Synthesis, Preclinical Evaluation, and a Pilot Clinical PET Imaging Study of (68)Ga-Labeled FAPI Dimer

Cancer-associated fibroblasts (CAFs) are crucial components of the tumor microenvironment. Fibroblast activation protein (FAP) is overexpressed in CAFs. FAP-targeted molecular imaging agents, including the FAP inhibitors (FAPIs) 04 and 46, have shown promising results in tumor diagnosis. However, these molecules have a relatively short tumor-retention time for peptide-targeted radionuclide therapy applications. We aimed to design a (68)Ga-labeled FAPI dimer, (68)Ga-DOTA-2P(FAPI)(2), to optimize the pharmacokinetics and evaluate whether this form is more effective than its monomeric analogs. Methods: (68)Ga-DOTA-2P(FAPI)(2) was synthesized on the basis of the quinoline-based FAPI variant (FAPI-46), and its binding properties were assayed in CAFs. Preclinical pharmacokinetics were determined in FAP-positive patient-derived xenografts using small-animal PET and biodistribution experiments. The effective dosimetry of (68)Ga-DOTA-2P(FAPI)(2) was evaluated in 3 healthy volunteers, and PET/CT imaging of (68)Ga-FAPI-46 and (68)Ga-DOTA-2P(FAPI)(2) was performed on 3 cancer patients. Results: (68)Ga-DOTA-2P(FAPI)(2) was stable in phosphate-buffered saline and fetal bovine serum for 4 h. The FAPI dimer showed high affinity and specificity for FAP in vitro and in vivo. The tumor uptake of (68)Ga-DOTA-2P(FAPI)(2) was approximately 2-fold stronger than that of (68)Ga-FAPI-46 in patient-derived xenografts, whereas healthy organs showed low tracer uptake and fast body clearance. The effective dose of (68)Ga-DOTA-2P(FAPI)(2) was 1.19E−02 mSv/MBq, calculated using OLINDA. Finally, the PET/CT scans of the 3 cancer patients revealed higher intratumoral uptake of (68)Ga-DOTA-2P(FAPI)(2) than of (68)Ga-FAPI-46 in all tumor lesions (SUV(max), 8.1–39.0 vs. 1.7–24.0, respectively; P < 0.001). Conclusion: (68)Ga-DOTA-2P(FAPI)(2) has increased tumor uptake and retention properties compared with (68)Ga-FAPI-46, and it could be a promising tracer for both diagnostic imaging and targeted therapy of malignant tumors with positive expression of FAP.