Lung cancer remains the leading cause of cancer-related death worldwide. Despite the success of immune checkpoint inhibitors (ICIs), particularly those targeting PD-1/PD-L1 pathways, a significant number of patients exhibit either primary resistance or acquire resistance over time. While genomic and epigenetic mechanisms contribute to this resistance, increasing evidence points to the pivotal role of immunometabolism—the interface of cellular metabolism and immune regulation. This review focuses on how altered metabolic states in both immune cells and cancer stem-like cells (CSCs) within the lung tumor microenvironment contribute to ICI resistance. We discuss key metabolic pathways involved in immune suppression, the metabolic plasticity of CSCs, and how these factors interact to shape a metabolically hostile tumor niche for immune effector cells. Finally, we explore emerging therapeutic strategies targeting immunometabolism to enhance immunotherapy efficacy in lung cancer.

BACKGROUND: Stem-like gastric cancer (GC) is an aggressive molecular subtype marked by poor prognosis and limited response to immune checkpoint blockade (ICB). The spatial mechanisms driving this resistance remain unclear. METHODS: We conducted spatially resolved single-cell transcriptomic profiling of diffuse-type GC tissues to uncover the spatial architecture and functional diversity of tumor and stromal populations. Cellular heterogeneity and region-specific signaling pathways were characterized using integrative bioinformatics analyses. RESULTS: We identified transcriptionally diverse, high-entropy cell populations predominantly localized in the deep tumor regions. These included unique endothelial and fibroblast subsets enriched for pro-tumorigenic and immune-regulatory signaling. A notable finding was the engagement of deep-region endothelial cells in VISFATIN (extracellular NAMPT) signaling through the ITGA5-ITGB1 integrin axis, associated with immune evasion and poor prognosis. This endothelial signaling program is distinct from and functionally independent of cancer-associated fibroblast (CAF)-mediated pathways. Elevated expression of the NAMPT-ITGA5-ITGB1 axis was observed in ICB non-responders and correlated with reduced overall survival. CONCLUSIONS: Our study delineates spatially defined cellular programs that contribute to immune escape in stem-like GC, highlighting a novel VISFATIN-integrin signaling axis as a potential biomarker and therapeutic target in immunotherapy-resistant tumors.

, resulting in intracellular glutathione (GSH) depletion, accumulation of lipid peroxidation products such as malondialdehyde and 4-hydroxynonenal, and increased lipid reactive oxygen species (ROS). Although SLC7A11 mRNA levels were upregulated, its protein levels were reduced due to increased ubiquitination and proteasomal degradation. Further analysis revealed that dRib suppressed the expression of OTUB1, a deubiquitinating enzyme that directly interacts with SLC7A11 and inhibits its ubiquitination. Overexpression of either OTUB1 or SLC7A11 restored cystine transport, replenished GSH levels, reduced lipid ROS and cell death, supporting their protective roles against ferroptosis. Transmission electron microscopy confirmed that dRib induced mitochondrial morphological changes consistent with ferroptosis, such as shrinkage and cristae loss. These findings demonstrate that the loss of OTUB1 promotes ferroptosis in pancreatic β-cells through destabilization of SLC7A11. Targeting the OTUB1-SLC7A11 pathway may offer a novel therapeutic approach for preserving β-cell survival and preventing the progression of diabetes.

