Jinghua Ren

Microbial cell therapies hold considerable promise as programmable and versatile modalities for targeted interventions in complex biological environments. Here, we developed a living bacterial delivery platform that could leverage its endogenous metabolism to synchronize the release of surface-anchored targeted protein degradation (TPD) chimeras and the secretion of immune-modulatory nanobodies (Nbs) for enhanced antitumor efficacy. By means of metabolic labeling coupled with bioorthogonal click chemistry, transferrin (Tf)-CD24 antibody chimeras (TransCACs) were covalently displayed on the surface of nonpathogenic Escherichia coli (E. coli) K12. In parallel, this strain was equipped with a constitutive expression module for the in situ biosynthesis of PD-L1-blocking nanobodies. Capitalizing on the natural tumor tropism of bacteria, our engineered E. coli K12 achieved tumor-targeted CD24 degradation, thereby augmenting macrophage-mediated phagocytosis and synergizing with PD-L1 blockade to elicit robust tumor-specific CD8+ T cell immunity. In vivo administration of engineered microbes led to marked tumor growth inhibition in both subcutaneous breast and orthotopic hepatocellular carcinoma models, along with prolonged animal survival, driven by remodeling of the suppressive tumor microenvironment through coordinated crosstalk between M1-like macrophages and tumor-resident memory (TRM)-like CD8+ T cells. Altogether, this integrated genetic engineering and metabolic labeling of bacteria (InGeM) opens avenues for the development of next-generation microbe-based cancer immunotherapies.

Background/Objectives: Ovarian cancer is the most lethal gynecologic malignancy, largely due to late diagnosis and the high prevalence of malignant ascites, a hallmark of advanced disease that is difficult to control and contributes to immune suppression and treatment failure. Despite advances in standard care, durable responses are rare. This study investigates a novel immunotherapeutic strategy designed to overcome the suppressed peritoneal microenvironment using an oncolytic vaccinia virus engineered to express a decoy-resistant IL-18 mutein. Methods: We generated a vaccinia virus (vvDD-nsmDR-18) expressing a non-secreted, mature, decoy-resistant IL-18. Viral expression was validated via RT-qPCR and fluorescence microscopy, while cytotoxicity was confirmed using CCK-8 assays. The antitumor efficacy of vvDD-nsmDR-18 was evaluated in the aggressive murine ID8a ovarian cancer model. The underlying mechanisms of action were investigated using flow cytometry and transcriptional profiling. Results: Treatment with vvDD-nsmDR-18 significantly prolonged survival and was associated with reduced abdominal distension consistent with decreased ascites burden. Immune analyses indicated enhanced T cell activation across multiple anatomical compartments, including tumors, peritoneal cavity, and spleens, the latter recently suggested to serve as a reservoir for tumor-reactive T cells. This systemic activation was characterized by increased IFN-γ and perforin expression. In addition, vvDD-nsmDR-18 treatment was associated with expansion of CD39⁺CD103⁺CD8⁺ tumor-reactive T cells and a shift toward a lower PD-1 expression phenotype within this population. Conclusions: These findings demonstrate that nsmDR-18-expressing oncolytic viruses can remodel the immunosuppressive landscape of advanced ovarian cancer, suggesting this approach is a promising candidate for further clinical development.

