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Fluorophorea??Labeled Cyclic Nucleotides as Potent Agonists of Cyclic Nucleotidea??Regulated Ion Channels
摘要： High-affinity fluorescent derivatives of cyclic adenosine and guanosine monophosphate are powerful tools for investigating their natural targets. Cyclic nucleotide-regulated ion channels belong to these targets and are vital for many signal transduction processes, such as vision and olfaction. The relation of ligand binding to activation gating is still challenging, and there is a need for fluorescent probes that enable the process to be broken down to the single-molecule level. This inspired us to prepare fluorophore-labeled cyclic nucleotides, which are composed of a bright dye and a nucleotide derivative with a thiophenol motif at position 8 that has already been shown to enable superior binding affinity. These bioconjugates were prepared by a novel cross-linking strategy that involves substitution of the nucleobase with a modified thiophenolate in good yield. Both fluorescent nucleotides are potent activators of different cyclic nucleotide-regulated ion channels with respect to the natural ligand and previously reported substances. Molecular docking of the probes excluding the fluorophore reveals that the high potency can be attributed to additional hydrophobic and cation-π interactions between the ligand and the protein. Moreover, the introduced substances have the potential to investigate related target proteins, such as cAMP- and cGMP-dependent protein kinases, exchange proteins directly activated by cAMP or phosphodiesterases.
关键词： fluorescent probes,nucleotides,ion channels
更新于2025-09-23 15:21:01
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Novel fluorescent triazinobenzimidazole derivatives as probes for labelling human A1 and A2B adenosine receptor subtypes
摘要： The expression levels and the subcellular localization of adenosine receptors (ARs) are affected in several pathological conditions as a consequence of changes in adenosine release and metabolism. In this respect, labelled probes able to monitor the AR expression could be a useful tool to investigate different pathological conditions. Herein, novel ligands for ARs, bearing the fluorescent 7-nitrobenzofurazan (NBD) group linked to the N1 (1,2) or N10 (3,4) nitrogen of a triazinobenzimidazole scaffold, were synthesized. The compounds were biologically evaluated as fluorescent probes for labelling A1 and A2B AR subtypes in bone marrow-derived mesenchymal stem cells (BM-MSCs) that express both receptor subtypes. The binding affinity of the synthetized compounds towards the different AR subtypes was determined. The probe 3 revealed a higher affinity to A1 and A2B ARs, showing interesting spectroscopic properties, and it was selected as the most suitable candidate to label both AR subtypes in undifferentiated MSCs. Florescence confocal microscopy showed that compound 3 significantly labelled ARs on cell membranes and the fluorescence signal was decreased by the cell pre-incubation with the A1 AR and A2B AR selective agonists, R-PIA and BAY 60-6583, respectively, thus confirming the specificity of the obtained signal. In conclusion, compound 3 could represent a useful tool to investigate the expression pattern of both A1 and A2B ARs in different pathological and physiological processes. Furthermore, these results provide an important basis for the design of new and more selective derivatives able to monitor the expression and localization of each different ARs in several tissues and living cells.
关键词： triazinobenzimidazole derivatives,Adenosine receptors,fluorescent probes,mesenchymal stem cells,A2B AR subtype
更新于2025-09-23 15:21:01
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Advances and perspectives in neara??infrared fluorescent organic probes for surgical oncology
摘要： Surgical resection of solid tumors is currently the most efficient and preferred therapeutic strategy for treating cancer. Despite significant medical, technical, and scientific advances, the complete treatment of this lethal disease is still a challenging task. New imaging techniques and contrast agents are urgently needed to improve cytoreductive surgery and patient outcomes. Tumor-targeted probes are valuable for guiding a surgical resection of tumor from subjective judgments to visual inspection. Near-infrared (NIR) fluorescent imaging is a promising technology in preclinical and clinical tumor diagnosis and therapy. The rapid development in NIR fluorophores with improved optical properties, targeting strategies, and imaging devices has brought about prospective study of novel NIR nanomaterials for intraoperative tumor detection. In this review, we summarize the recent development in NIR-emitting organic fluorophores and cancer-targeting strategies that specifically target and accumulate in tumors for the molecular imaging of cancerous cells. We believe this technique utilizing new fluorescent probes with an intraoperative optical imaging capacity could provide a more sensitive and accurate method for cancer resection guidance, thereby resulting in better surgical outcomes.
