Identification of HuR-RNA Interfering Compounds by Dynamic Combinatorial Chemistry and Fluorescence Polarization.

The RNA binding protein HuR regulates the post-transcriptional process of different oncogenes and tumor suppressor genes, and its dysregulation is linked with cancer. Thus, modulating the complex HuR-RNA represents a promising anticancer strategy. To search for novel HuR ligands able to interfere with the HuR-RNA complex, the protein-templated dynamic combinatorial chemistry (pt-DCC) method was utilized. The recombinant RRM1+2 protein construct, which contains essential domains for ligand-HuR binding and exhibits enhanced solubility and stability compared to the native protein, was used for pt-DCC. Seven acylhydrazones with over 80% amplification were identified. The binding of the fragments to HuR extracted from DCC was validated using STD-NMR, and molecular modeling studies revealed the ability of the compounds to bind HuR at the mRNA binding pocket. Notably, three compounds effectively interfered with HuR-RNA binding in fluorescence polarization studies, suggesting their potential as foundational compounds for developing anticancer HuR-RNA interfering agents.

BOPC1 Enantiomers Preparation and HuR Interaction Study. From Molecular Modeling to a Curious DEEP-STD NMR Application.

The Hu family of RNA-binding proteins plays a crucial role in post-transcriptional processes; indeed, Hu-RNA complexes are involved in various dysfunctions (i.e., inflammation, neurodegeneration, and cancer) and have been recently proposed as promising therapeutic targets. Intrigued by this concept, our research efforts aim at identifying small molecules able to modulate HuR-RNA interactions, with a focus on subtype HuR, upregulated and dysregulated in several cancers. By applying structure-based design, we had already identified racemic trans-BOPC1 as promising HuR binder. In this Letter, we accomplished the enantio-resolution, the assignment of the absolute configuration, and the recognition study with HuR of enantiomerically pure trans-BOPC1. For the first time, we apply DEEP (differential epitope mapping)-STD NMR to study the interaction of BOPC1 with HuR and compare its enantiomers, gaining information on ligand orientation and amino acids involved in the interaction, and thus increasing focus on the in silico binding site model.