Immunotherapy is treatment that uses a patient’s immune system fight malignant tumors. In 1891, William B. Coley, a bone sarcoma surgeon, carried out the study of immunotherapy for the treatment of malignant tumors for the first time. Coley injected streptococcal organisms into a cancer patient to cause erysipelas and stimulate the immune system. The patient’s tumor disappeared, presumably because of an attack by the immune system. Since then, Coley began the lifelong study of immunotherapy, later making him known as the ‘‘Father of Immunotherapy’’ (39). However, the CNS had long been regarded as an immune privileged system for many years, referring to the lack of specialized lymphatic channels in the brain. The concept of immune privilege was based on the original experimental data reported by Peter Medawar 50 years ago, which showed that foreign cells implanted in rodent brains were successfully transplanted, whereas the same cells were eliminated by the host immune system when placed in peripheral tissues (40). Until 2015, Louveau et al. (17) found functional lymphatic vessels within the dural sinus in their search for channels for T cells to enter and exit the meninges. These structures show all the molecular characteristics of lymphatic endothelial cells. They are capable of carrying both fluid and immune cells from the CSF, and connecting deep cervical lymph nodes. This finding suggests that the current dogma on brain tolerance and immune privilege is being revisited, and casts new light to the treatment of gliomas. Although the brain is an immunologically privileged site, the immune microenvironment offers the possibility of implementing immunotherapy for brain tumors. TLS are ectopic lymphoid formations arising in inflamed, infected, or tumoral tissues. They exhibit all the characteristics of structures in the lymph nodes (LN) associated with the generation of an adaptive immune response, including a T cell zone with mature dendritic cells (DC), a germinal center with follicular dendritic cells (FDC) and proliferating B cells (41). In tumors, TLS is thought to provide an alternative to tumor-draining lymph nodes as a site for antigen expression and activation of nascent T cells. Recent studies have demonstrated that the immunostimulatory agonistic CD40 antibody (αCD40) induces the formation of TLS near meningeal tissues in preclinical glioma models. Studies have revealed that the TLS is present in human gliomas and is associated with an increasing number of intratumoral T cells in GBM, suggesting an association between TLS and the regulation of immune responses in glioma patients (18). TLS in the brain can be manipulated therapeutically, probably triggering or suppressing immune responses. As immunotherapy faces many difficulties and challenges, current research continues to explore the immune basis for effective treatment of gliomas, which provides a strong basis for the development of novel therapeutic strategies in the future.

Over the past decade, immunotherapy has dramatically changed the clinical outcome of tumors. However, this promising immunotherapy has encountered serious challenges when applied in the treatment of gliomas, mainly due to the presence of the BBB and the immunosuppressive character of the TME. The BBB serves as an important protective barrier for the brain, controlling the exchange of substances between the blood and the CNS, and maintaining homeostasis within the CNS (42). Meanwhile, the BBB also provides a physical and biochemical barrier for drugs to enter the brain. The tight junctions and adhesions between the endothelial cells of the brain capillaries prevent intercellular diffusion, and molecules from the blood can only enter the brain through the luminal and plasma membranes of the endothelial cells (43). This physical barrier obviously limits the efficiency of intracranial antitumor drug delivery and intra-tumor aggregation. Numerous studies have been conducted to develop novel strategies for delivering therapeutics across the BBB, including focused ultrasound (FUS) (44) and nanotherapeutic drug delivery systems (NDDS) (45). Therefore, priorities in future research should be put on the immune access of the CNS −BBB and new ways to enhance the delivery efficiency and intra-tumor aggregation of immunotherapeutic drugs and to improve the effectiveness of GBM immunotherapy.

