Synthesis of a Novel Fluorescent Ruthenium Complex with an Appended Ac4GlcNAc Moiety by Click Reaction

Abstract

The O-linked β-N-acetylglucosamine (O-GlcNAc) modification is an abundant post-translational modification in eukaryotic cells, which plays a fundamental role in the activity of many cells and is associated with pathologies like type II diabetes, Alzheimer’s disease or some cancers. However, the precise connexion between O-GlcNAc-modified proteins and their function in cells is largely undefined for most cases. Confocal microscopy is a powerful and effective tool for in-cell elucidation of the function of biological molecules. Chemical labeling of non-ultraviolet or non-fluorescent carbohydrates with fluorescent tag is an essential step that makes intra-cellular microscopic inspection possible. Here we report a strategy based on the 1,3-dipolar cycloaddition, called click chemistry, between unnatural N-acetylglucosamine (GlcNAc) analogues Ac4GlcNAc (substituted with an azido group) and the corresponding fluorescent tag Ru(bpy)2(Phen-alkyne)Cl2 (4) to synthesize the fluorescent dye Ru(bpy)2(Phen-Ac4GlcNAc)Cl2 (5) under mild and neutral reaction conditions. Moreover, 5 showed good stability, desirable fluorescence characteristics, and exhibited rather low levels of cytotoxicity against sensitive MCF-7 cells. Additionally, we have achieved successful fluorescent imaging of 5 transported in living MCF-7 cells. Cell images displayed that proteins are potentially labelled with 5 in the cytoplasm.

Detection of Ru potential metallodrug in human urine by MALDI-TOF mass spectrometry: Validation and options to enhance the sensitivity

“…The facts mentioned above emphasize the importance of carefully conducted toxicity studies, as well as further in vivo screenings, which can be done, either in patients treated with increasing concentrations of chemotherapeutics [16] or in models that mimic human tumour pathology [17]. However, the improvement of non-invasive methods for their detection and the metabolite characterization in biological fluids is an essential and challenging hot topic [18].…”

“…Methods that can be used in the detection and quantification of metallodrugs, such as Ru-compounds in urine and other biological fluids, often require pre-separation by liquid chromatography with mass spectrometric detection, such as inductively coupled plasma mass spectrometry (ICP MS) [19], radioactive [3] or fluorescence labelling [18]. Concerning the speed of sample preparation and analyses, matrix-assisted desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) offers alternatives to other approaches, which involve the derivatization of samples and further purification of biological matrix [20].…”

Switchable DNA Catalysts for Proximity Labeling at Sites of Protein–Protein Interactions

“…To create the Ru-DNA conjugate, we first synthesized a Ru(bpy) 2 (phenanthrolinealkyne) complex (Schemes S1 and S2). 63 Next, we installed the Ru complex onto a ssDNA oligo bearing an azide modification using copper-catalyzed azide−alkyne coupling, followed by high-performance liquid chromatography (HPLC) purification (Scheme S3). We confirmed the identity of the conjugated oligo using gel electrophoresis, HPLC, and mass spectrometry (Figures S6 and S7).…”

Shining New Light on Biological Systems: Luminescent Transition Metal Complexes for Bioimaging and Biosensing Applications

“…The high lysosome specificity of these complexes also offers opportunities to monitor the therapeutic outcomes in situ by real-time tracking the changes in lysosomal integrity during photoinduced cell death. Tan, Mao, and co-workers have conjugated a pH-sensitive ruthenium(II) β-carboline complex (285) to Cdots for lysosome-targeted imaging and therapy. 517 This modification results in increased cellular uptake of the complex and TPA cross-section (δ 810nm = 1,118 GM), which allows the imaging of cancer cells, MCTSs, and zebrafish at a higher penetration depth as well as the more effective inhibition of cancer cell growth in both 2D cells and 3D MCTSs upon two-photon excitation.…”