The proposed research involves the development of new drugs for the treatment of AIDS. The investigators plan to design small molecule ligands to target the binding sites for or the regulatory proteins involved in HIV RNA transcription. The targets will be the enhancer and basal promoter elements contained within the long terminal repeat region of the proviral DNA. Recent studies in the laboratory of Dr. P. Dervan at Cal Tech have shown that synthetic pyrrole -imidizole polyamides can be designed and synthesized to target a wide range of DNA sequences with subnanmolar affinities. Studies in Dr. Gottesfeld's laboratory at Scripps Research Institute have shown that these designed ligands inhibit specific transcription factor DNA interactions and are effective inhibitors of gene transcription both in vitro and in vivo. Polyamides will be designed to bind the enhancer region of the HIV LTR and, specifically, to the sequences flanking and including the binding sites for Ets-1, LEF1 and NF-kB transcription factors and the region immediately flanking the TATA box element upstream from the transcription initiation site. The investigators will determine whether these polyamides inhibit the binding of recombinant Ets-1, LEF1 and NFkB proteins to the HIV enhancer and binding of the TATA-box binding protein to the HIV TATA element. Reconstituted transcription assays will be used to monitor the effect of the polyamides on both basal and activator-dependent RNA polymerase II transcription in vitro. Reporter constructs containing the HIV enhancer and promoter will be used to monitor transcription in transfection experiments in lymphoid cell lines. Collaborative studies are planned to determine whether the designed polyamides will inhibit virus replication in lymphoid cells in culture and in human peripheral blood lymphocytes.