Dear Editor, We explored the heterogeneity and clinical implications of cancer-associated fibroblasts (CAFs) within the gastric cancer microenvironment to understand their contributions to tumour progression and therapeutic resistance. CAFs, essential components of the tumour microenvironment, regulate tumour growth, immune responses, and therapeutic outcomes. We classified CAFs into three subtypes: myofibroblastic (myoCAF), immune-regulatory (irCAF), and inflammatory (infCAF), based on gene expression signatures and functional characteristics (Figure 1A–C). Each CAF subtype was characterized by unique marker genes from a literature review (Tables S1 and S2).1 Gene ontology (GO) analysis revealed that transcription factors NFKB1, RELA, SP1, JUN, and transcriptional regulator HDAC2 are key in CAF subtype regulation (Figure 1B). Using the MCODE algorithm2 for protein–protein interaction, we found myoCAFs were involved in blood vessel development and extracellular matrix modelling, while irCAFs were associated with peptide ligand-binding and chemokine receptors, indicating distinct roles in immune modulation (Figure 1D,E). To further investigate CAF stemness, we employed the StemID tool on single-cell cohorts. Clusters 17 and 8 showed notably high entropy and stemness, indicating that these fibroblasts are highly activated and possess stem-like properties, contributing to tumour progression and treatment resistance (Figure 1F,G). These fibroblasts were enriched in pathways related to extracellular matrix organization,3 including the NABA core matrisome and elastic fibre formation (Figure 1H).4 Our analysis further revealed a strong correlation between irCAF signatures and stem-like signatures in bulk gastric cancer samples, suggesting that irCAFs play a key role in sustaining an aggressive tumour microenvironment (Figure 1I-L).5 We evaluated the impact of these CAF subtypes on patient prognosis using data from the Cancer Genome Atlas (TCGA) stomach adenocarcinoma (STAD) dataset. High expression of infCAF, irCAF, and myoCAF signatures was linked to poor prognosis (Figure 2A). A combined analysis of all three CAF signatures also indicated poor outcomes for the high-expression group (Figure 2B). In the Yonsei Cancer Hospital cohort of 497 gastric cancer patients (Y497 dataset), these CAF subtypes were enriched in the stem-like type (Figure 2C). While the infCAF signature was not significantly linked to adverse outcomes, high expression of both irCAF and myoCAF signatures was consistently linked to worse clinical outcomes (irCAF: p = .0022, myoCAF: p = .0053) (Figure 2D). We classified patients into nine groups based on the CAF subtype signature via gene set enrichment test in the Y497 cohort. (Figure 2E). Furthermore, we confirmed that the CAF subtype signatures found in TCGA pan-cancer datasets act differently across various cancer types. All three CAF signatures in BLCA, LGG, and STAD cancer types were significant for patient prognosis (Figure 2F). Using Y497 transcriptome profiling, we performed a deconvolution analysis to understand how different CAF subtypes contribute to the immune tumour microenvironment. Both infCAF and irCAF showed similar trends in stem-like type samples, being positively correlated with macrophage M2 and regulatory T (Treg) cells, while CD8+ T cells were more prevalent in the gastric molecular subtype (Figure 2G). To understand the influence of CAFs on immune response and therapy resistance, we conducted single-cell analysis on non-responders to immune checkpoint blockade (ICB) therapy. High-entropy, stem-like activated cells were predominantly in clusters 1 and 7 (Figure 3A,B). Cluster 1 had mostly endothelial cells, while cluster 7 included fibroblasts, T cells, macrophages, and others (Figure 3C). Clusters 8 and 10, with the lowest stemness and entropy, were largely B cells linked to adaptive immunity. GO analysis of highly expressed genes within endothelial cells showed enrichment in pathways related to blood vessel development, cell migration, and VEGFA-VEGFA2 signalling (Figure 3D). The prognostic relevance of these genes was confirmed in the TCGA STAD dataset, where high expression correlated with poor outcomes (Figure 3E). Additionally, these genes were enriched in the stem-like type within the Yonsei Cancer Hospital cohort, demonstrating a consistent pattern (Figure 3F). We further assessed the expression of signature genes linked to drug resistance using bulk RNA-seq data from non-responder and responder groups to ICB treatment. The signature genes were mainly overexpressed in nonresponders (Figure 3G). The correlation between stem-like signatures and CAF subtypes was validated using a gastric cancer patient-derived organoid (PDO) dataset from Yonsei Cancer