Highly compact viral genomes provide a limited number of protein targets for therapeutic intervention. Recent studies aiming to elucidate the secondary structure and epitranscriptomic modifications of viral RNA genomes and virus-encoded coding and non-coding transcripts greatly expanded the repertoire of drug targets. This presentation will cover the applications of biochemical and molecular techniques that enable a valuable estimation of structural information content for target viral RNAs.

As part of our efforts to improve small molecule targeting strategies and gain fundamental insights into small molecule-RNA recognition, we have analyzed patterns in both RNA-biased small molecule chemical space and RNA topological space privileged for differentiation. We have applied these principles to functionally modulate conformations of the 3’-triple helix of the long noncoding RNA MALAT1, an enterovirus (EV71) IRES structure, and regulatory RNA in SARS-CoV-2.

The genome of SARS-CoV-2 comprises of approximately 30,000 nucleotides. Besides the coding part of the genome, the 5'- and 3'-UTR contain multiple highly conserved structural elements. By NMR spectroscopy, we investigated these 15 elements and elucidated their secondary and tertiary structures. We further screened fragment libraries to investigate the general druggability of these elements. We translated these insights to guide medicinal chemistry campaigns.

Epitranscriptomic RNA modifications play important roles in biology; however, there remains a major gap in our understanding of the scope, regulation, and function of these modifications in biological systems. Here, we present RNA-mediated activity-based protein profiling (RNABPP), a chemoproteomic strategy to study and target RNA-modifying enzymes in their native context, and discuss its application to elucidate novel epitranscriptomic pathways for eukaryotic gene regulation.

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 number of 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.

Using novel mRNA technology, companies like BioNTech/Pfizer and Moderna successfully developed and validated the efficacy of mRNA-based vaccines in the fight against COVID-19. To support rapid early discovery research efforts for mRNA-based therapeutics, Codex DNA has developed a first-of-its-kind mRNA automation solution capable of synthesizing mRNA in less than 24hrs. In this talk, we’ll present the steps taken to fully automate the synthesis of mRNA to help accelerate early discovery research.

In fragile X syndrome, the most common genetic form of mental retardation, a CGG trinucleotide–repeat expansion adjacent to the fragile X mental retardation 1 (FMR1) gene promoter results in its epigenetic silencing. We have found that FMR1 gene silencing is mediated by the FMR1 mRNA. The FMR1 mRNA contains the transcribed CGG-repeat tract as part of the 5′ untranslated region, which hybridizes to the complementary CGG-repeat portion of the FMR1 gene to form an RNA·DNA duplex. Here we describe how small molecules that bind the CGG-repeat RNA region can prevent FMR1 gene silencing in a model of embryonic development. We also describe a strategy for selectively reversing the epigenetic silencing marks on the FMR1 locus in patient-derived cells. Thus, our data show that an RNA-binding small molecule can selectively control the epigenetic activation state of a single gene in the genome, providing novel therapeutic implications for the treatment of fragile X syndrome.

The RNA-based targets continue to generate excitement in the Pharmaceutical and Biotech community as a possibility of going after undruggable targets in unmet medical needs. Applying appropriate tools to advance the research on RNA targets requires novel approaches from medicinal chemistry, computational sciences, target biology, and translation research.

An update on current approaches to advance the small molecule drug discovery programs will be provided in the talk.

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 programmable RNA binding module and an effector module, which together alters the regulation of a target transcript.

Recent years have seen a remarkable increase in RNA as a target for small molecules.  Our laboratory has been approaching this problem from a structure-based perspective. I will discuss our laboratory's efforts to identify RNA-binding small molecules and define what makes a good RNA target. I will discuss our laboratory's  Small Molecule Microarrays screening efforts and focus on specific RNA targets of interest to the lab.

Anima Biotech has developed mRNA Lightning platform, a novel approach for the discovery of small molecules that selectively control mRNA translation against hard targets and undruggable proteins. This talk features Anima’s oncology strategy from the assessment of the drug discovery process through multiple novel mechanisms of action targetable by Anima’s mRNA translation modulators, including how they can achieve selectivity in c-Myc-addicted tumors and mKRAS-driven tumors.

DNA-Encoded library (DEL) technology is a screening platform used throughout the pharmaceutical industry to discover novel chemical matter. Despite the successful application in protein targets, it has been challenging to apply DEL to RNA targets. We will share recent successful DEL screening efforts against proof-of-concept RNA targets, including validation data using biophysical assays.

Effective on protein targets, few were reported for ligand identification using DEL on RNAs. Here we discuss the recent advances of HitGen's DEL Targeting RNA platform. Case studies will be given for how challenges of identifying small molecules for RNA using DEL were overcome experimentally and computationally, how double-digit nanomolar molecules for one RNA target were identified, and how this optimized screening strategy can be applied universally on other RNA targets. 

Utilizing small molecules to modulate splicing has emerged as a successful therapeutic approach to regulating protein expression. Here, three diseases where small-molecule splicing modulators can be utilized are described: spinal muscular atrophy; familial dysautonomia; and Huntington’s disease. For each of these indications, I will discuss the correlation between pharmacokinetics and pharmacodynamics, as well as the correlation between pharmacodynamics and efficacy.

Small RNAs regulate the expression of stress response and virulence factors in enteric Gram-negative bacteria. The RNA chaperone Hfq quickly matches a divergent family of small RNAs with their complementary mRNA targets, amidst a pool of similar candidates. Single-molecule fluorescence microscopy shows how Hfq overcomes mRNA secondary structure, and how a series of probabilistic steps can lead to stable Hfq-RNA complexes that may be targets for small molecule inhibitors.