Valkyrie Probes: A Novel Class of Enzyme-Activatable Photosensitizers based on Sulfur- and Seleno-Rosamines with Pyridinium Unit

The rational design of activatable photosensitizers (aPSs) uncaged by specific disease biomarkers is currently booming due to their positive attributes to achieve targeted photodynamic therapy (PDT). In this context, we present here the synthesis and detailed photophysical characterization of a novel class of hetero-rosamine dyes bearing sulfur or selenium as bridging heavy atom and 4-pyridyl meso-substituent as optically tunable group. The main feature of such photoactive platforms is the spectacular change of their spectral properties depending on the caging/decaging status of their 4-pyridyl moiety (cationic pyridinium vs. neutral pyridine). The preparation of two alkaline phosphatase (ALP)-responsive probes (named Valkyrie probes) was achieved through formal N-quaternarization with 4-phosphoryloxybenzyl, the traditional recognition moiety for this important diagnostic enzyme. Bio-analytical validations including fluorescence/singlet oxygen phosphorescence enzyme assays and RP-HPLC-fluorescence/-MS analyses have enabled us to demonstrate the viability and effectiveness of this novel photosensitizer activation strategy. Since sulfur-containing Valkyrie probe also retains high fluorogenicity in the orange-red spectral range, this study highlights meso-pyridyl-substituted S-pyronin scaffolds as valuable candidates for the rapid construction of molecular phototheranostic platforms suitable for combined fluorescence diagnosis and PDT. "Two-in-one" hetero-xanthene-based probes! Sulfur and selenium analogs of rosamines bearing 4-pyridyl moiety as meso-substituent, were identified as valuable scaffolds for facile designing enzyme-activatable photosensitizers (Valkyrie probes) through N-quaternarization of their pyridine unit. Decaging strategy that readily unveils both fluorescence and/or photosensitization properties of such hetero-xanthene dyes, was demonstrated with alkaline phosphatase, thus enriching the arsenal of tools suitable for the development of smart light-responsive molecules for targeted photodynamic therapy and phototheranostics.image