title: Systematic lactose-functionalization of amphiphilic octaamine macrocycle as a gene carrier. Optimization of the charge, size, toxicity, and receptor factors for hepatocyte targeting.
abstract: We investigated the transfection properties of singly and triply lactose-functionalized derivatives, Lac(1) and Lac(3), and non-glycosylated one, Lac(0), of calix[4]resorcarene-based amphiphilic octaamine 1 in light of those previously reported for more extensively glycosylated compounds Lac(5) and Lac(8). They all strongly bind to the luciferase-encoding plasmid DNA pCMVluc (7040 base-pairs) with a saturation stoichiometry of Lac(n)/P approximately =0.5 (n=0, 1, or 3) or 0.7 (n=5 or 8), where P stands for a phosphate moiety of the plasmid DNA. The resulting carrier-DNA conjugates are positively charged and monomeric (monomolecular with respect to DNA) as such when n=0 or 1, neutral and monomeric when n=3, or neutral and aggregated when n=5 or 8. Transfection of HeLa (uterine) and HepG2 (hepatic) cell lines shows a general trend of decreasing luciferase expression efficiencies (E) in lively cells as well as cytotoxicities with increasing n's. The cell selectivities for HepG2 over HeLa sharply increase with increasing n's; E(HepG2)/E(HeLa)=0.3, 0.6, 7, 14, and 120 for Lac(0), Lac(1), Lac(3), Lac(5), and Lac(8), respectively, as a result of specific receptor pathway involving the asialoglycoprotein receptors on the hepatic (HepG2) cell surfaces and clustering lactose moieties of the carrier-DNA conjugates. The toxicity-corrected, overall efficiency of gene delivery to hepatocytes is optimized at Lac(3), which forms compactly packed (approximately 40 nm), charge-masked (xi approximately = 0 mV), and less toxic virus-like particles capable of receptor-mediated hepatocyte targeting.
authors.affiliation: Department of Synthetic Chemistry and Biological Chemistry, Graduate School of Engineering, Kyoto University, Katsura, Nishikyo-ku, Kyoto 615-8510, Japan.
authors.forename: Shohei
authors.initials: S
authors.lastname: Horiuchi
authors.forename: Yasuhiro
authors.initials: Y
authors.lastname: Aoyama
mesh_terms: D037263:Asialoglycoprotein Receptor / Q000378:metabolism*; D002470:Cell Survival / Q000187:drug effects; D004247:DNA / Q000235:genetics / Q000378:metabolism*; D005260:Female; D017930:Genes, Reporter; D015316:Genetic Therapy / Q000379:methods; D006031:Glycosylation; D006367:HeLa Cells; D022781:Hepatocytes / Q000378:metabolism*; D006801:Humans; D007785:Lactose / Q000737:chemistry*; D008156:Luciferases; D047028:Macrocyclic Compounds / Q000737:chemistry / Q000378:metabolism* / Q000633:toxicity; D010316:Particle Size; D010957:Plasmids; D011073:Polyamines / Q000737:chemistry / Q000378:metabolism* / Q000633:toxicity; D014162:Transfection / Q000379:methods*; D014599:Uterus / Q000166:cytology / Q000378:metabolism
chemical_list: D037263:Asialoglycoprotein Receptor; D047028:Macrocyclic Compounds; D011073:Polyamines; C513577:octaamine 1; D004247:DNA; D008156:Luciferases; D007785:Lactose