RNA-targeting small molecule drug discovery has greatly benefitted from the available cellular models for disease. The use of cell culture in early development has become a requirement for assessing the basic efficacy of a drug candidate. Thus, more research groups have implemented these techniques in their programs. In a leading example, Al-Hashimi and coworkers identified six small molecules with high affinity for TAR of HIV-1 through a computational approach. They docked a library of small molecules onto RNA dynamic structures generated by NMR and Molecular Dynamics (MD) simulations. The hit molecules inhibited the Tat—TAR interaction in vitro. They arrived at lead molecule, netilmicin, that had the best selectivity for HIV-1 TAR and inhibited HIV-1 replication in cells with a low IC50. The Disney group has studied aminoglycoside derivatives in 2009 for their ability to inhibit interactions between repeat RNA and proteins. Using their prediction database INFORNA, they discovered that a compound could bind to 1 x 1 UU internal loops on an N-methyl peptide backbone. They confirmed that like other compounds that target DM1 r(CUG), they could inhibit the complex between r(CUG)-MBNL1, disrupt nuclear foci, and increase nucleocytoplasmic transport of the gene in patient-derived DM1 fibroblasts. In that study the Disney group also described several approaches to validate the RNA targets of small molecules. In the first approach termed chemical cross-linking and isolation by pull down (Chem-CLIP) and chemical cross-linking and isolation by pull down to map binding sites (Chem-CLIP-Map).
These studies showed in cells that small molecules can direct target disease-causing r(CUG) repeats in DM1 and that impressively the compound can discriminate against other RNAs with shorter repeats and also between the mutant and wild type allele of the DMPK mRNA that contains r(CUG) disease-causing repeats. In an additional approach, dubbed small–molecule nucleic acid profiling by cleavage applied to RNA (Ribo-SNAP) showed that small molecules can be used to cleave RNA targets in cells and also importantly demonstrated that designer small molecules target precisely disease causing RNA repeats and discriminate against RNAs that are not disease causing but have short repeats of r(CUG). Thus, targeting RNA structure with small molecules can have important selectively discrimination implications in cells.
In 2014, Chenowith and colleagues reported a cationic triptycene scaffold that targets RNA and DNA three-way junctions. Subsequent studies showed that these molecules exhibited favorable cellular uptake and cytotoxicity in human ovarian cancer cell lines. In 2017, the Xodo group reported anthrafurandione and anthrathiophenedione small molecules with aminoethyl side chains could bind to RNA G-quadraplexes at the 5’-UTR of certain mRNAs. Further, these compounds were shown to suppress the KRAS oncogene in pancreatic cancer cells and induce apoptosis by reducing the metabolic activity of the cells.