An organolutetium nanosensitizer synergizes with PARP inhibition to unleash STING-mediated immunity for low-dose radioimmunotherapy.

The normal microbial occupants of the mammalian intestine are crucial for maintaining gut homeostasis, yet the mechanisms by which intestinal cells perceive and respond to the microbiota are largely unknown. Intestinal epithelial contact with commensal bacteria and/or their products has been shown to activate noninflammatory signaling pathways, such as extracellular signal-related kinase (ERK), thus influencing homeostatic processes. We previously demonstrated that commensal bacteria stimulate ERK pathway activity via interaction with formyl peptide receptors (FPRs). In the current study, we expand on these findings and show that commensal bacteria initiate ERK signaling through rapid FPR-dependent reactive oxygen species (ROS) generation and subsequent modulation of MAP kinase phosphatase redox status. ROS generation induced by the commensal bacteria Lactobacillus rhamnosus GG and the FPR peptide ligand, N-formyl-Met-Leu-Phe, was abolished in the presence of selective inhibitors for G protein-coupled signaling and FPR ligand interaction. In addition, pretreatment of cells with inhibitors of ROS generation attenuated commensal bacteria-induced ERK signaling, indicating that ROS generation is required for ERK pathway activation. Bacterial colonization also led to oxidative inactivation of the redox-sensitive and ERK-specific phosphatase, DUSP3/VHR, and consequent stimulation of ERK pathway signaling. Together, these data demonstrate that commensal bacteria and their products activate ROS signaling in an FPR-dependent manner and define a mechanism by which cellular ROS influences the ERK pathway through a redox-sensitive regulatory circuit.

Penetrative biomimetic nanovehicle boosts immunotherapy in triple-negative breast cancer via SOS1 blockade

Background: Triple-negative breast cancer (TNBC) has a unique tumor microenvironment (TME) that contributes to tumor progression and affects responses to immunotherapy. c-Jun N-terminal kinase (JNK), part of the MAPK pathway, plays a crucial role in inflammation and tumor progression and has been targeted in clinical trials for treating inflammation-related diseases and solid tumors. Indeed, JNK inhibitor (JNKi) BMS-986360 is currently being tested alone and in combination with chemotherapy or nivolumab in advanced solid tumors (NCT05625412). Our recent study showed that JNK promoted TNBC tumor growth and metastasis by creating an immunosuppressive TME via macrophage-derived CCL2 in preclinical models. Therefore, in this study, we tested our hypothesis that JNKi enhances the antitumor efficacy of macrophage-targeting agents in TNBC. Methods: The correlation of JNK signaling activity with the immune status of the TME in TNBC was analyzed using the TCGA dataset (n = 191). The antitumor efficacy of JNKi (BMS-986360, a reversible orally active pan-JNKi; or JNK-IN-8, a covalent pan-JNKi) combined with macrophage-targeting agents (pexidartinib or sotuletinib, both of which are orally active CSF-1R inhibitors) was assessed using immunocompetent syngeneic models of E0771 (M1 macrophage-enriched TME, sensitive to immune checkpoint inhibitors [ICI]), PyMT-M (M2-macrophage-enriched TME, partially sensitive to ICI), and 4T1.2 (neutrophil-enriched TME, resistant to ICI). The treatments’ effects on immune cell tumor infiltration were determined by flow cytometry. Results: To investigate the role of JNK signaling in regulating the immune landscape of the TME in cancer, we have developed a phospho-JNK (pJNK) gene signature to infer JNK phosphorylation status by gene expression. Using this pJNK gene signature and TCGA dataset, we found that high JNK (JNKhi) signaling activity was associated with high counts of Tregs and cancer-associated fibroblasts and low counts of CD8+ T cells, M1 macrophages, and neutrophils in TNBC tumors. TNBC tumors with JNKhi signaling activity were enriched with pathways regulating angiogenic and fibrotic stroma, e.g., Wnt and TGF-β signaling, which are the major inflammatory pathways involving tissue fibrosis and tumor progression. These results demonstrate a strong correlation between JNKhi signaling activity and an immunosuppressive TME in TNBC. Since JNK promoted an immunosuppressive TME in TNBC through macrophage-derived CCL2, we next investigated whether inhibiting JNK signaling enhances the antitumor efficacy of macrophage-targeting agents in mouse models. Compared with monotherapies, BMS-986360 or JNK-IN-8 plus pexidartinib or sotuletinib significantly reduced tumor growth in immunocompetent syngeneic TNBC models (E0771, PyMT-M, and 4T1.2; P < 0.01). Furthermore, compared with monotherapies, JNKi combined with pexidartinib reduced counts of Tregs and M2 macrophages and increased counts of M1 macrophages and cytotoxic CD8+ T cells in both E0771 and PyMT-M tumors. These results suggest that combination of JNKi with macrophage-targeting agents reduces tumor growth by inducing an immunoactive TME. Conclusion: JNKi can potentiate macrophage-targeting agents in TNBC by inducing an immunoactive TME. This finding highlights the need to conduct further studies of combining JNKi with immunotherapy to capitalize on JNK’s immune regulation of the TME and the potency of JNKi in synergistically improving the efficacy of immunotherapy for TNBC. Citation Format: Xuemei Xie, Takashi Semba, Young J. Gi, Yujia Qin, Ba Thong Nguyen, Bharat S. Kautal, Youping Deng, Jangsoon Lee, Naoto T. Ueno. Inhibiting JNK signaling enhances the antitumor efficacy of macrophage-targeting agents in triple-negative breast cancer by inducing an immunoactive tumor microenvironment [abstract]. In: Proceedings of the San Antonio Breast Cancer Symposium 2024; 2024 Dec 10-13; San Antonio, TX. Philadelphia (PA): AACR; Clin Cancer Res 2025;31(12 Suppl):Abstract nr P4-03-22.

