A Small-Molecule Probe with a Dual Function of miRNA Inhibition and Target identification.

By virtue of their key roles in pathologies, miRNAs represent a promising class of therapeutic targets. While high-fidelity small-molecule modulators of miRNAs can be identified via high-throughput screening using cellular reporter systems, their modes of action are elusive due to the lack of proper tools. Here, we report a small-molecule probe, 1 a, that is capable of elucidating its biological target along miRNA inhibition. Derived from norathyriol, a nature product, 1 a possessed a bioorthogonal alkyne moiety for subsequent labeling via copper-catalyzed azide-alkyne cycloaddition chemistry. We demonstrated that 1 a inhibited a panel of different miRNAs by blocking their loading onto argonaute 2 (AGO2), which is the key protein responsible for miRNA function. With the alkyne handle, we successfully identified AGO2 as an intracellular target of 1 a. Therefore, this work presents a novel small-molecule tool for suppressing and probing miRNA regulatory pathways.

Metal- and Strain-Free Bioorthogonal Cycloaddition of o-Diones with Furan-2(3H)-one as Anionic Cycloaddend.

The development of new bioorthogonal reactions with mutual orthogonality to classic bioorthogonal reactions such as the strain-promoted azide-alkyne click reaction and the inverse-electron-demand Diels-Alder reaction is of great importance in providing chemical tools for multiplex labelling of live cells. Here we report the first anionic cycloaddend-promoted bioorthogonal cycloaddition reaction between phenanthrene-9,10-dione and furan-2(3H)-one derivatives, where the high polarity of water is exploited to stabilize the highly electron-rich anionic cycloaddend. The reaction is metal- and strain-free, which proceeds rapidly in aqueous solution and on live cells with a second-order rate constant up to 119 M-1 s-1 . The combined utilization of this reaction together with the two other widely used bioorthogonal reactions allows for mutually orthogonal labelling of three types of proteins or three groups of living cells in one batch without cross-talking. Such results highlight the great potential for multiplex labelling of different biomolecules in live cells.