Periodic Reporting for period 4 - ViroPedTher (Oncolytic viruses for the treatment of pediatric brain tumors: An integrated clinical and lab approach)
Reporting period: 2023-09-01 to 2025-02-28
The overarching goal of our research is to develop new therapeutic strategies to improve the prognosis of children with high-risk brain tumors, such as Diffuse Midline Gliomas (DMGs). These tumors are among the most devastating diagnoses in childhood, with almost universal mortality and no effective therapies — a reality that represents both a scientific challenge and a moral imperative. To tackle this, we translated laboratory discoveries into the clinic, launching a first-in-children Phase I trial of the oncolytic adenovirus DNX-2401 (Delta-24-RGD) in combination with radiotherapy for children with DIPG. Published in the New England Journal of Medicine, this trial demonstrated that the intratumoral injection of DNX-2401 is safe and well-tolerated, extending the median overall survival to 17.8 months and inducing a measurable immune response, characterized by increased T cell infiltration and a transient pro-inflammatory shift. For the first time, families confronting DIPG could see evidence that an experimental therapy might alter the disease course. These findings highlighted both the promise and the limitations of the parental virus, motivating two complementary strategies: (i) combining Delta-24-RGD with small molecules (e.g. ONC201), immunomodulatory antibodies (e.g. CD40 agonists), and radiotherapy; and (ii) engineering next-generation adenoviruses with enhanced immune-stimulatory features. This effort led to the development of Delta-24-ACT, a derivative encoding an immune-activating ligand, which has shown superior efficacy to Delta-24-RGD in preclinical models of DIPG as well as in adult glioma and osteosarcoma. Delta-24-ACT is now being advanced toward clinical testing. In parallel, we expanded our preclinical toolkit by creating immunocompetent orthotopic mouse models of other lethal pediatric brain tumors, including atypical teratoid/rhabdoid tumors (AT/RTs) and embryonal tumors with multilayered rosettes (ETMRs), enabling rigorous testing of viral candidates across entities. Our search for additional immune vulnerabilities led us to identify TIM-3 as a non-canonical checkpoint upregulated in DMGs. In studies published in Cancer Cell, we demonstrated that TIM-3 blockade, alone or in combination with radiotherapy, significantly improves survival and reshapes the tumor immune landscape in murine models. Building on these insights, we are now focused on engineering more potent and selective oncolytic viruses, incorporating advanced immune-modulatory features to amplify antitumor activity while preserving safety. By combining clinical translation with cutting-edge immunovirology, and by expanding both preclinical models and therapeutic strategies, this ERC-supported project has laid the groundwork for transformative therapies. These next-generation immune-activating viruses hold the potential to change the therapeutic landscape for pediatric brain tumors and, most importantly, to bring hope of durable benefit to children and families facing diseases once deemed untreatable.
High-risk pediatric brain tumors such as diffuse midline gliomas (DMGs, formerly DIPGs), atypical teratoid/rhabdoid tumors (AT/RTs), and embryonal tumors with multilayered rosettes (ETMRs) remain some of the deadliest childhood cancers. Median survival for DMG patients is less than one year, and no curative therapies exist. This ERC Consolidator project set out to change this landscape by combining clinical insights, advanced virotherapy, and novel preclinical models to develop new treatment strategies.
Objectives
The project pursued three main aims:
1. Clinical and immune characterization, to study the immune response and tumor microenvironment in children treated with the oncolytic virus DNX-2401 (Delta-24-RGD).
2. Development of improved immune-viruses, to engineer and test next-generation adenoviruses armed with immune-stimulatory or immune-checkpoint–blocking molecules.
3. Extension to other lethal pediatric brain tumors , to establish new immunocompetent models and evaluate therapeutic potential beyond DMG.
Main Results
• Clinical trial outcomes: We conducted the world’s first phase I trial of DNX-2401 plus radiotherapy in children with DIPG (NEJM, 2022). Intratumoral virus injection was safe and feasible, extending median survival to 17.8 months versus 12 months with radiotherapy alone. Immune analysis showed T-cell activation and remodeling of the tumor microenvironment, with TIM-3 identified as a key resistance pathway (Cancer Cell, 2023).
• Next-generation viruses: Building on these findings, we created Delta-24-ACT, armed with the co-stimulatory ligand 4-1BBL, which improved survival and induced immune memory in DMG models (JCI Insight, 2022). New derivatives now combine 4-1BBL with TIM-3 blockade or other immune-modulating strategies. Combination therapies, such as DNX-2401 with ONC201 (Neuro-Oncology, 2024) or CD40 (Cell Reports Med 2025) agonists, also showed superior preclinical efficacy.
