The current state of immunotherapy for gliomas: an eye toward the future

JNSPG 75th Anniversary Invited Review Article

Free access

The last decade has seen a crescendo of FDA approvals for immunotherapies against solid tumors, yet glioblastoma remains a prominent holdout. Despite more than 4 decades of work with a wide range of immunotherapeutic modalities targeting glioblastoma, efficacy has been challenging to obtain. Earlier forms of immune-based platforms have now given way to more current approaches, including chimeric antigen receptor T-cells, personalized neoantigen vaccines, oncolytic viruses, and checkpoint blockade. The recent experiences with each, as well as the latest developments and anticipated challenges, are reviewed.

ABBREVIATIONS ALT = adoptive lymphocyte transfer; APC = antigen-presenting cell; BBB = blood-brain barrier; CAR = chimeric antigen receptor; CMV = cytomegalovirus; CTLA-4 = cytotoxic T-lymphocyte–associated protein 4; DAMP = damage-associated molecular pattern; DC = dendritic cell; EGFRvIII = epidermal growth factor receptor type III variant; GBM = glioblastoma; GM-CSF = granulocyte-macrophage colony-stimulating factor; HLA = human leukocyte antigen; MHC = major histocompatibility complex; OS = overall survival; PD-1 = programmed death 1; TAA = tumor-associated antigen; TCR = T-cell receptor; TSA = tumor-specific antigen.

The last decade has seen a crescendo of FDA approvals for immunotherapies against solid tumors, yet glioblastoma remains a prominent holdout. Despite more than 4 decades of work with a wide range of immunotherapeutic modalities targeting glioblastoma, efficacy has been challenging to obtain. Earlier forms of immune-based platforms have now given way to more current approaches, including chimeric antigen receptor T-cells, personalized neoantigen vaccines, oncolytic viruses, and checkpoint blockade. The recent experiences with each, as well as the latest developments and anticipated challenges, are reviewed.

ABBREVIATIONS ALT = adoptive lymphocyte transfer; APC = antigen-presenting cell; BBB = blood-brain barrier; CAR = chimeric antigen receptor; CMV = cytomegalovirus; CTLA-4 = cytotoxic T-lymphocyte–associated protein 4; DAMP = damage-associated molecular pattern; DC = dendritic cell; EGFRvIII = epidermal growth factor receptor type III variant; GBM = glioblastoma; GM-CSF = granulocyte-macrophage colony-stimulating factor; HLA = human leukocyte antigen; MHC = major histocompatibility complex; OS = overall survival; PD-1 = programmed death 1; TAA = tumor-associated antigen; TCR = T-cell receptor; TSA = tumor-specific antigen.

The last decade has seen a crescendo of FDA approvals for immunotherapies against solid tumors, including melanoma and carcinomas of the lung, breast, prostate, bladder, and kidney (renal cell). Many of these approvals herald checkpoint blockade therapies (antibodies to programmed death 1 [PD-1] and/or cytotoxic T-lymphocyte–associated protein [CTLA-4]), which have revolutionized both our expectations and our capacities for the treatment of advanced and even metastatic cancers. Glioblastoma (GBM), however, remains a prominent holdout.

It may seem, then, that GBM is simply a latecomer to the immunotherapeutic stage; however, the opposite is true. While, admittedly, the very first cancer vaccines were pitted against sarcomas by William Coley in the 1890s,16 GBM has a long history of study within the realm of tumor immunology. Pioneers such as Brooks and Roszman were some of the first to examine the interactions of GBM with the immune system even in the early 1970s.8,9 Likewise, some of the earliest predecessors to a variety of current-day immunotherapeutic modalities were trialed in the 1980s with GBM as their target.45,66 Focus later erupted on the identification of appropriate antigens for targeting. Glioma-associated antigens currently identified include IL13Rα2, HER2, gp100, TRP2, EphA2, survivin, WT1, SOX2, SOX11, MAGE-A1, MAGE-A3, AIM2, SART1, and cytomegalovirus (CMV) proteins. Epidermal growth factor receptor type III variant (EGFRvIII) and the IDH-1 mutant (R132H) represent truly tumor-specific targets.

Immunotherapeutic modalities for GBM and other cancers now run the gamut, ranging from antibodies to adoptive cell transfers to vaccines to virally based treatments to immune checkpoint blockade. The current status for many of these will be reviewed below, with a focus on T-cell–based platforms.

Adoptive Lymphocyte Transfers

Strategies for the direct enlistment of T cells, most simply, have included adoptive lymphocyte transfer (ALT), whereby autologous T cells are harvested; trained, expanded, and activated ex vivo against tumor; and transferred back into patients. In its first renditions,32,66 ALT encompassed the transfer of a variety of immune populations, not just T cells, which are now more increasingly the focus.

