Sequence specific DNA binding small molecules that can permeate human cells could potentially regulate transcription of specific genes. When one considers the fact that within the next few years the entire human genome will be mapped and sequenced and, coupled to the remarkable discoveries in biology and human medicine which link human disease to specific genes, fundamental research on these DNA binding ligands could lead to reagents for research in functional genomics and, importantly, a new class of human therapeutics. Our objective has been to elucidate chemical principles for the design of small molecules which bind predetermined double-helical DNA sequences with the affinity and specificity of proteins in order to target predetermined sites within the human genome. The Py-Im-Hp polyamides, the result of a 20-year chemistry program to understand the physical organic principles for DNA recognition, are cell permeable ligands only a few percent the size of a protein which have the affinity and specificity of transcription factors. These synthetic DNA binding ligands have been shown to penetrate human cells, traffic to the nucleus, find the promoter DNA sequences of transcriptionally active genes and inhibit gene expression. In this next funding period, the scope and limitations of this approach with regard to polyamide configuration, size, cell types, and different families of transcription factors will be examined. Cancer and viral genes important in human health will be targeted. With regard to specific aims, polyamides will be designed and synthesized to: (1) inhibit transcription of human breast cancer oncogenes (Her-2/neu), (2) inhibit transcription of Herpes Simplex Virus (HSV), (3) regulate c-fos promoter activity in a mouse, (4) up-regulate transcription by creation of polyamide-peptide conjugates as artificial transcription activators, (5) inhibit DNA replication in E coli and yeast, (6) chemically modify coding regions of genes by the design of polyamide-mitomycin and nitrogen mustard conjugates, (7) inhibit viral integration of murine leukemia virus into a host chromosome, and (8) enhance cell uptake further by polyamides with membrane translocation peptide sequences.
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