Formulation and Preclinical Testing of Tc-99m-Labeled HYNIC-Glc-FAPT as a FAP-Targeting Tumor Radiotracer.

Molecular imaging and targeted radiotherapy with radiolabeled fibroblast activation protein inhibitor (FAPI) targeting peptide probes hold great potential for enhancing the clinical management of patients with FAP-expressing cancers. However, the high cost of PET probes has prompted us to search for new FAP-targeting single-photon imaging agents. In this study, HYNIC-Glc-FAPT is synthesized and radiolabeled with technetium-99m using tricine/EDDA or dimer tricine as coligands to produce [Tc]Tc-tricine/EDDA-HYNIC-Glc-FAPT and [Tc]Tc-tricine-HYNIC-Glc-FAPT. Both [Tc]Tc-tricine/EDDA-HYNIC-Glc-FAPT and [Tc]Tc-tricine-HYNIC-Glc-FAPT were effectively synthesized with an excellent radiochemistry yield (both >97%, = 6) in a single-step technique, and their stability in PBS and human serum was satisfactory. Compared to [Tc]Tc-tricine-HYNIC-Glc-FAPT, [Tc]Tc-tricine/EDDA-HYNIC-Glc-FAPT exhibited a more hydrophilic nature with a log  of -3.53 ± 0.12. In vitro cellular uptake and blocking assays, internalization, efflux experiments, and affinity experiments all suggested a mechanism with high FAP-specificity and affinity. SPECT imaging and biodistribution of [Tc]Tc-tricine/EDDA-HYNIC-Glc-FAPT demonstrated sustained high tumor uptake in BALB/c nude mice bearing U87MG tumors for 6 h. It demonstrated a long-range retention characteristic and more rapid clearance ability from nontarget organs. Collectively, we successfully synthesized [Tc]Tc-tricine/EDDA-HYNIC-Glc-FAPT and [Tc]Tc-tricine-HYNIC-Glc-FAPT, and the excellent targeting properties of [Tc]Tc-tricine/EDDA-HYNIC-Glc-FAPT suggest a potential diagnostic value in future clinical studies for advanced-stage FAP-expressing malignancies, especially in prognostic evaluation of tumors for it low price and convenient source.