Preparation and Bioevaluation of <sup>99m</sup>Tc-Labeled FAP Inhibitors as Tumor Radiotracers to Target the Fibroblast Activation Protein
Qing Ruan, Junhong Feng, Yuhao Jiang, Xuran Zhang, Xiaojiang Duan, Qianna Wang, Guangxing Yin, Di Xiao, Junbo Zhang
Abstract
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-C5-FAPI and CN-PEG4-FAPI) were synthesized and radiolabeled with 99mTc to obtain [99mTc][Tc-(CN-C5-FAPI)6]+ and [99mTc][Tc-(CN-PEG4-FAPI)6]+ in high yields (>95%). They showed good stability in saline and mouse serum. The partition coefficient (log P) values of [99mTc][Tc-(CN-C5-FAPI)6]+ and [99mTc][Tc-(CN-PEG4-FAPI)6]+ were −0.86 ± 0.03 and −2.38 ± 0.07, respectively, indicating that they were good hydrophilic complexes. The low nanomolar IC50 values of CN-C5-FAPI and CN-PEG4-FAPI indicated that they had specificity to FAP. In vitro cellular uptake and blocking experiments implied a FAP-targeted uptake mechanism. The nanomolar Kd 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. [99mTc][Tc-(CN-PEG4-FAPI)6]+ showed a higher tumor uptake and a higher tumor/nontarget ratio than [99mTc][Tc-(CN-C5-FAPI)6]+. The results of micro-single-photon emission computed tomography (SPECT) imaging studies of [99mTc][Tc-(CN-C5-FAPI)6]+ and [99mTc][Tc-(CN-PEG4-FAPI)6]+ were in accordance with the biodistribution results, suggesting that [99mTc][Tc-(CN-PEG4-FAPI)6]+ is a promising tumor imaging agent for targeting FAP.