Cyclin D1 contributes to 15-40% of breast cancers through DNA amplification and overexpression. It is associated with estrogen receptor positive tumors, and predicts both a poor prognosis for ER+ tumors and resistance to anti-estrogen treatments. MMTV-cyclin D1 mice develop ER positive adenocarcinomas whose development is not blocked by inhibition of cyclin dependent kinase 4/6 using an MMTV-p16 transgene. Cyclin D1 is also downstream of multiple mitogenic pathways including erbB2, and its function is required for erbB2 transformation. We propose additional studies of the role of cyclin D1 in breast cancer as both a direct cause of ER positive tumors and as a mediator of erbB2's oncogenic function.
Aim 1 will evaluate Cdk 4-independent activities of cvclin D1, including interactions between cyclin D1 and the estrogen receptor, that contribute to breast cancer. MMTV-cyclin D1 will be crossed with MMTV-AIB1 transgenic mice to genetically test the role of cyclin D1/ER interactions. Mutant cyclin D1s incapable of Cdk 4 interactions (mu/KE) and lacking steroid coactivator interactions (mu/LALA) will be expressed in model breast cell lines and using MMTV in transgenic mice to evaluate the role of those activities in D1 oncogenesis. We will determine whether cyclin D1 regulates specific estradiol-regulated genes or directly alters ER coactivator/corepressor promoter binding in breast cancer cells.
Aim 2 will evaluate the proposed erbB2 ? cvclin D1 pathway in breast cancer. Expression patterns of erbB2 and cell cycle regulators will be evaluated in invasive breast cancers to confirm our preliminary observation that erbB2 is expressed non-redundantly with p16 and pRb loss. Cell cycle functions of erbB2 will be tested in transgenic models to determine whether cyclin D1 can rescue erbB2-deficiency phenotypes and establish whether erbB2 expression is sufficient to deregulate cell cycle progression without need for additional loss of p16 or gain of cyclin D1.
Aim 3 will use microarray analysis of laser capture microdissected hyperplasia and tumors from cyclin D1 and erbB2 transgene-induced tumors to identify genes involved in neoplastic progression from hvperlasia to cancer. Our initial screen has identified six genes whose altered regulation is seen in both human and mouse invasive adenocarcinoma. They are predominately genes involved in cell death regulation and we show several are estradiol regulated. These candidate genes involved in neoplastic progression will be validated by analysis of human cancers, in functional assays, and by assessment of their regulatory controls.
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