Design And Synthesis Of Small Molecule Inhibitors Of Fibroblast Activation Protein (FAP) And Its Application As A Tumor Imaging Probe

Fibroblast activation protein（FAP）is overexpressed in tumor-associated fibroblasts and is involved in various pro-tumor effects,such as angiogenesis,matrix remodeling,chemotherapy resistance,and immune suppression.FAP is upregulated in over 90%of human epithelial malignancies and has no significant expression in normal adult tissues,making it a promising target for tumor diagnosis and treatment.To improve the imaging efficacy of existing FAP imaging probes,this dissertation has developed a series of FAP small-molecule inhibitors with high affinity and applied them to tumor diagnosis.The specific research is as follows:（1）In order to systematically evaluate the structure-activity relationship of FAP small-molecule inhibitors,this dissertation designed and synthesized18 small-molecule inhibitors targeting the quinoline C6 site to investigate the effects of substituent type,configuration,push-pull electronic ability,and other factors on FAP affinity.This dissertation used the GP-AMC substrate to test the IC₅₀ values of these small-molecule inhibitors and after several rounds of screening,and identified a high-affinity structure,QI-18,which had an affinity 6.4 times higher than the classic structure UAMC-1110.The improvement in affinity helps to improve its efficacy in tumor diagnosis and treatment and lays a good foundation for subsequent functional studies.（2）Based on the research foundation of FAP small-molecule inhibitors,this dissertation designed and synthesized a targeted FAP radiopharmaceutical probe for tumor diagnosis.The probe[⁶⁸Ga]Ga-CY03 has excellent FAP affinity and can quickly accumulate in the tumor area,with high tumor uptake values and tumor-to-non-target ratios.Compared with the classic probe[⁶⁸Ga]Ga-FAPI-04,the probe[⁶⁸Ga]Ga-CY03 also has significant advantages in imaging,with high application value and clinical translation potential.（3）This dissertation further explored the possibility of using small-molecule inhibitors as FAP fluorescence probes.Two FAP fluorescence probes were designed based on our previous study.Among them,the activatable FAP molecular rotor probe FR-01 showed a 10-fold activation ratio in vitro and is a potential FAP diagnostic tracer.In summary,this dissertation has developed a series of small-molecule inhibitors with high FAP affinity and applied them to tumor diagnosis.By systematically studying the structure-activity relationship of small-molecule inhibitors,this dissertation has screened inhibitors at the picomolar level.FAP imaging probes prepared with this targeting group have excellent tumor diagnostic capabilities.These achievements provide guidance for the further development of FAP diagnostic and therapeutic probes.