Biological function of RNA domains may depend on the presence of a persistent 3D structure, conformational state changes, or the process of folding itself. Target engagement by small molecule ligands can capitalize or interfere with any of these, thereby modifying the function of an RNA. I will discuss types of different RNA targets, consequences for ligand molecular recognition and mechanism, and validation of targeting approaches.

Riboswitches are cis-acting gene regulatory elements that are located in the 5' untranslated region of mRNAs. Their widespread occurrence in bacteria makes them potential targets for new antibiotics. I will communicate our efforts to exploit riboswitches as drug targets. In particular, I will report on a druggability prediction method for RNAs, the discovery of riboswitch ligands via virtual screening and the development of an assay suitable for high-throughput screening.

I will speak about our efforts in clarifying and identifying RNA binding proteins as targets in oncology and neurodegeneration. My lab uses focused pooled CRISPR inhibition and knock out approaches to identify RBPs as vulnerabilities in different diseases, initially in oncology. We then use cutting-edge genomic methods to study the molecular pathways by which RBPs affect gene expression. Small molecule discovery approaches are then applied to modulate these in cancer systems, like avatars.

Discovery of new chemical entities modulating RNA function has been an emerging area in the past few years. Although successful against protein targets, there are a few challenges of applying DNA-encoded library (DEL) technology to discover RNA targeting molecules. Here we’d like to share the most recent successes in developing DEL screening methodology against RNA targets.

Androgen receptor (AR) signaling plays a dominant role in castration-resistant prostate cancers (CRPC) that have developed resistance to AR antagonists such as enzalutamide. We have identified a long non-coding RNA (lncRNA), NXTAR, that is located convergent with the AR gene and is repressed in human prostate tumors and cell lines. AR bound the NXTAR promoter, and inhibition of AR by the ACK1/TNK2 small molecule inhibitor (R)-9b excluded AR from the NXTAR promoter. Pharmacological restoration of NXTAR using (R)-9b abrogated enzalutamide-resistant prostate xenograft tumor growth. The restoration of NXTAR could serve as a new therapeutic modality for drug-resistant patients.

RNAs are invariably bound to and often modified by RNA-binding proteins (RBPs), which regulate many aspects of coding and non-coding RNA biology. Disruption of this network of RNA-protein interactions (RPIs) has been implicated in a many human diseases and targeting RPIs has arisen as a new frontier in RNA-targeted drug discovery. This talk will highlight newly developed technologies for validating and screening RPIs to enable RBP-targeted drug discovery.

RNA transglycosylation at guanosine (RNA-TAG) enables site-specific and covalent conjugation of fluorophores, affinity tags, or translational regulators, onto an RNA of interest. RNA-TAG utilizes a bacterial tRNA guanine transglycosylase (TGT) to exchange a guanine nucleobase within a specific 17-nucleotide RNA stem loop structure with a modified analog of the natural substrate preqeuosine1 (preQ1). RNA-TAG has been adapted to image RNA in fixed cells, regulate mRNA translation, and study RNA-protein interactions.

RNA-binding proteins (RBPs) provide for post-transcriptional and co-translational regulation of RNA metabolism. Oncogenic drivers that regulate the translational machinery, and interactions between RNAs and proteins at the level of translation can drive cancer pathogenesis. For example, MUSASHI (MSI) family has been identified as a translational regulator that is highly expressed in the most aggressive solid cancers and hematopoietic malignancies. Our laboratory has been exploring inhibitors to RNA regulators.

Anima Biotech is advancing mRNA Lightning, a novel platform for the discovery of selective small molecule mRNA drugs and their mechanisms of action. Our differentiated approach combines high scale automated phenotypic screening in live biology with AI mRNA image analysis that elucidates the mechanism of action. Anima’s pipeline projects will be presented, demonstrating how mRNA-regulation target space is amenable to selective modulation by small molecules.

Gene expression is heavily regulated at RNA level, including its stability, transport, protein binding, chemical state, and translation. These critical post-transcriptional gene expression regulatory processes open up opportunities for therapeutic intervention as a method to alter gene expression. I will present biotechnologies our lab is developing that utilize a “bifunctional design," with a programable RNA binding module and an effector module, which together alters the regulation of a target transcript.

Utilizing small molecules to modulate splicing has emerged as a successful therapeutic approach to regulating protein expression. Evrysdi (risdiplam), recently approved for the treatment of Spinal Muscular Atrophy, increases SMN protein levels. We review preclinical studies that indicate that drugs such as, risdiplam will be optimally therapeutic when given as early as possible after diagnosis and potentially these drugs will be required for the life of an SMA patient.

The past twenty years have seen an explosion of interest in the structure and function of RNA and DNA. While some 80% of the human genome is transcribed into RNA, just ~3% of those transcripts code for protein sequences. Here, we discuss our group’s efforts to target RNA and DNA with drug-like small molecules using a small-molecule microarray (SMM) screening platform and the molecular basis for these interactions.