In vivo activation of FAP-cleavable small molecule-drug conjugates for the targeted delivery of camptothecins and tubulin poisons to the tumor microenvironment

Small molecule -drug conjugates (SMDCs) are increasingly considered as a therapeutic alternative to antibodydrug conjugates (ADCs) for cancer therapy. OncoFAP is an ultra -high affinity ligand of Fibroblast Activation Protein (FAP), a stromal tumor -associated antigen overexpressed in a wide variety of solid human malignancies. We have recently reported the development of non -internalizing OncoFAP-based SMDCs, which are activated by FAP thanks to selective proteolytic cleavage of the -GlyPro- linker with consequent release of monomethyl auristatin E (MMAE) in the tumor microenvironment. In this article, we describe the generation and the in vivo characterization of FAP-cleavable OncoFAP-drug conjugates based on potent topoisomerase I inhibitors (DXd, SN-38, and exatecan) and an anti-tubulin payload (MMAE), which are already exploited in clinical -stage and approved ADCs. The Glycine-Proline FAP-cleavable technology was directly benchmarked against linkers found in AdcetrisTM, EnhertuTM, and TrodelvyTM structures by means of in vivo therapeutic experiments in mice bearing tumors with cellular or stromal FAP expression. OncoFAP-GlyPro-Exatecan and OncoFAP-GlyPro-MMAE emerged as the most efficacious anti -cancer therapeutics against FAP-positive cellular models. OncoFAPGlyPro-MMAE exhibited a potent antitumor activity also against stromal models, and was therefore selected for clinical development.