Hospital, showing that the immune-regulatory signature had the strongest correlation with the stem-like phenotype (Figure 3H,I). At the single-cell level, irCAFs demonstrated the highest expression of stem-like PDO signature genes, indicating their potential role in drug resistance (Figure 3J). To decipher the communication patterns of CAFs within the tumour microenvironment, we employed Cellchat6 analysis to identify key signalling pathways (Figure 4A). We identified eight cell types that clustered into three main communication patterns, predicting the involvement of fibroblast-derived ligands such as VEGF, PTN, and ANGPT (Figure 4B,C). Fibroblasts, as crucial components of the PTN signalling network, acted as both senders and receivers, particularly influencing mesenchymal stem cells, proliferative cells, peritoneal mesothelial cells, and tumour cells (Figure 4D). Within the CAF subtypes, PTN signalling was mediated by both irCAF and infCAF (Figure 4E). Predicted analysis of ligand-receptor pairs identified the PTN ligand and syndecan-2 (SDC2) receptor as critical components of the signalling pathway (Figure 4F). SDC2, a transmembrane proteoglycan involved in glycosaminoglycan metabolism, was found to be highly expressed in myoCAFs and cancer stem cells, correlating with poor prognosis in both the TCGA STAD dataset and the Y497 cohort (Figure 4H–K). Elevated SDC2 expression was particularly noted in ICB non-responder groups within both the Samsung Medical Center (SMC) and Y497 cohorts7 (Figure 4L,M). A strong correlation (R = .8, p = 0) was observed between SDC2 and the ACTA2 gene, a marker associated with resistance to ICB therapy (Figure 4N).8 Furthermore, SDC2 expression was notably enriched in the stem-like type of gastric cancer, potentially promoting tumorigenesis via glycosaminoglycan biosynthesis pathways such as heparan sulfate and chondroitin sulfate (Figure 4O,P).9 The transition from anti-tumorigenic syndecans to tumorigenic SDC2 may influence the invasive capacity and metastatic potential of highly aggressive tumour cells.10 In conclusion, our study highlights the heterogeneity of CAF subtypes in gastric cancer and their roles in prognosis and therapy resistance. Targeting CAFs could provide new avenues for personalised treatment of gastric cancer. JYS was responsible for the conceptualisation, methodology development, data analysis, manuscript drafting, wrote, review, editing, interpretation of results and supervision of the study. JHC contributed to the generation of PDO data and manuscript review and contributed to the interpretation of results. KS performed contributed to manuscript review.ETK provided funding acquisition, manuscript review and editing, and contributed to the interpretation of results. The authors have nothing to report. This work was funded by the National Research Foundation of Korea (NRF) with grants from the Korean Ministry of Science and ICT (RS-2024-00352590), Ministry of Education (RS-2023-00270936), and the Korea Health Industry Development Institute (KHIDI) funded by the Ministry of Health & Welfare (RS-2022-KH129726, RS-2024-00438990). The authors declare no conflict of interest. This study was approved by the Institutional Review Board of Yonsei Cancer Hospital (IRB no. 4-2017-0106). Informed consent was obtained from all participants. The datasets generated and/or analyzed during this study are not publicly available. Researchers interested in the organoid RNA-seq data for gastric cancer may contact Dr. Ji-Yong Sung ([email protected]). Please note: The publisher is not responsible for the content or functionality of any supporting information supplied by the authors. Any queries (other than missing content) should be directed to the corresponding author for the article.

Lung cancer remains the leading cause of cancer-related death worldwide. Despite the success of immune checkpoint inhibitors (ICIs), particularly those targeting PD-1/PD-L1 pathways, a significant number of patients exhibit either primary resistance or acquire resistance over time. While genomic and epigenetic mechanisms contribute to this resistance, increasing evidence points to the pivotal role of immunometabolism—the interface of cellular metabolism and immune regulation. This review focuses on how altered metabolic states in both immune cells and cancer stem-like cells (CSCs) within the lung tumor microenvironment contribute to ICI resistance. We discuss key metabolic pathways involved in immune suppression, the metabolic plasticity of CSCs, and how these factors interact to shape a metabolically hostile tumor niche for immune effector cells. Finally, we explore emerging therapeutic strategies targeting immunometabolism to enhance immunotherapy efficacy in lung cancer.

