Emily Tang
Jake Y. Chen- 1North Carolina School of Science and Mathematics, Durham, NC, United States
- 2Systems Pharmacology Artificial intelligence (AI) Research Center and Department of Biomedical Informatics and Data Science, School of Medicine, The University of Alabama at Birmingham, Birmingham, AL, United States
Chimeric antigen receptor–natural killer (CAR-NK) cell therapy has shown favorable results in treating hematological malignancies but with limited efficacy against solid tumors, including glioblastomas, which is partly due to the immunosuppressive microenvironment of solid tumors. This mini review focuses on the various immunosuppressive strategies employed by the glioblastoma microenvironment for immune evasion, including stromal barriers, hypoxic conditions, immunosuppressive cytokines, downregulation of activating ligands, and upregulation of immune checkpoints. A range of emerging strategies has been proposed to counteract these inhibitory effects, such as genetic engineering of NK cells and molecular targeting of the stroma in combination with oncolytic virus therapy. Future single-cell spatiotemporal omics studies are expected to further enable a personalized and dynamic approach to treating glioblastoma with improved outcomes.
1 Introduction
Glioblastomas are among the most aggressive and treatment-resistant brain tumors despite advancements in surgery, chemotherapy, and radiation therapy. One promising immunotherapy for glioblastomas is chimeric antigen receptor T cell (CAR-T) therapy to target tumor cells with the immune cell’s innate cytotoxic activity. However, CAR-T therapy has shown limited success and a high rate of recurrence in clinical studies (1). The emergence of chimeric antigen receptor natural killer (CAR-NK) therapy provides a way to counteract the limitations of previous treatments. Compared with CAR-T, CAR-NK therapy has off-the-shelf potential and better safety, such as reduced risk of cytokine release syndrome (CRS) and graft versus host disease (GvHD) (2, 3). However, the efficacy of CAR-NK therapy is compromised by the tumor microenvironment (TME), which limits its application in various tumors (2). The goal of this article is to explore how the glioblastoma TME affects natural killer cells and ways of targeting the TME to enhance CAR-NK therapy in glioblastomas.
2 Characteristics of glioblastomas and its immunosuppressive microenvironment
Glioblastomas are characterized by their immunosuppressive microenvironment, which contains endothelial cells, astrocytes, immune effector cells like microglia, myeloid-derived suppressor cells (MDSCs), tumor-associated macrophages (TAMs), regulatory T-cells (Tregs) and other noncellular components (4, 5). This environment is a complex interplay of tumor, immune, and molecular features. The cellular components are known to contribute to the suppressive environment by fostering tumor evasion, progression, and angiogenesis (4). For example, MDSCs are known to promote tumor growth by secreting certain factors, such as tumor necrosis factor (TNF)-α and vascular endothelial growth factor (VEGF) (6). Immunosuppression may be exacerbated by hypoxia, cytokines (IL4, IL10, and TGF-β), and varying expression of MHC class I molecules (5, 7), which aids in impairment of recognition by immune cells and immune invasion (7). The blood-brain barrier (BBB) is another anti-immune mechanism utilized by glioblastomas. The BBB is a membrane composed of microvascular endothelial cells that separate blood from brain interstitial fluid. For treatment to be administered, the BBB must be penetrated, posing a challenge to many therapies (8). The BBB is among one of a glioblastoma’s characteristics that presents a hurdle to treatment, and it is known that the BBB may be disrupted in certain pathological states that allow immune traffic to more easily infiltrate the central nervous system (9). Clinically, intratumoral/resection cavity and intracerebroventricular (ICV) administration, as well as focused ultrasound and convection-enhanced delivery, have been explored for treating glioblastoma with varying results (10, 11). Overall, the glioblastoma microenvironment contributes to reduced treatment efficacy through various immunosuppressive/evasive mechanisms and the BBB as a physical barrier. This tumor microenvironment represents a potential target for further modulation to enhance antitumor therapy.
3 Biology and function of NK cells in the central nervous system
Natural killer (NK) cells make up a small subset of immune cells in the brain and glioblastomas, consisting of about 1.5% of the total immune cells in a healthy CNS. In the healthy brain, NK cells perform a variety of immune functions: they participate in immune surveillance, work as cytotoxic effectors, and regulate inflammation (12). scRNA-seq analysis has revealed five NK subtypes: CD56bright, early CD56dim, intermediate CD56dim, late CD56dim and adaptive, all named to reflect their maturation. The two different cell states, CD56 bright and dim are notably distinct due to their differential expression of the CD56 neural cell adhesion molecule gene (NCAM1) (13). CD56 bright NK cells are considered precursors to the more mature CD56 dim NK cells. Current studies support that bright cells play a bigger cytokine effector role while dim cells are known for their cytotoxic ability (14, 15). The infiltration of NK cells into the central nervous system is poorly investigated, but research shows that higher NK migration occurs during BBB breakdown (12). In cerebral small vessel disease (CSVD), proteomic analyses revealed that CD56 dim NK cells may contribute to BBB breakdown through secretion of cathepsin D (CTSD), a molecule that participates in protein degradation. This allows the NK cells to further infiltrate and disrupt the neural environment, suggesting that NK cells may intensify damage in cerebrovascular diseases (16, 17). Indeed, NK cell trafficking is very complex and is influenced by many different factors, such as adhesion and cytokine networks secreted by NK, glial, vascular, and other CNS cells (12, 18, 19).
