We report a new strategy to regulate microRNAs (miRNAs) biogenesis by using bi-functional small molecules that consist of a pre-miRNA binding unit connected by a linker to a Dicer inhibiting unit. In this effort, fluorescence polarization-based screening was used to identify neomycin as a pre-miR-21 binding ligand. Although neomycin cannot inhibit miR-21 maturation, linking it to the RNase inhibitor 1 forms the bi-functional conjugate 7A, which inhibits the production of miR-21. We expect that this strategy will be applicable to design other molecules for miRNA regulation.

miR-155 plays key promoting roles in several cancers and emerges as an important anticancer therapeutic target. However, the discovery of small molecules that target RNAs is challenging. Peptidomimetics have been shown to be a rich source for discovering novel ligands to regulate cellular proteins. However, the potential of using peptidomimetics for RNA targeting is relatively unexplored. To this end, we designed and synthesized members of a novel 320 000 compound macrocyclic peptidomimetic library. An affinity-based screening protocol led to the identification of a pre-miR-155 binder that inhibits oncogenic miR-155 maturation in vitro and in cell and induces cancer cell apoptosis. The results of this investigation demonstrate that macrocyclic peptidomimetics could serve as a new scaffold for RNA targeting.

Background Alteration in hemodynamic shear stress at atheroprone sites promotes endothelial paracellular pore formation and permeability. The molecular mechanism remains unknown. Methods and Results We show that Nck (noncatalytic region of tyrosine kinase) deletion significantly ameliorates disturbed flow‐induced permeability, and selective isoform depletion suggests distinct signaling mechanisms. Only Nck1 deletion significantly reduces disturbed flow‐induced paracellular pore formation and permeability, whereas Nck2 depletion has no significant effects. Additionally, Nck1 re‐expression, but not Nck2, restores disturbed flow‐induced permeability in Nck1/2 knockout cells, confirming the noncompensating roles. In vivo, using the partial carotid ligation model of disturbed flow, Nck1 knockout prevented the increase in vascular permeability, as assessed by Evans blue and fluorescein isothiocyanate dextran extravasations and leakage of plasma fibrinogen into the vessel wall. Domain swap experiments mixing SH 2 (phosphotyrosine binding) and SH 3 (proline‐rich binding) domains between Nck1 and Nck2 showed a dispensable role for SH 2 domains but a critical role for the Nck1 SH 3 domains in rescuing disturbed flow‐induced endothelial permeability. Consistent with this, both Nck1 and Nck2 bind to platelet endothelial adhesion molecule‐1 ( SH 2 dependent) in response to shear stress, but only Nck1 ablation interferes with shear stress–induced PAK 2 (p21‐activated kinase) membrane translocation and activation. A single point mutation into individual Nck1 SH 3 domains suggests a role for the first domain of Nck1 in PAK recruitment to platelet endothelial cell adhesion molecule‐1 and activation in response to shear stress. Conclusions This work provides the first evidence that Nck1 but not the highly similar Nck2 plays a distinct role in disturbed flow‐induced vascular permeability by selective p21‐activated kinase activation.

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miRNAs are key cellular regulators and their dysregulation is associated with many human diseases. They are usually produced locally in a spatiotemporally controlled manner to target mRNAs and regulate gene expression. Thus, developing chemical tools for manipulating miRNA with spatiotemporal precise is critical for studying miRNA. Herein, we designed a strategy to control miRNA biogenesis with light controllable inhibitor targeting the pre-miRNA processing by Dicer. By conjugating two non-inhibiting units, a low affinity Dicer inhibitor and a pre-miRNA binder, through a photocleavable linker, the bifunctional molecule obtained could inhibit miRNA production. Taking advantage of the photocleavable property of the linker, the bifunctional inhibitor can be fragmented into separate non-inhibiting units and therefore be deactivated by light. We expect that this strategy could be applied to generate chemical biological tools that allow light-mediated spatiotemporal control of miRNA maturation and contribute to the study of miRNA function.