Collagen remodeling and tumor progression

描述（由申请人提供）：本研究的总体目标是阐明间质胶原重塑与乳腺肿瘤进展的相关性。乳腺肿瘤的进展伴随着结缔组织增生反应，其特征是间质胶原的显著重塑，并与组织的进行性硬化有关。渐进式转化的乳腺上皮细胞（MECs）对细胞外基质（ECM）硬度敏感，这意味着动态进化的微环境和渐进式异常的乳腺上皮之间的对话是肿瘤进展的关键。ecm降解基质金属蛋白酶（MMPs）在基质中表达，可切割间质胶原，并参与肿瘤进展的发病机制，MMP表达升高可预测人类乳腺病变的肿瘤进展。然而，MMP抑制剂的临床试验失败，表明ECM重塑的其他参数调节肿瘤行为。赖氨酸氧化酶和赖氨酸羟化酶家族也在基质中表达，在基质中它们交联胶原原纤维以增强组织的机械完整性。这些酶在肿瘤中也会升高，并导致组织纤维化。事实上，在乳房x光检查显示乳房致密的女性中，胶原蛋白和基质蛋白的水平会升高，从而增加患乳腺癌的风险。这些数据强调了胶原交联以及mmp降解对乳腺癌进展的重要性。然而，我们对胶原交联在肿瘤进展中的作用的理解是有限的。这一建议的目标是解剖，在分子和细胞生物学的细节，胶原交联，使ECM硬化，如何促进乳腺癌进展为恶性表型。鉴于肿瘤进展是通过癌基因的水平/活性升高和肿瘤抑制因子的水平/活性降低来介导的，我们认为胶原重塑/硬化通过改变关键癌基因或肿瘤抑制因子的水平/活性来影响肿瘤进展。一致地，我们发现ECM交联和硬度增强整合素和生长因子受体（GFR）信号传导并调节肿瘤抑制因子PTEN的水平。因此，我们假设胶原交联通过增强整合素- gfr信号和降低肿瘤抑制因子（如PTEN）的水平使组织变硬，从而促进乳房转化。该项目将采用跨学科的方法，利用工程方法在培养和体内测量和操纵ECM材料的特性和拓扑结构，结合分子细胞生物学方法和遗传小鼠模型，研究胶原重塑和交联是否通过硬化乳房组织来促进肿瘤进展，从而增强整合素和GFR信号和/或损害PTEN的表达/功能。从胶原蛋白丰度和结构的高风险和低风险女性的肿瘤前乳腺组织进行存档和新鲜活检将建立临床相关性。我们将利用有机型培养物的二维和三维活细胞成像，在天然和合成基质中修改交联、刚度和拓扑结构，并将使用经过药理学或抗体修饰的小鼠来改变胶原结构，以询问胶原交联和刚度是否通过增强PI3激酶信号来促进趋化性和硬性，从而影响MEC的运动和侵袭。这些实验将深入了解胶原蛋白丰度和交联如何促进肿瘤前病变向浸润性乳腺癌的进展。这些知识将有助于确定和表征驱动乳腺肿瘤进展到侵袭的分子机制的方法发展。本研究将为进一步阐明组织ECM在转移和治疗反应中的作用奠定基础。这项工作最终将有助于实现找到治疗乳腺癌的长期目标，并与其他类型的肿瘤直接相关。

DESCRIPTION (provided by applicant): The global objective of this research is to clarify the relevance of interstitial collagen remodeling in breast tumor progression. Neoplastic progression in the breast is accompanied by a desmoplastic response which is characterized by significant remodeling of interstitial collagen and is associated with a progressive stiffening of the tissue. Progressively transformed mammary epithelial cells (MECs) become sensitized to extracellular matrix (ECM) stiffness, implying that it is the dialogue between a dynamically evolving microenvironment and a progressively aberrant mammary epithelium that is key for neoplastic progression. ECM-degrading matrix metalloproteinases (MMPs) expressed in the stroma cleave interstitial collagen and contribute to the pathogenesis of tumor progression, and elevated MMP expression is predictive of neoplastic progression of human breast lesions. Yet clinical trials with MMP inhibitors failed, suggesting other parameters of ECM remodeling modulate tumor behavior. The lysyl oxidase and lysyl hydroxylase family of enzymes are also expressed in the stroma where they cross-link collagen fibrils to enhance the mechanical integrity of the tissue. These enzymes are also elevated in tumors and contribute to tissue fibrosis. Indeed, the levels of collagen and matrix proteins that enhance ECM stiffness are elevated in women with mammographically dense breasts, who are at higher risk for breast cancer. These data emphasize the importance of collagen cross-linking as well as MMP-degradation to breast cancer progression. Nevertheless, our understanding of the role of collagen cross linking to tumor progression is limited. The goal for this proposal is to dissect, in molecular and cell biological detail, how collagen cross-linking, which stiffens the ECM, contributes to breast cancer progression to a malignant phenotype. Given that tumor progression is mediated through elevated levels/activity of oncogenes and reduced levels/activity of tumor suppressors we suggest that collagen remodeling/stiffening influences tumor progression by modifying the levels/activity of key oncogenes or tumor suppressors. Consistently, we found that ECM cross-linking and stiffness enhance integrin and growth factor receptor (GFR) signaling and regulate levels of the tumor suppressor PTEN. Accordingly, we hypothesize that collagen cross-linking stiffens the tissue to promote breast transformation by enhancing integrin-GFR signaling and reducing the levels of tumor suppressors such as PTEN. The project will take an interdisciplinary approach, using engineering methods to measure and manipulate ECM materials properties and topology in culture and in vivo, together with molecular cell biology methods and genetic mouse models to investigate whether collagen remodeling and cross-linking promote tumor progression by stiffening the breast tissue to enhance integrin and GFR signaling and/or compromising PTEN expression/function. Archived and fresh biopsies of pre-neoplastic breast tissue from women at high and low risk for collagen abundance and structure will establish clinical relevance. We will exploit live cell imaging of organotypic cultures in two- and three-dimensions in natural and synthetic matrices with modified cross-linking, stiffness and topology and will use mice that have been pharmacologically or antibody-modified to have altered collagen structures to interrogate whether collagen cross-linking and stiffness influence MEC motility and invasion by enhancing PI3 kinase signaling to promote chemotaxis and durotaxis. These experiments will give insight into how collagen abundance and cross-linking contribute to the progression of pre-neoplastic lesions to invasive breast cancer. This knowledge will assist in the development of approaches to identify and characterize molecular mechanisms driving breast tumor progression to invasion. The studies will lay the groundwork for future studies aimed at clarifying the role of the tissue ECM in metastasis and treatment response. The work will eventually help to achieve the long-term goal of finding cures for breast cancer and is directly pertinent for other tumor types.