Novel ⁶⁸Ga-Labeled Pyridine-Based Fibroblast Activation Protein-Targeted Tracers with High Tumor-to-Background...

Compared to quinoline-based fibroblast activation protein (FAP)-targeted radiotracers, pyridine-based FAP-targeted tracers are expected to have faster pharmacokinetics due to their smaller molecular size and higher hydrophilicity, which we hypothesize would improve the tumor-to-background image contrast. We aim to develop ⁶⁸Ga-labeled pyridine-based FAP-targeted tracers for cancer imaging with positron emission tomography (PET), and compare their imaging potential with the clinically validated [⁶⁸Ga]Ga-FAPI-04. Two DOTA-conjugated pyridine-based AV02053 and AV02070 were synthesized through multi-step organic synthesis. IC₅₀(FAP) values of Ga-AV02053 and Ga-AV02070 were determined by an enzymatic assay to be 187 ± 52.0 and 17.1 ± 4.60 nM, respectively. PET imaging and biodistribution studies were conducted in HEK293T:hFAP tumor-bearing mice at 1 h post-injection. The HEK293T:hFAP tumor xenografts were clearly visualized with good contrast on PET images by [⁶⁸Ga]Ga-AV02053 and [⁶⁸Ga]Ga-AV02070, and both tracers were excreted mainly through the renal pathway. The tumor uptake values of [⁶⁸Ga]Ga-AV02070 (7.93 ± 1.88%ID/g) and [⁶⁸Ga]Ga-AV02053 (5.6 ± 1.12%ID/g) were lower than that of previously reported [⁶⁸Ga]Ga-FAPI-04 (12.5 ± 2.00%ID/g). However, both [⁶⁸Ga]Ga-AV02070 and [⁶⁸Ga]Ga-AV02053 showed higher tumor-to-background (blood, muscle, and bone) uptake ratios than [⁶⁸Ga]Ga-FAPI-04. Our data suggests that pyridine-based pharmacophores are promising for the design of FAP-targeted tracers. Future optimization on the selection of a linker will be explored to increase tumor uptake while maintaining or even further improving the high tumor-to-background contrast.