关键词： cancer detection,intraoperative optical imaging,near-infrared fluorescent probes,surgical resection
更新于2025-09-23 15:19:57
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Synthesis of novel small-molecule fluorescently labeled probes for the in vitro imaging of KCa3.1 channels
摘要： In order to facilitate the in vitro visualization of KCa3.1 channel-expressing cells, novel small-molecule imaging probes were designed and developed. Senicapoc showing high affinity and excellent selectivity towards the KCa3.1 channels was selected as targeting component. Different BODIPY dyes (15 - 20) were synthesized and connected by a Cu-catalyzed azide alkyne [3+2]cycloaddition with propargyl ether derivative 8 of senicapoc yielding fluorescently labeled ligands 21 - 26 targeting KCa3.1 channels. The novel dimethylpyrrole-based imaging probes 25 and 26 allow staining of KCa3.1 ion channels in NSCLC cells following a simple, fast and efficient protocol. The specificity was shown by removing the punctate staining pattern by pre-incubation with senicapoc. The density of KCa3.1 channels detected with fluorescent probe 25 and by immunostaining was identical. The punctate structure of the labeled channels could be observed in living cells as well. Molecular modeling studies showed binding of the senicapoc targeting component towards the binding site within the ion channel and orientation of the linker with the dye along the inner surface of the ion channel.
关键词： labeled ligands,KCa3.1 channel,fluorescent probes,non-small cell lung cancer cells,molecular modelling,imaging agents,cycloaddition
更新于2025-09-23 15:19:57
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A new-infrared probe targeting mitochondria via regulation of molecular hydrophobicity
摘要： Herein, we developed a near-infrared (NIR) fluorescent probe for mitochondrial staining based on the NIR fluorochrome, silicon-rhodamine. The hydrophobicity of the fluorescent core was systematically modified by conjugation with 10 different commercial amines. The resulting fluorescent compounds exhibited similar photophysical properties but diverse hydrophobicity. We identified the optimal level of hydrophobicity associated with high mitochondrial targeting efficiency. In particular, the SiR-Mito 8 probe provided excellent mitochondrial staining and successfully differentiated the live Hep3B cancer cells from normal L02 cells in vitro.
关键词： Mitochondria,silicon rhodamine,fluorescent probes,near-IR
更新于2025-09-23 15:19:57
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Facile Fabrication of Fluorescent Inorganic Nanoparticles with Diverse Shapes for Cell Imaging
摘要： In the present work, we describe a facile and general method of fabricating fluorescent inorganic nanoparticles with diverse shapes for cell imaging application. The hematite (α-Fe2O3) nanoparticles (HNPs) with three different shapes (i.e., spindle shape, ellipsoidal shape and quasi-spherical shape) were first prepared as model systems in consideration of good biocompatibility and the controllable morphology of α-Fe2O3. Three fluorescent HNPs with different shapes were achieved via one-pot sol-gel reaction of AIEgen-functionalized siloxane (AIEgen-Si(OCH3)3) and TEOS in the presence of PVP-stabilized HNPs. Due to the fluorescence originating from the thin AIEgens-contained SiO2 shell around the HNPs, their photoluminescent intensities can be tuned by changing the concentrations of TEOS and AIEgen-Si(OCH3)3 in feed prior to the sol-gel reaction. When the as-prepared fluorescent products were dispersed in water, they gave intense green light emission upon excitation at 360 nm with relatively high fluorescence quantum yield. Further, fluorescent HNPs exhibited low cytotoxicity and excellent photostability and, thus, were used as optical probes to preliminarily explore the effect of nanoparticle shapes on their cellular uptake behaviors. This work should open a facile way to prepare various fluorescent inorganic nanoparticles with specific morphology for various biological applications.
