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Lipocalin 2 (LCN2) is upregulated in many cancers, including pancreatic ductal adenocarcinoma (PDAC), and contributes to tumor development. LCN2 regulates microbial composition, which can influence PDAC outcomes, and has been implicated in ferroptosis resistance in several cancer types. However, the role of tumor-derived LCN2 in mediating ferroptosis and the tumor microbiome in PDAC remains unclear. Here, we show that human PDAC tumors with high LCN2 expression have altered expression of genes involved in ferroptosis, oxidative stress responses, and microbial regulation. Loss of LCN2 in PDAC cells dysregulates ferroptosis-associated pathways, increases lipid peroxidation, and sensitizes cells to ferroptosis induction. In orthotopic PDAC models, the ferroptosis inducer imidazole ketone erastin (IKE) promoted tumor growth in Lcn2-expressing tumors, whereas this effect was absent in tumors lacking LCN2. In addition, tumor-derived Lcn2 expression and ferroptosis induction were associated with distinct changes in the pancreatic tumor microbiome. Overall, these findings identify tumor-derived LCN2 as an important modulator of ferroptotic stress responses in PDAC and suggest that inhibition of tumor-derived LCN2 may enhance susceptibility to ferroptosis-based therapeutic strategies.

Pancreatic ductal carcinoma (PDAC) is characterized by a highly immunosuppressive, extracellular matrix-rich microenvironment, yet tumours display marked heterogeneity1-4. This raises the question of whether immune resistance is a global tumour property or is organized within spatially restricted niches. Here, using Perturb-map spatial functional genomics, we determine how different genes shape the growth and cellular environments of PDAC clones across space and time. This analysis revealed early gene-driven remodelling of local immune neighbourhoods preceding late-stage spatial clonal dominance. We identify SERPINE1 (encoding plasminogen activator inhibitor 1 (PAI1)) and SERPINB2 (encoding PAI2) as dominant regulators of tumour microenvironment control and immune evasion. These serpins promote stabilization of fibrin-rich extracellular matrix niches that spatially retain and programme macrophages towards immunosuppressive states while excluding cytotoxic T cells. Loss of Serpine1 or Serpinb2, or pharmacological inhibition of PAI1 or CD18, improves tumour control in mice and synergizes with anti-PD-1. Multimodal spatial analysis of patient tumours revealed that immunosuppressive niches form around rare SERPINB2- and SERPINE1-expressing PDAC subpopulations, dominated by SPP1+/MARCO+ macrophages. These findings identify cancer-derived SERPINE1 and SERPINB2 as local spatial organizers of immune suppression, linking tumour-intrinsic heterogeneity to local microenvironmental control and immunotherapy resistance in PDAC.

Protein-drug conjugates (PDCs) constitute a rapidly growing group of precise anticancer agents. Receptor-mediated endocytosis is a critical step in PDC action, which ensures the delivery of PDCs into the interior of cancer cells. Here, we report TriFHS-MMAE, the first multivalent PDC specifically targeting heparan sulfate proteoglycans (HSPGs) overexpressed by pancreatic cancer cells, which induces ultrafast and highly efficient aggregation-dependent endocytosis (ADE) of HSPGs. Using high-content screening with a near-kinome-wide library of inhibitors, we identified signaling pathways that govern ADE of HSPGs and discovered cascades that selectively operate in pancreatic cancer cells versus healthy cells. We show that priming cells with identified endocytic chemical modulators improves the targeting of the TriFHS-MMAE conjugate in vitro and in vivo. Overall, these findings provide insights into the interplay among signaling, endocytosis, and PDCs and support the development of specific therapies for pancreatic cancer.

