Synthesis and <i>In Vivo</i> and <i>In Vitro</i> Properties of ^99m Tc-Labeled Fibroblast Activation Protein Inhibitors

Fibroblast activation protein (FAP) is expressed in more than 90% of tumor-associated fibroblasts in epithelial cancers, providing an excellent target for nuclear medicine diagnostics and therapy. Given the high cost of PET-CT, developing SPECT probes targeting FAP is necessary. A novel FAP inhibitor derived from UAMC1110 was synthesized and conjugated to DOTA via PEG chains. This resulted in a series of inhibitors with good targeting specificity, tumor uptake, and pharmacokinetics. In this study, UAMC1110 derivatives were used as FAP-targeting pharmacophores; PEG chains of varying lengths were employed for pharmacokinetic modification, and HYNIC was used as a bifunctional chelator. The derivatives were labeled with ^99mTc using different coligand combinations to explore how PEG chain length and coligand composition affect the <i>in vivo</i> and <i>in vitro</i> properties of ^99mTc-labeled FAPI. Four UAMC1110 derivatives (P4, P6, P8, P12) with different PEG chain lengths were synthesized, and a series of hydrophilic ^99mTc complexes were prepared. Stability and specificity studies demonstrated that these complexes exhibited good <i>in vitro</i> and <i>in vivo</i> stability and FAP-targeting specificity. In micro-SPECT imaging, these tracers showed rapid tumor accumulation, with ^99mTc-TE-P12 and ^99mTc-TT-P12 showing promising tumor uptake, low nontarget organ uptake, and high T/NT ratios, indicating potential as SPECT probes.