4 CAR-NK therapy: current landscape and glioblastomas-specific challenges
4.1 CAR-NK therapy and its cellular sources
The CAR construct is composed of a few domains: an extracellular antigen-binding domain, transmembrane domains, intracellular signaling domains, and hinge regions (20). Majority of NK cells used in therapy are sourced from the NK92 cell line due to its “off the shelf” potential, lower manufacturing cost, and reduced sensitivity to freeze/thaw cycles. However, the drawbacks of this line include its tumorigenic potential and loss of expansion due to lethal irradiation prior to infusion (21, 22). Other prospective cell lines include peripheral blood mononuclear cells (PBMCs), which have been used in many clinical studies; umbilical cord blood (UCBs) due to the advantage of selecting donors with HLA compatibility and desired NK receptor traits; CD34+ hematopoietic progenitor cells (HPCs), which can be obtained in large amounts and exhibit high cytotoxicity in certain cancers such as leukemias; and induced pluripotent stem cells (iPSCs) for creating “off the shelf products (23)”.
4.2 CAR-NK immunotherapy success in hematological malignancies
The focus for treatment in glioblastomas has shifted to CAR-NK therapy due to successes in hematological malignancies. Several clinical trials (ClinicalTrials.gov) have been conducted to evaluate the efficacy of CAR-NK therapy in hematological tumors, such as leukemias, and has shown clinical significance (20). For instance, a study of CNTY-101 (an iPSC-derived anti-CD19 CAR-NK cell product) in CD19-Positive B-Cell malignancies demonstrated initial safety and efficacy (NCT05336409). Another first-in-human clinical trial of CD19-CAR-UCB-NK for relapsed or refractory CD19-positive non-Hodgkin’s lymphoma or chronic lymphocytic leukemia (CLL) was conducted in 11 patients. The results were mostly positive, with eight out of 11 patients responding to the treatment: seven achieved complete remission, and one had a partial remission. (NCT03056339).
4.3 CAR-NK challenges in solid tumors
While CAR-NK therapy provides many benefits, evident in hematological malignancies, its effectiveness is limited by certain barriers in solid tumors. Table 1 lists some recently completed as well as ongoing clinical trials on CAR-NK therapy for hematological malignancies and solid tumors. However, no outcomes have been reported for glioblastoma treatment yet. Several mechanisms are known to contribute to the reduced effect in solid tumors: hypoxia and other metabolite factors, NK-tumor interactions, cytokines, and exosomes (2).
Table 1. Clinical trials treating hematological malignancies and solid tumors with CAR-NK therapy.
Hypoxia is the process involving abnormal changes in metabolism due to insufficient oxygen supply. In the TME, hypoxia reduces ERK and STAT3 phosphorylation through protein tyrosine phosphatase 1 (SHP-1) dependent manner, diminishing NK cell cytotoxicity by up to 40% as measured by flow cytometry in an in vitro system (24). In addition, hypoxia promotes angiogenesis, metabolic reprogramming, immune evasion, inflammation, and genomic instability (25).
NK-tumor cell interaction also plays a role in reducing the efficacy of CAR-NK therapy in solid tumors. NKG2D is a surface receptor of NK cells and can stimulate immune effectors without antigen presentation (26). As a result, NKG2D plays a crucial role in immune activation; however, it may be downregulated due to the production of soluble NKG2DLs by tumor cells, therefore preventing NK-tumor cell association and diminishing the efficacy of CAR-NK therapy (2, 27).
Cytokines are signaling proteins that mediate communication within the immune system. Transforming growth factor-β (TGF-β) regulates cell growth and differentiation. It has been found that TGF-β is mainly produced by tumor cells and is related to the poor prognosis in solid tumors like lung, gastric, liver, and pancreatic cancer (28–30). TGF-β suppresses NK cell activation and function by downregulating activating receptors and inhibiting the mTOR pathway (31).
Exosomes, vesicles that contain various lipids and proteins, mediate communication and play an important role in disease prognosis (32). Tumor derived exosomes (TDEs) differ from normal exosomes in their biological function: they are involved in tumor formation, metastasis, angiogenesis, and are known to reprogram immune cells via inhibitory proteins, cytokines, RNA, and other factors, such as in non-small cell lung cancer, leukemia, and hepatocellular carcinoma (33–36). Glioblastoma releases exosomes containing 4IgB7H3, which can suppress NK-mediated tumor lysis (37). TDEs also negatively affect CAR-NK therapy by dysregulating NK cell function and reducing their antitumor activity (38).