Background: Given the heterogeneity of TNBC, identification of biologically and clinically distinct TNBC subtypes with unique therapeutic vulnerabilities is critically needed. Preclinically, DNMT3A protein expression is predictive of sensitivity to DNMT inhibitors (DNMTi). Additionally, treatment with DNMTi enhances antitumor immune responses. Ongoing (NCT05673200) and completed (NCT02957968) trials are evaluating the addition of DNMTi to chemoimmunotherapy for TNBC. However, it is unknown whether DNMT3A expression is associated with distinct TNBC biology or clinical outcomes. Methods: We measured DNMT3A protein using immunohistochemistry in early-stage TNBC tumors and evaluated its association with clinicopathologic characteristics, gene expression (RNASeq), spatial tumor immune microenvironment features (52 proteins using Nanostring GeoMx® Digital Spatial Profiler [DSP]), and recurrence-free survival (RFS). DNMT3A expression was categorized as negative (no staining) or positive (high: moderate/strong staining in >25% of nuclei; low/intermediate: weak staining in any % nuclei or moderate/strong staining in ≤25% of nuclei). Results: Among 345 patients, the median age was 54 years; most tumors were > 2 cm (53%) and N0 (62%). 281 (81%) were DNMT3A+ (high: 127 [37%], low/intermediate: 154 [45%]). DNMT3A+ TNBC were more often grade 3 (94% vs 86%, p = 0.037), with Ki-67 >15% (85% vs 67%, p = 0.002), and N+ (40% vs 26%, p= 0.043). There were no differences in age, menopausal status, tumor size, PD-L1 or stromal tumor-infiltrating lymphocytes (sTILs) between DNMT3A+ or DNMT3A- TNBC. To assess the impact of DNMT3A on the natural history of early TNBC, we focused our survival analyses on a subset of 103 (30%) patients who did not receive systemic therapy. DNMT3A expression was associated with worse RFS on a multivariable model adjusting for tumor size, nodal status, and sTILs (aHR 4.6, 95% CI 1.05-19.96, p=0.043; univariable results: HR 3.3; 95% CI: 0.79, 13.99; p = 0.102). The 5-year RFS (95% CI) of DNMT3A+ TNBC without chemotherapy was 66% (53-76), compared with 93% (59-99) for DNMT3A- TNBC. While TNBC with sTILs ≥ 30% had favorable 5-year RFS regardless of DNMT3A expression (86% DNMT3A+ vs 100% DNMT3A-), TNBC with sTILs <30% had particularly poor outcomes when also expressing DNMT3A (5y RFS 54% [38-67] for sTIL low and DNMT3A+ TNBC vs 90% [47-99] for sTIL low and DNMT3A- TNBC). RNA-Seq data (available for 154 [63%] of 345 tumors, regardless of receipt of chemotherapy) revealed differential expression of 234 genes between DNMT3A+ (n=126) and DNMT3A- (n=28) TNBC (e.g. MUC19, CEACAM5, and ZNF606, among others). GeoMx® DSP data (available for 289 [84%] tumors) showed that DNMT3A+ TNBC exhibited lower expression of CD3, CD4, CD8 (T cells) and granzyme B in the tumor compartment; lower expression of HLA-DR (antigen presentation), CD56 (NK cell), FAP alpha, SMA (fibroblasts) and CTLA4 (immune checkpoint) in the stromal compartment; and lower expression of CD127 (memory T cell), CD11c, CD14, CD68 (myeloid/macrophages), CD34 (stem cell), EpCAM and PanCK (epithelium), fibronectin (extracellular matrix) and TIM3 (immune checkpoint) in both the tumor and stromal compartments. Expression of Ki-67 was higher in both the tumor and stromal compartments of DNMT3A+ TNBC. Conclusions: DNMT3A expression was associated with worse clinical outcomes in TNBC (particularly in sTIL-low tumors), higher tumor proliferation, and diverging gene expression and tumor immune microenvironment profiles. While no differences in sTILs were noted by H&E quantification according to DNMT3A status, GeoMX DSP analyses revealed an immunosuppressed tumor and stromal microenvironment in DNMT3A+ tumors, characterized by decreased T, NK, myeloid, and antigen presentation markers. These data support targeting DNMT along with immunotherapy in DNMT3A+ TNBC as is currently being evaluated in NCT05673200. Citation Format: Roberto Leon-Ferre, David M Zahrieh, Sarah K Reed, Jodi M Carter, Saba Yasir, David Hillman, Judy C Boughey, Krishna Kalari, Peter C Lucas, Saranya Chumsri, Jennifer M Kachergus, Yi Liu, E. Aubrey Thompson, Harry D Bear, Fergus J Couch, James N Ingle, Liewei Wang, Matthew P Goetz. DNA methyltransferase 3A (DNMT3A) protein expression in triple-negative breast cancer (TNBC): Impact on clinical outcomes, gene expression, and tumor microenvironment [abstract]. In: Proceedings of the San Antonio Breast Cancer Symposium 2024; 2024 Dec 10-13; San Antonio, TX. Philadelphia (PA): AACR; Clin Cancer Res 2025;31(12 Suppl):Abstract nr PS12-07.

