Malignant tumors have long been typified as “immune cold” due to their ability to evade immune detection and suppress immune activity, rendering many conventional treatments ineffective. This immune evasion has posed significant challenges in oncology, as patients with immune-cold tumors often experience poor responses to chemotherapy and immunotherapy, culminating in dire prognoses. The Johns Hopkins team’s novel approach aims to reverse this phenomenon by transforming these tumors into “immune hot” environments that actively recruit and stimulate immune cells to attack cancer.

Central to this transformative approach are tertiary lymphoid structures (TLSs), which are lymph node-like aggregates that naturally form in sites afflicted by chronic inflammation, including certain tumors responsive to the immune system. TLSs serve as immunological hubs within tumors and have been strongly correlated with improved patient prognoses and responsiveness to therapy. Understanding the factors that foster TLS formation in tumors has been a pivotal goal in harnessing their anti-cancer potential.

Leveraging previous insights in breast cancer immunology, the researchers hypothesized that enhancing the local tumor milieu with specific immune-activating signals could fortify TLS development and functionality. They meticulously studied the complex cellular and molecular landscape of TLS-rich tumors to identify the critical stimuli driving their formation and activity. This reverse-engineering approach provided a blueprint for inducing TLS presence in otherwise TLS-deficient tumors.

The experimental intervention centered on simultaneously activating two key immune signaling pathways: the stimulator of interferon genes (STING) and the lymphotoxin-β receptor (LTβR). STING is a cytosolic DNA sensor that initiates robust innate immune responses, including the production of type I interferons and other inflammatory cytokines, thereby shaping adaptive immunity. LTβR signaling is essential for lymphorganogenesis and maintaining the structural integrity of lymphoid tissues. By delivering agonists that engage both STING and LTβR, the researchers engineered a highly stimulatory tumor environment conducive to immune cell recruitment and activation.

This dual activation regime precipitated a swift and powerful infiltration of cytotoxic CD8⁺ T cells into the tumor microenvironment, directly contributing to pronounced tumor growth inhibition. Notably, the treatment induced the formation of high endothelial venules (HEVs)—specialized blood vessels that function as selective gateways permitting lymphocyte extravasation from the bloodstream into the tumor stroma. The emergence of HEVs effectively opened the floodgates, enabling massive recruitment of both B cells and T cells to forge new TLS in situ.

Within these newly formed TLS, B lymphocytes exhibited hallmark germinal center reactions, a sophisticated immune process whereby B cells proliferate, undergo somatic hypermutation, and mature into plasma cells capable of producing high-affinity, tumor-specific antibodies. These plasma cells not only sustained local antibody production but also migrated to the bone marrow to establish a reservoir of long-lived memory cells. The presence of tumor-specific IgG antibodies and persistent plasma cells underscores the generation of durable systemic immunity capable of long-term tumor surveillance and relapse prevention.

Concurrently, the immunotherapy elevated populations of helper CD4⁺ T cells and memory CD8⁺ T cells, thereby orchestrating a balanced enhancement of humoral and cellular immunity. This comprehensive immune orchestration ensures that both antibody-mediated mechanisms and direct cytotoxic effects contribute synergistically to tumor eradication. Modulating immune signaling balance within the tumor bed appears critical for sustaining sustained anti-tumor activity.

These findings illuminate a mechanistically rich paradigm in which early, dual-pathway immune activation not only exerts immediate cytotoxic effects on tumor cells but also fosters the maturation and persistence of TLS that amplify and sustain anti-cancer immune responses over time. TLS maturation thereby operates as an immunological amplifier system, extending the reach and durability of immune-mediated tumor control well beyond the initial treatment window.

Dr. Masanobu Komatsu, principal investigator and senior scientist at the Johns Hopkins All Children’s Cancer & Blood Disorders Institute, emphasizes the transformative potential of this approach. “By constructing the appropriate immune architecture within tumors, we can potentiate both T cell and B cell defenses against cancer progression, relapse, and metastasis,” he states. This dual-pronged, immune-structural remodeling strategy promises to overcome the entrenched immunosuppressive barriers characteristic of many aggressive cancers.

