Preclinical evaluation of [18F]AlF-FAPI-74 for PET imaging to study cancer-associated fibroblast responses to radiotherapy

Background Cancer-associated fibroblasts (CAFs) are influential elements of the tumor microenvironment with significant roles in tumor progression and therapy resistance. However, if and how CAF-mediated responses to radiotherapy (RT) affects clinical outcomes remains undetermined. Here, we aimed to investigate impact of RT on CAFs using antigen-specific, non-invasive, molecular PET-imaging. The FAP-specific radiotracer [ ¹⁸ F]AlF-FAPI-74 was applied to monitor CAF dynamics following external beam RT in two syngeneic subcutaneous murine tumor models (LLC and CT26). Tumors were irradiated using two radiation regimens (1x12 Gy or 2x6 Gy), and PET/MR imaging was performed 7 days post-RT. Additionally, dynamics of FAP + CAFs in tumors was quantified ex vivo using flow cytometry and immunohistochemistry. Results Biodistribution studies of [ ¹⁸ F]AlF-FAPI-74 showed radiotracer signal in joint/bone structures and intestines in both mouse strains. Tumor-targeted irradiation led to significant reduction in tumor size. Uptake of [ ¹⁸ F]AlF-FAPI-74 in subcutaneous tumors was low but significantly above muscle-background values. Quantification of standardized uptake values (SUV) from static PET-images revealed two-fold increased PET signal in LLC tumors irradiated with 2x6 Gy. Ex vivo analysis confirmed low abundance of FAP + cells in tumors and demonstrated similar RT-induced changes in CAFs across the different models. Conclusions Our findings suggest that CAFs represent a relatively sparse cell population in subcutaneously transplanted tumor models, and that radiotherapy may induce a moderate increase in FAP ⁺ cells in LLC tumors. Additionally, we demonstrate that [ ¹⁸ F]AlF-FAPI-74 is a reliable biomarker for evaluating levels of FAP + stromal cells in tumors and for addressing potential therapy-induced changes in CAFs.