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Polyamine Analogues As Novel Anti-Estrogen Receptor Alpha Agents
Tumor Cell Biology II
Background. Polyamines are essential for cell growth. Synthetic polyamine analogues can mimic natural polyamines in their self-regulation but are unable to substitute for natural polyamines to promote cell growth. A novel family of longer chain analogues termed oligoamines has been developed. Our preliminary studies indicate that oligoamines down-regulate polyamine biosynthesis and inhibit breast cancer cell growth by induction of apoptosis. Importantly oligoamines suppress the expression and transcriptional activity of estrogen receptor alpha (ER); they appear to specifically inhibit the ER minimal promoter element bound by Sp1 and other regulatory factors. Moreover, ER is an important regulator of NF-kB in response to oligoamine cytotoxicity. Objective/Hypothesis. The objective of this project is to elucidate the effect of polyamines on regulation of estrogen signaling and determine the role of ER in oligoamine-induced cytotoxicity in human breast cancer. We hypothesize that 1) natural polyamines are important regulators of ER; 2) down-regulation of the ER signaling pathway by oligoamines contributes to oligoamine-induced cell cycle arrest and apoptosis; 3) oligoamines may be a novel class of agents to overcome endocrine resistance . Specific Aims and Study Design: Aim 1) To determine the role of the polyamine metabolism pathway in ER expression and function. Knockdown of key polyamine regulatory enzymes by siRNAs in human breast cancer cells will determine the precise role of natural polyamines in ER expression. Aim 2) To investigate the mechanisms by which oligoamines repress ER transcription. DNA affinity precipitation assays and mass spectrometry will be used to identify regulatory factors recruited at the ER minimal promoter. The effect of oligoamines on DNA binding activity of the newly identified factors at the ER promoter will be delineated by chromatin immunoprecipitation assay. Aim 3) To determine the mechanistic role of ER in oligoamine cytotoxicity. Cells with constitutive ER expression will be established to assess the role of ER in determining oligoamine cytotoxicity. The effects of oligoamines in endocrine-resistant breast cancer cells will be examined to determine if oligoamines can overcome hormone resistance. Potential Outcomes. These studies will enhance our understanding of polyamine analogue action and ER regulation and potentially provide defined molecular targets for polyamine analogues in breast cancer therapy and prevention.
Intracellular polyamines play an important role in cell differentiation and proliferation. Polyamine analogues can mimic natural polyamine regulation but are biologically inactive or have altered functions. Recently, by innovative techniques, a family of new polyamine analogues called oligoamines has been developed for cancer treatment. These novel analogues have been shown to be effective against the growth of human breast cancer cells through induction of cell death. Our preliminary studies indicate that oligoamines significantly inhibit synthesis and increase breakdown of natural polyamines, thereby depleting polyamine levels. They specifically suppress expression and activity of the estrogen receptor alpha (ER), a principal determinant of growth and differentiation in human breast cancer cells. Several lines of evidence suggest that polyamine analogues may interact with other regulatory proteins like Sp1 and NF-kB to mediate ER expression and oligoamine toxicity in breast cancer cells. These data indicate a new path for regulation of estrogen signaling by these novel analogues. These findings suggest a relationship between polyamines and ER expression in breast cancer cells and underscore the rationale of targeting polyamine metabolism as a potential approach to breast cancer therapy or prevention. In this proposal, we will pursue three lines of investigation to identify the precise role of polyamines in estrogen signaling as well as the molecular mechanisms by which oligoamines induce apoptosis through interference with the function of ER. The first aim will define the role of the polyamine pathway in regulation of the expression and activity of ER in human breast cancer cells. In the second aim, the mechanisms by which oligoamines suppress ER gene transcription will be investigated. The third aim will explore how ER contributes to oligoamine-induced programmed cell death and whether oligoamines can overcome resistance to endocrine therapy. The information gained from these translational studies will provide a foundation for early clinical trials in women with breast cancer and promote the design of new polyamine analogues that might be even more effective agents in treatment and prevention of breast cancer.