BACKGROUND: Stem-like gastric cancer (GC) is an aggressive molecular subtype marked by poor prognosis and limited response to immune checkpoint blockade (ICB). The spatial mechanisms driving this resistance remain unclear. METHODS: We conducted spatially resolved single-cell transcriptomic profiling of diffuse-type GC tissues to uncover the spatial architecture and functional diversity of tumor and stromal populations. Cellular heterogeneity and region-specific signaling pathways were characterized using integrative bioinformatics analyses. RESULTS: We identified transcriptionally diverse, high-entropy cell populations predominantly localized in the deep tumor regions. These included unique endothelial and fibroblast subsets enriched for pro-tumorigenic and immune-regulatory signaling. A notable finding was the engagement of deep-region endothelial cells in VISFATIN (extracellular NAMPT) signaling through the ITGA5-ITGB1 integrin axis, associated with immune evasion and poor prognosis. This endothelial signaling program is distinct from and functionally independent of cancer-associated fibroblast (CAF)-mediated pathways. Elevated expression of the NAMPT-ITGA5-ITGB1 axis was observed in ICB non-responders and correlated with reduced overall survival. CONCLUSIONS: Our study delineates spatially defined cellular programs that contribute to immune escape in stem-like GC, highlighting a novel VISFATIN-integrin signaling axis as a potential biomarker and therapeutic target in immunotherapy-resistant tumors.

, resulting in intracellular glutathione (GSH) depletion, accumulation of lipid peroxidation products such as malondialdehyde and 4-hydroxynonenal, and increased lipid reactive oxygen species (ROS). Although SLC7A11 mRNA levels were upregulated, its protein levels were reduced due to increased ubiquitination and proteasomal degradation. Further analysis revealed that dRib suppressed the expression of OTUB1, a deubiquitinating enzyme that directly interacts with SLC7A11 and inhibits its ubiquitination. Overexpression of either OTUB1 or SLC7A11 restored cystine transport, replenished GSH levels, reduced lipid ROS and cell death, supporting their protective roles against ferroptosis. Transmission electron microscopy confirmed that dRib induced mitochondrial morphological changes consistent with ferroptosis, such as shrinkage and cristae loss. These findings demonstrate that the loss of OTUB1 promotes ferroptosis in pancreatic β-cells through destabilization of SLC7A11. Targeting the OTUB1-SLC7A11 pathway may offer a novel therapeutic approach for preserving β-cell survival and preventing the progression of diabetes.

