In This Issue Cite This: ACS Med. Chem. Lett. 2018, 9, 515−516
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A NOVEL ASSAY FOR THE STUDY OF MICRORNA−PROTEIN INTERACTIONS IMPLICATED IN HUMAN DISEASES MicroRNAs are short noncoding RNA molecules involved in gene silencing and post-translational regulation of gene expression. Changes to microRNA expression are implicated in various human diseases, such as cancer. The interaction of binding proteins with microRNAs can lead to microRNA inhibition and reduced gene silencing, which enables aberrant protein expression. This interaction between proteins and microRNAs thus provides a therapeutic target for the development of inhibitors of the microRNA−protein complex. In This Issue, Garner and colleagues (DOI: 10.1021/ acsmedchemlett.8b00126) report in their Featured Letter a modification of a high throughput screening technology developed in their laboratory called catalytic enzyme-linked click chemistry assay (cat-ELCCA) to facilitate the discovery of microRNA−protein complex inhibitors. The authors focused on the prelet-7-Lin28 RNA−protein interaction for this work, where Lin 28 is an inhibitor of let-7 maturation by attaching to prelet-7. After immobilization of Lin 28, pre-microRNA containing a click chemistry handle and various small molecule libraries were added and incubated with the protein for high throughput screening. Binding of a labeled horseradish peroxidase to the click chemistry handle provided chemiluminescent output. Follow-up assays and subsequent SAR studies identified compounds that inhibit the prelet-7-Lin28 complex formation. Thus, the authors demonstrated the applicability of this new version of cat-ELCCA to the analysis of RNA−protein interactions and discovered a new class of inhibitors.
Their activity is regulated through various mechanisms, including palmitic acid attachment to a conserved serine residue. This modification is accomplished by the porcupine acyltransferase, while the serine hydrolase NOTUM removes the fatty acid from Wnts. NOTUM has emerged as a potential therapeutic target for degenerative diseases, but only few inhibitors of this enzyme have been identified. Herein, Cravatt and colleagues (DOI: 10.1021/acsmedchemlett.8b00191) present irreversible inhibitors of NOTUM. The authors screened the enzyme by activity-based profiling (ABPP) using various serine hydrolase-directed inhibitors containing ureas and carbamates. Follow-up studies with two N-hydroxycarbamates led to the optimized compound ABC99, which demonstrated low nanomolar potency and selectivity for NOTUM. The authors showed that ABC99 sustains Wntmediated cell signaling in the presence of NOTUM using a cellular assay. In order to investigate the NOTUM activity in native biological systems, the authors also synthesized a modified version of ABC99 containing an alkyne group to obtain a clickable probe. Future studies will involve experiments to further investigate the activity of ABC99.
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APPLICATION OF BIOLUMINESCENCE TO A THERMAL SHIFT ASSAY Target engagement measurements are critical for the characterization of protein−ligand interactions in preclinical studies. Thermal shift assays (TSAs) exploit the change in a target protein’s melting temperature that occurs upon ligand binding, and the observed shift tends to correlate with ligand concentration and affinity. However, the applicability of TSAs has various limitations, such as the availability of purified proteins. In the present issue, Dart et al. (DOI: 10.1021/acsmedchemlett.8b00081) report a modified version of a TSA in which NanoLuc luciferase, an enzyme with high thermostability that produces an exceptionally bright luminescent signal, was genetically coupled to the folded state of a target protein. The authors tested this new assay system with protein mitogen-activated protein kinase 14 (MAPK14), bromodomains, and histone deacetylases and relevant ligands. Further experiments demonstrated the applicability of this assay to library screening and compound profiling. This work has paved the way for additional studies with this versatile assay system, which will investigate miniaturization
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DEVELOPMENT OF IRREVERSIBLE ENZYME INHIBITORS THROUGH ACTIVITY-BASED PROTEIN PROFILING Wnt proteins are signaling molecules involved in embryonic development, tissue homeostasis and stem cell development. © 2018 American Chemical Society
Special Issue: Med Chem Tech: Driving Drug Development Published: June 14, 2018 515
DOI: 10.1021/acsmedchemlett.8b00250 ACS Med. Chem. Lett. 2018, 9, 515−516
ACS Medicinal Chemistry Letters
In This Issue
into well plates and use of CRISPR to tag proteins with Nluc.
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DOI: 10.1021/acsmedchemlett.8b00250 ACS Med. Chem. Lett. 2018, 9, 515−516
