Design and Characterization of Prodrugged Anti-CTLA-4 Antibodies

Therapeutic antibodies are widely used to treat diseases like cancer and inflammatory conditions by binding with high specificity to their molecular targets. Masking is a strategy to mitigate undesirable activity in nontarget tissues, improving safety and pharmacokinetic (PK) profiles by reducing target-mediated drug disposition. In this article, masking of an anti-CTLA-4 antibody by conjugating a large PEG molecule to specific sites on the antibody is explored. While anti-CTLA-4 immunotherapy benefits solid tumor treatment, its adverse events limit its utility. Multiple conjugation sites within the complementarity-determining regions and adjacent framework regions are evaluated to attenuate CTLA-4 binding. The optimal site for maximizing masking efficiency is identified, allowing for efficient bioconjugation and functional restoration upon exposure to a cleaving enzyme in the tumor microenvironment. The prodrugged antibody exhibits reduced binding to CTLA-4 and Fc gamma receptors, high stability, and an extended half-life in a mouse model. This technology has the potential to improve the PK profile and safety attributes of therapeutic antibodies.