Background: Cancer immunotherapy has revolutionized treatment, offering new hope for patients with previously hard-to-treat cancers. However, many solid tumors are classified as non-inflamed, meaning they are less responsive to immunotherapies. This underscores the need for novel strategies to transform these non-inflamed tumors into immune-inflamed ones. Oncolytic viruses, particularly when engineered, hold promise for stimulating immune responses and transforming these tumors. Cytokines, in particular, are promising for arming these viruses. Immune checkpoint inhibition, especially targeting the programmed cell death protein-1 (PD-1)/programmed death-ligand 1 (PD-L1) pathway, has been groundbreaking. In this study, we hypothesize that an oncolytic vaccinia virus (oVV) expressing a fusion protein of the PD-1 ectodomain and interleukin (IL)-2 (vvDD-PD-1ecto-R-IL-2) could mimic the combined effects of anti-PD-L1 antibodies and IL-2, transforming non-inflamed tumors and enhancing antitumor effects while minimizing the toxicity associated with systemic IL-2. Methods: We generated the vvDD-PD-1ecto-R-IL-2 oncolytic virus and confirmed transgene expression through reverse transcription quantitative PCR (RT-qPCR), ELISA and flow cytometry. The antitumor efficacy and safety of vvDD-PD-1ecto-R-IL-2 were evaluated in murine tumor models. The underlying mechanisms of action were investigated using RT-qPCR, ELISA and flow cytometry. Results: Our data demonstrate that the vvDD-PD-1ecto-R-IL-2 elicited significant antitumor effects, though the PD-1 ectodomain did not fully mimic the function of anti-PD-L1 antibodies, likely due to the elevated PD-L1 levels following oVV infection. However, PD-1 successfully directed IL-2 to associate with the cell membrane via PD-1ecto-R-IL-2/PD-L1 interaction, minimizing systemic toxicity. The accumulated IL-2 in the tumor microenvironment led to an increase in antitumor factors, including interferon (IFN)-γ, granzyme B, and perforin, while reducing protumor factors such as vascular endothelial growth factor and prostaglandin-endoperoxide synthase 2. Furthermore, treatment with the vvDD-PD-1ecto-R-IL-2 promoted the expansion of tumor-infiltrating stem-like CD8 ⁺ T cells (CD8 ⁺ TCF-1 ⁺ ), as well as functional CD8 ⁺ T-cell subsets, including IFN-γ ⁺ CD8 ⁺ , GzmB ⁺ CD8 ⁺ , CD107a ⁺ CD8 ⁺ , and perforin ⁺ CD8 ⁺ , and reduced the presence of severely exhausted PD-1 ⁺ TIM-3 ⁺ CD8 ⁺ T cells, PD-1 ⁺ LAG-3 ⁺ CD8 ⁺ T cells and PD-1 ⁺ TIGIT ⁺ CD8 ⁺ T cells, thereby transforming immunologically non-inflamed tumors into immune-inflamed tumors. Conclusions: Our findings suggest that PD-1-directed IL-2 delivered via oVV is both safe and effective, making it a promising candidate for clinical translation.

Glucose consumption by tumors induces metabolic restriction of T cells, which results in immune evasion and tumor progression. Regulating cellular metabolism represents a promising strategy to enhance cancer immunotherapy; however, redirecting glucose utilization from tumor cells to T cells is challenging. Herein, the activation of cytotoxic T cells using engineered peptide coacervates (PCs) containing interferon alpha (IFNα) and membranized with metal–phenolic networks (MPNs) (PC‐IFNα@MPNs), which promote glucose uptake and glycolysis, is reported. PC‐IFNα@MPNs modulate the molecular conformation of the co‐stimulatory lymphocyte function‐associated antigen 1 on CD8 ⁺ T cells, while suppressing tumor cell glycolysis through the sustained release of IFNα, thereby increasing the energy supply for T cells. Furthermore, PC‐IFNα@MPNs suppress tumor progression in preclinical orthotopic tumor mouse models by facilitating T cell infiltration and activation. When combined with immune checkpoint blockade (ICB), PC‐IFNα@MPNs further improve therapeutic outcomes (99% inhibition of tumor growth), even in ICB‐insensitive tumor models. Notably, PC‐IFNα@MPNs exert a robust immune‐memory effect (a 4.8‐fold increase in memory T cells) and provide long‐lasting anti‐tumor activity (over 74 days), thereby preventing postsurgical tumor recurrence. The present study offers insights into metabolic intervention mechanisms mediated by glucose modulation and provides a rational design for metal–organic materials in cancer therapy.

Radiotherapy is a common treatment option for patients with glioblastoma multiforme. However, tumor heterogeneity causes varying responses to radiation among different tumor subpopulations. Cancer cells that endure radiotherapy exhibit radioresistance, resulting in the ineffectiveness of radiation therapy and eventual tumor relapse. In this study, we discovered that the fibroblast growth factor-inducible 14 (Fn14)-positive tumor cells were enriched in tumor residual foci after radiation, ultimately leading to treatment failure. Fn14-expressing glioma cells survived ionizing radiation through preferential activation of DNA damage checkpoint response. We have thus engineered an Fn14-targeting and NIR-II responsive plasmonic gold nanosystem named Fn14-AuNPs, which can precisely internalize into Fn14-overexpressed glioma cells and have an excellent BBB-crossing capability. As gold nanoparticles, by inhibition of DNA repair processes and induction of G₂/M cells cycle arrest, Fn14-AuNPs nanoparticles improved the radiosensitivity of tumor cells. Meanwhile, Fn14-AuNPs induced localized heat under NIR-II photoirradiation, thus impeding RT-induced DNA damage checkpoint response. This versatile nanosensitizer, combined with NIR-II laser photoirradiation, can eradicate radioresistant subpopulations of glioblastoma and improve the therapeutic effect of radiotherapy. This finding presents an effective radiosensitization strategy by targeting radioresistant subpopulations, which can efficiently overcome the constraints imposed in clinical radiotherapy and offer a hopeful avenue to enhance the treatment effectivity of radiotherapy in glioblastoma.