Dear Editor, We explored the heterogeneity and clinical implications of cancer-associated fibroblasts (CAFs) within the gastric cancer microenvironment to understand their contributions to tumour progression and therapeutic resistance. CAFs, essential components of the tumour microenvironment, regulate tumour growth, immune responses, and therapeutic outcomes. We classified CAFs into three subtypes: myofibroblastic (myoCAF), immune-regulatory (irCAF), and inflammatory (infCAF), based on gene expression signatures and functional characteristics (Figure 1A–C). Each CAF subtype was characterized by unique marker genes from a literature review (Tables S1 and S2).1 Gene ontology (GO) analysis revealed that transcription factors NFKB1, RELA, SP1, JUN, and transcriptional regulator HDAC2 are key in CAF subtype regulation (Figure 1B). Using the MCODE algorithm2 for protein–protein interaction, we found myoCAFs were involved in blood vessel development and extracellular matrix modelling, while irCAFs were associated with peptide ligand-binding and chemokine receptors, indicating distinct roles in immune modulation (Figure 1D,E). To further investigate CAF stemness, we employed the StemID tool on single-cell cohorts. Clusters 17 and 8 showed notably high entropy and stemness, indicating that these fibroblasts are highly activated and possess stem-like properties, contributing to tumour progression and treatment resistance (Figure 1F,G). These fibroblasts were enriched in pathways related to extracellular matrix organization,3 including the NABA core matrisome and elastic fibre formation (Figure 1H).4 Our analysis further revealed a strong correlation between irCAF signatures and stem-like signatures in bulk gastric cancer samples, suggesting that irCAFs play a key role in sustaining an aggressive tumour microenvironment (Figure 1I-L).5 We evaluated the impact of these CAF subtypes on patient prognosis using data from the Cancer Genome Atlas (TCGA) stomach adenocarcinoma (STAD) dataset. High expression of infCAF, irCAF, and myoCAF signatures was linked to poor prognosis (Figure 2A). A combined analysis of all three CAF signatures also indicated poor outcomes for the high-expression group (Figure 2B). In the Yonsei Cancer Hospital cohort of 497 gastric cancer patients (Y497 dataset), these CAF subtypes were enriched in the stem-like type (Figure 2C). While the infCAF signature was not significantly linked to adverse outcomes, high expression of both irCAF and myoCAF signatures was consistently linked to worse clinical outcomes (irCAF: p = .0022, myoCAF: p = .0053) (Figure 2D). We classified patients into nine groups based on the CAF subtype signature via gene set enrichment test in the Y497 cohort. (Figure 2E). Furthermore, we confirmed that the CAF subtype signatures found in TCGA pan-cancer datasets act differently across various cancer types. All three CAF signatures in BLCA, LGG, and STAD cancer types were significant for patient prognosis (Figure 2F). Using Y497 transcriptome profiling, we performed a deconvolution analysis to understand how different CAF subtypes contribute to the immune tumour microenvironment. Both infCAF and irCAF showed similar trends in stem-like type samples, being positively correlated with macrophage M2 and regulatory T (Treg) cells, while CD8+ T cells were more prevalent in the gastric molecular subtype (Figure 2G). To understand the influence of CAFs on immune response and therapy resistance, we conducted single-cell analysis on non-responders to immune checkpoint blockade (ICB) therapy. High-entropy, stem-like activated cells were predominantly in clusters 1 and 7 (Figure 3A,B). Cluster 1 had mostly endothelial cells, while cluster 7 included fibroblasts, T cells, macrophages, and others (Figure 3C). Clusters 8 and 10, with the lowest stemness and entropy, were largely B cells linked to adaptive immunity. GO analysis of highly expressed genes within endothelial cells showed enrichment in pathways related to blood vessel development, cell migration, and VEGFA-VEGFA2 signalling (Figure 3D). The prognostic relevance of these genes was confirmed in the TCGA STAD dataset, where high expression correlated with poor outcomes (Figure 3E). Additionally, these genes were enriched in the stem-like type within the Yonsei Cancer Hospital cohort, demonstrating a consistent pattern (Figure 3F). We further assessed the expression of signature genes linked to drug resistance using bulk RNA-seq data from non-responder and responder groups to ICB treatment. The signature genes were mainly overexpressed in nonresponders (Figure 3G). The correlation between stem-like signatures and CAF subtypes was validated using a gastric cancer patient-derived