5 How the glioblastomas TME impairs NK function
5.1 Suppression by astrocytes and cancer associated fibroblasts
Glioblastomas are known for their suppressive TME and BBB that impedes immune infiltration. Astrocytes and microglia have been shown to alter cellular arrangement and contribute to a physical barrier in 3D models. Spheroid models were used to model the tumor, and pure spheroid models had loosely packed cells that exhibited large pores on the surface. The introduction of astrocytes and microglia caused the model to become more compact and structured in certain areas. The nuclear density was higher in these models and supports that these cells may alter the glioblastoma morphology to reduce infiltration (39). In addition, cancer associated fibroblasts (CAFs) are known to exhibit immunosuppressive properties and promote tumor progression. CAFs are a common characteristic of the TME and may impair the cytotoxic function of NK cells (40). This occurs through ferroptosis, a cell death process caused by build-up of iron-catalyzed lipid peroxides (41). A study in gastric cancer found that CAFs increase the intracellular reactive oxygen species (ROS) and malondialdehyde (MDA) lipids within NK cells, which has been shown to consistently contribute to CAF-induced death in NK92 cells. As expected, cell death was reversed by ferroptosis inhibitor Ferrostatin-1 (Fer-1) and cytotoxicity was partially repaired by Fer-1 and Lip-1, another ferroptosis inhibitor (42). In glioblastoma, enhanced ferroptosis was shown to attenuated antitumor cytotoxic killing of immune cells (43). Lastly, it should be noted that CAFs in glioblastoma may differ from those in epithelial tumors, as the fibroblast-like stromal cells or perivascular fibroblasts in glioblastoma may act as potential glioblastoma-initiating cells within the TME (44).
5.2 Impairment by indoleamine 2,3-dioxygenase, TGF-β, and adenosinergic processes
Indoleamine 2,3-dioxygenase (IDO) is an enzyme that facilitates the metabolism of tryptophan (Trp) and is controlled in the glioblastoma microenvironment to suppress immune activity. IDO-1 in the glioblastoma TME reduces tryptophan levels, which has an immunosuppressive effect and leads to CD8+ T cell exhaustion (45). In thyroid cancer, cancer cells produce kynurenine using IDO, resulting in NK dysfunction. Kynurenine enters the NK cell via the aryl hydrocarbon surface receptor and disrupts NK receptor expression through modulation of the STAT1 and STAT3 pathways (46). A recent study using the kynurenine inhibitor PVZB3001 demonstrated promising results, as this compound restored NK cell viability and function in A172 glioblastoma cell cultures (47). In addition to IDO/kynurenine, TGF-β and activin A inhibit NK cell function by reducing its efficacy in tissue homing and residency. Also, CD73-mediated production of adenosine drives reprogramming of the TME and contributes to immune evasion in solid tumors. Increased activity of CD39 and CD73 drives the adenosinergic process by converting ATP to AMP and then to adenosine, which suppresses immune responses (48). NK cells can be engineered to target the CD73-adenosine axis and block the immunosuppressive effect (49).
5.3 Downregulation of activating ligands
The NKG2D ligand family acts as activating receptors for immune responses in NK cells and some T cell. It has been a target in potentially improving cancer immunotherapy because of its selective expression and strong NK cell activating potency (50). However, glioblastomas stem cells (GSCs) in glioblastomas downregulate NKG2D ligands, which impairs NK cell-driven killing (51).
5.4 Upregulation of immune checkpoints
PD-L1 is an immunosuppressive receptor and is mainly located on immune cells such as macrophages, CD3+/CD8+ T cells, and NK cells. The expression of PD-1/PD-L1 in glioblastomas is upregulated and often contribute to immune escape. PD-L1 expression is elevated at the edge of glioblastomas tumor cells compared to the tumor core, suggesting the presence of a PD-L1-mediated barrier at the tumor margins that impedes immune cell infiltration (52). This is another line of evidence that the glioblastomas TME plays a significant role in impairing immune cell function.
6 Opportunities: reprogramming the TME to enhance CAR-NK therapy
6.1 Engineering CAR-NK cells to resist TGF-β and hypoxia
Recently, there have been many strategies proposed to address the limitations of CAR-NK immunotherapy. Mentioned previously, TGF-β impairs the antitumor activity of NK cells; now, studies show that NK cells engineered with the CRISPR-Cas9 system to knock SMAD4 exhibit increased resistance to TGF-β suppression (53, 54). The canonical TGF-β signaling pathway in NK cells is mediated by SMAD2 and SMAD3 phosphorylation, subsequently followed by the phosphorylated SMAD2/3 forming a complex with SMAD4. SMAD4 plays a central role in TGF-β signaling and enables the SMAD2/3 complex to influence gene expression in the nucleus. The knockout of the SMAD4 gene prevents the SMAD2/3 complex from forming a functional transcriptional complex and blocks the signaling pathway to the nucleus. Experimental data shows that NK cells with reduced or absent SMAD4 are less susceptible to TGF-β-induced downregulation of activating receptors and retain higher cytotoxic function (54–56).
Hypoxia is another hurdle in CAR-NK efficacy. Studies found that HER1-overexpressing NK cells genetically engineered with catalase, called HER1-CAR-CAT-NK cells, were more tolerant towards hypoxia and high levels of oxidative stress. Intratumoral delivery of HER1-CAR-CAT-NK cells resulted in sustained attenuation of tumor hypoxia and improved retention and antitumor activity of the engineered NK cells (57). Other strategies for overcoming the hypoxic TME include direct oxygen delivery to tumors, increasing intratumoral oxygen levels, and combinatorial approaches targeting pH, angiogenesis, and immune dysfunction (58).
