Development and evaluation of 18F-labeled novel radiopharmaceuticals for PET imaging of fibroblast activation protein expressing tumors.

This study aimed to develop and evaluate two novel 18F-labeled radiopharmaceuticals, [18F]AlF-NOTA-De-FAPI and [18F]AlF-NOTA-Glu-FAPI, for Positron Emission Tomography (PET) imaging of tumors expressing fibroblast activation protein. Molecular docking simulations predicted the binding affinities of NOTA-De-FAPI and NOTA-Glu-FAPI with FAP. The radiotracers were synthesized and evaluated for radiochemical yield, purity, and molar activity. Surface plasmon resonance analysis measured binding kinetics. In vitro and in vivo stability and distribution coefficients were assessed. PET/CT imaging and ex vivo biodistribution studies were conducted in U87MG tumor-bearing mice. A pilot clinical study compared [18F]AlF-NOTA-De-FAPI with [68Ga]Ga-DOTA-FAPI-04 and [18F]FDG in a liver cancer patient. Molecular docking simulations showed that NOTA-De-FAPI had a slightly higher binding affinity for FAP. The radiotracers were synthesized with high purity and molar activity. SPR analysis confirmed higher binding affinity of NOTA-De-FAPI (KD = 86.35 pM) compared to NOTA-Glu-FAPI (KD = 187 pM). In PET/CT imaging, [18F]AlF-NOTA-De-FAPI demonstrated higher tumor uptake in U87MG tumor-bearing mice, with a peak SUVmax of 2.71 ± 0.39 at 1 h post-injection. Ex vivo biodistribution studies showed that [18F]AlF-NOTA-De-FAPI had a tumor uptake of 9.16 ± 0.49 %ID/g at 1 h post-injection, significantly higher than [18F]AlF-NOTA-Glu-FAPI (6.60 ± 0.82 %ID/g). In the clinical study, [18F]AlF-NOTA-De-FAPI showed strong uptake in the primary tumor but exhibited higher physiological uptake in salivary glands, thyroid, and pancreas compared to [68Ga]Ga-DOTA-FAPI-04. [18F]AlF-NOTA-De-FAPI demonstrated potential as a FAP-targeting tracer with high specific uptake and favorable tumor-to-normal tissue ratios in preclinical models. However, clinical evaluation revealed limitations, such as high physiological uptake in certain glands and lower tumor uptake compared to [68Ga]Ga-DOTA-FAPI-04. Further optimization and clinical validation are needed.