With the negative results reported in the CheckMate 143 trial in nivolumab-treated patients with first recurrence of GBM (46), many observers have been convinced that GBM is an immunologically typical “cold tumor”, characterized by low T-cell infiltration. To investigate the immunobiology of GBM in the clinical setting, Hao C et al. (47) found that immunohistochemistry in GBM showed sparse T lymphocyte infiltrates and abundant microglia. This phenomenon suggests an “immunosuppressed state” in GBM and may be the reason for the therapeutic failure of immunotherapy in such tumors. The immunosuppressive properties of the GBM TME significantly inhibits T-cell infiltration and activation, and such a unique microenvironment stimulates tumor cell growth and invasion. On the one hand, immunosuppressive factors, such as PD-1 and indoleamine 2,3-dioxygenase (IDO), are highly expressed in glioma cells, which limit antigen presentation. Gliomas express immunosuppressive ligands on the cell surface, including the co-stimulatory molecule B7-homolog 1 (B7-H1), also known as programmed death ligand 1 (PD-L1) (48). Loss of the tumor suppressor phosphatase and tensin homolog (PTEN) enhances phosphoinositide-3 kinase (PI3K) activity and increases surface expression of B7-H1 in glioma cells (49). This ligand can bind to and stimulate the PD-1 receptor on activated T cells resulting in T cell quiescence and apoptosis (50, 51). Several studies have shown that PD-L1 is highly expressed in GBM cells (52), and combined checkpoint blockade immunotherapy has shown good efficacy in preclinical GBM mouse models (53). However, uncertainty remains regarding the clinical efficacy of PD-1/PD-L1 checkpoint blockade in GBM. Previous studies have shown that the PD-1/PD-L1 pathway plays a role in the malignant biological behavior of GBM, but other molecular signaling networks also play an indispensable role. As demonstrated by a study of Wainwright DA et al. (19), GBM cells express IDO enzymes that can catalyze the rate-limiting step in the catabolism of tryptophan to kynurenine−a pathway that is involved in T cell immune tolerance and immunosuppression. Therefore, there is an urgent need to explore effective sub-targeted combination therapies in TME to improve the clinical response to immunotherapy for GBM.

On the other hand, the glioma microenvironment contains a large population of immunosuppressive cells, mainly including tumor-associated macrophages (TAMs) and regulatory T cells (Tregs). In GBM, TAMs include resident parenchymal microglia, perivascular macrophages and peripheral monocyte-derived cells that are recruited by GBM to release growth factors and cytokines that affect tumors (54), and TAM recruited to tumor sites can be reprogrammed by GBM cells, leading to ineffective antitumor responses (55). Past evidence suggests that context-dependent symbiotic interactions between cancer cells and TAMs are critical for GBM tumor growth through the regulation of different cytokines, chemokines, metabolites, and other factors (56). In addition, TAMs secrete interleukin-10 (IL-10) (57) and transforming growth factor β (TGF-β) (58), which reduce the activity of immune cells in the organism and provide a favorable environment for tumor growth. Tregs have been identified as a pro-tumor subpopulation of CD4+ T cells in GBM tumor tissues and the circulatory system, which can direct cytotoxic T lymphocytes (CTLs) by suppressing tumor cell immune responses (59). Moreover, gliomas exhibit a low tumor mutational burden (TML) (60) and high intra-tumor heterogeneity (61), which are also barriers to immunotherapy for GBM. Such an immune environment leads to the inability of immunotherapy to achieve similar outcomes as other tumors in the treatment of GBM. Decades of efforts to target immunotherapy for GBM have yielded limited outcomes. Further research should focus on the immune microenvironment of GBM, constantly explore new tumor-associated and tumor-specific antigens, and address the immunosuppressive properties of tumors, thus advancing future immunotherapy for GBM.

There are currently more than 80 clinical trials available to evaluate the effects of immunotherapies for GBM (62), and several relevant studies have shown the potential of these immunotherapies for GBM. ICIs represent the most widely studied category of immunotherapies for GBM, including the PD-1, PDL-1 and CTLA-4 signaling pathways, which have shown promising responses in a variety of tumors (63). PD-L1 is highly expressed in GBM, making it an attractive potential target for immunotherapy trials (52). The CheckMate 143 trial was the first extensive evaluation of the efficacy of PD-1/PD-L1 immunotherapy for GBM, which assessed the efficacy of nivolumab (an anti-PD-1 monoclonal antibody) or bevacizumab in 369 patients with rGBM (64). The results showed that there was no significant difference in median OS between the two groups (9.8 months for nivolumab vs. 10.0 months for bevacizumab), but the objective response rate to treatment was higher in the bevacizumab group than that in the nivolumab group. Two recent trials, namely, CheckMate 548 trial and Checkmate 498 trial evaluated the role of nivolumab in newly diagnosed GBM. Patients received standard therapy (RT+TMZ) or standard therapy + nivolumab in Checkmate 548 trial (65), received RT+TMZ or RT + nivolumab in Checkmate 498 trial (66). The results of these two trials manifested that combination immunotherapy did not prolong the survival of GBM patients. In addition, some preclinical and clinical data show that the administration of dexamethasone alongside anti-PD-1 therapy decreases the survival of GBM patients in a dose-dependent manner (67). Dexamethasone reduces T-lymphocyte count by promoting apoptosis, in addition to decreasing lymphocyte functional capacity. Although some GBM patients are receiving long-term dexamethasone therapy for tumor invasion or radiation therapy-related brain edema, the physiological effects of steroids must be addressed in future clinical trials.

