PT - JOURNAL ARTICLE AU - Garate-Soraluze, Eneko AU - Serrano-Mendioroz, Irantzu AU - Fernández-Rubio, Leticia AU - De Andrea, Carlos E AU - Barrio-Alonso, Celia AU - Herrero, Claudia del Pilar AU - Teijeira, Alvaro AU - Luri-Rey, Carlos AU - Claus, Christina AU - Tanos, Tamara AU - Klein, Christian AU - Umana, Pablo AU - Rullan, Antonio AU - Simón, Jon Ander AU - Collantes, María AU - Sánchez-Mateos, Paloma AU - Melero, Ignacio AU - Rodriguez-Ruiz, Maria E TI - 4-1BB agonist targeted to fibroblast activation protein α synergizes with radiotherapy to treat murine breast tumor models AID - 10.1136/jitc-2024-009852 DP - 2025 Feb 01 TA - Journal for ImmunoTherapy of Cancer PG - e009852 VI - 13 IP - 2 4099 - http://jitc.bmj.com/content/13/2/e009852.short 4100 - http://jitc.bmj.com/content/13/2/e009852.full SO - J Immunother Cancer2025 Feb 01; 13 AB - Background Ionizing radiation (IR) is a double-edged sword for immunotherapy as it may have both immunosuppressive and immunostimulatory effects. The biological effects of IR on the tumor microenvironment (TME) are a key factor for this balance. Fibroblast activation protein (FAP) is expressed on the surface of cancer-associated fibroblasts (CAF) in many cancer types and its abundance is associated with the poor immune response to immune-checkpoint-blockade in patients. We hypothesized that IR increases FAP expression in CAFs, therefore the combination of IR with targeted immunomodulators such as an agonistic anti-FAP-4-1BBL fusion protein could enhance the immune-mediated antitumoral effects of these treatments.Methods The murine transplantable TS/A tumor-cell-line co-engrafted with CAFs was used to investigate increases in FAP expression in tumors following irradiation using immunohistochemistry, real-time polymerase chain reaction (RT-PCR) and multiplex tissue immunofluorescence. One lesion of bilateral tumor-bearing mice was only locally irradiated or combined with weekly injections of the bispecific muFAP-4-1BBL fusion protein (a mouse surrogate for RG7826). Tumor sizes were followed over time and TME was assessed by flow cytometry. Selective monoclonal antibody (mAb)-mediated depletions of immune cell populations, neutralizing interferon alpha/beta receptor 1 (IFNAR-I) IFNAR and interferon (IFN)-γ mAbs and gene-modified mice (4-1BB−/−) were used to delineate the immune cell subsets and mechanisms required for efficacy. 67Ga labeled muFAP-4-1BBL tracked by SPECT-CT was used to study biodistribution. In human colorectal carcinoma samples, the inducibility of FAP expression following radiotherapy was explored by multiplex immunofluorescence.Results Irradiation of TS/A+CAF tumors in mice showed an increase in FAP levels after local irradiation. A suboptimal radiotherapy regimen in combination with muFAP-4-1BBL attained primary tumor control and measurable abscopal effects. Immune TME landscape analyses showed post-treatment increased infiltration of activated immune cells associated with the combined radioimmunotherapy treatment. Efficacy depended on CD8+ T cells, type I IFN, IFN-γ and ability to express 4-1BB. Biodistribution studies of muFAP-4-1BBL indicated enriched tumor targeting to irradiated tumors. Human colorectal cancer samples pre and post irradiation showed enhanced FAP expression after radiotherapy.Conclusion Increased FAP expression in the TME as a result of radiotherapy can be exploited to target agonist 4-1BB immunotherapy to malignant tumor lesions using an FAP-4-1BBL antibody fusion protein.All data relevant to the study are included in the article or uploaded as supplementary information.