In-cell analysis and elaboration of RNA-targeting fragment ligands

The function and regulation of both coding and non-coding RNAs depend on their structures. A subset of RNAs contain pockets capable of binding ligands, and targeting RNA with small molecules can potentially modulate gene expression and cell state. To date, identifying and progressing these ligands within cells has posed a formidable challenge. I have developed a technology for transcript-specific detection of ligand binding in cells that is scalable for high throughput screens. This method simplifies the complex RNA ligand measurement into a manageable and efficient PCR-based strategy, which I used to discover a first-site fragment ligand that binds the 5’-UTR of MYC, a conventionally difficult-to-ligand target. In Aim 1, I propose to use my in-cell screening platform to discover molecules that exhibit enhanced binding to MYC mRNA when bound by the first-site ligand. Potential hits will be chemically linked to create ligands with substantially higher affinity than either ligand alone. My current first-site binder reduces c-MYC protein levels without affecting mRNA levels, suggesting translational regulation. In Aim 2, I will use functional studies to address a critical gap in our understanding of how small- molecule ligands can target RNA, to produce meaningful biological effects, focusing on how RNA structure affects ribosome recruitment to the mRNA. This project is designed to provide a comprehensive understanding of ligand-RNA interactions in cells, informing the development of RNA-targeted tool compounds and, ultimately, therapeutics. The overarching vision of this research and training program integrates RNA screening technologies, bioinformatics and cheminformatics, mechanistic studies, and mentoring opportunities. Through this program, I will gain a strong foundation for a career as an independent researcher and leader in RNA chemical biology and therapeutics. Project Number: 1F32GM157941-01A1 | Fiscal Year: 2025 | NIH Institute/Center: National Institute of General Medical Sciences (NIGMS) | Principal Investigator: Omar Hedaya | Institution: UNIV OF NORTH CAROLINA CHAPEL HILL, CHAPEL HILL, NC | Award Amount: $75,052 | Activity Code: F32 | Study Section: Special Emphasis Panel[ZRG1 F04B-S (20)] View on NIH RePORTER: https://reporter.nih.gov/project-details/11244705