Abstract 2928: TROP2 ProTriTAC™, a protease-activated T cell engager prodrug targeting TROP2 for the treatment of solid tumors

Tumor-associated calcium signal transducer 2 (Trop2) is a cell surface glycoprotein that promotes cell renewal, proliferation and tumorigenesis. Trop2 is homogenously overexpressed in numerous solid tumor types, including breast, head and neck, prostate, bladder, and non-small cell carcinoma; however, Trop2 is also expressed in several normal tissues, including skin, respiratory tract, and female reproductive organs. Although Trop2-targeted antibody drug conjugates (ADCs) have demonstrated clinical activity, with regulatory approvals in subsets of breast and bladder cancers, their broader utility is still hampered by off-target payload-mediated toxicities. To address this issue, and to avoid on-target off-tumor toxicities, we developed a Trop2-targeting ProTriTAC™ (Protease-activated Tri-specific T cell Activating Construct), a T cell engager prodrug which is engineered to be preferentially activated in tumor tissue. ProTriTACs consists of three humanized antibody-derived binding domains on a single polypeptide chain: anti-albumin for half-life extension, anti-CD3 for T cell engagement, and anti-target antigen for tumor cell engagement. The anti-albumin domain, bearing a masking moiety and a protease-cleavable linker, keeps the prodrug inert by inhibiting binding of the adjacent anti-CD3 domain to T cells. Cleavage of the linker by tumor-associated proteases removes the anti-albumin domain along with the masking moiety to reveal a potent active drug that directs T cell killing within the tumor. As designed, in vitro binding and functional assays show that the protease-activated T cell engager has >1000-fold improved binding to primary human T cells and ~100-fold improved T cell killing compared to its intact masked prodrug. In immunodeficient mice reconstituted with human T cells, anti-Trop2 ProTriTAC demonstrated robust anti-tumor activity in multiple xenograft tumor models, including HCC70 (breast), CAL27 (head and neck), and HPAFII (pancreatic), with complete tumor regression as low as 30 μg/kg. To determine the potential toxicity and toxicokinetic characteristics of anti-Trop2 ProTriTAC, an intra-animal dose escalation was conducted in cynomolgus monkeys until individual maximum tolerated dose (MTD). The starting dose was 20 μg/kg with weekly 3-fold dose escalations until MTD. The one-month study revealed that anti-Trop2 ProTriTAC was tolerated at the 180 μg/kg dose level, but not at the next higher dose level of 540 μg/kg. Toxicokinetics, gross pathol