KLF4 regulation of epithelial/mesenchymal transition in breast cancer

描述（由申请人提供）：播散性疾病是乳腺癌死亡的主要原因，确定控制转移形成和生长的分子机制对于开发挽救生命的治疗至关重要。 转移是一个多步骤的过程，通常涉及肿瘤细胞在上皮和间充质状态之间的转变。 这种可塑性已在体外乳腺癌细胞模型和肿瘤中观察到。 此外，表达间充质基因签名的乳腺癌是高度转移性的。 因此，识别控制上皮/间充质性质和上皮向间充质转化（EMT）的机制应该为有助于转移性疾病的过程提供重要的见解。 我们的研究表明，KLF 4（Kr <$ppel样因子4）是人类乳腺上皮细胞中上皮/间充质状态的主要调节因子。 KLF 4的持续表达对于维持非转化细胞中的上皮表型是必要的，而KLF 4过表达诱导乳腺癌细胞中从间充质状态转换为上皮状态。 KLF 4刺激上皮决定因子E-钙粘蛋白基因的转录，并抑制间充质表型诱导因子Snail的表达。 这些数据表明，KLF 4是调节乳腺上皮细胞的上皮/间充质状态的枢纽。支持这种可能性，转化生长因子（TGF）诱导的EMT伴随着KLF 4表达的抑制。 KLF 4增强上皮细胞表型的能力与KLF 8形成对比，KLF 8驱动间充质细胞表型，其表达受TGF β刺激。 因此，Krppel样因子决定乳腺上皮细胞的上皮或间质状态，并可能成为转移能力的基础。 该提案的重点是描述KLF 4在决定乳腺细胞上皮状态中的作用机制。 由于KLF 4在TGF β诱导的EMT过程中受到抑制，我们将首先确定KLF 4是否抑制TGF β诱导的EMT，以及这是否涉及KLF 4下调Snail。 这将揭示KLF 4的精确水平是否对确定TGF β诱导的EMT的程度以及可能的转移至关重要。 其次，我们将评估KLF 8和KLF 4在相同细胞内是否具有拮抗性。 如果是这样的话，这将表明这两种蛋白质的相对浓度决定了乳腺上皮细胞的上皮/间质状态，这种可能性将通过检查KLF 4和KLF 8在人类肿瘤中的相对表达及其与E-钙粘蛋白表达的相关性来进一步评估。 在第三个目标中，我们将评估KLF 4过表达对转移的影响，使用实验转移模型以及转基因模型的本地乳腺癌。 最后，我们预计KLF 4通过调节一系列基因的转录来维持上皮细胞的特征。 我们将使用基于基因组的方法来识别这些基因，并评估它们对上皮表型的贡献。 总之，这些研究将揭示由KLF 4控制的调节上皮表型的分子电路，并对指定转移潜力具有重要意义。 公共卫生相关性：播散性疾病是乳腺癌患者死亡的主要原因，确定控制转移形成和生长的分子机制对于开发挽救生命的治疗方式至关重要。 这项研究将确定KLF 4（一种控制基因活性的蛋白质）如何调节乳腺细胞从原发肿瘤中迁移出来并在另一个部位形成转移的能力。 通过这项工作确定的KLF 4所使用的途径可以作为预防乳腺癌转移的有用的治疗靶点。

DESCRIPTION (provided by applicant): Disseminated disease is the primary cause of death in breast cancer and identifying molecular mechanism(s) that control formation and growth of metastases is critical for developing life-saving treatments. Metastasis is a multi-step process that often involves transitions of tumor cells between epithelial and mesenchymal states. This plasticity has been observed in breast cancer cell models in vitro and in tumors. In addition, breast cancers that express a mesenchymal gene signature are highly metastatic. Thus, discerning mechanism(s) that control epithelial/mesenchymal properties and epithelial to mesenchymal transition (EMT) should provide significant insights into the processes that contribute to metastatic disease. Our studies revealed that KLF4 (Kr¿ppel-like Factor 4) is a dominant regulator of the epithelial/mesenchymal status in human breast epithelial cells. Sustained expression of KLF4 is necessary to maintain an epithelial phenotype in non-transformed cells, while KLF4 overexpression induces a switch from a mesenchymal to epithelial state in breast cancer cells. KLF4 stimulates transcription of the E-cadherin gene, an epithelial determinant, and inhibits expression of Snail, an inducer of the mesenchymal phenotype. These data indicate that KLF4 is a hub for regulating the epithelial/mesenchymal states of breast epithelia. Supporting this possibility, EMT induced by Transforming Growth Factor-¿ (TGF¿) is accompanied by a repression of KLF4 expression. The ability of KLF4 to enforce the epithelial phenotype is contrasted by KLF8 which drives a mesenchymal phenotype and whose expression is stimulated by TGF¿. Hence, Kr¿ppel-like factors determine the epithelial or mesenchymal states of breast epithelial cells and may underlie metastatic capacity. This proposal focuses on delineating mechanism(s) of action of KLF4 in dictating the epithelial state of breast cells. Since KLF4 is repressed during TGF¿-induced EMT, we will first determine if KLF4 inhibits TGF¿-induced EMT and if this involves KLF4 downregulation of Snail. This will reveal whether the precise level of KLF4 is critical for determining the extent of EMT induced by TGF¿ and possibly, metastasis. Secondly, we will assess whether KLF8 and KLF4 are antagonistic within the same cells. If so, this would indicate that the relative concentrations of these two proteins defines the epithelial/mesenchymal status of breast epithelial cells and this possibility will be further assessed by examining the relative expression of KLF4 and KLF8 in human tumors and their correlation with E-cadherin expression. In the third aim, we will assess the impact of KLF4 overexpression on metastasis using an experimental metastasis model as well as a transgenic model of autochthonous breast cancer. Lastly, we expect that KLF4 maintains epithelial characteristics by regulating transcription of a spectrum of genes. We will use genome-based approaches to identify these genes and assess their contributions to the epithelial phenotype. Together, these studies will reveal the molecular circuitry controlled by KLF4 that modulates the epithelial phenotype and has significant implications for specifying metastatic potential. PUBLIC HEALTH RELEVANCE: Disseminated disease is the primary cause of death in breast cancer patients and identifying the molecular mechanism(s) that control formation and growth of metastases is critical for developing life-saving treatment modalities. The studies proposed will determine how KLF4, a protein that controls the activity of genes, regulates the ability of breast cells to migrate out of a primary tumor and form metastases in another site. The pathways used by KLF4 that are identified through this work may then serve as useful therapeutic targets to prevent breast cancer metastasis.