Acidic‐pH‐Activatable Fluorescence Probes for Visualizing Exocytosis Dynamics

Live imaging of exocytosis dynamics is crucial for a precise spatiotemporal understanding of secretion phenomena, but current approaches have serious limitations. We designed and synthesized small‐molecular fluorescent probes that were chemically optimized for sensing acidic intravesicular pH values, and established that they can be used to sensitively and reliably visualize vesicular dynamics following stimulation. This straightforward technique for the visualization of exocytosis as well as endocytosis/reacidification processes with high spatiotemporal precision is expected to be a powerful tool for investigating dynamic cellular phenomena involving changes in the pH value.     

Alkene–Tetrazine Ligation for Imaging Cellular DNA

5‐Vinyl‐2′‐deoxyuridine (VdU) is the first reported metabolic probe for cellular DNA synthesis that can be visualized by using an inverse electron demand Diels–Alder reaction with a fluorescent tetrazine. VdU is incorporated by endogenous enzymes into the genomes of replicating cells, where it exhibits reduced genotoxicity compared to 5‐ethynyl‐2′‐deoxyuridine (EdU). The VdU–tetrazine ligation reaction is rapid (k≈0.02 M ^?1 s^?1) and chemically orthogonal to the alkyne–azide “click” reaction of EdU‐modified DNA. Alkene–tetrazine ligation reactions provide the first alternative to azide–alkyne click reactions for the bioorthogonal chemical labeling of nucleic acids in cells and facilitate time‐resolved, multicolor labeling of DNA synthesis.     

Lysosomal pH Rise during Heat Shock Monitored by a Lysosome‐Targeting Near‐Infrared Ratiometric Fluorescent Probe

Heat stroke is a life‐threatening condition, featuring a high body temperature and malfunction of many organ systems. The relationship between heat shock and lysosomes is poorly understood, mainly because of the lack of a suitable research approach. Herein, by incorporating morpholine into a stable hemicyanine skeleton, we develop a new lysosome‐targeting near‐infrared ratiometric pH probe. In combination with fluorescence imaging, we show for the first time that the lysosomal pH value increases but never decreases during heat shock, which might result from lysosomal membrane permeabilization. We also demonstrate that this lysosomal pH rise is irreversible in living cells. Moreover, the probe is easy to synthesize, and shows superior overall analytical performance as compared to the existing commercial ones. This enhanced performance may enable it to be widely used in more lysosomal models of living cells and in further revealing the mechanisms underlying heat‐related pathology.     

Stepwise Assembly of Turn-on Fluorescence Sensors in Multicomponent Metal–Organic Frameworks for in Vitro Cyanide Detection

The controlled synthesis of multicomponent metal–organic frameworks (MOFs) allows for the precise placement of multiple cooperative functional groups within a framework, leading to emergent synergistic effects. Herein, we demonstrate that turn-on fluorescence sensors can be assembled by combining a fluorophore and a recognition moiety within a complex cavity of a multicomponent MOF. An anthracene-based fluorescent linker and a hemicyanine-containing CN⁻-responsive linker were sequentially installed into the lattice of PCN-700. The selective binding of CN⁻ to hemicyanine inhibited the energy transfer between the two moieties, resulting in a fluorescence turn-on effect. Taking advantage of the high tunability of the MOF platform, the ratio between anthracene and the hemicyanine moiety could be fine-tuned in order to maximize the sensitivity of the overall framework. The optimized MOF-sensor had a CN⁻-detection limit of 0.05 μm , which is much lower than traditional CN⁻ fluorescent sensors (about 0.2 μm ).     

An E1-Catalyzed Chemoenzymatic Strategy to Isopeptide-N-Ethylated Deubiquitylase-Resistant Ubiquitin Probes

Triazole-based deubiquitylase (DUB)-resistant ubiquitin (Ub) probes have recently emerged as effective tools for the discovery of Ub chain-specific interactors in proteomic studies, but their structural diversity is limited. A new family of DUB-resistant Ub probes is reported based on isopeptide-N-ethylated dimeric or polymeric Ub chains, which can be efficiently prepared by a one-pot, ubiquitin-activating enzyme (E1)-catalyzed condensation reaction of recombinant Ub precursors to give various homotypic and even branched Ub probes at multi-milligram scale. Proteomic studies using label-free quantitative (LFQ) MS indicated that the isopeptide-N-ethylated Ub probes may complement the triazole-based probes in the study of Ub interactome. Our study highlights the utility of modern protein synthetic chemistry to develop structurally and new families of tool molecules needed for proteomic studies.     

External-Radiation-Induced Local Hydroxylation Enables Remote Release of Functional Molecules in Tumors

Radiation-induced cleavage for controlled release in vivo is yet to be established. We demonstrate the use of 3,5-dihydroxybenzyl carbamate (DHBC) as a masking group that is selectively and efficiently removed by external radiation in vitro and in vivo. DHBC reacts mainly with hydroxyl radicals produced by radiation to afford hydroxylation at para/ortho positions, followed by 1,4- or 1,6-elimination to rescue the functionality of the client molecule. The reaction is rapid and can liberate functional molecules under physiological conditions. This controlled-release platform is compatible with living systems, as demonstrated by the release of a rhodol fluorophore derivative in cells and tumor xenografts. The combined benefits of the robust caging group, the good release yield, and the independence of penetration depth make DHBC derivatives attractive chemical caging moieties for use in chemical biology and prodrug activation.     