Early ALT approaches looked to overcome limited numbers of tumor-specific T cells by supplying them genetically modified antigen-specific T-cell receptor (TCR)–α and –β chains.36 Efforts to in turn bypass major histocompatibility complex (MHC) restriction, and to allow for targeting of nonprotein antigens, led to the development of chimeric antigen receptor (CAR) T cells.26 CARs consist of an extracellular domain resembling an antibody that recognizes the tumor-associated antigen (TAA) of choice fused to intracellular T-cell signaling components (Fig. 1). CAR T cells targeting CD19 proved successful at inducing remission in several hematological cancers, which has resulted in FDA approval of two CAR T-cell therapies in those cancers to date.47

FIG. 1.
FIG. 1.

CAR T-cell therapy. T cells transfected with CARs are capable of recognizing tumor cell surface antigens in MHC-independent fashion to instigate tumor cell killing. Copyright Duke University. Published with permission.

The majority of studies with CAR T cells in GBM target EGFRvIII,34 IL13Rα2,11 or HER2.1 EGFRvIII is expressed on approximately 30% of primary human GBMs and, importantly, is absent from normal cells, making EGFRvIII a promising CAR target.72 A phase 1 study by O’Rourke et al. concluded that a single infusion of EGFRvIII CAR T cells was safe based on the absence of off-target toxicities and lack of cytokine release syndrome.53 Significantly, 4 patients who underwent tumor resection following CAR infusion had detectable levels of intratumoral CAR T cells, despite receiving them intravenously.

IL13Rα2 is expressed by more than 60% of GBMs and is not significantly expressed on normal brain tissue19 (although it is expressed on normal kidney). In contrast to the intravenously administered EGFRvIII CAR T cells above, Brown et al. injected IL13Rα2 CAR T cells intracranially, which proved safe in the 3 patients receiving treatment.11 In a subsequent study, the same group achieved demonstrable regression of tumors following intracranial CAR T-cell administration.10

HER2 is a TAA that is overexpressed in a number of cancers, including about 80% of GBMs.49 Importantly, HER2 is also expressed by numerous healthy tissues, so off-target toxicity by HER2-targeting CARs represents a serious risk.52 Nonetheless, successful preclinical studies with HER2-specific CARs led to a phase 1 trial that resulted in a tolerable safety profile.1 It remains to be seen if CARs targeting EGFRvIII, IL13Rα2, and HER2 will have sufficient antitumor capabilities in the clinic.

Recent advances in CAR T-cell biology aim to overcome some of the weaknesses suffered by current approaches. Fourth-generation, or armored, CARs are equipped to overexpress cytokines that potentiate the function and/or persistence of CARs in vivo, such as IL-15.40

Perhaps the foremost limitation impeding CAR therapy for GBM, however, is profound tumor heterogeneity, even at the single-cell level.57 None of the CAR target antigens discussed above are universally expressed, permitting outgrowth of antigen-negative clones. Several strategies have aimed to bypass tumor heterogeneity by engineering CARs with multiple specificities. Hegde et al. generated a bispecific CAR consisting of an HER2-targeting scFv joined to an IL13Rα-targeting IL-13 mutein.27 Not only were these CAR T cells able to efficiently target and kill HER2- or IL13Rα2-expressing tumor cells, but they displayed potentiated activation, without exhaustion, when bound to HER2 and IL13Rα2 simultaneously. Taking this approach one step further, the same group developed a trivalent CAR that targets HER2, IL13Rα2, and EphA2.4 Compared with monovalent and bivalent CARs, the trivalent CAR exhibited superior antitumor activity in a preclinical model of GBM and in vitro when cocultured with primary human GBM cells.

Newer approaches aimed at countering heterogeneity focus on bestowing on CARs the capacity for eliciting epitope spread and stimulating endogenous tumor-specific immunity. A prominent and very recent example (albeit not in GBM) is found in the work of Kuhn et al., who engendered CAR T cells with CD40 ligand (CD40 L). CD40 L+ CAR T cells skirted tumor immune escape through induction of a sustained, endogenous immune response against mouse leukemia/lymphoma that produced superior antitumor efficacy. These CARs were capable of credentialing host antigen-presenting cells (APCs) to prime endogenous T cells against a variety of unidentified tumor antigens, circumventing antigen loss variants.41

One of the largest lessons learned from studies with ALT has been the antitumor benefit obtained with myeloconditioning prior to adoptive transfer.21,60 Lymphodepletion using total body irradiation or high-dose chemotherapy in mice and humans has leveraged the induction of cytokines and consequent homeostatic T-cell proliferation.21 Lymphodepletion induces a massive T-cell proliferation and amplifies tumor-specific immune responses, often by replacing the T-cell pool with only those T cells that can propagate an immune response against the targeted antigen.21

Vaccine-Based Strategies

Much of the immunotherapeutic work in GBM has involved the employment of vaccines. Although the classic image that the word “vaccine” conjures is one of preventative measure, most current tumor vaccines are intended as therapeutic modalities, initiated after tumor detection (and, typically, after failed standards of care as well). Exceptions include cervical and hepatocellular carcinomas, where human papilloma virus and hepatitis B etiologies confer the ability to vaccinate prophylactically against a virus, and therefore, indirectly, a cancer.

