OLIGONUCLEOTIDE ANALOGS AS GENE REGULATION AGENTS

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The overall goal of this Program Project is to develop nuclease resistant, nonionic oligonucleotide analogs which can be used to control gene expression in a sequence specific manner at either the mRNA or DNA level. This revised application consists of the following projects: Project I, "Chemistry of Oligonucleoside Methylphosphonates", Paul S. Miller, P>I>; Project II, "Physiochemistry and Molecular Cytology", Dr. Paul O. P. Ts'o, P.I., Dr. Stephen A. Lesko and Dr. Lou-sing Kan, Co-P.I.; Project III, "Oligonucleotide Analogs as Antiviral/Anticancer Agents", Dr. Laure Aurelian, P.I.; and Project IV, "Modulation of Tumor Growth In Vitro and In Vivo", Dr. Esther Chang, P.I. In addition there are two core components, the Scientific Core, Dr. Paul S. Miller, P.I. and the Administrative Core, Dr. Paul O. P. Ts'o, P.I. In Project I, new chemical approaches include the use of 2'-O-methylribose nucleotides in addition to deoxyribonucleotides, the synthesis of chiral (R and S) specific Matagens, the synthesis of deoxyribonucleotide triesters as potential pro-drugs, the use of deoxyuridine substituted oligomers and finally, the development of novel alkylating agents attached to Matagen in addition to the further development of matagen derivatized with psoralen. In Project II, the study of triplex formation will include both pyrimidine-pyrimidine-purine triplexes and purine-purine-pyrimideine triplexes, as well as analogs of cytidine and various types of novel backbone linkages. The investigation will include spectroscopic studies, NMR studies, protection assay, and calorimetry, but also attempt will be made to form triplex with the Matagen and the target segment of DNA in living cells. Through the computer microscopic imaging system, individual RNA and DNA will be visualized quantitatively with specific probes. In Project III, Matagen and psoralen derivatized Matagen will be used to investigate the function of immediate early genes (IEmRNA 4.5 encoding IE68K and IE12K), the regulatory transactivating genes (IE110, IE175), as well as the ribonucleotides reductase gene (RR1). These studies are important to the understanding of viral transactivation and virus growth in neural cells. In Project IV, Matagen and derivatized Matagen will be used to suppress the Ha-ras oncogene expression, (as measured by the p21) in correlation to neoplastic properties in various cellular and animal models.