Glioblastoma is notoriously difficult to treat, but this review points to a different strategy: instead of asking NK cells to recognize one perfect tumor antigen, block the molecular brakes that keep them from attacking. Glioblastoma multiforme is the most common type of primary malignant brain tumor and has a dismal overall survival rate.

To date, no GBM therapy has yielded successful results in survival for patients beyond baseline surgical resection, radiation, and chemotherapy. Natural killer cell-based immunotherapies are a rising candidate to treat GBM through monoclonal antibody therapy targeting tumor-associated antigens, immune checkpoint inhibitors, CAR-NK cell therapy, and Bi-specific killer cell engagers.

The review focuses on EGFR, HER2, CD155, and IL-13Rα2, and proposes investigating LLT1 and cell surface proliferating cell nuclear antigen as targets for NK cell-based immunotherapy. The passage identifies concurrent radiation therapy and TMZ chemotherapy as part of the treatment approach for GBM, without specifying additional components.

The incorporation of immunotherapy into GBM treatment has yielded disappointing results, amid challenges including the blood–brain barrier, the highly immunosuppressive tumor microenvironment, the immune privileged status of the central nervous system, and molecular heterogeneity.

The review focuses on using natural killer cells to target and destroy GBM cells through therapeutic means that utilize their innate anticancer activities. Unlike T cells, NK cells do not require prior sensitization to specific antigens or the presence of major histocompatibility complex molecules for activation.

NK cell activation is regulated by the balance of inhibitory and activating signals received from various receptors. This ability allows NK cells to identify and eliminate infected or transformed cells without prior exposure, MHC matching, or co-stimulatory signals.

In humans, natural killer cells constitute approximately five to twenty percent of the total circulating lymphocyte population, with CD56bright and CD56dim subsets carrying out cytokine-production and cytotoxic functions. NK cells do not possess a dominant antigen receptor for cytotoxic activation, unlike T and B cells, which possess a single receptor to govern most of their activation and development.

NK cells require the sum of activating signals to exceed the influence of inhibitory receptors to have an effective response. Agonist antibodies binding a single receptor type, other than CD16, are unable to elicit cytokine secretion or cytolytic activity from the NK cell.

Activation appears to require simultaneous cross-linking of different receptors, often with additive or synergistic effects. NK cells are effective recognizers of tumor cells lacking MHC class I on their surface, yet mostly spare tumor cells that do express MHC class I.

When exposed to tumor cells, NK cells trigger degranulation to induce cytolytic apoptosis and release cytokines to modulate the overall immune response. The cytokines discharged by NK cells mainly encompass IFN-gamma, TNF-alpha, and granulocyte–macrophage colony-stimulating factor.

Activated CD16-positive NK cells are associated with improved survival in GBM patients, and their presence correlates with lower-grade tumors. GBM’s immunosuppressive microenvironment, marked by PD-L1, IL-8, and TGF-beta, hinders NK function.

Low numbers of NK cells infiltrate GBM, representing approximately two percent of the total immune cells. However, their scarcity does not diminish their efficacy, as evidenced by their strong impact on GBM and potential to prevent systemic metastasis.

Some NK immunotherapies target NK interactions with GBM cells, while others directly modify NK cells to target GBM. Immune checkpoint inhibitors target molecular interactions between GBM and NK cells that inhibit antitumor functions, most often using monoclonal antibodies against an inhibitory ligand or receptor.

When inhibitory immune checkpoints are blocked, this pushes the NK cell towards activation in the sum of its activity. Relevant checkpoints include PD-1 and PD-L1, TIGIT and CD155, CD47, and B7-H3, although advanced studies have not yielded successful results so far.

Ex vivo activation involves bathing NK cells in cytokines such as IL-2, IL-12, IL-15, IL-18, or IL-21, leading to an increase in CD16 expression and antitumor cytokine production. Genetic modification of NK cells involves changing protein expression through engineering, most commonly with chimeric antigen receptors.

CARs are genetically constructed receptors designed to target a specific antigen, leading to activation of the CAR-expressing NK cell. There are multiple studies evaluating CAR-NK cells in malignant brain tumors, and the cells are being explored through intravenous, intratumoral, intrathecal, or intracranial administration.

EGFR is a receptor tyrosine kinase and one of the most common oncogenic mutations found in GBM. EGFR activation and subsequent intracellular signaling leads to cell proliferation and survival, benefitting GBM cancer cells. EGFR variant three contains a deletion that leads to constitutive activity of the EGFR pathway without ligand binding, and it often occurs alongside EGFR amplification and overexpression in GBM.