The majority of cancers do not have an identified viral precipitant, and the same ability for these cancers is therefore not similarly afforded. In the case of GBM, the detection of tumor-borne CMV antigens has sparked debate regarding whether CMV might be similarly etiologic, or whether, in perhaps more palatable fashion, GBMs situated in a locally immunosuppressed milieu might simply express reactivated viral antigens (that can in turn serve as therapeutic targets).15,51 In either event, vaccination against most tumors, then, represents an effort to stimulate host immunity to established disease.

Vaccination remains one of the earliest developed immunotherapeutic modalities, dating back to Coley’s aforementioned injections of Streptococcus pyogenes and Serratia marcesens toxins into patients with sarcoma.16 Vaccines, as active forms of immunotherapy, elicit and amplify the host’s immunity by the introduction of foreign antigens or APCs, typically combined with adjuvant. Currently, cancer vaccines being employed for GBM can be divided into two main categories: peptide vaccines and cell-based vaccines (Fig. 2).

FIG. 2.
FIG. 2.

Vaccine modalities employed in GBM (peptide, neoantigen, DC). Methods for synthesis of each vaccine modality are depicted, including a technique for tumor whole-exome sequencing and neoantigen identification (upper). Peptide and neoantigen vaccines are delivered with the goal of uptake by endogenous DCs, while DC vaccines are loaded ex vivo with antigen prior to administration. Either vaccine modality has the aim of stimulating DCs priming of CD8+ and CD4+ T cells to effect cellular and humoral immune responses against tumor (lower). Copyright Duke University. Published with permission.

Peptide Vaccines

Tumors are typically characterized by a significant number of mutations, yet GBM is notorious for possessing a relatively low mutational burden.30 Encoded proteins/peptides from mutant genes give rise to antigens that are found only in tumor cells but absent from normal cells. These antigens are called tumor-specific antigens (TSAs), and those TSAs that have not been previously described are termed “neoantigens.” Only a minority of mutations are processed into neoepitopes that can be presented by APCs on human leukocyte antigen (HLA) to elicit a T-cell response. Likewise, many potential tumor antigens derive not from mutations but rather from mis- or overexpressed normal proteins that remain expressed in other tissues (TAAs). Targeting such shared antigens carries the risk of collateral autoimmunity, including encephalitis.5 Ultimately, the relative lack of specific and highly expressed antigenic epitopes in GBM is a limiting factor for the development of vaccine-based strategies.

EGFRvIII, to date, remains the single most relevant and noncontroversial TSA for GBM, found in 20%–30% of tumors. A peptide vaccine, Rindopepimut (CDX-110), was designed to span and foster immune recognition of the mutated sequence in the late 1990s. Rindopepimut demonstrated excellent preclinical efficacy in murine brain tumor models, including the induction of humoral and cytotoxic T-cell responses.28

Promising early-phase clinical data64 led to a multicenter, double-arm phase III clinical trial (ACT IV), enrolling 745 patients with newly diagnosed GBMs. Patients receiving rindopepimut demonstrated decent humoral immune responses but exhibited no significant improvement to median overall survival (OS) when compared to the control arm.71 Of note, survival in the control group exceeded expectations. Likewise, an unusually low cutoff value for EGFRvIII positivity (an inclusion criterion) may have affected trial results. ACT IV’s disappointing outcome decelerated the development of the EGFRvIII-targeted peptide vaccine, although testing of other vaccine modalities targeting EGFRvIII, such as ADU-623, is ongoing (registration no. NCT01967758, clinicaltrials.gov).

When present, mutations in isocitrate dehydrogenase (IDH) are found in nearly 100% of tumor cells and not in the body, making it a rare, truly homogeneously expressed TSA.75 Approximately 80% of low-grade gliomas have mutations in IDH, the most common being the R132H mutation in IDH1 (70% of all IDH mutations).75 In GBM, mutations in IDH1 suggest that the tumor has secondarily developed from a lower-grade glioma, and IDH1 mutations are rarely found in de novo GBM. In preclinical murine models, a polypeptide targeting R132H elicited antigen-specific CD4+ T-cell and humoral responses following presentation on MHC class II (class I epitopes were lacking).63 Phase I clinical trials investigating peptide vaccines in IDH1R132H-mutated grade III and IV gliomas (registration no. NCT02454634, clinicaltrials.gov) and recurrent grade II gliomas (registration no. NCT02193347, clinicaltrials.gov) are ongoing.

