Wild-type p53 tumor suppressor protein (wtp53) promotes cell cycle arrest and apoptosis and inhibits VEGF-dependent angiogenesis, which is required to support rapid tumor growth. In contrast, mutant p53 (mtp53) fails to inhibit VEGF-dependent angiogenesis and thereby permits rapid tumor growth. However, apoptosis can be restored in tumor cells by co-expression of wtp53 and mtp53 or by treatment with PRIMA-1 (p53 reactivation and induction of massive apoptosis), a small molecule activator of mtp53. We recently observed that in many tumor cells, the effect of mtp53 on VEGF is mediated by progestins and the progesterone receptor. Importantly, PRIMA-1 also blocks this reaction. These data suggest that mtp53, which is often expressed at a high level in tumor cells, might be a useful molecular target for anti-tumor drugs, if its wild-type function could be efficiently re-activated in vivo. Furthermore, such an approach might reduce breast cancer risk in women exposed to progestins (for example, during hormone replacement therapy). This proposal will explore the potential of this idea. The proposed research will test the following hypothesis: PRIMA-1 re-activates mtp53 and restores wtp53 function, thereby inhibiting cell proliferation, expression of VEGF, angiogenesis and metastasis, and stimulating apoptosis in p53-deficient breast tumor cells.
The specific aims of the proposed research are: (1) Determine the mechanism by which PRIMA-1 induces cell-cycle arrest and/or apoptosis in p53-defective human breast tumor cells. (2) Determine the mechanism by which PRIMA-1 regulates expression of VEGF in p53-defective human breast tumor cells. (3) Characterize the effects of PRIMA-1 on incidence and progression of human breast tumor xenografts in nude mice. (4) Characterize the effects of PRIMA-1 on metastasis of human breast cancer cells in nude mice. The methods to be used include FACS, ELISA, Western blotting, immunohistochemistry, real-time RT-PCR, gelshift, chromatin immunoprecipitation and in vivo xenograft tumor mouse models. It is expected that the proposed research will yield valuable insights into the role of wtp53 and mtp53 in carcinogenesis. These studies should lead to the development of a novel approach for treating progestin-dependent and progestinindependent breast cancer in humans. Furthermore, we contend that the research will provide critical data relevant to minimizing breast cancer risk and metastasis in post-menopausal women and other women exposed to exogenous progestins.
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