6.2 Combination treatments, genetic engineering, and oncolytic viruses
Other advancements have come out by genetic manipulation and co-administering drugs to ameliorate the functional defects of NK cells in the TME. Since activation of the CXCL12/CXCR4 axis restricts autophagy, an essential cellular process in NK cells, inhibition of CXCR4 and C/EBPβ restores NK functionality (59). This can be seen as a strategy that can be employed when developing NK products for CAR-NK therapy to improve efficacy and resistance to the TME.
Hypoxic tumors overexpress proteins like VEGF, which influence angiogenesis and promotes immune escape. Combination therapy with anti-angiogenic drugs targeting VEGF may be considered to normalize vascular structure and improve the efficacy of CAR-NK therapy. However, it is known that monotherapy with anti-angiogenic drugs can exacerbate the immunosuppressive TME and result in poor prognosis (2). Recently, studies have found that administering oncolytic viruses engineered to secrete immunostimulatory cytokines improves the efficacy of CAR-NK therapy in neuroblastoma and other cancers. The modified herpes simplex virus C021 expresses interleukin-21 (IL-21), which boosts NK cell proliferation and persistence. In addition, the combination of C021 with anti-ROR1 CAR-NK cells significantly increased cytotoxicity against neuroblastoma cells (60). The C021 virus effectively reprograms the TME from a cold (immunosuppressive) to hot (immunostimulatory) environment and enhances immune infiltration. As a result, the integration of cytokine-secreting oncolytic viruses with CAR-NK cells provides a synergistic and novel immunotherapeutic approach for enhancing anti-tumor efficacy (61).
6.3 Targeting CAFs and HER2 expressing cells
Some recent developments have turned attention to stromal players that contribute to immunosuppression in the TME. The presence of CAFs in the TME are associated with poor prognosis and research has shown that combination therapy with CAF inhibitors enhanced CAR-NK cytotoxicity (62). CAR-NK-cP6 cells modified to continuously produce the P6 peptide disrupt the interaction between TGF-β1 and its receptor TGF-βR1. CAFs are known to secrete TGF-β1, a major immunosuppressive cytokine which creates a physical and molecular barrier for immune surveillance (63). The modified cells block TGF-β1 signaling in CAFs through a paracrine effect, therefore allowing CAR-NK-cP6 cells to display cytotoxic efficacy in a CAF-rich environment. Targeting TGF-β can provide synergistic effects for eliminating CAFs, cancer cells, and their immunosuppressive effect on immune cells (64).
HER2 is expressed in many tumors, such as glioblastomas and breast cancers. Moderate to high HER2 expression has been detected in 41% of primary glioblastoma samples and in the majority of investigated glioblastoma cell lines, as shown by immunohistochemistry (65). As a result, HER2 has been a target for CAR-NK therapy in glioblastomas; however, monotherapy is not enough to overcome the immunosuppressive TME in advanced-stage glioblastomas. Combination therapy of NK-92/5.28.z CAR-NK cells and anti-PD-1 checkpoint inhibition demonstrated robust anti-tumor activity and was able to selectively lyse HER2-expressing glioblastomas cells in vitro. Checkpoint blockade PD-1 has a considerable effect due its ability to block the pathways of immune response. Recent studies have identified PD-1+ NK cell subsets with impaired immune functions in tumors such as high-grade serous ovarian cancer and B-cell chronic lymphocytic leukemia (B-CLL), making them suitable targets for combined immune checkpoint blockade therapy. Administration of PD-1 inhibitors allows both CAR-NK cells and endogenous T cells to proliferate and maintain their cytotoxic functions (66–68). In mouse models, the modified CAR-NK cells were capable of eradicating smaller HER2 tumors and developed durable immunity against glioblastomas rechallenge (69).
7 Discussion
CAR-NK immunotherapy relies on the innate cytotoxic and targeting ability of NK cells in tumors. The key challenges in glioblastomas include the stromal barriers, hypoxia environment, immune-inhibitory cytokines, downregulation of activating ligands, and upregulation of immune checkpoints. Recent advancements have begun addressing these limitations with promising outcomes, such as modifying NK cells to resist hypoxic environments, modulating stromal elements, and combination treatments.
Future work is needed to further improve the field of CAR-NK therapy. Given the heterogeneity of the glioblastomas TME, patient-specific profiling using single-cell and spatial tools will be crucial for identifying immune escape mechanisms and elucidating novel combination strategies. These high-resolution tools can reveal spatial architecture of stromal cells, ligand-receptor interactions, and immune infiltrates, allowing for clear-cut design of CAR constructs tailored for TME landscapes. More specifically, upcoming efforts could include: (i) pairing spatial ligand–receptor maps to select CAR antigens and cytokine payloads; and (ii) using patient-matched organoid or spheroid co-cultures with astrocytes and microglia to benchmark NK cell persistence and motility under hypoxia. Overall, integrating CAR-NK therapy with TME reprogramming will be essential for overcoming immunosuppression and enhancing infiltration, persistence, and cytotoxicity of NK cells. With ongoing biological and therapeutic progress, CAR-NK therapy will transition from a generalized strategy to a personalized and dynamic approach for treating glioblastomas.
Author contributions
ET: Formal Analysis, Writing – review & editing, Writing – original draft, Conceptualization, Investigation. JC: Writing – original draft, Supervision, Writing – review & editing.