organoid (PDO) dataset from Yonsei Cancer Hospital, showing that the immune-regulatory signature had the strongest correlation with the stem-like phenotype (Figure 3H,I). At the single-cell level, irCAFs demonstrated the highest expression of stem-like PDO signature genes, indicating their potential role in drug resistance (Figure 3J). To decipher the communication patterns of CAFs within the tumour microenvironment, we employed Cellchat6 analysis to identify key signalling pathways (Figure 4A). We identified eight cell types that clustered into three main communication patterns, predicting the involvement of fibroblast-derived ligands such as VEGF, PTN, and ANGPT (Figure 4B,C). Fibroblasts, as crucial components of the PTN signalling network, acted as both senders and receivers, particularly influencing mesenchymal stem cells, proliferative cells, peritoneal mesothelial cells, and tumour cells (Figure 4D). Within the CAF subtypes, PTN signalling was mediated by both irCAF and infCAF (Figure 4E). Predicted analysis of ligand-receptor pairs identified the PTN ligand and syndecan-2 (SDC2) receptor as critical components of the signalling pathway (Figure 4F). SDC2, a transmembrane proteoglycan involved in glycosaminoglycan metabolism, was found to be highly expressed in myoCAFs and cancer stem cells, correlating with poor prognosis in both the TCGA STAD dataset and the Y497 cohort (Figure 4H–K). Elevated SDC2 expression was particularly noted in ICB non-responder groups within both the Samsung Medical Center (SMC) and Y497 cohorts7 (Figure 4L,M). A strong correlation (R = .8, p = 0) was observed between SDC2 and the ACTA2 gene, a marker associated with resistance to ICB therapy (Figure 4N).8 Furthermore, SDC2 expression was notably enriched in the stem-like type of gastric cancer, potentially promoting tumorigenesis via glycosaminoglycan biosynthesis pathways such as heparan sulfate and chondroitin sulfate (Figure 4O,P).9 The transition from anti-tumorigenic syndecans to tumorigenic SDC2 may influence the invasive capacity and metastatic potential of highly aggressive tumour cells.10 In conclusion, our study highlights the heterogeneity of CAF subtypes in gastric cancer and their roles in prognosis and therapy resistance. Targeting CAFs could provide new avenues for personalised treatment of gastric cancer. JYS was responsible for the conceptualisation, methodology development, data analysis, manuscript drafting, wrote, review, editing, interpretation of results and supervision of the study. JHC contributed to the generation of PDO data and manuscript review and contributed to the interpretation of results. KS performed contributed to manuscript review.ETK provided funding acquisition, manuscript review and editing, and contributed to the interpretation of results. The authors have nothing to report. This work was funded by the National Research Foundation of Korea (NRF) with grants from the Korean Ministry of Science and ICT (RS-2024-00352590), Ministry of Education (RS-2023-00270936), and the Korea Health Industry Development Institute (KHIDI) funded by the Ministry of Health & Welfare (RS-2022-KH129726, RS-2024-00438990). The authors declare no conflict of interest. This study was approved by the Institutional Review Board of Yonsei Cancer Hospital (IRB no. 4-2017-0106). Informed consent was obtained from all participants. The datasets generated and/or analyzed during this study are not publicly available. Researchers interested in the organoid RNA-seq data for gastric cancer may contact Dr. Ji-Yong Sung ([email protected]). Please note: The publisher is not responsible for the content or functionality of any supporting information supplied by the authors. Any queries (other than missing content) should be directed to the corresponding author for the article.

The Schlafen gene family encodes for proteins involved in various biological tasks, including cell proliferation, differentiation, and T cell development. Schlafens were initially discovered in mice, and have been studied in the context of cancer biology, as well as their role in protecting cells during viral infection. This protein family provides antiviral barriers via direct and indirect effects on virus infection. Schlafens can inhibit the replication of viruses with both RNA and DNA genomes. In this review, we summarize the cellular functions and the emerging relationship between Schlafens and innate immunity. We also discuss the functions and distinctions of this emerging family of proteins as host restriction factors against viral infection. Further research into Schlafen protein function will provide insight into their mechanisms that contribute to intrinsic and innate host immunity.

