The hypothesis was put forth recently that the ability of a widely used drug to eradicate a specific diseased cell population could be attributed in part to the ability of that drug to form a special type of DNA adduct. The adduct attracted proteins selectively expressed in the diseased cell. It was reasoned that adducts engaged in tight complexes with proteins would be difficult to repair. The concealed adducts would persist in and selectively kill the diseased cells. The adduct-shielding model suggested a new paradigm by which aberrantly expressed proteins might be exploited to increase the specificity of genotoxic compounds. Bifunctional molecules were designed in which a DNA damaging warhead was tethered to a recognition domain for the estrogen receptor (ER), which is over expressed in many tumors. DNA adducts were formed that were strongly attracted to the ER in vitro and selectively killed cells that over expressed the ER. DNA damaging molecules were also designed that successfully attracted the androgen receptor (AR) and progesterone receptor (PR) to DNA adducts, and these molecules also caused selective cell killing. In the only case thus far examined, one compound was also shown to kill tumors in mouse xenografts. The goal of the proposed work is to elucidate the mechanism by which selective killing occurs. Our hypothesis is that the ER, AR and PR bind to the toxin-DNA adducts and preclude access to the adducts by repair enzymes. In proposed studies, the removal of DNA lesions engaged in complexes with the steroid receptors will be assessed; these studies will determine whether the receptors can indeed hinder repair of the DNA lesions. Synthetic methods will be applied to vary the architecture and properties of the novel DNA adduct complexes in order to identify the molecular features responsible for selective toxicity. Incorporation of protein recognition domains with a range of affinities for hormone receptors will establish whether DNA adducts engaged in tighter adduct-binding complexes are more difficult to repair (and hence more toxic) than those in weaker complexes. Variation of the warhead will be used to direct damage to different helical surfaces. Alterations in the linker connecting the warhead and protein recognition domain will establish the optimal molecular arrangements for adduct-receptor interactions, repair shielding and selective toxicity.
