Preclinical characterization of a novel FAP-targeted PET tracer, [⁶⁸Ga]Ga-CB6NT-FAPI

Fibroblast activation protein (FAP) is an established target for molecular imaging of the tumor stroma. In this study, we report the development and preclinical evaluation of a novel FAP-targeted positron emission tomography (PET) tracer, [⁶⁸Ga]Ga-CB6NT-FAPI. [⁶⁸Ga]Ga-CB6NT-FAPI was efficiently radiolabeled under mild conditions (room temperature, 10 min), yielding high radiochemical purity (>99.9%) and demonstrating excellent in vitro stability. In vitro studies showed significantly higher uptake in FAP-positive cells than in FAP-negative cells, which was reduced by excess unlabeled ligand, confirming FAP-specific binding. In vivo PET/MR imaging in a U87MG xenograft model demonstrated clear tumor uptake with favorable contrast. Blocking studies resulted in a marked reduction in tumor signal, and ex vivo biodistribution confirmed specific tumor accumulation with an approximately 99.9% reduction under blocking conditions. The tracer exhibited partial hepatobiliary involvement in addition to renal clearance. These findings indicate that CB6NT-FAPI represents a pharmacokinetically differentiated FAPI tracer that expands the diversity of FAP-targeted PET imaging by providing an alternative balance between renal and hepatobiliary clearance.