DESIGN & SYNTHESIS OF OLIGONUCLEOTIDE ANALOG ANTIVIRALS

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A long-range goal of this research program will be the development of modified oligonucleotides, which will inhibit the ability of human cytomegalovirus (HCMV) to infect and replicate in human cells. The basic concept is to construct oligonucleotide derivatives, which are complementary to viral mRNA sequences that are essential for viral replication. The oligonucleotides would be designed to function by catalytically cleaving their complementary sequence. Three types of modifications are to be investigated. These include: 1) oligonucleotides modified at the phosphodiester backbone by stable lipophilic metal complexes designed to prevent nuclease cleavage and to enhance transport; 2) transition metal ligand complexes designed to cleave RNA by a hydrolytic mechanism; and 3) redox active transition metal complexes designed to cleave DNA and RNA through activation of dioxygen leading to radical abstraction of hydrogen atoms at the C-l' and C-4'positions of ribose or deoxyribose. On the basis of molecular modeling, modification at N-2 of dG appears to provide an optimal location for attaching ligands that are to be positioned in the minor groove in a DNA-DNA or DNA-RNA double helix. The nucleases designed to operate on RNA by a hydrolytic mechanism will require two types of modification. N-2 of deoxyguanosine will serve as a site for tethering an imidazole to be positioned adjacent to the 2'-OH of the complementary cytidine. And, C-5 of deoxyuridine will be modified by ligands designed to position a metal complex against the 3'-phosphate destined for cleavage. In the initial phases of the research, it will be necessary to use a relatively simple model system in order to test the concept of sequence-specific cleavage of mRNA. Model studies will be carried out initially with the sequences 5'-ATCT*CAGTG*G*TA-3' and 5'-GAAT*T*CTTTG*CC-3', which are complementary to residues 489-500 and 411-422 of rabbit beta-globin mRNA. Modifications are introduced at N-2 of G* and C-5 of T* spaced either five or six nucleotide units apart. Studies are planned to determine how these modifications affect the specificity and strength of hybridization to complementary DNA and RNA sequences. Subsequently additional modified oligonucleotides complementary to a number of different regions in the rabbit globin mRNA will be synthesized and binding and cleavage experiments carried out. Ultimately, the approach could be applicable to the control of a broad range of viral infections, including most of the viruses frequently associated with AIDS (e.g., herpes viruses, hepatitis viruses, and papillomaviruses). The modular nature of the approach gives it potentially broad applicability to a broad range of viruses. If the model compounds developed during this grant period function as anticipated, then we will be in a position to design oligonucleotides targeted to specific HCMV sequences.