Funding
The author(s) declare that no financial support was received for the research, and/or publication of this article.
Conflict of interest
The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.
The author(s) declared that they were an editorial board member of Frontiers, at the time of submission. This had no impact on the peer review process and the final decision.
Generative AI statement
The author(s) declare that no Generative AI was used in the creation of this manuscript.
Any alternative text (alt text) provided alongside figures in this article has been generated by Frontiers with the support of artificial intelligence and reasonable efforts have been made to ensure accuracy, including review by the authors wherever possible. If you identify any issues, please contact us.
Publisher’s note
All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.
References
1. Begley SL, O’Rourke DM, and Binder ZA. CAR T cell therapy for glioblastoma: A review of the first decade of clinical trials. Mol Ther. (2025) 33:2454–61. doi: 10.1016/j.ymthe.2025.03.004
2. Wang W, Liu Y, He Z, Li L, Liu S, Jiang M, et al. Breakthrough of solid tumor treatment: CAR-NK immunotherapy. Cell Death Discov. (2024) 10:40. doi: 10.1038/s41420-024-01815-9
3. Xiong Q, Zhu J, Zhang Y, and Deng H. CAR-NK cell therapy for glioblastoma: what to do next? Front Oncol. (2023) 13:1192128. doi: 10.3389/fonc.2023.1192128
4. Lin H, Liu C, Hu A, Zhang D, Yang H, and Mao Y. Understanding the immunosuppressive microenvironment of glioma: mechanistic insights and clinical perspectives. J Hematol Oncol. (2024) 17:31. doi: 10.1186/s13045-024-01544-7
5. Sharma P, Aaroe A, Liang J, and Puduvalli VK. Tumor microenvironment in glioblastoma: Current and emerging concepts. Neurooncol Adv. (2023) 5:vdad009. doi: 10.1093/noajnl/vdad009
6. He S, Zheng L, and Qi C. Myeloid-derived suppressor cells (MDSCs) in the tumor microenvironment and their targeting in cancer therapy. Mol Cancer. (2025) 24:5. doi: 10.1186/s12943-024-02208-3
7. Burster T, Gärtner F, Bulach C, Zhanapiya A, Gihring A, and Knippschild U. Regulation of MHC I molecules in glioblastoma cells and the sensitizing of NK cells. Pharm (Basel). (2021) 14:1–15. doi: 10.3390/ph14030236
8. Noorani I and de la Rosa J. Breaking barriers for glioblastoma with a path to enhanced drug delivery. Nat Commun. (2023) 14:5909. doi: 10.1038/s41467-023-41694-9
9. Morimoto T, Nakazawa T, Maeoka R, Nakagawa I, Tsujimura T, and Matsuda R. Natural killer cell-based immunotherapy against glioblastoma. Int J Mol Sci. (2023) 24:2111. doi: 10.3390/ijms24032111
10. Chaichana KL, Pinheiro L, and Brem H. Delivery of local therapeutics to the brain: working toward advancing treatment for Malignant gliomas. Ther Delivery. (2015) 6:353–69. doi: 10.4155/tde.14.114
11. Paldor I, Chaichana KL, Brem H, and Tyler BM. Targeted local therapy for management of intracranial high-grade gliomas. Prog Neurol Surg. (2018) 32:159–71. doi: 10.1159/000469688
12. Ning Z, Liu Y, Guo D, Lin WJ, and Tang Y. Natural killer cells in the central nervous system. Cell Commun Signal. (2023) 21:341. doi: 10.1186/s12964-023-01324-9
13. Netskar H, Pfefferle A, Goodridge JP, Sohlberg E, Dufva O, Teichmann SA, et al. Pan-cancer profiling of tumor-infiltrating natural killer cells through transcriptional reference mapping. Nat Immunol. (2024) 25:1445–59. doi: 10.1038/s41590-024-01884-z
14. Poli A, Michel T, Thérésine M, Andrès E, Hentges F, and Zimmer J. CD56bright natural killer (NK) cells: an important NK cell subset. Immunology. (2009) 126:458–65. doi: 10.1111/j.1365-2567.2008.03027.x
15. Weber S, Menees KB, Park J, Agin-Liebes J, Lin C-C, Alcalay RN, et al. Distinctive CD56dim NK subset profiles and increased NKG2D expression in blood NK cells of Parkinson’s disease patients. NPJ Parkinson’s Dis. (2024) 10:36. doi: 10.1038/s41531-024-00652-y
16. Yu D, Cai W, Chen X, Lu D, Hu M, Lu T, et al. Natural killer cells disrupt nerve fibers by granzyme H in atheriosclerotic cerebral small vessel disease. J Gerontol A Biol Sci Med Sci. (2023) 78:414–23. doi: 10.1093/gerona/glac173
17. Seo SU, Woo SM, Im S-S, Jang Y, Han E, Kim SH, et al. Cathepsin D as a potential therapeutic target to enhance anticancer drug-induced apoptosis via RNF183-mediated destabilization of Bcl-xL in cancer cells. Cell Death Dis. (2022) 13:115. doi: 10.1038/s41419-022-04581-7
18. Garofalo S, Cocozza G, Porzia A, Inghilleri M, Raspa M, Scavizzi F, et al. Natural killer cells modulate motor neuron-immune cell cross talk in models of Amyotrophic Lateral Sclerosis. Nat Commun. (2020) 11:1773. doi: 10.1038/s41467-020-15644-8
19. Kveštak D, Juranić Lisnić V, Lisnić B, Tomac J, Golemac M, Brizić I, et al. NK/ILC1 cells mediate neuroinflammation and brain pathology following congenital CMV infection. J Exp Med. (2021) 218:1–14. doi: 10.1084/jem.20201503
20. Pang Z, Wang Z, Li F, Feng C, and Mu X. Current progress of CAR-NK therapy in cancer treatment. Cancers (Basel). (2022) 14:1–24. doi: 10.3390/cancers14174318
21. Gong JH, Maki G, and Klingemann HG. Characterization of a human cell line (NK-92) with phenotypical and functional characteristics of activated natural killer cells. Leukemia. (1994) 8:652–8.
