Optimizing Endosomal Escape for Effective Aptamer-Based Ubiquitin-Mediated Degradation of c-Met.

Endosomal escape is a significant challenge in the delivery of nucleic acid therapeutics, which also limits the cytosolic activity of aptamers that bind membrane proteins and enter cells by endocytosis. Here, we systematically mapped the colocalization of the c-Met targeting aptamer SL1 with early, recycling, and late endosomes following endocytosis, revealing predominant endosomal retention with minimal cytosolic release. We further evaluated two endosomal escape agents, the small molecule chloroquine and the peptide GALA, and found that chloroquine more effectively facilitates aptamer endosomal escape. To functionally exploit enhanced cytosolic delivery, we developed an aptamer-based PROTAC that promotes the interaction of the membrane protein c-Met with an E3 ligase, leading to its ubiquitination. With enhanced endosomal escape, this system achieved ubiquitin-mediated degradation of c-Met, inhibited proliferation and migration of target cells, and suppressed tumor growth in a xenograft model. These results link optimized endosomal escape to intracellular aptamer function and establish a strategy for membrane protein degradation via the ubiquitin-proteasome pathway.