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Identification of HuR-RNA Interfering Compounds by Dynamic Combinatorial Chemistry and Fluorescence Polarization
Serena Della Volpe1,2, Roberta Listro1, Francesca Alessandra Ambrosio3
1Department of Drug Sciences, University of Pavia, Via Taramelli 12, 27100 Pavia, Italy.
Abstract:
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.
Insights
Researchers identified novel compounds targeting the HuR-RNA complex, a key factor in cancer development. These potential anticancer agents show promise in interfering with HuR-RNA interactions for therapeutic strategies.
Area of Science:
- Biochemistry
- Molecular Biology
- Oncology
Background:
- The RNA-binding protein HuR plays a critical role in post-transcriptional regulation of genes involved in cancer.
- Dysregulation of HuR is frequently observed in various cancers, making it a potential therapeutic target.
- Modulating the HuR-RNA complex is a promising strategy for developing novel anticancer agents.
Purpose of the Study:
- To identify novel HuR ligands capable of interfering with the HuR-RNA complex.
- To explore the potential of protein-templated dynamic combinatorial chemistry (pt-DCC) for discovering HuR modulators.
Main Methods:
- Utilized protein-templated dynamic combinatorial chemistry (pt-DCC) with a recombinant RRM1+2 HuR protein construct.
- Identified and amplified potential ligands using DCC.
- Validated ligand binding to HuR using Saturation Transfer Difference Nuclear Magnetic Resonance (STD-NMR).
- Performed molecular modeling studies to elucidate binding modes.
- Assessed interference with HuR-RNA binding using fluorescence polarization assays.
Main Results:
- Seven acylhydrazone compounds demonstrated significant amplification (>80%) in the pt-DCC screening.
- STD-NMR and molecular modeling confirmed that identified compounds bind to HuR within the mRNA binding pocket.
- Three compounds effectively inhibited HuR-RNA complex formation in fluorescence polarization assays.
Conclusions:
- The study successfully identified novel HuR ligands using pt-DCC.
- These compounds bind to the HuR mRNA binding site and interfere with HuR-RNA interactions.
- The identified compounds serve as promising foundational agents for the development of new anticancer therapies targeting HuR.
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