Because the abundance of TLS has been positively associated with outcomes across diverse tumor types, this dual activation of STING and LTβR may offer a broadly applicable therapeutic avenue. It holds potential to substantially boost the efficacy of existing modalities, including checkpoint inhibitor immunotherapies, which often falter in “immune cold” cancers, as well as traditional chemotherapeutic regimens. Enhancing the tumor’s inherent immune competence could therefore represent a universal adjunct to improve cancer treatment paradigms.

Ongoing research efforts by Komatsu’s team are delving deeper into the complex molecular mechanisms underlying TLS induction and function following STING and LTβR stimulation. Furthermore, preparations are underway to translate these preclinical findings into clinical trials involving both adult and pediatric cancer patients. These future studies aim to validate safety, optimize dosing, and determine the most effective combination regimens to integrate TLS induction with current immuno-oncology standards.

Funded principally by NIH/National Cancer Institute grants alongside support from the Department of Defense’s Congressionally Directed Cancer Research Program and the Florida Department of Health Bankhead Coley Cancer Research Program, this research reflects a significant multidisciplinary collaboration. The work’s potential to fundamentally alter cancer immunotherapy highlights the critical role of federally supported science in pushing the boundaries of medical innovation.

As immunotherapy revolutionizes cancer care, the ability to deliberately engineer tumor microenvironments to foster TLS formation marks a bold and exciting frontier. This therapeutic blueprint exemplifies how reprogramming immune system architecture within tumors can unmask new vulnerabilities in cancer. Ultimately, such innovations stand to shift the paradigm from merely treating tumors to empowering the body’s own immune machinery to deliver durable, systemic tumor control and improve patient survival worldwide.

Subject of Research: Enhancing anti-tumor immunity by inducing tertiary lymphoid structures via dual STING and LTβR activation in immune-cold tumors

Article Title: Therapeutic induction of tertiary lymphoid structures promotes durable anti-cancer immunity in immune-cold tumors

News Publication Date: September 2, 2025

Web References:
https://www.nature.com/articles/s41590-025-02259-8?fromPaywallRec=false#Sec11

References:
Johns Hopkins All Children’s Hospital research publication in Nature Immunology, 2025

Image Credits:
Nature Immunology

Keywords:
Cell lines, Cancer cells

Glioblastoma is notoriously difficult to treat due to its highly invasive and malignant nature, compounded by its classification as an immunologically “cold” tumor. This means the tumor microenvironment actively suppresses immune activity, thwarting conventional immunotherapies’ effectiveness. Nearly all GBM patients experience tumor recurrence within six to nine months post-treatment, and recurrent tumors often develop resistance to chemotherapy and radiation, leading invariably to patient mortality. Current therapeutic strategies address symptoms and slow progression but fail to offer curative outcomes, thus underscoring the urgent need for innovative solutions.

The researchers sought to address these challenges by designing a fusion molecule that simultaneously delivers the cytotoxic effects necessary to kill tumor cells and the immunomodulatory signals required to activate the body’s immune defenses. This FSK is composed of an enhanced form of Interleukin-24 (IL-24S), renowned for its tumor-selective cytotoxicity, coupled with Interleukin-15 (IL-15), a potent immune-stimulating cytokine known to activate natural killer (NK) cells and T lymphocytes. The fusion aims to overcome the immunosuppressive microenvironment of GBM, effectively converting a “cold” tumor into an immunologically active battlefield.

Testing this molecule in an immunocompetent mouse model of glioblastoma revealed striking therapeutic outcomes. The FSK demonstrated superior tumor regression and prolonged survival compared to treatments involving either IL-24S or IL-15 alone. Crucially, the therapy not only induced direct tumor cell death but also enhanced infiltration of key immune cells—including T cells, dendritic cells, macrophages, and NK cells—within the tumor microenvironment. This suggests the treatment orchestrates a coordinated immune assault, improving both local control and potentially systemic antitumor immunity.

Delivering therapeutic agents effectively to the brain has been a longstanding hurdle due to the blood-brain barrier (BBB), a highly restrictive physiologic interface that prevents most molecules and viruses from reaching CNS tumors. To circumvent this challenge, the team engineered a delivery system that utilizes a type 5 adenovirus vector to express the fusion superkine. Not stopping there, they innovatively paired this vector with a noninvasive, targeted delivery technique employing focused ultrasound (FUS) combined with intravenously infused microbubbles (MBs). This focused ultrasound double microbubble (FUS-DMB) method transiently and safely opens the BBB, allowing the adenovirus vector carrying the FSK to penetrate the brain’s protective barrier and deliver its payload directly to the tumor.

