Orthogonal Light‐ and Enzyme‐Triggered and‐Gate Prodrugs for Senolytic Therapy

ABSTRACT Senescence‐associated β‐galactosidase (SA‐β‐gal) is widely used to target senescent cells, but its activity in non‐senescent tissues can limit selectivity. Here, we developed a light–enzyme AND‐gate strategy in which an o‐nitrobenzyl photocage blocks β‐galactoside cleavage until 390 nm irradiation. The strategy was evaluated using a fluorescent probe (LS‐C), a doxorubicin prodrug (LS‐D), and an ARV‐771‐based PROTAC (LS‐A). LS‐C exhibited dual‐trigger‐dependent fluorescence activation, while LS‐D preferentially induced apoptosis and LS‐A promoted BRD4 degradation in senescent cells following light activation. Across multiple senescence models, LS‐D and LS‐A showed higher senolytic indices than their corresponding parent compounds. In senescent lung cancer patient‐derived organoids, LS‐D combined with light increased organoid death, whereas either treatment alone had minimal effects. These results provide proof‐of‐concept evidence for sequential light–enzyme AND‐gating as a strategy for controlling payload activation in senescent cells under the tested in vitro and ex vivo conditions.