Although GBM is known for a relatively low mutational load,30 tumor heterogeneity is still an obstacle, especially under the selective pressure of single-targeted therapies. Such therapies can lead to antigen escape, the state in which a tumor loses expression of the targeted antigen.59 Creating a paradigm to identify and combine multiple neoantigen candidates, as well as to predict their ability to be presented by HLA, is an evolving priority.

Two recently published prominent trials highlight the trend toward personalized cancer vaccines targeting neoantigens.29,35 In the first study, a personalized cancer vaccine strategy was prepared from neoantigens derived by comparing whole-exome sequencing data from the resected tumor with those from matched normal tissues.35 For each patient, 7–20 peptides with high-affinity HLA class I binding predictions were selected for vaccine development.35 The second study combined both neoantigens with unmutated TAAs to increase the number of actionable epitopes.29 Vaccines consisting of 9 unmutated peptides (APVAC1) were administered to patients followed by a 20-peptide neoantigen pool (APVAC2).29 Both studies reflect phase I clinical trials that produced favorable numbers of infiltrating tumor-reactive T cells with memory phenotypes and neoantigen-specific clonal expansion.29,35

Cell-Based Vaccines

Cell-based antitumor vaccines come predominantly in two varieties: tumor cell vaccines and dendritic cell vaccines. Early vaccines frequently pursued the use of killed or inactivated tumor cells, much the same way an antiviral vaccine might be constructed. After fairly poor success rates, various groups in the late 1980s began genetically engineering tumor cells to elaborate a number of immune-stimulating cytokines, most famously, granulocyte-macrophage colony-stimulating factor (GM-CSF). Versions of GM-CSF secreting tumor cell vaccines have been employed for GBM, with technical difficulties often revealed.56 In recent years, a phase I trial was completed, employing the most current generation: autologous irradiated tumor cells accompanied by an allogeneic tumor cell line (K-562) secreting GM-CSF. Vaccination was associated with systemic T-cell activation and antitumor immunity.18

More commonly, vaccine-based therapies for GBM (and other cancers) employ dendritic cells (DCs). Championed by Steinman in 1973, the DC and its central role in choreographing immune responses was established by the early 1990s.67 The expansion, loading, and activation of DCs with tumor antigens ex vivo was soon advocated as a rational antitumor vaccine strategy and, most prominently, forms a mainstay of sipuleucel-T, the first FDA-approved cancer vaccine. With regard to gliomas, a large variety of clinical trials employing DCs have been published out of the US, Europe, and Japan.24 Definitive phase III evidence for efficacy remains lacking, however, and production is decidedly labor intensive and expensive. DCVax, a DC vaccine project developed by Northwest Biotherapeutics based on the work of Linda Liau and colleagues,43 has now entered phase III clinical trials (registration nos. NCT00045968 and NCT02146066, clinicaltrials.gov). Results are awaited.

Recent advances to DC vaccines include preconditioning of the vaccine site. In a high-profile study, DCs loaded with CMV phosphoprotein 65 (pp65) RNA experienced significant improvements to lymph node homing and prolonged patient OS after the vaccine site was preconditioned with tetanus/diphtheria.50 These phenomena are now being better studied in phase I and II clinical trials (registration nos. NCT00639639, NCT02465268, and NCT02366728, clinicaltrials.gov).

A Word on Viral-Based Therapy

There have been many iterations of viral-based therapy for GBM throughout the last 30 years, and, although some have achieved efficacy in subsets of patients, unmitigated success remains elusive. Viral-based therapies are not a mode of immunotherapy per se but deserve mention here. Recent renditions have taken the form of tumor-lysing oncolytic viruses, which aim to initiate an endogenous secondary immune response on tumor cell lysis (Fig. 3).

FIG. 3.
FIG. 3.

Oncolytic virus therapy. Viruses such as PVS-RIPO are administered locally with the aim of tumor-specific uptake/infection. Lysis of tumor cells instigates release of antigen and DAMP, which stimulate a secondary immune response. Copyright Duke University. Published with permission.