CRISPR-Cas systems have transformed viral genetics by enabling precise and efficient manipulation of large DNA virus genomes. This review provides a practical framework for applying CRISPR technology to herpesviruses and other large DNA viruses as an alternative and complement to traditional BAC recombination. Key considerations include nuclease choice; sgRNA design that minimizes cut-to-edit distance and prevents re-cutting; donor template configuration and homology arm length; and synchronized delivery of Cas complexes and donor DNA. Strategies to promote HDR efficiency, such as the use of small-molecule modulators, are also summarized. In addition, practical workflows for clone selection, genotypic validation, and phenotypic confirmation are summarized. Case studies in herpes simplex virus type 1 and human cytomegalovirus illustrate how optimized CRISPR designs achieve reproducible, scarless knock-ins and conditional gene manipulation at essential loci without complementing cell lines. Together, these approaches establish CRISPR as a flexible, scalable platform for functional genomics, antiviral target discovery, and translational virology, enabling direct editing of clinical isolates previously inaccessible with bacterial artificial chromosome-based methods.

Objective: Prototype foamy virus (PFV) is an attractive gene delivery platform due to its large cargo capacity and favorable safety profile; however, the structural basis of its interaction with heparan sulfate (HS), a critical attachment factor for viral entry, remains undefined. The objective of this study was to identify the structural determinants of HS recognition within PFV Env and to evaluate whether rational, structure-guided engineering could enhance viral entry and gene transfer efficiency. Methods: structural modeling, molecular docking, and systematic mutagenesis of the PFV Env receptor-binding domain (RBD), targeted residue substitutions, and combinatorial mutations spanning the upper domain (UD) and lower domain (LD) were generated and evaluated using quantitative cell-based transduction assays. In addition, Tet-On-inducible Env-expressing stable producer cell lines were established to provide a reproducible platform for functional validation. Results: Alanine substitutions at R298, R440, and E446 in the UD abolished infectivity, confirming their essential roles in HS-mediated attachment. In contrast, selective substitutions at adjacent positions, Q296R and G403F in the UD, and E232N, I330F, and I334F in the LD, enhanced transduction efficiency by up to 1.32-fold relative to the wild type. Combinatorial variants integrating beneficial UD and LD mutations exhibited synergistic effects, achieving a transduction efficiency of 68.9%, corresponding to a 1.55-fold increase over the wild type (44.4%). Interspecies domain replacement with simian foamy virus Env reduced infectivity, underscoring the context-specific nature of PFV-HS interactions. In the inducible stable cell system, the LD var6 mutant achieved 8.6% transduction compared to 4.4% for the wild type, representing up to a 1.95-fold increase. Discussion: These findings define the structural determinants of HS recognition in PFV Env and demonstrate that residue-level, structure-guided engineering can enhance PFV transduction efficiency. This study provides experimentally validated insight into PFV Env-HS interactions and establishes a rational framework for further optimization of PFV-based gene delivery technologies.

Herpes simplex virus type 1 (HSV-1) infection remains a major global health challenge, yet the mechanisms underlying strain-specific innate immune responses are poorly understood. Here, we show that distinct HSV-1 strains differentially activate the absent in melanoma 2 (AIM2) inflammasome. The HF strain robustly induces AIM2-dependent inflammasome activation, whereas the F and KOS strains elicit minimal responses despite comparable infection efficiency. We demonstrate that this difference is driven by viral genomic features rather than replication capacity. Genomic analyses identify a poly(T) DNA sequence within the UL25-UL26 intergenic region that is enriched in the HF strain. Deletion of a 14-mer poly(T) sequence markedly impairs inflammasome activation, cytokine release, and host protection in vivo, whereas introduction of a poly(T) tract into the F strain is sufficient to confer AIM2 activation and enhanced host defense. Furthermore, poly(T)-mediated AIM2 activation is length-dependent, conserved in human macrophages, and requires a cGAS-STING-IRF1 licensing axis. Together, these findings identify viral poly(T) DNA as a key determinant of strain-specific AIM2 inflammasome activation and reveal how viral genomic variation shapes innate immune recognition.

-induced skin damage.