22. Schönfeld K, Sahm C, Zhang C, Naundorf S, Brendel C, Odendahl M, et al. Selective inhibition of tumor growth by clonal NK cells expressing an ErbB2/HER2-specific chimeric antigen receptor. Mol Ther. (2015) 23:330–8. doi: 10.1038/mt.2014.219
23. Xie G, Dong H, Liang Y, Ham JD, Rizwan R, and Chen J. CAR-NK cells: A promising cellular immunotherapy for cancer. EBioMedicine. (2020) 59:102975. doi: 10.1016/j.ebiom.2020.102975
24. Teng R, Wang Y, Lv N, Zhang D, Williamson RA, Lei L, et al. Hypoxia impairs NK cell cytotoxicity through SHP-1-mediated attenuation of STAT3 and ERK signaling pathways. J Immunol Res. (2020) 2020:4598476. doi: 10.1155/2020/4598476
25. Meng W, Hao Y, He C, Li L, and Zhu G. Exosome-orchestrated hypoxic tumor microenvironment. Mol Cancer. (2019) 18:57. doi: 10.1186/s12943-019-0982-6
26. Wensveen FM, Jelenčić V, and Polić B. NKG2D: A master regulator of immune cell responsiveness. Front Immunol. (2018) 9:441. doi: 10.3389/fimmu.2018.00441
27. Zhang C, Röder J, Scherer A, Bodden M, Pfeifer Serrahima J, Bhatti A, et al. Bispecific antibody-mediated redirection of NKG2D-CAR natural killer cells facilitates dual targeting and enhances antitumor activity. J Immunother Cancer. (2021) 9:1–15. doi: 10.1136/jitc-2021-002980
28. Cruz-Bermúdez A, Laza-Briviesca R, Vicente-Blanco RJ, García-Grande A, Coronado MJ, Laine-Menéndez S, et al. Cancer-associated fibroblasts modify lung cancer metabolism involving ROS and TGF-β signaling. Free Radic Biol Med. (2019) 130:163–73. doi: 10.1016/j.freeradbiomed.2018.10.450
29. Zeng D, Li M, Zhou R, Zhang J, Sun H, Shi M, et al. Tumor microenvironment characterization in gastric cancer identifies prognostic and immunotherapeutically relevant gene signatures. Cancer Immunol Res. (2019) 7:737–50. doi: 10.1158/2326-6066.CIR-18-0436
30. Gough NR, Xiang X, and Mishra L. TGF-β Signaling in liver, pancreas, and gastrointestinal diseases and cancer. Gastroenterology. (2021) 161:434–452.e15. doi: 10.1053/j.gastro.2021.04.064
31. Viel S, Marçais A, Guimaraes FS, Loftus R, Rabilloud J, Grau M, et al. TGF-β inhibits the activation and functions of NK cells by repressing the mTOR pathway. Sci Signal. (2016) 9:ra19. doi: 10.1126/scisignal.aad1884
32. Wang JS, Schellenberg SJ, Demeros A, and Lin AY. Exosomes in review: A new frontier in CAR-T cell therapies. Neoplasia. (2025) 62:101147. doi: 10.1016/j.neo.2025.101147
33. Liu Z, Chen Z, Zhang J, Liu J, Li B, Zhang Z, et al. Role of tumor-derived exosomes mediated immune cell reprograming in cancer. Gene. (2024) 925:148601. doi: 10.1016/j.gene.2024.148601
34. Boyiadzis M, Chang-Sook H, and Whiteside TL. Biologically-active exosomes in plasma of AML patients inhibit innate immunity and promote leukemia progression. J Immunother Cancer. (2015) 3:278. doi: 10.1186/2051-1426-3-S2-P278
35. Chen T, Liu Y, Li C, Xu C, Ding C, Chen J, et al. Tumor-derived exosomal circFARSA mediates M2 macrophage polarization via the PTEN/PI3K/AKT pathway to promote non-small cell lung cancer metastasis. Cancer Treat Res Commun. (2021) 28:100412. doi: 10.1016/j.ctarc.2021.100412
36. Huang M, Huang X, and Huang N. Exosomal circGSE1 promotes immune escape of hepatocellular carcinoma by inducing the expansion of regulatory T cells. Cancer Sci. (2022) 113:1968–83. doi: 10.1111/cas.15365
