title: N-Acetylgalactosamine Block-co-Polycations Form Stable Polyplexes with Plasmids and Promote Liver-Targeted Delivery.
abstract: The liver is an ideal target for nucleic acid therapeutic applications (i.e., siRNA, gene therapy, and genome editing) due to its ability to secrete proteins into the blood. In this work, we present the first synthesis of a novel monomer derived from N-acetyl-d-galactosamine (GalNAc) and its polymerization as a facile route to create multivalent delivery vehicles with exceptional targeting efficiency to asialoglycoprotein receptors (ASGPRs) on liver hepatocytes. A series of cationic diblock GalNAc glycopolymers composed of a GalNAc-derived block of fixed length (n = 62) and cationic 2-aminoethylmethacrylamide (AEMA) blocks of varying lengths (n = 19, 33, and 80) have been synthesized and characterized. In addition, nontargeted control polymers consisting of either glucose or polyethylene glycol-derived neutral blocks with an AEMA cationic block were also created and examined. All polymeric vehicles were able to bind and encapsulate plasmids (pDNA) into polymer-pDNA complexes (polyplexes). The GalNAc-derived polyplexes were colloidally stable and maintained their size over a period of 4 h in reduced-serum cell culture media. The GalNAc-derived homopolymer effectively inhibited the uptake of Cy5-labeled asialofetuin (a natural ligand of ASGPRs) by cultured hepatocyte (HepG2) cells at lower concentrations (IC50 = 20 nM) than monomeric GalNAc (IC50 = 1 mM) and asialofetuin (IC50 = 1 μM), suggesting highly enhanced ASGPR binding due to multivalency. These polymers also showed cell type-specific gene expression in cultured cells, with higher protein expression in ASGPR-presenting HepG2 than HeLa cells, which lack the receptor. Biodistribution studies in mice show higher accumulation of pDNA and GalNAc-derived polymers in the liver compared with the glucose-derived nontargeted control. This study demonstrates the first facile synthesis of a multivalent GalNAc-derived block copolymer architecture that promotes enhanced delivery to liver and offers insights to improve targeted nanomedicines for a variety of applications.
authors: Department of Chemical Engineering and Materials Science, and Center for Genome Engineering, ‡Department of Chemistry and Center for Genome Engineering, and §Department of Genetics, Cell Biology and Development, and Center for Genome Engineering, University of Minnesota , Minneapolis, Minnesota 55455, United States.
authors: Yogesh K
authors: YK
authors: Dhande
authors: Department of Chemical Engineering and Materials Science, and Center for Genome Engineering, ‡Department of Chemistry and Center for Genome Engineering, and §Department of Genetics, Cell Biology and Development, and Center for Genome Engineering, University of Minnesota , Minneapolis, Minnesota 55455, United States.
authors: Bharat S
authors: BS
authors: Wagh
authors: Department of Chemical Engineering and Materials Science, and Center for Genome Engineering, ‡Department of Chemistry and Center for Genome Engineering, and §Department of Genetics, Cell Biology and Development, and Center for Genome Engineering, University of Minnesota , Minneapolis, Minnesota 55455, United States.
authors: Bryan C
authors: BC
authors: Hall
authors: Department of Chemical Engineering and Materials Science, and Center for Genome Engineering, ‡Department of Chemistry and Center for Genome Engineering, and §Department of Genetics, Cell Biology and Development, and Center for Genome Engineering, University of Minnesota , Minneapolis, Minnesota 55455, United States.
authors: Dustin
authors: D
authors: Sprouse
authors: Department of Chemical Engineering and Materials Science, and Center for Genome Engineering, ‡Department of Chemistry and Center for Genome Engineering, and §Department of Genetics, Cell Biology and Development, and Center for Genome Engineering, University of Minnesota , Minneapolis, Minnesota 55455, United States.
authors: Perry B
authors: PB
authors: Hackett
authors: Department of Chemical Engineering and Materials Science, and Center for Genome Engineering, ‡Department of Chemistry and Center for Genome Engineering, and §Department of Genetics, Cell Biology and Development, and Center for Genome Engineering, University of Minnesota , Minneapolis, Minnesota 55455, United States.
authors: Theresa M
authors: TM
authors: Reineke
mesh_terms: D000116:Acetylgalactosamine / Q000737:chemistry*; D000818:Animals; D037263:Asialoglycoprotein Receptor / Q000235:genetics / Q000378:metabolism; D004247:DNA / Q000008:administration & dosage; D004337:Drug Carriers / Q000009:adverse effects / Q000138:chemical synthesis / Q000737:chemistry*; D005021:Ethylamines / Q000737:chemistry; D015316:Genetic Therapy / Q000379:methods; D006367:HeLa Cells; D056945:Hep G2 Cells; D022781:Hepatocytes / Q000187:drug effects / Q000378:metabolism; D006801:Humans; D008099:Liver / Q000187:drug effects / Q000378:metabolism*; D008689:Methacrylates / Q000737:chemistry; D051379:Mice; D008810:Mice, Inbred C57BL; D010957:Plasmids / Q000008:administration & dosage*; D011073:Polyamines / Q000737:chemistry*; D000071228:Polyelectrolytes
chemical_list: D000116:Acetylgalactosamine; D037263:Asialoglycoprotein Receptor; D004247:DNA; D004337:Drug Carriers; D005021:Ethylamines; D008689:Methacrylates; D011073:Polyamines; D000071228:Polyelectrolytes; C514459:aminoethyl methacrylate