The overall goal of this project is to elucidate the mechanism of action of polyamine analogues in facilitating the uptake and function of antisense oligonucleotides (ODNs). The hypothesis is that the selectivity of polyamine analogues in DNA condensation and/or stabilization of RNA.DNA hybrids can be utilized to facilitate the cellular uptake and function antisense ODNs for breast cancer therapy. Antisense ODNs targeted to c-myc and HER-2 genes will be studied. To evaluate the structural specificity effects of polyamine analogues on RNA.DNA hybrid stabilization, association constants will be determined using a molecular beacon technique. The role of ODN nanoparticles in facilitating cellular uptake of ODNs will be quantified by dynamic light scattering as well as electron, scanning force and polarizing spectroscopy. Cellular uptake and function of ODNs will be measured using MCF-7 and SK-BR-3 breast cancer cells. ODN uptake will be determined using radioactive and fluorescent ODNs. Effects of polyamine analogue/ODN combinations on target gene expression, cell growth arrest, and apoptosis will be examined to demonstrate enhanced function of ODNs. Subsequently, HER-2 gene targeted ODN and selected polyamine analogues will be examined for in vivo ODN delivery and function using a transgenic mouse model of breast cancer. The effects of polyamine analogues and ODNs on natural polyamines will be quantified by HPLC. These studies will advance our knowledge of the mechanism of cellular uptake of ODNs, and help to develop novel approaches for breast cancer therapy.