Programmed Instability of Ligand Conjugation Manifold for Efficient Hepatocyte Delivery of Therapeutic Oligonucleotides.

Ligand-targeted drug delivery (LTDD) has gained more attention in the field of nucleic acid therapeutics. To further elicit the potential of therapeutic oligonucleotides by means of LTDD, we newly developed ( R )- and ( S )-3-amino-1,2-propanediol (APD) manifold for ligand conjugation. N -acetylgalactosamine (GalNAc)/asialoglycoprotein receptor (ASGPr) system has been shown to be a powerful and robust paradigm of LTDD. Our novel APD-based GalNAc (GalNAc _APD ) was shown to have intrinsic chemical instability that could play a role in better manipulation of active drug release. The APD manifold also enables facile production of conjugates through an on-support ligand cluster synthesis. We showed in a series of in vivo studies that while the knockdown activity of antisense oligonucleotides (ASOs) bearing 5'-GalNAc _APD was comparable to the conventional hydroxy-L-prolinol-linked GalNAc (GalNAc _HP ), 3'-GalNAc _APD elicited ASO activity by more than twice as much as the conventional 3'-GalNAc _HP . This was ascribed partly to the GalNAc _APD 's ideal susceptibility to nucleolytic digestion, which is expected to facilitate cytosolic internalization of ASO drugs. Moreover, an in vivo / ex vivo imaging study visualized the enhancement effect of monoantennary GalNAc _APD on liver localization of ASOs. This versatile manifold with chemical and biological instability would benefit therapeutic oligonucleotides that target both the liver and extrahepatic tissues.