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.