• Extension to other lethal pediatric brain tumorsWe demonstrated the therapeutic valute of oncolytic viruses for other aggressive pediatric brain tumors such as AT/RT and ETMR (Clin Can Res, 2019).
• Clinical translation: Preparations are underway for a new phase II multicenter trial of DNX-2401 in recurrent or refractory high-grade pediatric brain tumors (Princess Máxima Center, Utrecht, EudraCT-2024-515009-24-00).
Exploitation and Dissemination
Results have been widely disseminated through high-impact publications (NEJM, Cancer Cell, JCI Insight, Neuro-Oncology, Clin Cancer Res, Mol Cancer Ther), invited conference presentations, and international workshops. Several of our immune-virus designs are protected by intellectual property and form the basis of an ERC Proof-of-Concept project (POPCORN, 101212926), ensuring continuity toward translation. Our novel tumor models are being shared with the wider research community to accelerate discovery.
Impact
This ERC project has delivered the first clinical evidence that oncolytic virotherapy can remodel the immune landscape of pediatric DMGs and extend survival. It has produced a pipeline of next-generation immune-viruses and unique preclinical models for testing them. By integrating clinical data, novel therapeutics, and translational readiness, the project has laid the foundation for future trials and created new opportunities for children facing currently incurable brain tumors.
Objectives
The project pursued three main aims:
1. Clinical and immune characterization, to study the immune response and tumor microenvironment in children treated with the oncolytic virus DNX-2401 (Delta-24-RGD).
2. Development of improved immune-viruses, to engineer and test next-generation adenoviruses armed with immune-stimulatory or immune-checkpoint–blocking molecules.
3. Extension to other lethal pediatric brain tumors , to establish new immunocompetent models and evaluate therapeutic potential beyond DMG.
Main Results
• Clinical trial outcomes: We conducted the world’s first phase I trial of DNX-2401 plus radiotherapy in children with DIPG (NEJM, 2022). Intratumoral virus injection was safe and feasible, extending median survival to 17.8 months versus 12 months with radiotherapy alone. Immune analysis showed T-cell activation and remodeling of the tumor microenvironment, with TIM-3 identified as a key resistance pathway (Cancer Cell, 2023).
• Next-generation viruses: Building on these findings, we created Delta-24-ACT, armed with the co-stimulatory ligand 4-1BBL, which improved survival and induced immune memory in DMG models (JCI Insight, 2022). New derivatives now combine 4-1BBL with TIM-3 blockade or other immune-modulating strategies. Combination therapies, such as DNX-2401 with ONC201 (Neuro-Oncology, 2024) or CD40 (Cell Reports Med 2025) agonists, also showed superior preclinical efficacy.
• Extension to other lethal pediatric brain tumorsWe demonstrated the therapeutic valute of oncolytic viruses for other aggressive pediatric brain tumors such as AT/RT and ETMR (Clin Can Res, 2019).
• Clinical translation: Preparations are underway for a new phase II multicenter trial of DNX-2401 in recurrent or refractory high-grade pediatric brain tumors (Princess Máxima Center, Utrecht, EudraCT-2024-515009-24-00).
Exploitation and Dissemination
Results have been widely disseminated through high-impact publications (NEJM, Cancer Cell, JCI Insight, Neuro-Oncology, Clin Cancer Res, Mol Cancer Ther), invited conference presentations, and international workshops. Several of our immune-virus designs are protected by intellectual property and form the basis of an ERC Proof-of-Concept project (POPCORN, 101212926), ensuring continuity toward translation. Our novel tumor models are being shared with the wider research community to accelerate discovery.
Impact
This ERC project has delivered the first clinical evidence that oncolytic virotherapy can remodel the immune landscape of pediatric DMGs and extend survival. It has produced a pipeline of next-generation immune-viruses and unique preclinical models for testing them. By integrating clinical data, novel therapeutics, and translational readiness, the project has laid the foundation for future trials and created new opportunities for children facing currently incurable brain tumors.
This ERC project has pushed the boundaries of pediatric neuro-oncology by demonstrating, for the first time, the feasibility and safety of delivering an oncolytic adenovirus to children with diffuse midline gliomas. Our Phase I clinical trial showed that intratumoral administration of DNX-2401 was well-tolerated and biologically active, remodeling the tumor microenvironment and providing signs of positive therapeutic activity in this lethal disease. Building on these insights, we engineered a new generation of immune-armed adenoviruses, such as Delta-24-ACT and others, that integrate direct tumor lysis with immune modulation. This dual strategy represents a significant advancement over existing approaches, as checkpoint inhibitors alone have proven ineffective in DMG, and earlier viruses lacked sufficient immunological potency. By proving feasibility in patients and developing more potent viral candidates, this project has defined a new therapeutic paradigm for pediatric brain tumors.