title: PEG-appended beta-(1-->3)-D-glucan schizophyllan to deliver antisense-oligonucleotides with avoiding lysosomal degradation.
abstract: Schizophyllan is a natural beta-(1-->3)-d-glucan existing as a triple helix in water and as a single chain in dimethylsulfoxide (DMSO). As we already reported, when a homo-polynucleotide [e.g., poly(dA) or poly(C)] is added to the schizophyllan/DMSO solution and subsequently DMSO is exchanged for water, the single chain of schizophyllan forms a complex with the polynucleotide. One of the potential applications for this novel complex is an antisense-oligonucleotide (AS ODN) carrier. The present paper describes a modification technique that enabled us to introduce PEG only to the side chain of schizophyllan. This technique consisted of periodate oxidation of the glucose side chain and subsequent reaction between methoxypolyethylene glycol amine and the formyl terminate, followed by reduction with NaBH4. Subsequently, we made a complex from PEG-appended schizophyllan and an AS ODN sequence, and carried out an in vitro antisense assay, administrating the AS ODN complex to depress A375 c-myb mRNA of A375 melanoma cell lines. The PEG-SPG/AS ODN complex showed more enhanced antisnese effect than naked AS ODN dose, i.e., the same level as that of RGD-appended SPG. Here, the RGD system has been shown one on the most effective AS ODN carrier (Science 261 (1993) 1004-1012). When we added nigericin to the assay system, the antisense effect was not affected in the PEG-SPG system, on the other hand, it was almost eliminated in the RGD system. Nigericin is well known to interrupt transport from endosome to lysosome. Therefore, the difference between the PEG and RGD complexes indicates that, in the PEG system, AS ODN was able to escape from lysosomal degradation. The present work has thus proposed a new strategy to delivery AS ODN using schizophyllan as a new carrier.
authors.affiliation: Department of Chemical Process & Environments, The University of Kitakyushu, 1-1, Hibikino, Wakamatu-ku, Kitakyushu, Fukuoka 808 0135, Japan.
authors.forename: Ryouji
authors.initials: R
authors.lastname: Karinaga
authors.forename: Kazuya
authors.initials: K
authors.lastname: Koumoto
authors.forename: Masami
authors.initials: M
authors.lastname: Mizu
authors.forename: Takahisa
authors.initials: T
authors.lastname: Anada
authors.forename: Seiji
authors.initials: S
authors.lastname: Shinkai
authors.forename: Kazuo
authors.initials: K
authors.lastname: Sakurai
mesh_terms: D001673:Biodegradation, Environmental; D045744:Cell Line, Tumor; D020099:Coated Materials, Biocompatible / Q000737:chemistry; D016503:Drug Delivery Systems / Q000379:methods*; D020868:Gene Silencing; D018390:Gene Targeting / Q000379:methods*; D015316:Genetic Therapy / Q000379:methods; D006801:Humans; D008247:Lysosomes / Q000378:metabolism*; D008422:Materials Testing; D008545:Melanoma / Q000235:genetics* / Q000378:metabolism*; D016376:Oligonucleotides, Antisense / Q000008:administration & dosage* / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D011092:Polyethylene Glycols / Q000737:chemistry*; D012566:Sizofiran / Q000737:chemistry*; D014162:Transfection / Q000379:methods*
chemical_list: D020099:Coated Materials, Biocompatible; D016376:Oligonucleotides, Antisense; D011092:Polyethylene Glycols; D012566:Sizofiran