Antisense oligonucleotides (ASOs) can modulate gene expression at the mRNA level, providing the ability to tackle conventionally undruggable targets and usher in an era of personalized medicine. A key mode of action for ASOs relies upon RNase H-engagement in the nucleus, however, most mature mRNA is present in the cytoplasm. This disconnect limits the efficacy and biomedical applications of ASOs. In this paper, we have established a new mechanism of action for achieving potent and targeted mRNA knockdown by leveraging a lysosomal degradation pathway. To achieve this, we employ autophagosome-tethering compound (ATTEC) technology that utilises bifunctional small molecules for lysosomal trafficking. In this manner, to induce degradation of target mRNA located in the cytoplasm, we conjugated an ATTEC warhead, ispinesib, to RNase H-inactive ASOs. These fully 2′-O-Methylated RNase H-inactive ASOs have higher chemical stability and tighter mRNA binding than conventional ‘gapmer’ sequences, but cannot be recognised by RNase H. Using our lysosomal trafficking antisense oligonucleotide (LyTON) technology, we show significant lysosome-dependent knockdown of multiple molecular targets in various cell lines, via transfection and gymnotic uptake. The LyTON modification is also able to boost the knockdown efficacy of RNase H-active ‘gapmer’ ASOs. Engineered to degrade mRNA independent of RNase H recognition, LyTONs will enable gene silencing using oligonucleotide chemistries with higher chemical stability, tighter mRNA binding affinity, and improved cell delivery profiles. This will enable us to target a wider range of disease-relevant mRNA, potentially leading to the development of new therapies.