The clinical potency of anti-programmed death-ligand 1 (PD-L1) therapy in metastatic triple-negative breast cancer (TNBC) is modest primarily because of the intrinsic low immunogenicity and an immunosuppressive tumor microenvironment (TME). Photodynamic therapy (PDT), an inducer of immunogenic cell death (ICD), has the potential to enhance antitumor immune response and improve PD-L1 blockade efficacy. DTP, a novel photosensitizer developed previously, has demonstrated potent ROS-dependent photocytotoxicity, yet its immunomodulatory effects remain unexplored. This study investigated the induction of ICD and dendritic cell (DC) maturation following DTP-PDT in vivo and in vitro. A bilateral TNBC model was developed to assess the efficacy of DTP-PDT combined with α-PD-L1 therapy on untreated distant tumors and to explore its potential immunological mechanisms. The results showed that DTP-PDT effectively induced ICD, demonstrated by calreticulin membrane exposure, high mobility group box 1 protein release, and increased secretion of interferon-γ and tumor necrosis factor-α, resulting in DC maturation. The combination of DTP-PDT and α-PD-L1 significantly inhibited distant tumor growth. This effect was associated with increased CD8+ and CD4+ T cells infiltration, and reduced numbers of regulatory T cells, in the distant tumor and spleen. In conclusion, DTP-PDT enhanced TNBC sensitivity to α-PD-L1 by inducing ICD, and its combination withα-PD-L1 could remodel the immunosuppressive TME and enhance systemic immunity, resulting in a therapeutic effect against distant metastasis. This study provides experimental validation for a combined strategy of DTP-PDT and α-PD-L1, proposing a potential therapeutic approach for metastatic TNBC.