37. Afshar Y, Sharifi N, Kamroo A, Yazdanpanah N, Saleki K, and Rezaei N. Implications of glioblastoma-derived exosomes in modifying the immune system: state-of-the-art and challenges. Rev Neurosci. (2025) 36:315–25. doi: 10.1515/revneuro-2024-0095
38. Hosseini R, Asef-Kabiri L, Yousefi H, Sarvnaz H, Salehi M, Akbari ME, et al. The roles of tumor-derived exosomes in altered differentiation, maturation and function of dendritic cells. Mol Cancer. (2021) 20:83. doi: 10.1186/s12943-021-01376-w
39. Heinrich MA, Huynh NT, Heinrich L, and Prakash J. Understanding glioblastoma stromal barriers against NK cell attack using tri-culture 3D spheroid model. Heliyon. (2024) 10:e24808. doi: 10.1016/j.heliyon.2024.e24808
40. De Jaeghere EA, Denys HG, and De Wever O. Fibroblasts fuel immune escape in the tumor microenvironment. Trends Cancer. (2019) 5:704–23. doi: 10.1016/j.trecan.2019.09.009
41. Li D and Li Y. The interaction between ferroptosis and lipid metabolism in cancer. Signal Transduct Target Ther. (2020) 5:108. doi: 10.1038/s41392-020-00216-5
42. Yao L, Hou J, Wu X, Lu Y, Jin Z, Yu Z, et al. Cancer-associated fibroblasts impair the cytotoxic function of NK cells in gastric cancer by inducing ferroptosis via iron regulation. Redox Biol. (2023) 67:102923. doi: 10.1016/j.redox.2023.102923
43. Liu T, Zhu C, Chen X, Guan G, Zou C, Shen S, et al. Ferroptosis, as the most enriched programmed cell death process in glioma, induces immunosuppression and immunotherapy resistance. Neuro Oncol. (2022) 24:1113–25. doi: 10.1093/neuonc/noac033
44. Ah-Pine F, Khettab M, Bedoui Y, Slama Y, Daniel M, Doray B, et al. On the origin and development of glioblastoma: multifaceted role of perivascular mesenchymal stromal cells. Acta Neuropathol Commun. (2023) 11:104. doi: 10.1186/s40478-023-01605-x
45. Zhou Y, Yao L, Ma T, Wang Z, Yin Y, Yang J, et al. Indoleamine 2,3-dioxygenase-1 involves in CD8(+)T cell exhaustion in glioblastoma via regulating tryptophan levels. Int Immunopharmacol. (2024) 142:113062. doi: 10.1016/j.intimp.2024.113062
46. Park A, Yang Y, Lee Y, Kim MS, Park YJ, Jung H, et al. Indoleamine-2,3-dioxygenase in thyroid cancer cells suppresses natural killer cell function by inhibiting NKG2D and NKp46 expression via STAT signaling pathways. J Clin Med. (2019) 8:1–17. doi: 10.3390/jcm8060842
47. Yoshioka S, Ikeda T, Fukuchi S, Kawai Y, Ohta K, Murakami H, et al. Identification and characterization of a novel dual inhibitor of indoleamine 2,3-dioxygenase 1 and tryptophan 2,3-dioxygenase. Int J Tryptophan Res. (2022) 15:11786469221138456. doi: 10.1177/11786469221138456
48. Chambers AM and Matosevic S. Immunometabolic dysfunction of natural killer cells mediated by the hypoxia-CD73 axis in solid tumors. Front Mol Biosci. (2019) 6:60. doi: 10.3389/fmolb.2019.00060
49. Chambers AM, Lupo KB, Wang J, Cao J, Utturkar S, Lanman N, et al. Engineered natural killer cells impede the immunometabolic CD73-adenosine axis in solid tumors. Elife. (2022) 11:1–22. doi: 10.7554/eLife.73699.sa2
50. Liu H, Wang S, Xin J, Wang J, Yao C, and Zhang Z. Role of NKG2D and its ligands in cancer immunotherapy. Am J Cancer Res. (2019) 9:2064–78.
