One Ligand, Two Imaging Modalities: [ 99m Tc]Tc-FAPI-74 and [ 18 F]AlF-FAPI-74

Fibroblast activation protein (FAP) is highly expressed on cancer-associated fibroblasts, making it an attractive target for oncological imaging and therapy. FAPI-74, a FAP inhibitor in phase II clinical evaluation with 18 F as a positron emission tomography tracer, has exhibited high tumour uptake and specificity. Development of single photon emission computed tomography (SPECT) analogue [ 99m Tc]Tc-FAPI-74 offers advantages of lower cost and accessibility through established SPECT infrastructure. Herein, we report the radiolabeling and characterization of FAPI-74 with 99m Tc and [ 18 F]AlF, and comparative evaluation of their binding properties and biodistribution in FAP-positive and FAP-negative xenograft mouse models. FAPI-74 was radiolabeled with [ 99m Tc]Tc-tricarbonyl, achieving radiochemical conversion (RCC) of 20% and 0.32 GBq/µmol molar activity (Am). Comparatively, [ 18 F]AlF labeling resulted in 75% RCC, and 3.33 GBq/µmol Am. Both tracers exhibited >99% radiochemical purity. In vitro binding to FAP+ cells was comparable between [ 99m Tc]Tc-FAPI-74 and [ 18 F]AlF-FAPI-74 (29.9 ± 4.7% vs. 31.4 ± 11.0% binding and K D values of 14.6 nM vs. 5.09 nM). In vivo, tumour uptake in HT29 xenografts was similar (2.14 ± 0.29 vs. 1.59 ± 0.12 %ID/g) with a %ID/g of