Oncolytic viruses are typically delivered intratumorally or postsurgically into the resection cavity. The goal of therapy is to infect and lyse tumor cells, instigating immunogenic cell death pathways. Immune responses are initiated through the release of damage-associated molecular patterns (DAMPs) and TAAs/TSAs. DAMPs recruit immune cells to the site by interacting with pattern-recognition receptors on APCs, such as DCs.46 In addition to instigating DAMP release, oncolytic viruses themselves harbor pathogen-associated molecular patterns that are recognized by the innate immune system, potentially instigating an additional mode of attack.46

Two oncolytic viruses (DNX-2401 and PVS-RIPO) have been granted a fast-track designation by the FDA for expedited drug review. DNX-2401 is a replication-competent adenovirus that was engineered with a specific mutation to restrict viral replication. This virus contains an arginine/glycine/aspartic acid motif to target integrins on GBM, in order to increase infective specificity for tumor cells.42

PVS-RIPO is a replication-competent, live attenuated poliovirus vaccine/human rhinovirus chimera. This virus enters cells expressing the receptor CD155, which is upregulated on malignant cells and also expressed on APC. In vitro, PVS-RIPO is cytotoxic to tumor cells and promotes chronic, sublethal infection of APCs, resulting in proinflammatory cytokine production.12 In a phase I study, 61 patients with recurrent grade IV malignant glioma received PVS-RIPO intratumorally and had an OS rate of 21% at both 24 and 36 months. Comparatively, historical control groups exhibited OS rates of 14% at 24 months and 4% at 36 months. PVS-RIPO is currently in phase II clinical testing.20

Checkpoint Blockade

Immune checkpoints are molecules on the surface of activated T cells that serve as “brakes” to prevent an uncontrolled inflammatory response. Signaling through the classical immune checkpoints CTLA-4 and PD-1 leads to inactivation and even apoptosis of activated T-cells, respectively. Blockade or antagonism of these molecules allows for sustained T-cell activation (Fig. 4).55

FIG. 4.
FIG. 4.

Checkpoint blockade. Activated T cells upregulate checkpoints such as CTLA-4 and PD-1, which can bind B7 and PD-L1 on tumor cells, respectively. Signaling through checkpoints leads to T-cell inactivation, exhaustion, or even apoptosis. Checkpoint blockade therapy involves the administration of antibodies designed to interfere with ligand binding to checkpoints, preserving and perpetuating T-cell activated states. Copyright Duke University. Published with permission.

CTLA-4 has been the most extensively studied immune checkpoint receptor. It dampens T-cell activation by competing with the co-stimulatory molecule CD28 for binding of B7 on APCs. Blockade of CTLA-4 with drugs such as ipilimumab leads to increased availability of CD28, which amplifies T-cell responses. Ipilimumab was first FDA approved for metastatic melanoma in 2010 and is now an accepted therapy in several cancer types. Toxicity has limited its applicability in GBM.

Similarly to CTLA-4, PD-1 moderates the immune response under physiological conditions, and its presence on T cells and other immune cells may be exploited by various cancers. PD-1 is a member of the CD28 family expressed on activated T cells, B cells, dendritic cells, and macrophages. Binding of PD-1 to its ligand PD-L1 (often expressed by tumor cells) leads to impaired T-cell activation through decreased TCR signaling and decreased induction of key transcription factors such as activator protein 1 (AP-1) and nuclear factor of activated T cells (NFAT).65 Early trials of anti-PD-1 treatment in other cancers promoted durable antitumor responses, which led to FDA approvals of nivolumab (anti-PD-1, Bristol-Meyers-Squibb) and pembrolizumab (anti PD-1, Merck).

To date, PD-1 monotherapy has been used to treat various cancers, including melanoma, non–small cell lung cancer, breast cancer, renal cell carcinoma, and some pediatric tumors. Despite successes with checkpoint blockade in many cancers, little efficacy has been seen thus far in GBM. After encouraging preclinical studies,2,69 the first large phase III clinical trial (registration no. NCT02017717, clinicaltrials.gov) of PD-1 checkpoint blockade in recurrent GBM compared nivolumab monotherapy with standard of care bevacizumab (anti-VEGF).58 In early 2017, the trial was closed after it failed to meet the primary OS endpoint.

Focus has now shifted to uncovering and addressing contributors to treatment failures. GBM has a notoriously low tumor mutational burden, which is a documented contributor to checkpoint blockade resistance.30 While this is not a feature of GBM that is alterable per se, there are case reports of successful checkpoint blockade therapy in rare GBM patients bearing germline defects in mismatch repair and resultant hypermutated states.7,33 The argument persists that subsets of GBM patients with specific and/or increased numbers of mutations may be more amenable to treatment.

Two recent exciting early-phase clinical trials explored anti–PD-1 therapy in the neoadjuvant setting in GBM, with promising results.14,61 Schalper et al. conducted a single-arm phase II trial in 30 patients who received presurgical and postsurgical nivolumab.61 No clinical benefit was observed, although biopsies before and after treatment revealed increases to chemokine transcription (CXCL10, CCL4, and CCL3L1), T-cell diversity, and immune cells.

