Molecular Basis of Slits as Tumor Suppressors Controlling CXCR4/SDF1 in Breast

描述（由申请人提供）：癌症生物学的目标是了解细胞致瘤时发生的遗传和表观遗传变化的进展。 许多研究表明，CXCR 4在引导乳腺癌细胞转移到其配体SDF 1（CXCL 12）水平高的部位中起着关键作用。 相比之下，相对较少的研究已经探索了在病变发展的早期使该途径失调的后果，即使大部分（>90%）的人乳腺肿瘤在转化的非常早期阶段显示出不适当的CXCR 4表达。 CXCR 4在疾病过程的早期被上调的观察结果，结合最近的研究表明，癌相关的成纤维细胞作为其配体SDF 1的局部来源，已经引发了CXCR 4也可能在原发性肿瘤生长中发挥关键作用的猜测。 我们已经在乳腺增生性病变中发现了这种疾病特征，这些病变在Slit 2和Slit 3或其Robo 1受体中具有功能丧失，这为我们提供了独特的机会来了解CXCR 4和SDF 1在动物模型的综合生理学中乳腺转化早期阶段的功能。 我们的初步数据表明，Slits或其Robo 1受体的功能丧失突变导致组织组织丧失，增殖增加和微环境的许多变化，包括血管生成增加。 基于我们的初步数据，本申请的总体假设是，乳腺中SLIT/ROBO 1信号传导的缺失导致SDF 1/CXCR 4上调，这反过来有助于上皮转化和肿瘤微环境的产生。 为了解决这一假设，本提案的具体目标有三个方面。 在目的I中，我们建议通过使用功能获得和功能丧失方法确定CXCR 4在Slit 2-/-、Slit 3-/-和Robo 1-/-组织中的信号传导状态，来研究SDF 1/CXCR 4信号传导在早期病变发展中的作用。 我们还将评估人类乳腺肿瘤中Slits、Robo 1和Cxcr 4之间是否发生类似的表达变化。 在目标II中，我们建议通过产生腺体来定义乳腺上皮和间质之间发生的串扰，其中SLIT/ROBO 1信号在上皮或间质中被选择性消除。 在目的III中，我们研究了在不存在SLIT/ROBO 1信号传导的情况下产生的促血管生成环境。 我们建议阐明SDF 1的作用，单独，在促进新血管生成，消除VEGF激活Robo 1空背景。 我们还建议通过在乳腺癌细胞中重新表达Slit 2和Slit 3基因并评估对肿瘤生长和肿瘤血管生成的影响来探索Slit 2和Slit 3作为肿瘤抑制因子的作用。 总之，我们已经确定SDF 1/CXCR 4是体内SLIT/ROBO 1信号传导的关键下游调节靶点。 我们建议阐明这种趋化因子轴的失调如何协调细胞与其环境之间的不适当相互作用，导致组织和周围微环境的转变。 公共卫生相关性：乳腺癌的起源仍然难以捉摸。 尽管它被认为是一种遗传性疾病，但据估计，只有5-10%的人类乳腺癌与已知的基因突变有因果关系。 鉴定其突变允许病变进展并变成恶性的基因对于鉴定潜在的治疗靶点至关重要。 对于许多癌症，尤其是乳腺癌，在转移中具有确定作用的候选靶标是G蛋白偶联受体CXCR 4。 本申请的目的是了解CXCR 4如何使用乳腺作为模型系统促进肿瘤进展。

DESCRIPTION (provided by applicant): The goal of cancer biology is to understand the progression of genetic and epigenetic changes occurring in cells as they become tumorigenic. Many studies have demonstrated a critical role for CXCR4 in guiding metastasizing breast cancer cells to sites with high levels of its ligand SDF1 (CXCL12). In contrast, relatively few studies have explored the consequences of having this pathway deregulated early in lesion development, even though a large fraction (>90%) of human breast tumors display inappropriate CXCR4 expression at very early stages of transformation. The observation that CXCR4 is upregulated early in the disease process, combined with recent studies demonstrating that carcinoma associated fibroblasts serve as a local source of its ligand SDF1, have fueled speculation that CXCR4 may also play a pivotal role in primary tumor growth. We have identified such a disease signature in hyperplastic lesions of mammary glands harboring loss-of-function in Slit2 and Slit3 or its Robo1 receptor, giving us the unique opportunity to understand the function of CXCR4 and SDF1 in the early stages of breast transformation within the integrated physiology of an animal model. Our preliminary data demonstrate that loss-of-function mutations in Slits or their Robo1 receptor lead to loss of tissue organization, elevated proliferation and a host of changes in the microenvironment, including increased angiogenesis. Based on our preliminary data, the overall hypothesis of the application is that loss of SLIT/ROBO1 signaling in breast leads to upregulated SDF1/CXCR4, which, in turn, contributes to epithelial transformation and generation of the tumor microenvironment. To address this hypothesis, the Specific Aims of this proposal are three-fold. In Aim I, we propose to investigate the role SDF1/CXCR4 signaling plays in early lesion development by determining the signaling status of CXCR4 in Slit2-/-;Slit3-/- and Robo1-/- tissue using both gain- and loss-of-function approaches. We will also evaluate whether similar expression changes occur between Slits, Robo1 and Cxcr4 in human breast tumors. In Aim II, we propose to define the cross talk that occurs between mammary epithelia and stroma by generating glands in which SLIT/ROBO1 signaling is selectively eliminated in the epithelia or stroma. In Aim III, we investigate the pro-angiogenic environment that arises in the absence of SLIT/ROBO1 signaling. We propose to elucidate the role of SDF1, alone, in promoting neoangiogenesis by eliminating VEGF activation in the Robo1-null background. We also propose to explore the role of Slit2 and Slit3 as tumor suppressors by re-expressing the genes in breast cancer cells and evaluating the effects on tumor growth and tumor angiogenesis. In summary, we have identified SDF1/CXCR4 as key, downstream regulatory targets of SLIT/ROBO1 signaling in vivo. We propose to elucidate how misregulation of this chemokine axis orchestrates inappropriate interactions between cells and their environment, leading to transformation of the tissue and surrounding microenvironment. PUBLIC HEALTH RELEVANCE: The genesis of breast cancer has remained elusive. Even though it is considered a heritable disease, it is estimated that only 5-10% of all human breast cancers are causally linked to known genetic mutations. Identifying genes whose mutations allow a lesion to progress and become malignant is crucial for identifying potential therapeutic targets. For a number of cancers, prominently breast, a candidate target with an established role in metastasis is the G-protein coupled receptor CXCR4. The goal of this application is to understand how CXCR4 contributes to tumor progression using breast as a model system.