Background: Hepatocellular carcinoma (HCC) causes a significant mortality burden worldwide. Radiotherapy (RT) is the primary locoregional treatment modality for HCC. However, the efficacy of RT in HCC is limited by tumor microenvironment (TME) hypoxia, immunosuppression, and extracellular matrix (ECM) stiffness.Methods: We developed a novel RGD-modified liposomal platform (RGD@LP-Y) that encapsulates the ROCK inhibitor Y-27632 through thin-film hydration. We characterized the RGD@LP-Y by the transmission electron microscope (TEM), UV-Vis spectrophotometer, and dynamic light scattering instrument (DLS). A high-stiffness hydrogel co-culture system mimicking mechanical TME was established to explore the role of RGD@LP-Y on matrix stiffness remodeling. In vitro evaluations included cytotoxicity, reactive oxygen species (ROS) generation, mitochondrial function, immunogenic cell death (ICD) markers, and immune cell activation. Mechanistic investigations encompassed matrix stiffness regulation analysis, flow cytometry profiling of pro-inflammatory macrophages, dendritic cell (DC) maturation, transcriptome sequencing, and western blotting. In vivo validation used xenograft models treated with intravenous RGD@LP-Y and localized RT. Biosafety was confirmed through organ histology, serum biochemistry analysis, and hemolysis assay.Results: RGD@LP-Y downregulated matrix stiffness markers (YAP/COL1) and activated PI3K/AKT/NF-κB signaling to drive pro-inflammatory macrophage polarization and DC maturation. The synergistic effects were observed in combination with RT. The treatment of RGD@LP-Y and RT inhibited HCC proliferation, induced apoptosis, suppressed mitochondrial respiration, elevated intracellular ROS, and thus enhanced ICD. In vivo, RGD@LP-Y+RT demonstrated potent tumor suppression and immune activation without systemic toxicity.Conclusion: RGD@LP-Y enhances RT sensitivity by remodeling ECM stiffness, modulating the hypoxia and immunosuppressive conditions within TME, and enhancing the ICD. The study provides a safe combinatorial approach for HCC therapy.

Hypoxia is known to stimulate reactive oxygen species (ROS) generation. Because reduced glutathione (GSH) is compartmentalized in cytosol and mitochondria, we examined the specific role of mitochondrial GSH (mGSH) in the survival of hepatocytes during hypoxia (5% O2). 5% O2 stimulated ROS in HepG2 cells and cultured rat hepatocytes. Mitochondrial complex I and II inhibitors prevented this effect, whereas inhibition of nitric oxide synthesis with Nomega-nitro-L-arginine methyl ester hydrochloride or the peroxynitrite scavenger uric acid did not. Depletion of GSH stores in both cytosol and mitochondria enhanced the susceptibility of HepG2 cells or primary rat hepatocytes to 5% O2 exposure. However, this sensitization was abrogated by preventing mitochondrial ROS generation by complex I and II inhibition. Moreover, selective mGSH depletion by (R,S)-3-hydroxy-4-pentenoate that spared cytosol GSH levels sensitized rat hepatocytes to hypoxia because of enhanced ROS generation. GSH restoration by GSH ethyl ester or by blocking mitochondrial electron flow at complex I and II rescued (R,S)-3-hydroxy-4-pentenoate-treated hepatocytes to hypoxia-induced cell death. Thus, mGSH controls the survival of hepatocytes during hypoxia through the regulation of mitochondrial generation of oxidative stress.

Rationale: Iron-saturated Lf (Holo-Lactoferrin, Holo-Lf) exhibits a superior anticancer property than low iron-saturated Lf (Apo-Lf). Ferroptosis is an iron-dependent cell death characterized by the accumulation of lipid peroxidation products and lethal reactive oxygen species (ROS). Radiotherapy also exerts its therapeutic effect through ROS.Methods: The effect of different iron-saturated Lf on ferroptosis and radiotherapy were tested on triple-negative breast cancer (TNBC) cell line MDA-MB-231 and non-TNBC cell line MCF-7.Results: Holo-Lf significantly increased the total iron content, promoted ROS generation, increased lipid peroxidation end product, malondialdehyde (MDA), and enhanced ferroptosis of MDA-MB-231 cells. By contrast, Apo-Lf upregulated SLC7a11 expression, increased GSH generation and inhibited ferroptosis of MDA-MB-231 cells. However, non-TNBC MCF-7 cells were resistant to Holo-Lf-induced ferroptosis because MCF-7 cells have a higher redox balance capacity than MDA-MB-231 cells. More importantly, Holo-Lf downregulated HIF-1α expression, ameliorated the hypoxia microenvironment in subcutaneous MDA-MB-231 tumors, and promoted radiation-induced DNA damage to hypoxic MDA-MB-231 cells. Finally, the efficacy of radiotherapy to MDA-MB-231 tumors was enhanced by Holo-Lf.Conclusion: Holo-Lf could induce ferroptosis in MDA-MB-231 cells and sensitize MDA-MB-231 tumors to radiotherapy.