Cloughesy et al. enrolled 35 patients to a randomized open-label pilot study of pembrolizumab to determine whether neoadjuvant blockade would alter the tumor microenvironment and improve OS compared with adjuvant blockade alone.14 Patients in the neoadjuvant group reached a median OS of 417 days, while patients in the adjuvant-alone group had an OS of 228.5 days. PFS was also significantly increased in the neoadjuvant group (99.5 days vs 72.5 days). Neoadjuvant PD-1 therapy was associated with an increased IFN-γ responsive gene signature and decreased cell cycle–related gene signature in the tumor.

Challenges for the Future

GBM imbues a unique set of obstacles to successful immunotherapy. Hindrances readily ascribable to GBM include a low tumor mutational burden;30 broad tumor heterogeneity;57 restricted CNS drug/immune access;38 and, perhaps most saliently, rampant T-cell dysfunction.74 While a full discussion of each of these topics is beyond the scope of this review, brief attention to two particularly salient immunological issues is worthwhile: those of immune access and immune suppression.

Immune Access

The long-held paradigm of CNS immune privilege has powered the model of poor immune access to the intracranial compartment and to those tumors harbored within. The permissiveness of the brain for immune entry has been a subject of study and speculation for decades. Peter Medawar’s tissue engrafting studies of the 1940s did much to suggest the brain as being absent a lymphatic drainage system,48 and the blood-brain barrier (BBB) is a long-recognized restriction to CNS access,3 including to immune cells.54

Immunotherapy, however, is likely less hindered by CNS immune privilege than dogma has dictated.24 In the many years since Medawar’s experiments, newer studies have picked away at the concept of the brain as immunologically isolated. Routes of antigenic egress from the brain to the deep cervical lymph nodes (via the arachnoid sheath of the olfactory nerves, through the cribriform plate, and to the nasal mucosa) were uncovered beginning in the 1980s.17 In more recent years, the identification of the glial-lymphatic (glymphatic) pathway offered a mechanism for fluid and solute clearance from the brain, linking the parenchyma and interstitium to the CSF spaces.31 Most recently, the discovery of functional lymphatic vessels in the meninges provided a direct drainage pathway for CSF and immune cells from the brain into the cervical lymph nodes.44

Likewise, evidence for T-cell entry and immunosurveillance within the brain has been provided,54,62 marring the BBB’s reputation as a hermetic seal to immune cell entry. In any event, damage incurred to the BBB in the context of gliomas and other tumors likely provides more unrestrained immune access in the setting of cancer.70 Recent successes with immune checkpoint blockade against intracranial metastases suggest that treatment failures with GBM may be more tumor- than location-specific.68

Still, despite losing some of the protections of the BBB, tumors within the intracranial compartment are not without weapons for engendering unique modes of immune evasion. Tumors situated intracranially appear capable of usurping the brain’s capacities for stymieing immune responses to construct their own forms of privilege, keeping T cells out and putting up newly discovered barriers to immunotherapy. A recent high-profile paper, for instance, highlighted the bizarre and novel capacity of tumors within the brain, specifically, to sequester T cells away in the bone marrow where they can do no harm, fostering antigenic ignorance and immune evasion.13 Thus, while our construct of CNS immune privilege has faded some, the idiosyncrasies of the brain immune environment continue to offer distinct challenges to immune-based platforms.24

Immune Suppression

As noted, immune dysfunction is a hallmark of GBM, with reports dating to the 1970s.22 Unfortunately, the T cells required for effective antitumor responses are particularly victimized.74 Their plight is reflected in patient T-cell lymphopenia,13 as well as various characterized forms of classical T-cell dysfunction, such as anergy9 and regulatory T-cell–imposed tolerance.23,25 As highlighted above, the enigmatic sequestration of T cells in bone marrow is a newly discovered mode of T-cell dysfunction that contributes to AIDS-level T-cell counts in GBM patient blood and severely restricts the access of T cells to tumors in the CNS.13

As the emphasis shifts more and more onto immune checkpoint blockade and to deciphering treatment resistance, T-cell dysfunction needs to be front and center. T-cell exhaustion may be a particularly relevant mode of dysfunction limiting success. In other cancers, resistance to immune checkpoint blockade at the T-cell level is frequently marked by the upregulation of multiple alternative immune checkpoints,39 which can include TIM-3, LAG-3, BTLA, 2B4, CD160, CD39, and TIGIT. Mounting expression of these alternative checkpoints on T cells signals an exhausted state.6

While exhaustion may be reversible in early stages, it can rapidly progress beyond rescue. Woroniecka et al. recently demonstrated comparatively severe T-cell exhaustion among tumor-infiltrating lymphocytes isolated from patients and mice with GBM and the corresponding inability of mice with gliomas to respond to PD-1 blockade.73 The mounting expression of alternative immune checkpoints may indicate a state of terminal exhaustion that cannot be reversed by traditional checkpoint blockade alone. Preclinical work has, however, highlighted a possible synergy between PD-1, TIM-3, and LAG-3 blockade in mice with gliomas.37 Clinical trials targeting TIM-3 and LAG-3, either alone or in combination with anti-PD-1, are underway in GBM (registration nos. NCT02658981 and NCT02817633, clinicaltrials.gov). Likewise, studies focused on understanding tumor-induced exhaustion are needed.

