The asialoglycoprotein receptor is abundantly expressed on hepatocytes making it possible to develop hepatocyte-targeted therapies. Selective delivery of tri-antennary N-acetylgalactosamine (GalNAc) conjugated oligos to hepatocytes is attractive for the treatment of metabolic and cardiovascular diseases. However, the synthesis of multivalent GalNAc ligands, currently used in clinical trials, is time consuming and expensive due to their multistep synthesis. We report here design and synthesis of monomeric GalNAc units with minimal number of amide bonds, and symmetrical GalNAc clusters through novel ring structures in fewer synthetic steps. Monomeric GalNAc phosphoramidites are synthesized from commercially available penta-acetylated galactosamine, through a fused oxazoline intermediate. Ring opening of the oxazoline with various chain length alcohols gave β-anomers and the diol functionalities are introduced either through coupling of diol building blocks or via a dihydroxylation reaction on terminal alkenes. Additionally, the syntheses of clusters are carried out with polyphenol and cyclens for faster assembly via amidation reactions. Sequential protection and activation reactions afforded the GalNAc phosphoramidites. The monomeric GalNAc phosphoramidites have the advantage of forming multi-antennary GalNAc clusters in various formats on DNA/RNA synthesizer during the oligonucleotide synthesis. The new GalNAc units synthesized are conjugated to an ApoB100 gapmer antisense oligonucleotide selected as a tool oligo for evaluation in mice. The new GalNAc units conjugated to gapmers showed potent knockdown of ApoB protein in the mice plasma following subcutaneous administration. The knockdown efficiency and duration of effect of the new GalNAc-oligos are similar or better than the standard tri-antennary GalNAc moiety that is currently used in clinical trials. In summary, we synthesized new GalNAc moieties via facile chemical synthesis in fewer steps, conjugated to antisense oligos and demonstrated comparable liver-targeting efficacy in mice. These GalNAc building blocks could be of great value in developing less expensive liver targeted oligonucleotide-based drug candidates.

Design and synthesis of novel GalNAc linker moieties, conjugation to antisense oligonucleotides and evaluation in mice

ACS Spring Meeting 2021

The official archive and repository for all ACS Meetings & Expositions abstracts and select posters or presentations.