Smart Magnetic and Fluorogenic Photosensitizer Nanoassemblies Enable Redox-Driven Disassembly for Photodynamic Therapy

Stimuli-responsive smart photosensitizer (PS) nanoassemblies that allow enhanced delivery and controlled release of PSs are promising for imaging-guided photodynamic therapy (PDT) of tumors. However, the lack of high-sensitivity and spatial-resolution signals and fast washout of released PSs from tumor tissues have impeded PDT efficacy in vivo. Herein, we report tumor targeting, redox-responsive magnetic and fluorogenic PS nanoassemblies ( NP-RGD ) synthesized via self-assembly of a cRGD- and disulfide-containing fluorogenic and paramagnetic small molecule ( 1-RGD ) for fluorescence/magnetic resonance bimodal imaging-guided tumor PDT. NP-RGD show high r₁ relaxivity but quenched fluorescence and PDT activity; disulfide reduction by glutathione (GSH) promotes efficient disassembly into a small-molecule probe ( 2-RGD ) and an organic PS (PPa-SH), which could further bind with intracellular albumin, allowing prolonged retention and cascade activation of fluorescence and PDT to ablate tumors.     

A Novel Phenoxazine-based Fluorescent Probe for the Detection of HOCl in Living Cells

Hypochlorous acid (HOCl), one of the reactive oxygen species (ROS), is a key microbicidal agent which is used for natural defense. However, it is also linked to varieties of human diseases owing to the overproduction of HOCl. Much effort has been made to exploit selective fluorescent sensors for the detection of HOCl, but most of them have some disadvantages such as short excitation wavelength, low selectivity, and slow response and so on. These restrict the biological application of the probes. In this work, BR-O was designed and synthesized on the base of phenoxazine for the detection of HOCl. BR-O exhibited a violent fluorescence enhancement in the presence of HOCl, showing excellent selectivity and high sensitivity. More importantly, the probe BR-O was capable of detecting exogenous and endogenous HOCl in living cells.     

An Activatable Polymeric Reporter for Near-Infrared Fluorescent and Photoacoustic Imaging of Invasive Cancer

Discriminative detection of invasive and noninvasive breast cancers is crucial for their effective treatment and prognosis. However, activatable probes able to do so in vivo are rare. Herein, we report an activatable polymeric reporter (P-Dex) that specifically turns on near-infrared (NIR) fluorescent and photoacoustic (PA) signals in response to the urokinase-type plasminogen activator (uPA) overexpressed in invasive breast cancer. P-Dex has a renal-clearable dextran backbone that is linked with a NIR dye caged with an uPA-cleavable peptide substrate. Such a molecular design allows P-Dex to passively target tumors, activate NIR fluorescence and PA signals to effectively distinguish invasive MDA-MB-231 breast cancer from noninvasive MCF-7 breast cancer, and ultimately undergo renal clearance to minimize the toxicity potential. Thus, this polymeric reporter holds great promise for the early detection of malignant breast cancer.     

From 1D to 2D Cd(II) and Zn(II) Coordination Networks by Replacing Monocarboxylate with Dicarboxylates in Partnership with Azine Ligands: Synthesis,Crystal Structures,Inclusion, and Emission Properties

Eight mixed-ligand coordination networks, [Cd(2-aba)(NO₃)(4-bphz)_3/2]_n·n(dmf) (1), [Cd(2-aba)₂(4-bphz)]_n·0.75n(dmf) (2), [Cd(seb)(4-bphz)]_n·n(H₂O) (3), [Cd(seb)(4-bpmhz)]_n·n(H₂O) (4), [Cd(hpa)(3-bphz)]_n (5), [Zn(1,3-bdc)(3-bpmhz)]_n·n(MeOH) (6), [Cd(1,3-bdc)(3-bpmhz)]_n ·0.5n(H₂O)·0.5n(EtOH) (7), and [Cd(NO₃)₂(3-bphz)(bpe)]_n·n(3-bphz) (8) were obtained by interplay of cadmium nitrate tetrahydrate or zinc nitrate hexahydrate with 2-aminobenzenecarboxylic acid (H(2-aba)), three dicarboxylic acids, sebacic (decanedioic acid, H₂seb), homophthalic (2-(carboxymethyl)benzoic acid, H₂hpa), isophthalic (1,3-benzenedicarboxylic acid, H₂(1,3-bdc)) acids, bis(4-pyridyl)ethane (bpe) and with four azine ligands, 1,2-bis(pyridin-4-ylmethylene)hydrazine (4-bphz), 1,2-bis(1-(pyridin-4-yl)ethylidene) hydrazine (4-bpmhz), 1,2-bis(pyridin-3-ylmethylene)hydrazine (3-bphz), and 1,2-bis(1-(pyridin-3-yl) ethylidene)hydrazine (3-bpmhz). Compounds 1 and 2 are 1D coordination polymers, while compounds 3–8 are 2D coordination polymers. All compounds were characterized by spectroscopic and X-ray diffraction methods of analysis. The solvent uptakes and stabilities to the guest evacuation were studied and compared for 1D and 2D coordination networks. The de-solvated forms revealed a significant increase of emission in comparison with the as-synthesized crystals.