Pancreatic cancer faces the challenges of tumor progression and postoperative pancreatic fistula during the perioperative period, which severely affect patient prognosis. In this study, an injectable self-healing multifunctional hydrogel (CMCGel) was developed based on dynamic cross‑linking between manganese ions (Mn2+) and carboxymethyl chitosan loaded with chlorella extract (CE). This platform implements an integrated "Remove-Remodel-Repair" (3R) strategy for comprehensive perioperative management. CMCGel exhibits distinctive pH/laser dual-responsive behavior: In the acidic tumor microenvironment, the accelerated release of CE and Mn2+ enhances photodynamic therapy and chemodynamic therapy (CDT). This combination, activated by laser and gated by endogenous pH, achieves remarkable tumor suppression rates of 85.81% in vitro and 96.01% in vivo. As pH normalizes, Mn2+ shifts from CDT to activate the cyclic GMP-AMP synthase stimulator of interferon genes (cGAS-STING) pathway, remodeling the immune microenvironment. Crucially, CMCGel's adhesion strength in artificial pancreatic fluid at 48 h was 4703.80 ± 50.36 Pa, only 8.17% lower than that at 10 min. This enzyme-resistant property enables durable mechanical sealing at the fistula site, reducing amylase leakage by 55.71% and creating favorable conditions for tissue repair. By integrating pH/laser-triggered combination therapy, immunomodulation, and enzyme-resistant barrier functionality into a single platform, CMCGel presents a clinically promising strategy for perioperative management of pancreatic cancer.

As it is a tumour-associated antigen in epithelial cells, research on claudin18.2 (CLDN18.2) has focused on its role as a therapeutic target in pancreatic cancers and its part in maintaining tight junctions. We elucidate the role of trogocytosis-related CLDN18.2 in CD8+ T cells and pancreatic ductal adenocarcinoma (PDAC) progression. We constructed humanised hCD34+, Trp53R172HKrasG12DPdx1-cre (KPC), Cldn18.2 knockout (KO), and patient-derived xenograft/organoid mouse models. Flow cytometry, immunofluorescence, single-cell RNA-sequencing and immunoprecipitation-mass spectrometry (IP-MS) were performed. CLDN18.2+CD8+ T cells indicated poor pancreatic cancer prognosis and immunotherapeutic resistance. CD8+ T cells acquired CLDN18.2 from tumour cells via trogocytosis, inhibiting their activation and cytotoxicity. "Dressed" CLDN18.2 suppressed glucose uptake, glycolysis and cytotoxicity of tumour-infiltrating CD8+ T cells. Mechanically, trogocytosis-related CLDN18.2 induced GSK3β/CK1α-mediated β-catenin phosphorylation, promoting β-catenin ubiquitination and proteasome degradation in CD8+ T cells. CLDN18.2 interacted with β-catenin's N-terminal domain via its C-terminal domain, further strengthening the interaction between β-catenin and CK1α. Moreover, CLDN18.2+CD8+ T cells preferentially 'homed' to the bone marrow through the CXCL12/CXCR4 axis, skewed haematopoietic stem cell myeloid differentiation and induced systemic immune senescence via IL1α. Notably, preclinical mouse studies showed PC18.1 peptide sensitised immunotherapy and suppressed PDAC progression by disrupting the CLDN18.2/β-catenin interaction in CD8+ T cells. Trogocytosis-related CLDN18.2 inhibited the glucose uptake, glycolysis and cytotoxicity of tumour-infiltrating CD8+ T cells by promoting the ubiquitin-proteasomal degradation of β-catenin in PDAC. Therefore, targeting trogocytosis-related CLDN18.2+CD8+ T cells may be a promising therapeutic strategy to inhibit PDAC progression.

