Figure 2.

Summary of trafficking and mode of action of ASOs and conjugated siRNAs. (1) Oligonucleotides are first internalized into the endosomal pathway by gymnotic uptake or by receptor-mediated uptake, (2) in early endosomes, dissociation with the targeting moiety or ligand is initiated, and (3) maturation of the early endosome leads to late endosomes where oligonucleotides are found to escape in the cytosol. Late endosomes can either mature to (4) lysosomes, where most of the material is degraded by the low-pH and degradation enzymes, or (5) to Multivesicular bodies where they are (6) exported to the external environment. In the cytoplasm, (A.I) siRNAs are loaded into RISC, where (A.II) the passenger strand is degraded and the guide strand selected. After target recognition by the guide strand (A.III), the mRNA transcript is cleaved and degraded. For ASOs, they can either (B) degrade the target mRNA through the RNAse H1 enzyme or (C) block the translation machinery of the transcript. In the nucleus, ASOs can interfere with (D) the splicing machinery and generate a modified mature mRNA or (B) directly downregulate the target through RNAse H1. Created with BioRender.com, VP25FLPEXQ.