Active pH Modulation by Proton Channel‐Peptide Nucleic Acid Complex for Effective siRNA Endosomal Escape

Small interfering RNA (siRNA) has shown great potential for treating various genetic diseases, but lysosomal degradation limits its bioavailability. Here, we present the design of a Channel-PNA-siRNA (CPR) complex that facilitates endosomal escape by actively inducing osmotic rupture of the endosomes. Our design utilizes the transmembrane domain of the M2 proton channel (M2TM) to facilitate proton influx into the liposome while serving as an anchor for siRNA; this surface tethering is mediated by a peptide nucleic acid (PNA) linker that enables stable, non-covalent binding. Our results indicate that the CPR complex interrupts the endosomal pathway, effectively reducing the colocalization of delivered siRNA with lysosomes and enhancing gene regulation efficiency. Furthermore, we demonstrate dual-target gene knockdown using CPR-tethered liposomes by encapsulating additional siRNAs in the empty cavity of the liposome. These findings highlight the therapeutic potential of modulating the endosomal pH environment to prevent lysosomal degradation, suggesting that the CPR nanocarrier design could serve as a versatile platform for targeted RNA delivery.