e13139 Background: Triple-negative breast cancer (TNBC) is an aggressive subtype with no specific molecular targets and an immunosuppressive tumor microenvironment (TME). While immune checkpoint inhibitors (ICIs) show promise, their effectiveness is limited by immune evasion mechanisms. Photodynamic therapy (PDT) induces localized tumor cell death but has suboptimal effects on the immune system and TME. We developed AuC@SCOF-Mn, a novel nanoenzyme, to enhance PDT efficacy and activate antitumor immunity. Methods: AuC@SCOF-Mn is a multifunctional nanoenzyme composed of gold nanoclusters (AuNCs) integrated into the covalent organic framework (COF) with metallosalphen Mn centers. The AuNCs generate reactive oxygen species (ROS) under light exposure, enhancing PDT, while Mn centers catalyze ROS generation and modulate the immune response. In vitro , we evaluated the ability of AuC@SCOF-Mn to induce glutathione depletion, mitochondrial dysfunction, DNA damage, and apoptosis in 4T1 tumor cells. In vivo , its impact on tumor growth, immune cell infiltration, and recurrence was assessed in the TNBC mouse models. RNA-seq was conducted to reveal potential mechanisms. The potential for synergistic immune activation when combined with ICIs to against lung metastasis was also explored. Results: AuC@SCOF-Mn enhanced the ability of PDT to deplete GSH, inducing mitochondrial dysfunction as well as DNA damage, leading to extensive apoptosis of cancer cells in vitro . Besides, AuC@SCOF-Mn significantly inhibited tumor growth and enhanced CD8 + T cell and dendritic cell (DC) infiltration into the TME under light irradiation in vivo . It also polarized macrophages toward the pro-inflammatory M1 phenotype and reduced immunosuppressive regulatory T cells (T regs ) and myeloid-derived suppressor cells (MDSCs). RNA-seq revealed the significant upregulation of the cytokines-cytokines interaction signaling pathway to promote immune cell chemotaxis. The pro-inflammatory cytokine levels in both the TME and circulation were higher in the AuC@SCOF-Mn and PDT combined group than in the control and PDT-only groups, which helped inhibit tumor recurrence by long-term antitumor immunity. Moreover, combined with ICIs, AuC@SCOF-Mn suppressed lung metastasis via improving systemic immune responses in metastatic TNBC mice models. Furthermore, AuC@SCOF-Mn showed limited toxicity to the main organs and circulation system. Conclusions: The findings of this study suggest AuC@SCOF-Mn, as a promising nanoenzyme, can enhance PDT effects and activate the potent immune response to inhibit TNBC growth and recurrence with good biocompatibility. Its ability to promote immune cell infiltration and synergize with ICIs provides a novel strategy to improve metastatic TNBC outcomes with the potential for clinical translation.

