Preparation and Bioevaluation of 99m Tc-Labeled FAP Inhibitors as Tumor Radiotracers to Target the Fibroblast Activation Protein.

Fibroblast activation protein (FAP) is overexpressed in cancer-associated fibroblasts (CAFs) in a majority of human epithelial cancers. With low expression in normal organs, FAP has become a promising molecular target for tumor theranostics. To develop a lower cost and more widely available alternative to positron emission tomography (PET), two isocyanide-containing FAP inhibitors (CN-C 5 -FAPI and CN-PEG 4 -FAPI) were synthesized and radiolabeled with 99m Tc to obtain [ 99m Tc][Tc-(CN-C 5 -FAPI) 6 ] + and [ 99m Tc][Tc-(CN-PEG 4 -FAPI) 6 ] + in high yields (>95%). They showed good stability in saline and mouse serum. The partition coefficient (log  P ) values of [ 99m Tc][Tc-(CN-C 5 -FAPI) 6 ] + and [ 99m Tc ] [Tc-(CN-PEG 4 -FAPI) 6 ] + were -0.86 ± 0.03 and -2.38 ± 0.07, respectively, indicating that they were good hydrophilic complexes. The low nanomolar IC 50 values of CN-C 5 -FAPI and CN-PEG 4 -FAPI indicated that they had specificity to FAP. In vitro cellular uptake and blocking experiments implied a FAP-targeted uptake mechanism. The nanomolar K d values from the saturation binding assay indicated that they had significantly high target affinity to FAP. The biodistribution and blocking study in BALB/c nude mice bearing U87MG tumors showed that both exhibited specific tumor uptake. [ 99m Tc][Tc-(CN-PEG 4 -FAPI) 6 ] + showed a higher tumor uptake and a higher tumor/nontarget ratio than [ 99m Tc][Tc-(CN-C 5 -FAPI) 6 ] + . The results of micro-single-photon emission computed tomography (SPECT) imaging studies of [ 99m Tc][Tc-(CN-C 5 -FAPI) 6 ] + and [ 99m Tc ] [Tc-(CN-PEG 4 -FAPI) 6 ] + were in accordance with the biodistribution results, suggesting that [ 99m Tc][Tc-(CN-PEG 4 -FAPI) 6 ] + is a promising tumor imaging agent for targeting FAP.