Conclusions

Immunotherapeutic successes against GBM have been limited, despite decades of effort. The tumor and its CNS confines offer unique challenges to immunotherapy, including constraints to drug and immune access, marked heterogeneity, low mutational burden, and severe tumor-imposed T-cell and other immune dysfunction. Newly discovered challenges, such as tumor-directed sequestration of T cells in bone marrow,13 highlight our need to elucidate the brain’s complex mechanisms for limiting immune responses. Failures notwithstanding, we continue to better understand these limitations and to adapt our therapeutic strategies accordingly. Recent results from trials of neoadjuvant checkpoint blockade provide new cause for optimism as we work to decode mechanisms for treatment resistance. Renewed focus on preventing and reversing T-cell dysfunction is warranted in the efforts to deliver our therapies into a viable and functioning immune system.

Acknowledgments

We acknowledge Daniel Wilkinson, PhD; Pakawat Chongsathidkiet, MD; Cosette Champion, BS; Jessica Waibl-Polania, BS; Selena Lorrey, BS; Megan Llewellyn, MSMI; and Tecca Wright, MA, for their assistance with research, illustrations, and manuscript preparation.

Disclosures

J.H.S. has an equity interest in Annias Immunotherapeutics, which has licensed intellectual property from Duke related to the use of the PEP-CMV vaccine in the treatment of GBM. J.H.S. has an equity interest in Istari Oncology, which has licensed intellectual property from Duke related to the use of poliovirus and D2C7 in the treatment of GBM. J.H.S. has additional relationships with Celldex (intellectual property, royalties) and Medicenna Therapeutics (consulting). J.H.S. is an inventor on patents related to PEP-CMV DC vaccine with tetanus, as well as poliovirus vaccine and D2C7 in the treatment of GBM. Duke and certain Duke investigators could benefit financially if related therapies prove effective and are commercially successful.

Author Contributions

Conception and design: both authors. Acquisition of data: both authors. Analysis and interpretation of data: both authors. Drafting the article: both authors. Critically revising the article: both authors. Reviewed submitted version of manuscript: both authors. Administrative/technical/material support: both authors.

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Article Information

Contributor Notes

Correspondence John H. Sampson: Duke University Medical Center, Durham, NC. john.sampson@duke.edu.INCLUDE WHEN CITING DOI: 10.3171/2019.5.JNS181762.Disclosures J.H.S. has an equity interest in Annias Immunotherapeutics, which has licensed intellectual property from Duke related to the use of the PEP-CMV vaccine in the treatment of GBM. J.H.S. has an equity interest in Istari Oncology, which has licensed intellectual property from Duke related to the use of poliovirus and D2C7 in the treatment of GBM. J.H.S. has additional relationships with Celldex (intellectual property, royalties) and Medicenna Therapeutics (consulting). J.H.S. is an inventor on patents related to PEP-CMV DC vaccine with tetanus, as well as poliovirus vaccine and D2C7 in the treatment of GBM. Duke and certain Duke investigators could benefit financially if related therapies prove effective and are commercially successful.
Headings
Figures
  • View in gallery

    CAR T-cell therapy. T cells transfected with CARs are capable of recognizing tumor cell surface antigens in MHC-independent fashion to instigate tumor cell killing. Copyright Duke University. Published with permission.

  • View in gallery

    Vaccine modalities employed in GBM (peptide, neoantigen, DC). Methods for synthesis of each vaccine modality are depicted, including a technique for tumor whole-exome sequencing and neoantigen identification (upper). Peptide and neoantigen vaccines are delivered with the goal of uptake by endogenous DCs, while DC vaccines are loaded ex vivo with antigen prior to administration. Either vaccine modality has the aim of stimulating DCs priming of CD8+ and CD4+ T cells to effect cellular and humoral immune responses against tumor (lower). Copyright Duke University. Published with permission.

  • View in gallery

    Oncolytic virus therapy. Viruses such as PVS-RIPO are administered locally with the aim of tumor-specific uptake/infection. Lysis of tumor cells instigates release of antigen and DAMP, which stimulate a secondary immune response. Copyright Duke University. Published with permission.

