Copper-Free Click Reactions with Polar Bicyclononyne Derivatives for Modulation of Cellular Imaging

The ability of cells to incorporate azidosugars metabolically is a useful tool for extracellular glycan labelling. The exposed azide moiety can covalently react with alkynes, such as bicyclo[6.1.0]nonyne (BCN), by strain‐promoted alkyne–azide cycloaddition (SPAAC). However, the use of SPAAC can be hampered by low specificity of the cycloalkyne. In this article we describe the synthesis of more polar BCN derivatives and their properties for selective cellular glycan labelling. The new polar derivatives [amino‐BCN, glutarylamino‐BCN and bis(hydroxymethyl)‐BCN] display reaction rates similar to those of BCN and are less cell‐permeable. The labelling specificity in HEK293 cells is greater than that of BCN, as determined by confocal microscopy and flow cytometry. Interestingly, amino‐BCN appears to be highly specific for the Golgi apparatus. In addition, the polar BCN derivatives label the N‐glycan of the membrane calcium channel TRPV5 in HEK293 cells with significantly enhanced signal‐to‐noise ratios. Labelling in living systems: Polar variants of bicyclononyne (BCN) were synthetically prepared and evaluated for metabolic labelling of cell‐surface glycans. Significantly enhanced signal‐to‐noise ratios were obtained, and fluorescent derivatives of the new BCN probes were applied for selective imaging of the TRPV5 receptor. Selective Golgi staining was demonstrated with an amino variant of BCN.