CTLA-4, also known as CD152, is a high-affinity receptor for B7 that induces negative costimulatory signaling on activated T cells (68). The CTLA-4 inhibitor, ipilimumab, is currently in clinical trials in GBM including NCT04323046, NCT04396860, NCT04817254 (69). A phase I clinical trial investigated the intracerebral (IC) administration of ipilimumab (IPI) and nivolumab (NIVO) in combination with intravenous administration of NIVO (70). The results showed that IC administration of NIVO and IPI following maximal safe resection of rGBM was feasible, safe, and associated with a prolonged OS. It is evident that ICIs hold great promise in the treatment of primary and recurrent brain tumors and treatment-induced immune-related adverse events (IrAEs) (71). An interim result reported from a phase I/II clinical trial [NCT03174197] of the PD-L1 antagonist atezolizumab with TMZ and RT showed that more than half of the patients enrolled had Grade 3 or higher adverse events that were likely related to treatment (72). In the future, increased awareness of the risks associated with ICIs and combination therapies will be essential to support treatment decisions.

Cancer vaccines work by exposing tumor-associated antigens to antigen-presenting cells (APCs), which activate immune effector cells to achieve an anti-cancer immune response. GBM-specific targets are scarce, but several targets have been identified that are specifically expressed or abundant in tumor cells, including EGFRvIII, which is a mutant version of the EGFR receptor. The cytomegalovirus (CMV) tegument phosphoprotein 65 (pp65) and IDH1 (R132H)-mutant peptides are frequently and specifically expressed in GBM (73–75). Rindopepimut (CDX-110) is a peptide vaccine that targets EGFRvIII, consisting of a unique EGFRvIII peptide sequence conjugated to keyhole limpet hemocyanin that serves as an adjuvant and activates both the humoral and cellular immunity (76). The phase II trial (36) and phase III trial (77) on EGFRvIII-targeted peptide vaccine showed improved PFS and OS compared to historical cohorts, and patients could be safely treated with Rindopepimut for a longer period. The results of the recently reported double-blind randomized phase II trial of Rindopepimut with Bevacizumab for patients with relapsed EGFRvIII-expressing glioblastoma (ReACT) showed longer OS and better overall response rates in patients receiving the peptide vaccine compared with placebo (78), suggesting a possible synergy between Rindopepimut with Bevacizumab. In future studies, the therapeutic strategy that combines Rindopepimut with other immunotherapies to improve the efficacy of GBM can continue to attract wide attention. In addition, multi-targeted vaccines that initiate immune responses to multiple tumor-associated antigens may better address intra-tumor heterogeneity.

Adoptive cell therapy (ACT) is a highly personalized cancer therapy that involves the administration of immune cells with direct anti-cancer activity to a cancer-bearing host (79). Autologous CAR T-cell therapy was the first ACT therapy to enter clinical translation and commercialization, achieving significant improvements in patients with invasive B-cell malignancies. CAR T-cell therapy is a novel treatment modality that exploits the patient’s immune system to fight cancer (80). CAR T cells overcome the limitations of previous T cell-based immunotherapies by redirecting T cell responses to specific tumor antigens (81). First-generation CAR T cells, only containing a single CD3 ζ-signaling module, show poor proliferative responses and low cytotoxicity, resulting in poor antitumor efficacy (82). The currently approved CAR T-cell therapies are second-generation CARs containing CD28 or 4-1BB signaling domains (83). The design of the CAR structure affects the pharmacokinetic profile of CAR T cells, thereby affecting the efficacy and the likelihood of adverse events. Future research will focus on the development of next-generation CAR with the aim of improving the efficacy and safety of CAR T cells. Following the success of the paradigm shift in CAR-engineered adoptive T cell therapies and advances in technologies that can transform cells into powerful antitumor weapons, the interest in NK cells as immunotherapy candidates has grown exponentially (84). T-cell receptors (TCRs) guide NK-92 cells, which have recently been shown to mediate successful antitumor responses (85).