Neutrophil extracellular traps (NETs), which consist of chromatin DNA studded with granule proteins, are released by neutrophils in response to both infectious and sterile inflammation. Beyond the canonical role in defense against pathogens, the extrusion of NETs also contributes to the initiation, metastasis, and therapeutic response of malignant diseases. Recently, NETs have been implicated in the development and therapeutic responses of various types of tumors. Although extensive work regarding inflammation in tumors has been reported, a comprehensive summary of how these web‐like extracellular structures initiate and propagate tumor progression under the specific microenvironment is lacking. In this review, we demonstrate the initiators and related signaling pathways that trigger NETs formation in cancers. Additionally, this review will outline the current molecular mechanisms and regulatory networks of NETs during dormant cancer cells awakening, circulating tumor cells (CTCs) extravasation, and metastatic recurrence of cancer. This is followed by a perspective on the current and potential clinical potential of NETs as therapeutic targets in the treatment of both local and metastatic disease, including the improvement of the efficacy of existing therapies.

The success of cancer immunotherapy is largely associated with immunologically hot tumors. Approaches that promote the infiltration of immune cells into tumor beds are urgently needed to transform cold tumors into hot tumors. Oncolytic viruses can transform the tumor microenvironment (TME), resulting in immunologically hot tumors. Cytokines are good candidates for arming oncolytic viruses to enhance their function in this transformation. Here, we used the oncolytic vaccinia virus (oVV) to deliver interleukin-9 (IL-9) into the tumor bed and explored its antitumor effects in colon and lung tumor models. Our data show that IL-9 prolongs viral persistence, which is probably mediated by the up-regulation of IL-10. The vvDD-IL-9 treatment elevated the expression of Th1 chemokines and antitumor factors such as IFN-γ, granzyme B, and perforin. IL-9 expression increased the percentages of CD4⁺ and CD8⁺ T cells in the TME and decreased the percentage of oVV-induced immune suppressive myeloid-derived suppressor cells (MDSC), leading to potent antitumor effects compared with parental virus treatment. The vvDD-IL-9 treatment also increased the percentage of regulatory T cells (Tregs) in the TME and elevated the expression of immune checkpoint molecules such as PD-1, PD-L1, and CTLA-4, but not GITR. The combination therapy of vvDD-IL-9 and the anti-CTLA-4 antibody, but not the anti-GITR antibody, induced systemic tumor-specific antitumor immunity and significantly extended the overall survival of mice, indicating a potential translation of the IL-9-expressing oncolytic virus into a clinical trial to enhance the antitumor effects elicited by an immune checkpoint blockade for cancer immunotherapy.

Radiotherapy is identified as a crucial treatment for patients with glioblastoma, but recurrence is inevitable. The efficacy of radiotherapy is severely hampered partially due to the tumor evolution. Growing evidence suggests that proneural glioma stem cells can acquire mesenchymal features coupled with increased radioresistance. Thus, a better understanding of mechanisms underlying tumor subclonal evolution may develop new strategies. Herein, data highlighting a positive correlation between the accumulation of macrophage in the glioblastoma microenvironment after irradiation and mesenchymal transdifferentiation in glioblastoma are presented. Mechanistically, elevated production of inflammatory cytokines released by macrophages promotes mesenchymal transition in an NF‐κB‐dependent manner. Hence, rationally designed macrophage membrane‐coated porous mesoporous silica nanoparticles (MMNs) in which therapeutic anti‐NF‐κB peptides are loaded for enhancing radiotherapy of glioblastoma are constructed. The combination of MMNs and fractionated irradiation results in the blockage of tumor evolution and therapy resistance in glioblastoma‐bearing mice. Intriguingly, the macrophage invasion across the blood‐brain barrier is inhibited competitively by MMNs, suggesting that these nanoparticles can fundamentally halt the evolution of radioresistant clones. Taken together, the biomimetic MMNs represent a promising strategy that prevents mesenchymal transition and improves therapeutic response to irradiation as well as overall survival in patients with glioblastoma.