51. Poorva P, Mast J, Cao B, Shah MV, Pollok KE, and Shen J. Killing the killers: Natural killer cell therapy targeting glioma stem cells in high-grade glioma. Mol Ther. (2025) 33:2462–78. doi: 10.1016/j.ymthe.2025.02.043
52. Jiacheng D, Jiayue C, Ying G, Shaohua W, Wenhui L, and Xinyu H. Research progress and challenges of the PD-1/PD-L1 axis in gliomas. Cell Biosci. (2024) 14:123. doi: 10.1186/s13578-024-01305-6
53. Castriconi R, Cantoni C, Della Chiesa M, Vitale M, Marcenaro E, Conte R, et al. Transforming growth factor beta 1 inhibits expression of NKp30 and NKG2D receptors: consequences for the NK-mediated killing of dendritic cells. Proc Natl Acad Sci U.S.A. (2003) 100:4120–5. doi: 10.1073/pnas.0730640100
54. Rea A, Santana-Hernández S, Villanueva J, Sanvicente-García M, Cabo M, Suarez-Olmos J, et al. Enhancing human NK cell antitumor function by knocking out SMAD4 to counteract TGFβ and activin A suppression. Nat Immunol. (2025) 26:582–94. doi: 10.1038/s41590-025-02103-z
55. Cabo M, Santana-Hernández S, Costa-Garcia M, Rea A, Lozano-Rodríguez R, Ataya M, et al. CD137 costimulation counteracts TGFβ Inhibition of NK-cell antitumor function. Cancer Immunol Res. (2021) 9:1476–90. doi: 10.1158/2326-6066.CIR-21-0030
56. Thangaraj JL, Coffey M, Lopez E, and Kaufman DS. Disruption of TGF-β signaling pathway is required to mediate effective killing of hepatocellular carcinoma by human iPSC-derived NK cells. Cell Stem Cell. (2024) 31:1327–1343.e5. doi: 10.1016/j.stem.2024.06.009
57. Liu Y, Chen J, Tian J, Hao Y, Ma X, Zhou Y, et al. Engineered CAR-NK cells with tolerance to H2O2 and hypoxia can suppress postoperative relapse of triple-negative breast cancers. Cancer Immunol Res. (2024) 12:1574–88. doi: 10.1158/2326-6066.CIR-23-1017
58. Jaing TH, Hsiao YW, and Wang YL. Chimeric antigen receptor cell therapy: empowering treatment strategies for solid tumors. Curr Issues Mol Biol. (2025) 47:1–19. doi: 10.3390/cimb47020090
59. Portale F, Carriero R, Iovino M, Kunderfranco P, Pandini M, Marelli G, et al. C/EBPβ-dependent autophagy inhibition hinders NK cell function in cancer. Nat Commun. (2024) 15:10343. doi: 10.1038/s41467-024-54355-2
60. Rashidi A, Meybodi MA, Cao W, Chu H, Warlick ED, Devine S, et al. Myeloablative versus reduced-intensity hematopoietic cell transplantation in myelodysplastic syndromes: systematic review and meta-analysis. Biol Blood Marrow Transplant. (2020) 26:e138–41. doi: 10.1016/j.bbmt.2020.03.003
61. Chu Y, Tian M, Saini U, Ayala-Cuesta J, Klose K, Mendelowitz AS, et al. Combinatorial immunotherapy with anti-ROR1 CAR NK cells and an IL-21 secreting oncolytic virus against neuroblastoma. Mol Ther Oncol. (2025) 33:200927. doi: 10.1016/j.omton.2024.200927
62. Lee YE, Go GY, Koh EY, Yoon HN, Seo M, Hong SM, et al. Synergistic therapeutic combination with a CAF inhibitor enhances CAR-NK-mediated cytotoxicity via reduction of CAF-released IL-6. J Immunother Cancer. (2023) 11:1–15. doi: 10.1136/jitc-2022-006130
63. Boyd LNC, Andini KD, Peters GJ, Kazemier G, and Giovannetti E. Heterogeneity and plasticity of cancer-associated fibroblasts in the pancreatic tumor microenvironment. Semin Cancer Biol. (2022) 82:184–96. doi: 10.1016/j.semcancer.2021.03.006
64. Shin SH, Lee YE, Yoon HN, Yuk CM, An JY, Seo M, et al. An innovative strategy harnessing self-activating CAR-NK cells to mitigate TGF-β1-driven immune suppression. Biomaterials. (2025) 314:122888. doi: 10.1016/j.biomaterials.2024.122888
65. Zhang C, Burger MC, Jennewein L, Genßler S, Schönfeld K, Zeiner P, et al. ErbB2/HER2-specific NK cells for targeted therapy of glioblastoma. J Natl Cancer Inst. (2016) 108:1–12. doi: 10.1093/jnci/djv375
66. Yazdanpanah-Samani M, Ramezani A, Sheikhi A, Mostafavi-Pour Z, and Erfani N. Anti-PD-L1 chimeric antigen receptor natural killer cell: Characterization and functional analysis. Apmis. (2024) 132:1115–27. doi: 10.1111/apm.13471
67. Farhat M, Croft W, Parry HM, Verma K, Kinsella FAM, Xu J, et al. PD-1 expression contributes to functional impairment of NK cells in patients with B-CLL. Leukemia. (2024) 38:1813–7. doi: 10.1038/s41375-024-02271-1
68. Greppi M, Tabellini G, Patrizi O, Obino V, Bozzo M, Rutigliani M, et al. PD-1(+) NK cell subsets in high grade serous ovarian cancer: an indicator of disease severity and a target for combined immune-checkpoint blockade. J Exp Clin Cancer Res. (2025) 44:258. doi: 10.1186/s13046-025-03508-2
Keywords: glioblastomas, microenvironment, chimeric antigen receptor, natural killer cell, immunotherapy
Citation: Tang E and Chen JY (2025) Shaping the glioblastoma microenvironment to enhance CAR-NK immunotherapy. Front. Immunol. 16:1681966. doi: 10.3389/fimmu.2025.1681966
Received: 08 August 2025; Accepted: 18 November 2025; Revised: 14 October 2025;
Published: 01 December 2025.
Edited by:
Raul Silva García, Mexican Social Security Institute, MexicoReviewed by:
David Akhavan, University of Kansas Medical Center, United StatesCopyright © 2025 Tang and Chen. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.
*Correspondence: Emily Tang, dGFuZzI3ZUBuY3NzbS5lZHU=