Triple-negative breast cancer (TNBC), known for its therapeutic challenges and high metastatic potential, is commonly treated with ionizing radiation (IR). In the tumor microenvironment (TME) of TNBC, macrophages occupy a place and are reprogrammed into two main types, M1 and M2, in response to different stimuli such as IR or various cytokines. However, the detailed molecular mechanism remains to be elucidated. Using our cell model, we applied IR dose-dependently (0.5/8 Gy) to observe macrophage polarization and found that high-dose IR could increase the proportion of M1 macrophages. These trends are consistent with polarization by LPS (M1) and IL-4/IL-13 (M2), respectively. By integrating transcriptomics and cytokine array profiling, we demonstrated that IKAROS family zinc finger 1 (IKZF1), a transcription factor involved in DNA repair, downregulates secretion of the downstream target, CCL5, in macrophages under high-dose IR exposure. In the collected conditioned medium, the high-dose IR group could significantly inhibit the migration and proliferation of TNBC cells. Even, recombinant CCL5 protein was recruited to modulate the TME of TNBC, and the migration ability of TNBC cells was reversed. In the direct co-culture system, phagocytic event was enhanced by high-dose IR. In conclusion, under high-dose irradiation conditions, IKZF1 is inhibited and CCL5 secretion is reduced. The TME environment is conducive to polarization of the M1 subtype and enhances its ability to phagocytose TNBC cells, thereby inhibiting TNBC progression. This finding suggests that the benefits of radiotherapy in TNBC may also be linked to a new chapter in immunotherapy. Citation Format: Kuo Han-Hsi, Yu-Chan Chang. Radiation supports the polarization of M1 macrophages against triple-negative breast cancer cells via the IKZF1 - CCL5 axis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 3289.

Triple negative breast cancer (TNBC) exhibits limited responsiveness to immunotherapy owing to its immunosuppressive tumor microenvironment (TME). Here, a reactive oxygen species (ROS)-labile nanodrug encapsulating the photosensitizer Ce6 and Bcl-2 inhibitor ABT-737 was developed to provoke a robust immune response via the synergistic effect of photodynamic therapy (PDT) and the reversal of apoptosis resistance. Upon exposure to first-wave near-infrared laser irradiation, the generated ROS triggers PEG cleavage, facilitating the accumulation of the nanodrug at tumor region and endocytosis by tumor cells. Further irradiation leads to the substantial generation of cytotoxic ROS, initiating an immunogenic cell death (ICD) cascade, which prompts the maturation of dendritic cells (DCs) as well as the infiltration of T cells into the tumor site. Meanwhile, Bcl-2 inhibition counteracts apoptosis resistance, thereby amplifying PDT-induced ICD and bolstering antitumor immunity. As a result, the ROS-sensitive nanodrug demonstrates a potent inhibitory effect on tumor growth.

Background Triple-negative breast cancer (TNBC) is a subtype of breast cancer with limited therapeutic options. Eupalinolide O (EO) was reported to inhibit tumor growth. This study is aimed at exploring the role of EO on TNBC both in vivo and in vitro. Methods. In in vitro experiments, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and clonogenic assay were conducted to measure the impact of EO on TNBC cell growth at different concentrations and time points. Flow cytometry was conducted to evaluate cell apoptosis. Mitochondrial membrane potential (MMP) loss, caspase-3 activity, and reactive oxygen species (ROS) generation were assessed. The expressions of apoptosis-related mRNAs and Akt/p38 MAPK signaling pathway-related proteins were measured. In in vivo experiments, by injecting TNBC cells into the nude mice to induce xenograft tumor, mice were treated with EO for 20 days. Then, in vivo bioluminescence imaging system was utilized to monitor the growth and distribution of TNBC cells. Tumor volume and weight were also recorded. Hematoxylin-eosin (HE) staining and ELISA assay were applied to observe tumor tissue morphology and ROS levels. Furthermore, western blotting was conducted to observe the expression of apoptosis-related proteins and Akt/p38 MAPK signaling pathway-associated proteins. Results EO inhibited the cell viability and proliferation of TNBC cells but not normal epithelial cells. Furthermore, EO induced apoptosis, decreased MMP, and elevated caspase-3 activity and ROS content in TNBC cells. Meanwhile, the expression of apoptosis-related mRNAs and Akt/p38 MAPK pathway-related proteins was regulated by EO treatment. Besides, in vivo experiments demonstrated EO not only suppressed tumor growth, Ki67 expression, ROS generation, and Akt phosphorylation but also upregulated caspase-3 expression and p-38 phosphorylation. Conclusion EO may induce cell apoptosis in TNBC via regulating ROS generation and Akt/p38 MAPK pathway, indicating EO may be a candidate drug for TNBC.