Of all ACT-based immunotherapies currently in development for GBM, genetically engineered CAR T cells are at the forefront, with encouraging results reported in several clinical trials (86). O’Rourke et al. (34) reported 10 rGBM patients treated with a single dose of intravenous second-generation (i.e., 4-1BB, CD3ζ) EGFRvIII CAR T-cell therapy [NCT02209376] and found that the manufacture and infusion of CAR-modified T-cell (CART)-EGFRvIII cells were feasible and safe, without evidence of extratumoral toxicity or cytokine release syndrome. Ahmed N et al. (87) reported an open-label phase I dose-escalation trial, which demonstrated that the infusion of autologous HER2-specific CAR-modified virus-specific T cells (HER2-CAR VSTs) is safe and may be associated with clinical benefit in progressive GBM patients. Moreover, further evaluation of HER2-CAR VSTs as a single agent or in combination with other immunomodulatory methods for the treatment of GBM is warranted in phase 2b study. In addition, the prospect of combining ACT with other treatments for GBM currently under investigation, such as immune checkpoint blockade or oncolytic viruses, has been demonstrated (88, 89). CAR technology has emerged as a particularly attractive area of research related to GBM (90). The clinical trial results of ACT immunotherapy, particularly with CAR T cells, demonstrate a safe and feasible strategy for eliciting an effective immune response in GBM as well as great potential. However, ACT immunotherapy still faces many challenges before the full potential of ACT can be realized, and the need for continued familiarity with ACT in the future may contribute to a deeper understanding of the general mechanisms of cellular immunity and its role in GBM.

Engineered viruses constitute a promising therapeutic approach to addressing the immunosuppression of the GBM microenvironment by killing tumor cells through direct lysis and stimulation of anti-tumor immune responses (91). The most common viruses are herpesviruses, reoviruses, pox virus, or adenoviruses, which are subjected to varying degrees of genetic engineering (91, 92). It has been previously demonstrated that Zika virus (ZIKV) targets GBM stem cells and prevents death of mice with gliomas. In addition, Zika virus can be used to target GBM tissues, generating an immune-sensitized ZIKV strain that is effective alone or in combination with immunotherapy (93, 94). Hence, oncolytic ZIKV treatment can be adopted by immunotherapies, which may facilitate combination therapy for GBM. Desjardins A et al. (95) demonstrated that intratumoral infusion of PVSRIPO in rGBM patients had no neuroviral potential, and that patients treated with PVSRIPO had higher survival rates than historical controls at 24 months and 36 months. DNX-2401 (Delta-24-RGD; tasadenoturev) is a tumor-selective, replication-competent oncolytic adenovirus. Lang FF et al. (25) reported that treatment with DNX-2401 resulted in dramatic responses with long-term survival in recurrent high-grade gliomas, possibly due to the direct tumorolytic effect of the virus, which then stimulates an immune-mediated anti-glioma response. teserpaturev, as one of the genetically engineered oncolytic viruses (OVs) based on herpes simplex virus-1 (G47Δ), is the first oncolytic virus approved for the treatment of malignant gliomas (96). Several other clinical trials on the lysis of viral therapy for GBM are currently underway (97), and attention should be paid to the results of these trial data, as it is crucial to understand the current status of viral therapy for GBM. Several completed trials have demonstrated that OVs therapy is a safe and promising treatment modality for GBM patients, and further optimization of drug delivery and exploration of multimodal combination therapy options are needed to fully realize its therapeutic potential in the future.

Furthermore, gene therapy (98), TAM therapy (99), and recombinant cytokines such as IL-10 (51), interferon transforming growth factor-β (TGF-β) (100), colony- stimulating factor 1 receptor (CSF1R) (62) have been used in GBM clinical trials and have shown some therapeutic potential in specific glioma populations. As research delves deeper and data accumulate, more meaningful advances in immunotherapy regimens for gliomas will be achieved, but many obstacles and difficulties still need to be overcome before these regimens can be used in the clinic, especially difficulties in drug delivery, immune heterogeneity, and tumor heterogeneity. Additionally, the combination strategy of multiple immunotherapies, or the combination strategy of immunotherapy with targeted therapy and chemotherapy, may be an effective solution to these problems, which is worthy of further exploration in the future.