Proximity-Activated Photocatalytic Pharmacology: From Subcellular Compartments to Neural Circuits

Conspectus

Spatial precision of endogenous biological signaling is dictated by submicrometer microdomains. Traditional pharmacological modalities lack spatial resolution needed to interrogate discrete coordinates, whereas genetic methods often lack rapid kinetic control needed to capture dynamic signaling networks. While photochemical tools offer noninvasive, spatiotemporally resolved control, their biological translation is limited by UV phototoxicity, nonspecific ROS damage, and the difficulty to achieve true subcellular localization.

We summarize our development of proximity-activated photocatalytic pharmacology via spatially confined small-molecule release, focusing on mechanistic evolution of biocompatible, metal-free photocatalytic deboronative hydroxylation of organoboronates. We present a paradigm where reaction fidelity is governed not by intrinsic photochemical properties of substrates but by physical coordinates of the localized photocatalyst and short-lived reactive species generated within defined spatial boundaries. This platform operates through localized generation of transient intermediates, specifically superoxide anions (O2·–) and H2O2, whose intracellular diffusion is constrained by native cellular quenching machinery. By exploiting steep concentration gradients of species, we establish a high-resolution reaction sphere for photocatalytic bond cleavage to enable precise interrogation of biological microdomains.

Our research program progressed through three distinct generations to span increasingly complex biological scales:

• First generation (cellular scale): We established a visible-light-driven small-molecule release system using biocompatible organic xanthene dyes and physiological ascorbates. Operating through a single-electron transfer pathway, this manifold selectively reduces molecular oxygen to O2·– to drive bioorthogonal deboronative hydroxylation while bypassing cytotoxic energy transfer pathways.

• Second generation (subcellular scale): We achieved subcellular precision by engineering targeted chemical scaffolds and hybrid genetically targeted platforms (e.g., SNAP-FL) to restrict photocatalytic activity to specific organelle coordinates. By advancing to fully genetically encoded 12 kDa miniature photoenzymes (miniSOG), we eliminated exogenous chemical catalyst requirements. Together, these methods enabled us to map compartment-specific drug toxicities, drive selective mitochondrial depolarization, and achieve nucleus-specific epitranscriptomic remodeling of m6A RNA methylation.

• Third generation (circuit scale): We deployed these platforms to decipher complex architecture of the mammalian nervous system. By expressing membrane-anchored SNAP-FL at projection-specific synaptic terminals, we achieved optochemical neuromodulation in acute brain slices within seconds. Ultimately, this enabled cell-type-specific photocatalytic pharmacology in vivo, facilitating the selective release of GABABR agonists within NaV1.8+ terminals to functionally delineate peripheral neural circuits driving pruritus in mice.

By transitioning from bulk cellular activation to localized, genetically directed modulation, we provide a unified mechanistic framework that demonstrates how photocatalyst localization and confined reactive species dynamics dictate complex biological outcomes across multiple spatiotemporal scales.