The FUS-DMB technique operates by inducing oscillation and cavitation of microbubbles within cerebral blood vessels under ultrasound exposure, leading to reversible disruption of tight junctions in the endothelial cells forming the BBB. This temporary permeability boosts penetration of the viral vector without causing neurological damage or eliciting significant inflammation, a major advancement over invasive surgical delivery methods or systemic treatments with poor CNS bioavailability. The ability to precisely and safely shuttle gene therapy vectors into brain tissue could herald a new era for treating brain pathologies beyond glioblastoma—including metastases and neurodegenerative diseases.

Dr. Paul B. Fisher emphasized the novelty and transformative potential of this approach, expressing optimism about an upcoming clinical trial projected to launch in 2026. This trial will investigate the safety and efficacy of the IL-24 gene therapy and accompanying viral delivery methods in glioblastoma patients. According to Fisher, the fusion superkine and FUS-DMB delivery together could represent an unprecedented “knockout” solution for brain cancer, aiming to achieve what has so far proved elusive—the elusive “holy grail” of a cure for this devastating disease.

Complementing Fisher’s vision, Dr. Swadesh K. Das highlighted the fusion superkine as a differentiated platform that simultaneously accomplishes tumor cell eradication and localized immune activation. By merging gene therapy with advanced immunotherapy principles, the treatment is designed not only to attack established tumors but also to establish durable immune memory, potentially preventing relapse. Such immunological “education” of the host immune system is critical given glioblastoma’s notorious tendency to evade conventional therapies and redevelop.

Peers reviewing the study echoed its significance, noting that previous efforts to develop adenoviral vectors co-expressing multiple therapeutic genes have been hampered by technical hurdles such as impaired viral assembly or inadequate gene expression. The successful construction of the Ad5FSK vector, co-expressing IL-24S and IL-15 without compromising viral function, marks a major milestone in viral immunotherapy. Moreover, the noninvasive FUS-DMB delivery system further elevates the approach’s translational potential by overcoming delivery challenges unique to the brain’s anatomy.

Importantly, the FUS-DMB platform’s versatility extends beyond glioblastoma treatment. By enhancing delivery of viral and molecular therapeutics across the BBB, this technology could be adapted to target other intracranial tumors or neurological disorders requiring CNS gene delivery. The increased targeting precision and systemic administration route represent powerful advantages over localized, invasive delivery techniques traditionally employed in neuro-oncology and neurology.

Looking ahead, the research team plans to expand preclinical testing using clinical GBM tumor samples and to eventually transition into human trials. The long-term vision includes applying this combined fusion superkine and focused ultrasound delivery strategy to not only primary brain cancers but also secondary brain tumors arising from metastases outside the CNS. Such advancements could profoundly alter the treatment paradigm, moving from palliative interventions towards noninvasive cures.

This innovative study was supported by numerous funding entities, including the National Foundation for Cancer Research and the National Cancer Institute, and involved a multidisciplinary collaborative team spanning molecular biology, immunology, neurosurgery, and biomedical engineering. The authors disclosed relevant ties to InterLeukin Combinatorial Therapies, Inc., reflecting ongoing translational and commercialization paths for this promising technology.

In summary, the creation of a fusion superkine that couples the selective tumoricidal power of IL-24S with the immune mobilizing capacity of IL-15, delivered through an ingeniously designed noninvasive focused ultrasound microbubble platform, stands out as a pioneering breakthrough in glioblastoma immunotherapy. This multifaceted treatment not only achieves potent tumor cell killing but also harnesses and revitalizes the immune system’s ancient defenses within the brain’s hostile environment. If clinical trials validate these findings, patients suffering from glioblastoma may soon have access to a therapy with curative potential, breaking a long-standing impasse in brain cancer treatment that has persisted for decades.

Subject of Research: Animals

Article Title: Novel fusion superkine, IL-24S/IL-15, enhances immunotherapy of brain cancer

News Publication Date: 21-Jun-2025

Web References:

• Journal for ImmunoTherapy of Cancer Article
• DOI: 10.1136/jitc-2024-011198

Image Credits: VCU Massey Comprehensive Cancer Center

Keywords: Brain cancer, Glioblastomas, Blood brain barrier