Curcumin Enhanced Ionizing Radiation-Induced Immunogenic Cell Death in Glioma Cells through Endoplasmic Reticulum Stress Signaling Pathways.

Objective: Local radiotherapy may cause distant tumor regression via inducing immunogenic cell death (ICD). Here, we investigated the effect of curcumin on ionizing radiation-induced immunogenic cell death in normoxic or hypoxic glioma cells and its mechanism in vitro and vivo. Methods: Hypoxic or normoxic glioma cell apoptosis and the cell surface exposure of calreticulin (CRT) were detected by flow cytometry. Extracellular ATP and HSP70 were measured by chemiluminescence assay and ELISA, respectively. Endoplasmic reticulum (ER) stress protein levels were detected by western blot. Moreover, the induction of bona fide ICD was detected by vaccination assays in mice bearing glioma model. Spleen lymphocytes and tumor-infiltrating lymphocyte subsets were analyzed by flow cytometry and immunohistochemistry. Results: Curcumin incubation before X-ray irradiation significantly increased radiation-induced apoptosis rate in normoxic or hypoxic glioma cells. Curcumin enhanced radiation-induced CRT exposure, release of HSP70 and ATP, and ER stress signaling activity. After treatment with ER stress pathway inhibitors, cell apoptosis and CRT exposure induced by the combination treatment of curcumin and X-ray were reduced. In vaccination experiments, glioma cells irradiated by X-ray produced a strong immunogenic response rejecting tumor formation in 70% mice. In comparison, cells treated by curcumin and X-ray produced a stronger immune response rejecting tumor formation in 90% mice. The combination treatment increased the percentage of tumor-infiltrating CD4+, CD8+ T lymphocytes, and CD11c+ dendritic cells compared to X-ray irradiation alone. Conclusion: Ionizing radiation-induced normoxic or hypoxic glioma immunogenic cell death could be further enhanced by curcumin through activating the ER stress PERK-eIF2α and IRE1α-XBP1 signaling pathways.

Bip inhibition in glioma stem cells promotes radiation-induced immunogenic cell death.

Tumor regression in sites distant to the irradiated field are thought to be associated with emission of damage-associated molecular patterns (DAMPs) molecules and generation of immunogenic cell death (ICD). Glioma stem cells (GSCs) are resistant to high doses of radiation, and ultimately select the outgrowth of a more aggressive tumor. This study showed high-dose IR triggered fewer DAMPs molecules exposure and release in GSCs comparing to matched non-GSCs. Downregulation of binding immunoglobulin protein (Bip) promoted IR-mediated endoplasmic reticulum stress to generate DAMPs molecules by PERK and IRE1-α phosphorylation, and increased dendritic cells mature and effector T lymphocytes activation. GSCs treated with Bip knockdown and IR efficiently prevented tumor generation, and reduced post-radiotherapy tumor recurrence. These data suggest that Bip plays a critical role in inhibition of IR-induced ICD in GSCs, and Bip inhibition may be a promising strategy on adjuvant therapy by ameliorating tumor immune microenvironment.

MHC class I dysfunction of glioma stem cells escapes from CTL-mediated immune response via activation of Wnt/ß-catenin signaling pathway.

Glioma stem cells (GSCs) decrease T cells cognition and evade systemic immunosurveillance via downregulations or defects of major histocompatibility complex class I (MHC-I) molecule and antigen-processing machinery (APM) components. Improvement of tumor surface antigens of GSCs may be effective strategy to trigger an adaptive immune response and activate cytotoxic T cells (CTLs) to eliminate glioma. In this study, our data indicated that downregulations of MHC-I and APM components expressions were associated with Wnt pathway activation in GSCs. Histone deacetylases (HDAC) inhibition improved MHC-I and APM components expressions, which could be partly reverted by Wnt pathway activation. Blocking CTLs-mediated killing decreased the anti-tumor effect of tumor lysate vaccine. The enhancement of T cells immune response resulting from HDAC inhibition was dependent on CTLs cognition on tumor antigens presented by upregulated MHC-I molecule in GSCs. These data suggest that suppression of stemness pathway may be effective for GSCs-based immunotherapy against immune-escaped tumors.

Knockdown of cZNF292 suppressed hypoxic human hepatoma SMMC7721 cell proliferation, vasculogenic mimicry, and radioresistance.

Hypoxia is a classic feature of the tumor microenvironment, and has been established as a key epigenetic factor modulating the outcome of radiotherapy. Circular RNAs (circRNAs) are novel RNA molecules with covalently closed circular structures and are highly expressed in eukaryotic transcriptomes. Although previous analysis have shown that circRNA ZNF292 (cZNF292) was hypoxia-responsive and exhibited a proangiogenic function in vitro, the molecular mechanism of cZNF292's biological function is still unclear and deserves further exploration. In this study, we investigated the effect of cZNF292 on the vasculogenic mimicry (VM) and radiosensitivity of hypoxic hepatoma SMMC7721 cells and its mechanism. Our data indicated that cZNF292 could be induced by hypoxia in a time-dependent manner in hepatoma cells independent of hypoxia inducible factor (HIF)-1α. Knockdown of cZNF292 increased SRY (sex determining region Y)-box 9 (SOX9) nuclear translocation, subsequently reduced Wnt/ß-catenin pathway activity, leading to suppression of hypoxic hepatoma cell proliferation, VM, and radioresistance in vitro and in vivo. Our results delineated a novel mechanism of cZNF292 in enhancing hypoxic tumor cell radiosensitivity, which might provide valuable targets for radiation therapy for hepatoma.