  • View in gallery

    Checkpoint blockade. Activated T cells upregulate checkpoints such as CTLA-4 and PD-1, which can bind B7 and PD-L1 on tumor cells, respectively. Signaling through checkpoints leads to T-cell inactivation, exhaustion, or even apoptosis. Checkpoint blockade therapy involves the administration of antibodies designed to interfere with ligand binding to checkpoints, preserving and perpetuating T-cell activated states. Copyright Duke University. Published with permission.

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    Dudley MEWunderlich JRRobbins PFYang JCHwu PSchwartzentruber DJ: Cancer regression and autoimmunity in patients after clonal repopulation with antitumor lymphocytes. Science 298:8508542002

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    • Export Citation
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    • Export Citation
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    • Export Citation
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    Fecci PEMitchell DAWhitesides JFXie WFriedman AHArcher GE: Increased regulatory T-cell fraction amidst a diminished CD4 compartment explains cellular immune defects in patients with malignant glioma. Cancer Res 66:329433022006

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    • Export Citation
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    • Export Citation
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    Hegde MMukherjee MGrada ZPignata ALandi DNavai SA: Tandem CAR T cells targeting HER2 and IL13Rα2 mitigate tumor antigen escape. J Clin Invest 126:303630522016

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    • Export Citation
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    • Export Citation
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    • Export Citation
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    Hodges TROtt MXiu JGatalica ZSwensen JZhou S: Mutational burden, immune checkpoint expression, and mismatch repair in glioma: implications for immune checkpoint immunotherapy. Neuro Oncol 19:104710572017

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    • Export Citation
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    • Export Citation
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    • Export Citation
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    Johanns TMMiller CADorward IGTsien CChang EPerry A: Immunogenomics of hypermutated glioblastoma: a patient with germline POLE deficiency treated with checkpoint blockade immunotherapy. Cancer Discov 6:123012362016

    • Search Google Scholar
    • Export Citation
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    Johnson LAScholler JOhkuri TKosaka APatel PRMcGettigan SE: Rational development and characterization of humanized anti-EGFR variant III chimeric antigen receptor T cells for glioblastoma. Sci Transl Med 7:275ra222015

    • Search Google Scholar
    • Export Citation
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    Keskin DBAnandappa AJSun JTirosh IMathewson NDLi S: Neoantigen vaccine generates intratumoral T cell responses in phase Ib glioblastoma trial. Nature 565:2342392019

    • Search Google Scholar
    • Export Citation
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    • Export Citation
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    • Search Google Scholar
    • Export Citation
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    Kmiecik JPoli ABrons NHWaha AEide GEEnger PO: Elevated CD3+ and CD8+ tumor-infiltrating immune cells correlate with prolonged survival in glioblastoma patients despite integrated immunosuppressive mechanisms in the tumor microenvironment and at the systemic level. J Neuroimmunol 264:71832013

    • Search Google Scholar
    • Export Citation
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    Koyama SAkbay EALi YYHerter-Sprie GSBuczkowski KARichards WG: Adaptive resistance to therapeutic PD-1 blockade is associated with upregulation of alternative immune checkpoints. Nat Commun 7:105012016

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    • Export Citation
  • 40

    Krenciute GPrinzing BLYi ZWu MFLiu HDotti G: Transgenic expression of IL15 improves antiglioma activity of IL13Rα2-CAR T cells but results in antigen loss variants. Cancer Immunol Res 5:5715812017

    • Search Google Scholar
    • Export Citation
  • 41

    Kuhn NFPurdon TJvan Leeuwen DGLopez AVCurran KJDaniyan AF: CD40 ligand-modified chimeric antigen receptor T cells enhance antitumor function by eliciting an endogenous antitumor response. Cancer Cell 35:473488488.e1–488.e6 2019

    • Search Google Scholar
    • Export Citation
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    Lang FFConrad CGomez-Manzano CYung WKASawaya RWeinberg JS: Phase I study of DNX-2401 (Delta-24-RGD) oncolytic adenovirus: replication and immunotherapeutic effects in recurrent malignant glioma. J Clin Oncol 36:141914272018

    • Search Google Scholar
    • Export Citation
  • 43

    Liau LMPrins RMKiertscher SMOdesa SKKremen TJGiovannone AJ: Dendritic cell vaccination in glioblastoma patients induces systemic and intracranial T-cell responses modulated by the local central nervous system tumor microenvironment. Clin Cancer Res 11:551555252005

    • Search Google Scholar
    • Export Citation
  • 44

    Louveau ASmirnov IKeyes TJEccles JDRouhani SJPeske JD: Structural and functional features of central nervous system lymphatic vessels. Nature 523:3373412015

    • Search Google Scholar
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