EGFR also plays a role in promoting an immunosuppressive environment and evading immune responses from T cells and NK cells. EGFR influences GBM immunology in ways that favor tumor survival and proliferation, making EGFR a target of NK cell immunotherapy for GBM. Immunotherapies against EGFR have been constructed to use EGFR as an activation target, inhibit EGFR signaling by blocking ligand binding, or combine EGFR-targeting tyrosine kinase inhibitors with other immunotherapy strategies.

CAR-NK cells expressing CARs specific for EGFR and EGFR variant three have been studied in preclinical animal settings and are being considered for clinical studies. Figure one maps five proposed ways to stimulate NK-cell activity against glioblastoma: targeting EGFR, HER2, or IL-thirteen-R-alpha-two with CAR-based approaches, and blocking CD155/TIGIT or HLA-E/NKG2A checkpoints with antibodies.

The arrows highlight resulting NK functions, including cytotoxic molecules such as perforin and granzyme B, antibody-dependent cellular cytotoxicity, and cytokines including interferon gamma and tumor necrosis factor alpha. This matters because it organizes distinct target-and-mechanism strategies for countering GBM’s immunosuppressive environment.

HER2 is a receptor tyrosine kinase and an oncogenic driver in many cancers, including GBM. HER2 expression in GBM tumors supports tumor cell proliferation, survival, and angiogenesis, while HER2 overexpression and amplification are highly correlated with poor prognosis.

An immortalized cell line called NK-92/5.28.z expresses an HER2-specific CAR on NK cells and contains a second-generation CAR with a CD28 costimulatory domain and CD3-zeta signaling domain. NK-92/5.28.z is being tested in humans with glioblastoma in a Phase I clinical trial and has so far been shown to be safe.

CD155, also known as the Poliovirus Receptor or PVR, is an antigen overexpressed in GBM and has functions that favor tumor growth and proliferation. CD155 interacts with CD226 and CD96, which drive anti-tumor responses, but it also interacts with the inhibitory NK-cell receptor TIGIT.

When bound by CD155, TIGIT inhibits NK cell antitumor activities and benefits tumor-cell survival and proliferation. Blocking the CD155-TIGIT checkpoint has been shown to increase NK cell anticancer functioning, but targeting this checkpoint in GBM remains under evaluation in cell culture and animal models, with no clinical trials as of publication.

IL-13Rα2 is a subunit of the receptor for interleukin IL-13. GBM tumors overexpress IL-13Rα2 on their surfaces, and this subunit does not contain the same intracellular signaling cascade that induces antitumor immunity. IL-13Rα2 binds IL-13 more readily than IL-13Rα1, so GBM tumors potentially use it to bind available IL-13 and prevent immune responses by stimulating TGF-beta production.

HLA-E is a nonclassical HLA class I molecule that can benefit cancer progression through immune escape and mediate inhibitory signals to NK cells through the NKG2A receptor. The monoclonal antibody monalizumab targets NKG2A on NK cells and prevents interaction with HLA-E, thereby preventing inhibitory signals to the NK cell.

HLA-E is expressed on GBM cells and GBM stem cells, and its expression in gliomas correlates with more aggressive pathological tumor grades. Studies in GBM have so far been conducted in in vitro preclinical settings; preclinical in vivo studies and clinical patient studies have not yet been conducted.

PCNA normally functions in the nucleus in DNA replication, cell-cycle regulation, and DNA repair, and it is highly expressed in many cancers because of its role in rapidly dividing cells. PCNA can also be expressed on cell surfaces as a monomer, where it forms a complex with HLA class I molecules and mediates inhibitory interactions with NK cells through NKp44.

The review distinguishes PCNA expressed on the cell surface as csPCNA, and identifies the NKp44-csPCNA checkpoint as an inhibitor of NK cell anticancer activity. Blocking this checkpoint has been demonstrated as a potentially effective strategy in several cancers, including glioblastoma.

The review proposes csPCNA for further study using monoclonal antibody therapy, BiKEs, and CAR-NK cells. Figure two presents a proposed mechanism for how glioblastoma, or GBM, evades natural killer cell-mediated killing. In panel A, GBM displays LLT1 and cell-surface PCNA, which engage the NK-cell receptors NKR-P1A, also called CD161, and NKp44, delivering inhibitory signals that suppress anticancer functions.

Panel B proposes monoclonal antibodies against LLT1 and/or PCNA to block these interactions, potentially allowing antibody-dependent cellular cytotoxicity, NK-cell activation, cytokine secretion, and tumor-cell killing. The review describes NK cell-based immunotherapies as promising from multiple strategic angles: monoclonal antibody therapy targeting tumor-associated antigens, immune checkpoints, and pure ADCC; CAR-NK cell therapy and other adoptive NK therapies; and Bi-specific killer cell engagers.

The review argues that NK-cell therapy for glioblastoma may need several coordinated targets, combining tumor antigens with checkpoint blockade and engineered NK cells while the proposed csPCNA strategy remains a hypothesis for further study.