Molecular regulation of breast cancer growth

描述（由申请人提供）：我们的实验室长期以来一直对识别分子和理解调节肿瘤生长的机制感兴趣。我们最近在乳腺癌妇女的尿液中发现了NGAL（中性粒细胞明胶酶-相关脂钙蛋白）的存在，既可以单独存在，也可以与明胶酶MMP-9结合。NGAL是脂钙蛋白家族的一员，脂钙蛋白家族是一组小的分泌蛋白，可以运输和呈递配体，与细胞表面受体结合，形成大分子复合物。NGAL的存在与乳腺癌的发生和进展有关，并且已提出其作为人类乳腺癌生物标志物的潜力。我们最近证明，NGAL在人乳腺癌细胞中的过度表达可诱导EMT（上皮细胞向间充质细胞转化），这是癌症进展的标志。我们发现NGAL诱导上皮标记物如E-cadherin的下调，同时伴有间质标记物vimentin和纤维连接蛋白的上调，以及诱导与E-cadherin介导的细胞-细胞粘附丧失一致的散射表型的发展。我们还确定转录因子Slug的表达水平，已知可诱导EMT，在表达ngal的细胞中增加。此外，NGAL过表达可显著下调雌激素受体α (er- α)的表达。最后，我们已经证明NGAL过表达导致人类乳腺癌细胞的运动性和侵袭性显著增加。综上所述，这些数据表明NGAL可以诱导EMT，并可能调节乳腺癌的侵袭性。在我们提案的具体目标的背景下，我们将确定NGAL诱导乳腺癌细胞EMT的机制。这些机制研究将辅以一系列体内研究，在这些研究中，我们将使用三种互补的体内模型来确定NGAL对人乳腺癌体内进展的影响。这些研究还将评估NGAL作为乳腺癌治疗靶点的潜在作用。在本研究的第三个目的中，我们将确定NGAL单独或与其他癌症生物标志物联合是否可能成为乳腺癌的诊断和/或预后生物标志物。这些研究是在以下具体目标的背景下提出的：确定NGAL诱导乳腺癌细胞上皮细胞向间质细胞转化的机制；2. 研究NGAL在体内是否能促进人乳腺癌的进展；3. 在乳腺癌动物模型和人类乳腺癌患者中，确定NGAL单独存在或与其他尿癌生物标志物复合存在是否预测肿瘤的存在和治疗效果。通过系统地识别可能在侵袭性乳腺癌发展中发挥作用的新分子，并通过剖析这些分子可能发挥其作用的机制，我们将有机会开发新的和改进的治疗、诊断和预后策略，从而显著提高乳腺癌患者的生存率。

DESCRIPTION (provided by applicant): Our laboratory has had a long standing interest in identifying the molecules and understanding the mechanisms that regulate tumor growth. We have recently identified the presence of NGAL (Neutrophil Gelatinase- Associated Lipocalin) both alone, and complexed to the gelatinase MMP-9, in the urine of women with breast cancer. NGAL is a member of the lipocalin family of proteins, a group of small, secreted proteins that can transport and present ligands, bind to cell-surface receptors, and form macromolecular complexes. The presence of NGAL has been correlated with breast cancer development and progression and its potential as a biomarker of human breast cancer has been suggested. We have recently demonstrated that the overexpression of NGAL in human breast cancer cells induces an EMT (Epithelial to Mesenchymal Transformation), a hallmark of cancer progression. We have found that NGAL induces the down-regulation of epithelial markers such as E-cadherin, along with a concomitant upregulation of the mesenchymal markers vimentin and fibronectin as well as inducing the development or a scattering phenotype consistent with the loss of E-cadherin- mediated cell-cell adhesion. We have also determined that the expression level of the transcription factor Slug, known to induce EMT, is increased in NGAL-expressing cells. In addition, the expression of the estrogen receptor alpha (ERalpha) was significantly downregulated by NGAL over-expression. Finally, we have demonstrated that NGAL overexpression leads to significantly increased human breast cancer cell motility and invasivity. Taken together, these data demonstrate that NGAL can induce EMT and may regulate breast cancer aggressiveness. Within the context of the Specific Aims of our proposal, we will determine the mechanism(s) by which NGAL induces EMT in breast cancer cells. These mechanistic studies will be complemented by a series of in vivo ones in which we will determine the effects of NGAL on human breast cancer progression in vivo using three complementary in vivo models. These studies will also evaluate the potential role of NGAL as a therapeutic target in the treatment of breast cancer. In the third aim of this study, we will determine whether NGAL, alone or in combination with other cancer biomarkers, may be a diagnostic and/or prognostic biomarker for breast cancer. These studies are proposed within the context of the following Specific Aims: 1. To determine the mechanism(s) by which NGAL induces an epithelial to mesenchymal transition in breast cancer cells; 2. To determine whether NGAL can promote human breast cancer progression in vivo; 3. To determine whether the presence of NGAL, alone or multiplexed with other urinary cancer biomarkers, predicts tumor presence and therapeutic efficacy in animal models of breast cancer and in human patients with breast cancer. By systematically identifying novel molecules that may be playing a role in the development of aggressive breast cancer, and by dissecting the mechanisms by which such molecules may be exerting their effects, we will have the opportunity to develop new and improved therapeutic, diagnostic and prognostic strategies that could result in significantly improved breast cancer patient survival.