The tumor microenvironment (TME) of typical tumor types such as triple-negative breast cancer is featured by hypoxia and immunosuppression with abundant tumor-associated macrophages (TAMs), which also emerge as potential therapeutic targets for antitumor therapy. M1-like macrophage-derived exosomes (M1-Exos) have emerged as a promising tumor therapeutic candidate for their tumor-targeting and macrophage-polarization capabilities. However, the limited drug-loading efficiency and stability of M1-Exos have hindered their effectiveness in antitumor applications. Here, a hybrid nanovesicle is developed by integrating M1-Exos with AS1411 aptamer-conjugated liposomes (AApt-Lips), termed M1E/AALs. The obtained M1E/AALs are loaded withperfluorotributylamine (PFTBA)and IR780, as P-I, to construct P-I@M1E/AALs for reprogramming TME by alleviating tumor hypoxia and engineering TAMs. P-I@M1E/AAL-mediated tumor therapy enhances the in situ generation of reactive oxygen species, repolarizes TAMs toward an antitumor phenotype, and promotes the infiltration of T lymphocytes. The synergistic antitumor therapy based on P-I@M1E/AALs significantly suppresses tumor growth and prolongs the survival of 4T1-tumor-bearing mice. By integrating multiple treatment modalities, P-I@M1E/AAL nanoplatform demonstrates a promising therapeutic approach for overcoming hypoxic and immunosuppressive TME by targeted TAM reprogramming and enhanced tumor photodynamic immunotherapy. This study highlights an innovative TAM-engineering hybrid nanovesicle platform for the treatment of tumors characterized by hypoxic and immunosuppressive TME.

Pancreatic cancer (PC) remains difficult to treat due to its dense extracellular matrix (ECM), immunosuppressive tumor microenvironment (TME), and deep‐seated anatomy. To address these challenges, we developed IR&ZnPc@LNP‐NO, an ultrasound (US)‐responsive lipid nanosonosensitizer that synergizes sonodynamic therapy (SDT), chemotherapy, and immunotherapy for orthotopic PC. IR&ZnPc@LNP‐NO undergoes three key US‐activated responses: 1) size reduction, 2) controlled release of irinotecan (IR) and nitric oxide (NO), and 3) generation of reactive oxygen species (ROS). Under low‐dose US, IR&ZnPc@LNP‐NO reduces in size (from ∼120 to ∼40 nm), enhancing tumor penetration, and releases NO to remodel the TME by normalizing vasculature and degrading ECM. This enhances nanosonosensitizers accumulation and cytotoxic T cells (CTLs) infiltration. High‐dose US irradiation triggers the generation of cytotoxic ROS, which, in combination with IR‐mediated chemotherapy, induces immunogenic cell death (ICD) and enhances antitumor immunity. Additionally, combining IR&ZnPc@LNP‐NO with PD‐L1 antibody (αPD‐L1) immunotherapy significantly prolongs survival in orthotopic PC models. The cascade strategy—size reduction, TME remodeling, and multimodal therapy—effectively overcomes stromal and immunosuppressive barriers, offering a robust platform for treating deep‐seated PC.

Pancreatic cancer (PC) remains difficult to treat due to its dense extracellular matrix (ECM), immunosuppressive tumor microenvironment (TME), and deep-seated anatomy. To address these challenges, we developed IR&ZnPc@LNP-NO, an ultrasound (US)-responsive lipid nanosonosensitizer that synergizes sonodynamic therapy (SDT), chemotherapy, and immunotherapy for orthotopic PC. IR&ZnPc@LNP-NO undergoes three key US-activated responses: 1) size reduction, 2) controlled release of irinotecan (IR) and nitric oxide (NO), and 3) generation of reactive oxygen species (ROS). Under low-dose US, IR&ZnPc@LNP-NO reduces in size (from ∼120 to ∼40nm), enhancing tumor penetration, and releases NO to remodel the TME by normalizing vasculature and degrading ECM. This enhances nanosonosensitizers accumulation and cytotoxic T cells (CTLs) infiltration. High-dose US irradiation triggers the generation of cytotoxic ROS, which, in combination with IR-mediated chemotherapy, induces immunogenic cell death (ICD) and enhances antitumor immunity. Additionally, combining IR&ZnPc@LNP-NO with PD-L1 antibody (αPD-L1) immunotherapy significantly prolongs survival in orthotopic PC models. The cascade strategy-size reduction, TME remodeling, and multimodal therapy-effectively overcomes stromal and immunosuppressive barriers, offering a robust platform for treating deep-seated PC.