Synthesis and Preclinical Evaluation of Druglike ¹⁸F-Labeled Fibroblast Activation Protein (FAP) Inhibitors with Enhanced Tumor Retention

Fibroblast activation protein (FAP) is upregulated in cancer and fibrosis, making it an ideal target for imaging and therapy. Most FAP radioligands are large, highly polar, chelator-based molecules that suffer from limited tissue penetration and rapid tumor washout. In this study, we developed two covalently ¹⁸F-labeled, druglike FAP inhibitors, [¹⁸F]<b>5a</b> and [¹⁸F]<b>5b</b>, featuring quaternary ammonium moieties linked via PEG chains to enhance tumor retention while maintaining high selectivity and favorable pharmacokinetics. Both radiotracers showed high affinity and specific uptake <i>in vitro</i> and <i>in vivo</i>. Compared to the clinically used [¹⁸F]AlF-NOTA-FAPI-74, [¹⁸F]<b>5a</b> and [¹⁸F]<b>5b</b>, exhibited significantly improved tumor retention at 6 h p.i. ([¹⁸F]<b>5a</b>: 4.48 ± 0.34%IA/g; [¹⁸F]<b>5b</b>: 6.70 ± 0.22%IA/g and [¹⁸F]AlF-NOTA-FAPI-74:0.54 ± 0.08%IA/g). These findings highlight the importance of polarity tuning and the utility of quaternary ammonium groups for obtaining sustained tumor retention. They offer a valuable design strategy for novel radiotheranostic ligands that contain covalently bound radionuclides for imaging and treatment of FAP-positive tumors.