Reversible one-step acylation facilitates mitochondrial delivery of functional RNA

Mitochondria-targeted RNA therapeutics hold promise for treating mitochondrial disorders and cancer, yet effective mitochondrial gene modulation remains challenging due to the strong negative charge and intrinsic instability of RNA, which hinder transport across the mitochondrial double membrane. Herein, we demonstrate, for the first time, mitochondrial RNA delivery enabled by reversible 2'-hydroxyl (2'-OH) acylation chemistry. Installation of a triphenylphosphonium (TPP)-bearing acyl group at the RNA 2'-OH in a single step creates a plug-and-play platform that simultaneously enhances RNA stability and directs selective mitochondrial accumulation, while allowing spontaneous recovery of native RNA structure and activity. In cell-based assays, this strategy enables efficient and selective silencing of mitochondria-encoded genes with minimal off-target effects. Importantly, in vivo delivery of siND1 selectively suppresses MTND1 expression without affecting MTCO1 and significantly inhibits tumor growth in a xenograft model. Conclusively, this work establishes reversible RNA acylation as a simple and versatile chemical framework for mitochondrial RNA delivery and therapy.