Drug discovery and development scheme for liver-targeting bridged nucleic acid antisense oligonucleotides

Abstract

Antisense oligonucleotides (ASOs) containing bridged nucleic acids (BNAs) have been proven to be very powerful. However, ensuring a reliable discovery and translational development scheme for this class of ASOs with wider therapeutic windows remains a fundamental challenge. We here demonstrate the robustness of our scheme in the context of the selection of ASOs having two different BNA chemistries (2, 0 4 0 -BNA/ locked nucleic acid [LNA] and amido-bridged nucleic acid [AmNA]) targeting human proprotein convertase subtilisin/ kexin type 9 (PCSK9). The scheme features a two-step process, including (1) a unique and sensitive in vitro screening approach, called Ca 2+ enrichment of medium (CEM) transfection, and (2) a ligand-targeted drug delivery approach to better reach target tissues, averting unintended accumulation of ASOs. Using CEM screening, we identified a candidate ASO that shows >70% cholesterol-lowering action in monkeys. An N-acetylgalactosamine (GalNAc) ligand then was appended to the candidate ASO to further broaden the therapeutic margin by altering the molecule's pharmacokinetics. The Gal-NAc conjugate, HsPCSK9-1811-LNA, was found to be at least ten times more potent in non-human primates (compared with the unconjugated counterpart), with reduced nephrotoxicity in rats. Overall, we successfully showed that our drug development scheme is better suited for selecting clinically relevant BNA-based ASOs, especially for the treatment of liver-associated diseases.

Chemistry of Therapeutic Oligonucleotides That Drives Interactions with Biomolecules

“…Hence, we developed a monomeric GalNAc phosphoramidite unit, with which we can freely change the ligand valency, and introduced it into PS-ASO equipped with bridged nucleic acids (2 ,4 -BNA/LNA). Unexpectedly, a remarkable improvement in knockdown activity was observed with the introduction of only one GalNAc (Figure 2b,c) [45][46][47]. This suggests that the univalent GalNAc targets are clustered with ASGPR.…”

The Coming Age of Antisense Oligos for the Treatment of Hepatic Ischemia/Reperfusion (IRI) and Other Liver Disorders: Role of Oxidative Stress and Potential Antioxidant Effect

“…Thus far, the ASOs discussed have fallen into a single “activator” or “silencer” category. However, LNAs, specifically gapmers and mixmers, present relatively new and upcoming ASOs in the field for their ability to induce loss-of-function or gain-of-function expression changes [ 112 , 113 ]. In addition to their versatility, LNAs work with much greater specificity and stability in targeting a gene and do not require a delivery agent to enter the cell.…”

A Review of Progress on Targeting LDL Receptor-Dependent and -Independent Pathways for the Treatment of Hypercholesterolemia, a Major Risk Factor of ASCVD

“…In human hepatocytes, bempedoic acid showed inhibition of cholesterol synthesis as well as increases in LDLR activity [ 61 ]. Another protein, inducible degrader of LDL receptor (IDOL), also increases LDL receptor degradation by a different mechanism [ 178 ] which was shown to be regulated by LXR. IDOL knockdown in hepatocytes increased LDLR protein, and IDOL overexpression enhanced LDLR degradation [ 179 ].…”