Structurally Rigid 9‑Amino-benzo[<i>c</i>]cinnoliniums Make Up a Class of Compact and Large Stokes-Shift Fluorescent Dyes for Cell-Based Imaging Applications

Classic fluorescent dyes, such as coumarin, naphthalimide, fluorescein, BODIPY, rhodamine, and cyanines, are cornerstones of various spectroscopic and microscopic methods, which hold a prominent position in biological studies. We recently found that 9-amino-benzo­[<i>c</i>]­cinnoliniums make up a novel group of fluorophores that can be used in biological studies. They are featured with a succinct conjugative push–pull backbone, a broad absorption band, and a large Stokes shift. They are potentially useful as a small-molecule alternative to R-phycoerythrin to pair with fluorescein in multiplexing applications

GSH-Activated NIR Fluorescent Prodrug for Podophyllotoxin Delivery

Theranostic prodrug therapy enables the targeted delivery of anticancer drugs with minimized adverse effects and real-time <i>in situ</i> monitoring of activation of the prodrugs. In this work, we report the synthesis and biological assessment of the near-infrared (NIR) prodrug DCM-S-PPT and its amphiphilic copolymer (<i>m</i>PEG-DSPE)-encapsulated nanoparticles. DCM-S-PPT is composed of podophyllotoxin (PPT) as the anticancer moiety and a dicyanomethylene-4<i>H</i>-pyran (DCM) derivative as the NIR fluorescent reporter, which are linked by a thiol-specific cleavable disulfide bond. <i>In vitro</i> experiments indicated that DCM-S-PPT has low cytotoxicity and that glutathione (GSH) can activate DCM-S-PPT resulting in PPT release and a concomitant significant enhancement in NIR fluorescence at 665 nm. After being intravenously injected into tumor-bearing nude mice, DCM-S-PPT exhibited excellent tumor-activated performance. Furthermore, we have demonstrated that <i>m</i>PEG-DSPE as a nanocarrier loaded with DCM-S-PPT (<i>m</i>PEG-DSPE/DCM-S-PPT) showed even greater tumor-targeting performance than DCM-S-PPT on account of the enhanced permeability and retention effect. Its tumor-targeting ability and specific drug release in tumors make DCM-S-PPT a promising prodrug that could provide a significant strategy for theranostic drug delivery systems

Förster Resonance Energy Transfer Switchable Self-Assembled Micellar Nanoprobe: Ratiometric Fluorescent Trapping of Endogenous H₂S Generation via Fluvastatin-Stimulated Upregulation

H₂S produced in small amounts by mammalian cells has been identified in mediating biological signaling functions. However, the in situ trapping of endogenous H₂S generation is still handicapped by a lack of straightforward methods with high selectivity and fast response. Here, we encapsulate a semi-cyanine-BODIPY hybrid dye (<b>BODInD-Cl</b>) and its complementary energy donor (<b>BODIPY1</b>) into the hydrophobic interior of an amphiphilic copolymer (<i>m</i>PEG-DSPE), especially for building up a ratiometric fluorescent H₂S nanoprobe with extraordinarily fast response. A remarkable red-shift in the absorption band with a gap of 200 nm in the H₂S response can efficiently switch off the Förster resonance energy transfer (FRET) from <b>BODIPY1</b> to <b>BODInD-Cl</b>, subsequently recovering the donor fluorescence. Impressively, both the interior hydrophobicity of supramolecular micelles and electron-withdrawing nature of indolium unit in <b>BODInD-Cl</b> can sharply increase aromatic nucleophilic substitution with H₂S. The ratiometric strategy based on the unique self-assembled micellar aggregate <b>NanoBODIPY</b> achieves an extremely fast response, enabling in situ imaging of endogenous H₂S production and mapping its physiological and pathological consequences. Moreover, the amphiphilic copolymer renders the micellar assembly biocompatible and soluble in aqueous solution. The established FRET-switchable macromolecular envelope around <b>BODInD-Cl</b> and <b>BODIPY1</b> enables cellular uptake, and makes a breakthrough in the trapping of endogenous H₂S generation within raw264.7 macrophages upon stimulation with fluvastatin. This study manifests that cystathione γ-lyase (CSE) upregulation contributes to endogenous H₂S generation in fluvastatin-stimulated macrophages, along with a correlation between CSE/H₂S and activating Akt signaling pathway