Triplex Hybridization of siRNA with Bifacial Glycopolymer Nucleic Acid Enables Hepatocytetargeted Silencing

Abstract

We have submitted a new manuscript, entitled “Triplex hybridization of siRNA with bifacial glycopolymer nucleic acid enables hepatocyte-targeted silencing” for consideration as an article in ACS Chemical Biology. My co-authors, Xin Xia, Zhun Zhou, Chris DeSantis, John Rossi and I have agreed on the submission of this manuscript. In the current manuscript, we demonstrate the utility of polyacrylates modified with both an artificial base-triple and a sugar (GalNAc) to serve as an siRNA packaging and delivery platform to hepatocytes in culture and in vivo (mice). This work is based upon our previously published finding that base-triple modified polyacrylates can triplex hybridize with oligo T/U DNA/RNA and deliver luciferase-targeted siRNA to HeLa cells engineered to express luciferase (JACS 2015). Herein, we expand on the scope of this work by employing a well-established liver-targeting ligand (GalNAc) to silence an endogenous human target, ApoB, the primary protein component of low density lipoprotein. Our platform was found to be effective in both hepatocyte (HepG2) cell culture as well as in an animal model of cardiac disease that features elevated ApoB levels. This manuscript thus validates the use of a synthetic platform that is designed to integrate both packaging via triplex hybridization with tissue-targeting ligand display. This enabling method could be useful as a new, convenient siRNA platform that accessible through scalable synthesis. Further, this platform would improve the efficiency of screening siRNA sequences for silencing efficacy using the identical glycopolymer carrier, thus alleviating the need for covalent ligand modification of each RNA substrate as typical for competing technologies.

Synthesis of Bifacial Peptide Nucleic Acids with Diketopiperazine Backbones

“…21 22 23 Prior work has demonstrated that α-peptide bPNAs can intracellularly target oligo T/U internal loops in DNA and RNA 9 24 as well as unstructured oligos. 2 4 6 Triplex hybrid stems are functionally compatible with native nucleic acid architectures and can noncovalently target ligands to U-rich internal loops (URILs) in small interfering RNA duplexes, 25 as well as long noncoding RNAs (lncRNAs). 24 Recently, we discovered that backbone structured bPNAs containing just two base-tripling residues 26 27 can readily engage with unstructured oligos as well as structured URILs in lncRNA; moreover, affinity is increased when the dipeptide bPNAs are cyclized to a diketopiperazine (DKP) (Scheme 1 ).…”