Pancreatic ductal adenocarcinoma (PDAC) remains one of the most lethal malignancies. Tertiary lymphoid structures (TLS) have emerged as key components of the tumor immune microenvironment, associated with improved survival and enhanced immunotherapy response. However, non-invasive detection of TLS remains a significant clinical challenge. This multicenter study aimed to develop and validate a deep learning-based CT model to noninvasively predict TLS status and evaluate its prognostic and predictive value across surgical, non-surgical, and prospective immunochemotherapy cohorts. We enrolled 223 surgically resected patients with PDAC from two centers for model development and 82 non-surgical patients for clinical validation. A ResNet50-based deep learning network (DLN) was trained to predict TLS status from preoperative contrast-enhanced CT. The locked model was evaluated in 44 patients from a prospective immunochemotherapy trial (ChiCTR2000032293), with pathology-anchored verification, outcome assessment, and spatial immune profiling via cyclic multiplex tyramide signal amplification. The DLN achieved area under the curves (AUCs) of 0.987, 0.937, and 0.853 in training, internal, and external validation cohorts. In the non-surgical cohort (n=82), DLN-predicted TLS-positive patients had significantly longer overall survival (OS; median 19 vs 7 months; HR=3.611, p<0.0001). In the prospective cohort, pathology-anchored verification yielded an AUC of 0.929 (specificity=1.000). DLN-high patients demonstrated superior objective response rate (90.0% vs 26.5%, p=0.001), progression-free survival (PFS) (median 13.1 vs 5.3 months, p<0.001), and OS (median 17.6 vs 8.7 months, p<0.001). The DLN score independently predicted OS (HR=0.052, p<0.001) and PFS (HR=0.079, p<0.001) in multivariable analysis. The DLN score was orthogonal to programmed death-ligand 1 (PD-L1) expression (rho=0.011, p=0.946) but correlated with dendritic cell-T helper cell spatial interactions (p=0.037) and T helper-B cell spatial co-localization (rho=0.307, p=0.043)-two hallmarks of functional TLS architecture-and inversely with tumor cell density (rho=-0.399, p=0.007). This DLN-based CT model reliably predicts TLS status in PDAC with prospectively validated prognostic and predictive value in immunochemotherapy, biological grounding in TLS-associated spatial immune architecture, and orthogonal predictive value to PD-L1, supporting a complementary biomarker strategy for immunotherapy patient selection.

Fluorescence‑guided surgery using activatable probes is promising, but single‑channel probes often give false negatives-especially in pancreatic cancer due to tumor heterogeneity and limited imaging depth. Here, we built a universal charge-switching gated PET platform based on cyanine dyes bearing a meso-quinolinium moiety. This platform-by decoupling quenching from the recognition group-enabled the construction of two structurally analogous, hypoxia-activatable near-infrared (NIR) probes (Cy5-Ql-NTR and Cy7-Ql-NTR), which have comparable responsive kinetics (sensitive, fast-responding, and low-background), good water solubility and favorable renal clearance, but differ in tissue imaging depth: the former generates brighter signals in superficial regions, while the latter enables deeper tissue penetration and serves as a self-calibration reference. Notably, compared to a single probe, the combined metabolic distribution of the two probes covers a broader range of tumor target tissues. Through their dual-channel imaging, depth-dependent signal attenuation is corrected, and blind spots during probe delivery are reduced. Compared to single-channel fluorescence-guided surgery in subcutaneous pancreatic cancer and orthotopic pancreatic tumor, the dual-channel approach yielded clean margins, preserved significantly more normal tissue, and left no residual tumor on histology. This self-referenced detection strategy suppresses false negatives in single-channel detection, offering a practical and optimized solution for intraoperative guidance in pancreatic cancer.

Intrahepatic cholangiocarcinoma (iCCA) is a highly lethal malignancy with limited therapeutic options. The spatial architecture and functional diversity of tertiary lymphoid structures (TLSs) in iCCA remain unclear. Here, we present a multimodal spatial atlas of TLSs and identified intratumoral TLSs (iTLSs) as independent prognostic markers. Bulk proteomic profiling of 214 discovery and 155 validation cases identified a four-tier TLS-based tumor microenvironment classification system and supported development of a TLS-predictive random forest classifier. Imaging mass cytometry revealed that iTLS+ tumors harbor structured immune architectures, where M1-like tissue-resident macrophages (RTMs), dendritic cells, and CXCL13+ CD4+ T cells colocalize to form antigen-presenting neighborhoods (apc-CNs) spatially coupled to TLS core regions (TLScore-CNs). Single-cell spatial transcriptomics further resolved 61 TLSs into 14 spatial niches and defined a pseudotemporal maturation continuum: aggregated, activated, and postactivated. Intraniche communication, primarily mediated by ifnCAFs, iCAFs, and CXCL12+ macrophages, evolved dynamically with maturation. Single-nucleus RNA sequencing combined with Tangram-based spatial mapping revealed CXCL12+ macrophages and iCAFs forming a peripheral band in aggregated TLSs, whereas ifnCAFs infiltrated TLS interiors during activation. These findings define TLS heterogeneity and provide insights for stroma-directed immunotherapy.