Mammalian therapeutic riboswitches: Engineering and custom ligands.

Regulation with riboswitches, which induce or repress gene expression in response to a ligand, is moving from proof-of-concept studies to robust clinical-grade gene therapies. These initial stages in RNA-based precision medicine will soon benefit from the next generation of riboswitches, whose engineering results from the combination of deep learning, structural biology, and innovative selection technologies. Recent breakthroughs in these strategies now allow for the identification of riboswitches responsive to wisely chosen pre-selected ligands. Selection of the best RNA-friendly and clinically relevant ligands is within the focus of this review. We analyze natural riboswitches in bacteria and eukaryotes and their distinct ligands. We describe how the field of riboswitch design has moved from traditional to novel strategies for identifying switches that function to regulate expression in vivo with bespoke ligands. We provide a rational analysis of the key features the ideal ligands must hold to grant functionality and ease clinical translation. Future developments may allow autonomous regulators, controlled by disease-generated metabolites, capable of disease control with no human intervention.