Rational design and evaluation of 2-((pyrrol-2-yl)methylene)thiophen-4-ones as RNase L inhibitors.

Ribonuclease L (RNase L) plays a crucial role in an antiviral pathway of interferon-induced innate immunity by degrading RNAs to prevent viral replication. Modulating RNase L activity thus mediates the innate immune responses and inflammation. Although a few small molecule-based RNase L modulators have been reported, only limited molecules have been mechanistically investigated. This study explored the strategy of RNase L targeting by using a structure-based rational design approach and evaluated the RNase L-binding and inhibitory activities of the yielded 2-((pyrrol-2-yl)methylene)thiophen-4-ones, which exhibited improved inhibitory effect as determined by in vitro FRET and gel-based RNA cleavage assay. A further structural optimization study yielded selected thiophenones that showed >30-fold more potent inhibitory activity than that of sunitinib, the approved kinase inhibitor with reported RNase L inhibitory activity. The binding mode with RNase L for the resulting thiophenones was analyzed by using docking analysis. Furthermore, the obtained 2-((pyrrol-2-yl)methylene)thiophen-4-ones exhibited efficient inhibition of RNA degradation in cellular rRNA cleavage assay. The newly designed thiophenones are the most potent synthetic RNase L inhibitors reported to date and the results revealed in our study lay the foundation for the development of future RNase L-modulating small molecules with new scaffold and improved potency.