关键词： inorganic nanoparticles,fluorescent probes,cell imaging,shape
更新于2025-09-19 17:15:36
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Recent advances in fluorescent probes for peroxynitrite detection
摘要： Peroxynitrite (ONOO?), known as a kind of reactive nitrogen species, is a significant factor involved in a broad spectrum of physiological and pathological processes. However, their biological functions remain to be controversial or poorly characterized yet, mainly because of the lack of reliable methods for sensitive and specific detection of ONOO? in vitro and in vivo. Thus, approaches that enable noninvasive visualized imaging of ONOO? will be of enormous benefit in understanding the processes of related diseases and developing novel drugs. During the last decades, fluorescent probes, with high sensitivity and real-time spatial imaging capacity, receive more and more attention. Herein, we strive to summarize the current fluorescent probes for ONOO?, including small-molecular fluorescent probes, nanoparticles-based fluorescent probes, and genetically encoded fluorescent probes. Furthermore, the challenges and attempt to give an outlook on the possible further developments are discussed.
关键词： Fluorescent probes,Nanoparticles,Peroxynitrite,Imaging
更新于2025-09-19 17:15:36
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Development of a Prea??assembled TBET Fluorescent Probe for Ratiometric Sensing of Anticancer Platinum(ll) Complexes
摘要： Fluorescence microscopy has emerged as an attractive technique to probe intracellular processing of Pt-based anticancer compounds. Herein, we reported the first Through-Bond Energy Transfer (TBET) fluorescent probe NPR1 designed for sensitive detection and quantitation of PtII complexes. The novel TBET probe was successfully applied for ratiometric fluorescence imaging of anticancer PtII complexes such as cisplatin and JM118 in cells. Capitalizing on the ability of the probe to discriminate between PtII complexes and their PtIV derivatives, the probe was further applied to study the activation of PtIV prodrug complexes that are known to release active PtII species after intracellular reduction.
关键词： Platinum Drugs,Fluorescent Probes,Ratiometric,Cisplatin,Through Bond Energy Transfer
更新于2025-09-19 17:13:59
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Novel fluorescently labelled ATP analogues for direct monitoring of ubiquitin activation
摘要： Simple and robust assays to monitor enzymatic ATP cleavage with high efficiency in real-time are scarce. To address this shortcoming, we developed novel fluorescently labelled adenosine tri-, tetra- and pentaphosphate analogues of ATP. The novel ATP analogues bear – in contrast to earlier reports – only a single acridone-based dye at the terminal phosphate group. The dye’s fluorescence is quenched by the adenine component of the ATP analogue and is restored upon cleavage of the phosphate chain and dissociation of the dye from the adenosine moiety. Thereby the activity of ATP cleaving enzymes can be followed in real-time. We demonstrate this proficiency for ubiquitin activation by the ubiquitin-activating enzymes UBA1 and UBA6 which represents the first step in an enzymatic cascade leading to the covalent attachment of ubiquitin to substrate proteins, a process that is highly conserved from yeast to humans. We found that the efficiency to serve as cofactor for UBA1/UBA6 very much depends on the length of the phosphate chain of the ATP analogue: triphosphates are used poorly while pentaphosphates are most efficiently processed. Notably, the novel pentaphosphate-harbouring ATP analogue supersedes the efficiency of recently reported dual-dye labelled analogues and thus, is a promising candidate for broad applications.
关键词： ubiquitin,fluorescent probes,PET,UBA1,ATP
更新于2025-09-19 17:13:59
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Protein‐specific, multi‐color and 3D STED imaging in cells with DNA‐labeled antibodies
摘要： Photobleaching is a major challenge in fluorescence microscopy, in particular if high excitation light intensities are used. Signal-to-noise and spatial resolution may be compromised, which limits the amount of information that can be extracted from an image. Photobleaching can be bypassed with exchangeable labels, which transiently bind to and off a target and thereby replenish destroyed labels by intact ones from a reservoir. Here, we demonstrate confocal and STED microscopy with short, fluorophore-labeled oligonucleotides that transiently bind to complementary oligonucleotides attached to protein-specific antibodies. The constant exchange of fluorophore labels in DNA-based STED imaging bypasses photobleaching that occurs with covalent labels. We show that this concept is suitable for targeted, two-color STED imaging of whole cells.
关键词： multicolor imaging,DNA-PAINT,STED microscopy,fluorescence,fluorescent probes
更新于2025-09-19 17:13:59