The goals of this proposed work are to develop and optimize the properties of a promising new binding motif for single-stranded polynucleotides, and to utilize this information to construct molecules which may have anti- retroviral properties. The circular ligands described in this proposal are designed to block gene expression for specific sequences of viral RNA by binding exceptionally tightly to these sequences. It has been shown in published reports that linear oligonucleotides and their analogs possess some of these inhibitory properties. Circular oligonucleotides or their analogs may prove to have sufficient advantages over other oligonucleotides that they will eventually be useful as drugs against viral infection. These long-term goals will require that binding by these ligands be studied on a molecular basis before they can be tested in vitro and in vivo for inhibitory activity against expression of HIV-1 genes. Experiments planned for the term spanned by this proposal include both molecular and structural studies as well as applications of the most promising ligands. First, optimization of the yield of the reaction which converts linear precursors into circles will be carried out. Second, structural optimization of the ligands, including size and sequence and their effects on binding strength will be studied. Also examined will be the extent of target sequence variability in RNA strands, as well as the level of sequence specificity that can be achieved in binding. Studies of the nuclease susceptibility of oligonucleotides consisting of natural nucleotides and of synthetically modified ones will add important practical information. Finally, the ligands will be examined for their ability to inhibit transcription and translation of specific genetic sequences in vitro. The insight gained from these proposed experiments should allow the design of optimized molecules which have the potential to act as inhibitors of retroviral replication.
Showing the most recent 10 out of 11 publications
