Bendre_2023_Molecules_28_

Fibroblast activation protein alpha (FAP-alpha) is a cell-surface protein overexpressed on cancer-associated fibroblasts that constitute a substantial component of tumor stroma and drive tumorigenesis. FAP is minimally expressed by most healthy tissues, including normal fibroblasts. This makes it a promising pan-cancer diagnostic and therapeutic target. In the present study, we synthesized two novel tracers, [(68)Ga]Ga-SB03045 and [(68)Ga]Ga-SB03058, bearing a (2S,4S)-4-fluoropyrrolidine-2-carbonitrile or a (4R)-thiazolidine-4-carbonitrile pharmacophore, respectively. [(68)Ga]Ga-SB03045 and [(68)Ga]Ga-SB03058 were evaluated for their FAP-targeting capabilities using substrate-based in vitro binding assays, and in PET/CT imaging and ex vivo biodistribution studies in an HEK293T:hFAP tumor xenograft mouse model. The IC(50) values of (nat)Ga-SB03045 (1.59 +/- 0.45 nM) and (nat)Ga-SB03058 (0.68 +/- 0.09 nM) were found to be lower than those of the clinically validated (nat)Ga-FAPI-04 (4.11 +/- 1.42 nM). Contrary to the results obtained in the FAP-binding assay, [(68)Ga]Ga-SB03058 demonstrated a ~1.5 fold lower tumor uptake than that of [(68)Ga]Ga-FAPI-04 (7.93 +/- 1.33 vs. 11.90 +/- 2.17 %ID/g), whereas [(68)Ga]Ga-SB03045 (11.8 +/- 2.35 %ID/g) exhibited a tumor uptake comparable to that of [(68)Ga]Ga-FAPI-04. Thus, our data suggest that the (2S,4S)-4-fluoropyrrolidine-2-carbonitrile scaffold holds potential as a promising pharmacophore for the design of FAP-targeted radioligands for cancer diagnosis and therapy.