AKT MEDIATED TAMOXIFEN RESISTANCE IN BREAST CANCER CELLS

描述(申请人提供)：雌激素受体α(ERAlpha) 通过招募辅助激活因子或辅助抑制因子来增强或抑制转录 蛋白质到靶基因中的调控元件。癌症起源于 雌激素靶组织，如乳腺和子宫内膜，依赖于 雌激素--促进生长的雌激素。抗雌激素，如他莫昔芬，可阻断 招募共抑制子或阻止共激活子的ERpha活性 募集，抑制乳腺癌细胞的生长。然而，大多数 ERα阳性乳腺癌最终获得雌激素非依赖性生长 并对他莫昔芬产生抗药性。我们发现， 生长因子调节的丝氨酸/苏氨酸激酶AKT磷酸化ERa 在体外，并允许在没有雌激素的情况下激活ERAlpha。Ser167英寸 N-末端激活蛋白功能1(AF-1)的负调节区 ERAlpha结构域是AKT磷酸化和激活所必需的。PI3 AKT的上游激活剂KK也增加了AF-L的活性，该活性是 被肿瘤抑制基因PTEN抑制。我们还观察到AKT 过度表达足以将ERα阳性的MCF-7乳腺癌转化为 细胞来自他莫昔芬，对耐药表型敏感。 三苯氧胺耐药与雌激素非依赖性血管内皮细胞的表达 抗凋亡基因Bc l-2，减少细胞凋亡。从这些结果来看，我们 假设：1)AKT通过磷酸化ERAlpha，促进招募 在没有雌激素的情况下将共激活蛋白转化为ERα，从而导致 雌激素调节基因的雌激素非依赖性表达。2)AKT，至 ERpha依赖和ERpha非依赖性机制，保护癌细胞 抗他莫昔芬诱导的细胞凋亡。3)AKT的本构激活 肿瘤的生长因子过表达、基因扩增或缺失 抑癌基因PTEN参与雌激素靶向肿瘤的发生 纸巾。以下研究将检验这些假设：1)我们将确定 AKT如何调节共激活子和共抑制子：ERpha相互作用。2)由 利用显性-负性ERAlpha：共抑制物融合基因，我们将确定 ERpha依赖和非ERpha信号转导的相对重要性 AKT介导的他莫昔芬耐药事件。3)通过使PTEN失活，通过 显性负性突变体或反义RNA，我们将确定是否丢失 PTEN导致ERα活性增加和三苯氧胺耐药生长 乳腺癌细胞。

DESCRIPTION (provided by applicant): The estrogen receptor alpha (ERalpha) enhances or represses transcription by recruiting co-activator or co-repressor proteins to regulatory elements in the target genes. Cancers originating from estrogen-target tissues, such as breast and endometrium, are dependent on estrogen-ERalpha for growth. Anti-estrogens, such as tamoxifen, which block ERalpha activity by recruiting co-repressors or preventing co-activator recruitment, inhibit the growth of breast cancer cells. However, most of ERalpha-positive breast cancers eventually acquire estrogen-independent growth properties and become resistant to tamoxifen. We have discovered that the growth factor-regulated serine/threonine kinase AKT phosphorylates ERalpha in vitro and permits activation of ERalpha in the absence of estrogen. Ser167 in the negative regulatory region of the N-terminal activator function 1 (AF-1) domain of ERalpha is essential for phosphorylation and activation by AKT. PI3 kinase, the upstream activator of AKT, also increased AF-l activity, which was suppressed by the tumor suppressor gene PTEN. We also observed that AKT overexpression is sufficient to convert ERalpha-positive MCF-7 breast cancer cells from the tamoxifen-sensitive to the resistant phenotype. Tamoxifen-resistance correlated with estrogen-independent expression of the anti-apoptotic gene Bcl-2 and reduced apoptosis. From these results we hypothesize that: 1) AKT, by phosphorylating ERalpha, promotes recruitment of co-activator proteins to ERalpha in the absence of estrogen, which results in estrogen-independent expression of estrogen-regulated genes. 2) AKT, through ERalpha-dependent and ERalpha-independent mechanisms, protects cancer cells against tamoxifen-induced apoptosis. 3) Constitutive activation of AKT due to growth factor overexpression, gene amplification or loss of the tumor suppressor gene PTEN contributes to cancer development in estrogen-target tissues. The following studies will test these hypotheses: 1) we will determine how AKT modulates the co-activator and co-repressor:ERalpha interaction. 2) By using a dominant-negative ERalpha:co-repressor fusion gene, we will determine the relative importance of ERalpha-dependent and ERalpha-independent signaling events in AKT-mediated tamoxifen-resistance. 3) By inactivating PTEN, through a dominant-negative mutant or antisense RNA, we will determine whether loss of PTEN leads to increased ERalpha activity and tamoxifen-resistant growth of breast cancer cells.