastoma (GBM) establishes a highly immunosuppressive microenvironment through intricate molecular dialogues with tumor‐associated macrophages (TAMs), contributing to immunotherapy resistance. Methods: We integrated advanced biological network analyses with multimodal AI to identify CTSB as a key biomarker. Functional validation included GBM‐macrophage co‐culture systems, chromatin immunoprecipitation (ChIP), dual‐luciferase reporter assays, molecular docking, and co‐immunoprecipitation (Co‐IP). In vivo experiments employed orthotopic (GL261 and CT‐2A) and subcutaneous GBM models with lentivirus‐mediated CTSB knockdown and anti‐PD‐1 therapy. Tumor microenvironment dynamics were assessed via in‐house generated single‐cell RNA sequencing and multiplex immunofluorescence. Results: Cathepsin B (CTSB) was identified as a critical driver of aggressive GBM progression and poor outcomes. Mechanistically, macrophage‐derived IL‐6 activates STAT3 in tumor cells, upregulating CTSB expression. Structural and Co‐IP analyses revealed CTSB, secreted by tumor cells, binds the C‐terminus of macrophage S100A10, reinforcing IL‐6 secretion, forming a feedforward loop between CTSB+ GBM cells and S100A10+ macrophages. This loop enhances tumor growth, invasion, and immune evasion via PD‐L1 upregulation. In vivo, CTSB blockade reduced TAM activation, increased CD8+ T cell infiltration, and synergized with anti‐PD‐1 therapy. Conclusions: This study unveils a targetable GBM‐macrophage signaling axis, proposing CTSB inhibition as a strategy to enhance immunotherapy efficacy in GBM patients.
Hao Zhang, Nan Zhang, Xisong Liang et al.· Advancement of science· 0 citations
Aim: Despite the contribution of cancer stem-like cells (CSLCs) to acquired paclitaxel resistance in non-small cell lung cancer (NSCLC), the biomarkers and regulatory mechanisms sustaining their stemness under chemotherapy pressure remain poorly understood. This study aimed to identify the stemness-maintaining programs underlying CSLC-associated paclitaxel resistance.
Methods: Paclitaxel-resistant NSCLC cell models were established. RNA-seq data from resistant spheres and adherent resistant cells were integrated with Gene Ontology/Kyoto Encyclopedia of Genes and Genomes/gene set enrichment analysis and patient transcriptomic datasets to identify CSLC maintenance-associated candidate biomarkers. Inhibitors, sphere-forming assays, CD104-CD166+CD49fhi flow cytometry, reverse transcription quantitative polymerase chain reaction, western blotting, and ST6GAL1 knockdown or overexpression were used for functional and mechanistic validation. Sambucus nigra agglutinin lectin blotting was performed to assess epidermal growth factor receptor (EGFR) α2,6-sialylation. Clinical relevance was assessed using ST6GAL1 immunohistochemistry on 46 clinical lung tumor tissues.
Results: Paclitaxel-resistant NSCLC cells exhibited enhanced sphere formation and CD104-CD166+CD49fhi expansion. N-glycosylation was activated in resistant spheres. A seven-gene N-glycosylation signature was identified as a CSLC-associated candidate biomarker in acquired paclitaxel resistance. Inhibiting N-glycosylation suppressed the EGFR-mTOR-SOX2/BMI1 axis, decreased CSLCs, and restored paclitaxel sensitivity. ST6GAL1 regulated EGFR α2,6-sialylation. ST6GAL1 depletion also decreased EGFR abundance, suppressed mTOR-SOX2/BMI1 signaling, and sensitized paclitaxel-resistant spheres to paclitaxel rather than adherent cells. ST6GAL1 expression, which was higher in tumors from patients who underwent chemotherapy, showed a trend toward poorer survival among chemotherapy-treated patients. The seven-gene signature was associated with shorter disease-free survival but not overall survival in lung cancer patients.
Conclusions: ST6GAL1-mediated α2,6-sialylation of EGFR contributes to the maintenance of CSLC-associated paclitaxel resistance through mTOR-SOX2/BMI1 signaling. These findings identify ST6GAL1-dependent EGFR sialylation as a potential therapeutic target in CSLC-associated chemoresistance.
Yu-Xi Yang, Bing-Hui Liang, Wei-Jie Hong et al.· Cancer Drug Resistance· 0 citations
Nasopharyngeal cancer (NPC) is a malignant tumor that originates from mucosal epithelial cells of the nasopharynx. As an emerging cancer‐associated microRNA, miR‐138‐5p has been shown to modulate various malignant cell malignancies. However, its precise biological function and regulatory mechanism in NPC remain to be further elucidated. Human NPC cell lines (HK‐1 and C666‐1) were used for in vitro study, and nude mouse xenograft models were used for in vivo investigations. A bioinformatic method was utilized for the prediction of RNA expression patterns and molecular interactions. RT‐qPCR was performed for the detection of miR‐138‐5p and CENPN expression. Luciferase activity detection and RNA pulldown experiments were used for verification of gene interaction. Flow cytometry and colony formation assays were conducted to evaluate cell cycle distribution, proliferative capacity, and apoptosis rate. Furthermore, extracellular acidification rate, lactate generation, and glucose uptake were assessed to explore the regulatory effects of miR‐138‐5p and CENPN on glycolysis in NPC cells. Protein levels of CENPN and key molecules associated with cell cycle, apoptosis, glycolysis, and the PI3K signaling pathway were quantified by western blot analysis. CENPN expression is analyzed to be upregulated in head and neck cancer samples, and its high expression is associated with unfavorable clinical prognosis. In NPC cells, miR‐138‐5p was markedly downregulated and was verified to directly target the 3′ untranslated region of CENPN, thereby negatively regulating its expression. Functionally, overexpressing miR‐138‐5p obstructed malignant phenotype, glycolysis, and tumor growth of NPC cells, where these inhibitory effects were significantly counteracted by enforced CENPN expression. Moreover, miR‐138‐5p overexpression inactivated the PI3K/Akt/mTOR signaling by reducing CENPN expression in NPC cells. In conlusion, miR‐138‐5p inhibits cell growth, glycolysis, and tumorigenesis in NPC by directly targeting CENPN, thereby inhibiting the PI3K/Akt/mTOR signaling pathway. These findings highlight the potential utility of miR‐138‐5p as both a diagnostic indicator and a therapeutic intervention target for NPC management.
Hong-Xia Guan, Yang Jiang, Hui Li· Journal of biochemical and m...· 0 citations
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