Research Project
3506130
DESCRIPTION (Adapted from applicant's abstract): Antisense oligodeoxynucleotides (ODNs) have promise as sequence specific agents for treatment of viral diseases, but they suffer from low potency and high cost. A hepatocyte specific delivery system for ODNs will be developed to improve potency of 3'-modified ODNs for ultimate therapy of Hepatitis B (Phase III). The delivery system is designed around a galactose-specific membrane receptor in hepatocytes which rapidly internalizes bound ligands and delivers them to the lysosomes. A galactose-coated, macromolecular carrier system will improve uptake of attached ODNs. Protease sensitive peptide linkers are used which release the ODN from the carrier after delivery to the lysosomes. The released ODNs have 3'-modifications that will be optimized to prevent nuclease degradation and aid in transport to the cytosol/nucleus. The delivery system will be evaluated using two different antisense assays. The 3'-modifications will be optimized in microinjection Paramecium. This novel assay allows antisense effects to be studied separately from ODN uptake issues. A hepatoma cell-line (Hep G2) will be used to optimize the delivery system. Antisense inhibition of surface antigen synthesis will be quantitated using an ELISA assay. Various constructs of 3'-modified peptide-ODNs polyamine "carrier" molecules and galactose "targeting" ligands will be evaluated for improved antisense potency. ODN delivery by these conjugates will be studied by cell fractionation.
