OS02.5.A PRECLINICAL EVALUATION OF ALPHA-KETOAMIDE-BASED FAP RADIOLIGANDS FOR TARGETING BRAIN METASTASES

Abstract BACKGROUND Brain metastases are the most common malignant tumors of the central nervous system in adults, affecting approximately 20% of patients with solid cancers. Current therapeutic options remain limited. We have recently demonstrated that fibroblast activation protein (FAP) is expressed in brain metastases, making it a promising target for theranostic applications. We aimed to develop novel FAP-targeting radioligands based on alpha-ketoamide FAP inhibitors (FAPi) and assess their binding, internalization, and in vivo biodistribution. MATERIAL AND METHODS FAP expression in brain metastases and control non-tumorous brain tissue was determined by an enzymatic assay, ELISA, and immunohistochemistry. Novel alpha-ketoamide FAPi with variable linker lengths were radiolabeled with 99mTc and their binding and internalization was assessed in vitro. Biodistribution studies were conducted in mice bearing FAP-expressing subcutaneous tumors; intratumoral accumulation was visualized using SPECT imaging. The most promising candidate was further tested in an intracranial tumor model. RESULTS FAP expression was significantly higher in brain metastases compared to non-tumorous brain tissue both at the protein and enzymatic activity level. Among the tested compounds, 99mTc-labelled NS654 showed specific binding to FAP-expressing cells and efficient internalization in vitro. In vivo, it exhibited favorable pharmacokinetics, with selective accumulation in FAP-expressing subcutaneous tumors. Notably, the compound crossed the blood-tumor barrier and accumulated in intracranial tumors. CONCLUSION FAP is expressed in brain metastases of diverse solid tumors and represents a potential theranostic target. We developed a novel 99mTc-labeled radioligand based on an alpha-ketoamide FAP inhibitor that selectively accumulates in tumors and crosses the blood-tumor barrier. These findings support further development of alpha-ketoamide-based FAP-targeting compounds for imaging and potential therapeutic applications in brain metastases.