Radioactivity- and time-dependent cellular cytotoxicity of astatine-211-labeled FAP-targeting radiopharmaceuticals

Abstract Fibroblast activation protein (FAP), which is overexpressed in cancer-associated fibroblasts (CAFs) is an attractive stromal target for a-therapy. We previously reported moderate anti-tumor effects of 211 At-labeled FAP inhibitors (FAPI) in a PANC-1 xenograft mouse model, where insufficient tumor accumulation and the absence of FAP expression in tumor cells limited therapeutic efficacy. In this study, we performed in vitro mechanistic follow-up experiments using FAP-negative PANC-1 cells to clarify the radioactivity–time-dependent cytotoxicity of 211 At-labeled FAPI compared with free 211 At (non-conjugated 211 At). We found that, whereas free 211 At showed only limited cytotoxicity, 211 At-labeled FAPI exerted significant cytotoxic effects. Systematic variation of activity concentrations and exposure durations revealed that cytotoxicity depended primarily on sustained pericellular exposure rather than initial radioactivity levels. These findings provide mechanistic, hypothesis-generating insight into how CAF-targeted 211 At radiopharmaceuticals can exert effective cytotoxicity without direct tumor cell targeting and may inform future optimization of stromal-targeted a-therapy, while further in vivo and translational studies will be required.