IMPACT OF THE MICROENVIRONMENT ON THERAPEUTIC RESPONSE

乳腺癌在一个动态的微环境中进化。随着癌细胞发展到侵袭期，上皮癌细胞颠覆和招募其他宿主细胞，反过来，纤维肌腱、肌上皮细胞和内皮作用于癌细胞本身。我们现在知道，细胞外基质(ECM)和生长因子可以从根本上改变癌细胞/1的行为，这一背景可以强烈地影响癌细胞对治疗剂/2的反应。在肿瘤方面，细胞外基质是乳腺癌细胞和宿主细胞的产物，肿瘤细胞基因组和转录组的变化调节这种反应。我们假设乳腺癌对治疗药物的反应会因此而改变。 复杂的微环境。我们预测，微环境的细胞组成对于预测哪些药物将成功控制每一种给定的乳腺癌至关重要。我们还认为，未能考虑微环境的作用可能是我们未能成功治疗某些乳腺癌的部分原因。因此，我们认为，考虑到正常组织和特定恶性肿瘤的背景，测试新的治疗方法是重要的。 此外，我们假设治疗反过来将改变癌细胞和微环境。例如，我们已经证明，在乳腺癌治疗中广泛使用的电离辐射在三维培养(3D)/4中持续改变小鼠/3的间质成分以及人乳腺上皮细胞的细胞-细胞和细胞外基质相互作用。作为分割治疗的结果，经治疗的肿瘤组织是治疗之星的标准。因此，重要的是要记住这样一个事实，即肿瘤的表型总是 随着治疗的阶段和持续时间以及治疗方式的变化而变化。 这项建议的目的是批判性地检查微环境相互作用(即细胞-ECM、细胞-细胞-肌上皮和间质-基质相互作用)对治疗反应的影响，并利用这些信息来提高我们可以通过实验和计算对信号通路进行建模的准确性。我们将通过研究60个乳腺癌细胞系的代表性亚类来实现这一点，这些亚类构成了这个综合癌症生物学项目的总体重点。具体来说，我们会： 目的1.建立乳腺癌细胞与成纤维细胞和/或 来自正常或肿瘤组织的肌上皮细胞，将乳腺癌细胞通路逻辑的起始状态定义为共培养的函数。我们在这个目标中的目标是重新定义路径逻辑模型的初始条件，以便它们与共同文化中的条件相匹配。然后，修改后的模型可以用于预测在共培养中对Raf-MEK-ERK途径操作的反应，这可能不同于在项目2中2D和3D培养分析中观察到的反应。 目的2.通过测量体外生长反应、基因表达和表型，确定乳腺癌细胞对Raf-MEK-ERK模块抑制剂在Aim 1发展的特定微环境中的反应。这些研究将测试基于目标1的信息做出的路径逻辑模型预测。响应信息将被用于进一步完善模型。 目的3.建立人源化基质的体内模型，以测试具有微环境的制剂？基于目标2中定义的响应签名。这些研究将提供与体内发现的一致的路径逻辑模型初始条件。然后，对Raf-MEK-ERK扰动的体内反应的通路逻辑模型预测将得到实验验证。

Breast cancer evolves in a dynamic microenvironment. The epithelial cancer cells subvert and recruit other host cells as the cancer progresses to the invasive stage, in turn, fibrobtasts, myoepithetial cells and the endothelium act upon the cancer cells themselves. We now know that extracelUar matrix (ECM) and growth factors can radically alter the behavior of cancer cells/1, and that context can strongly influence how cancer cells respond to therapeutic agents/2. In the tumor context, the extracellular matrix is both a product of the breast cancer cells and the host cells, and that variation in the tumor cell genome and transcriptome modulate this response. We hypothesize that the response of breast cancer to therapeutic agents is modified by this complex microenvironment. We predict that the cellular composition of the microenvironment is crucial for predicting which agents will successfully control each given breast cancer. We also believe that the failure to consider the role of the microenvironment may partially account for our failure to successfully treat some breast cancers. We therefore posit that it is important to test new therapeutics considering the context of normal tissue and the particular malignant tumor. Further, we postulate that the therapy in turn will modify both the cancer cells and the microenvironment. We have shown for example that ionizing radiation, widely used in breast cancer therapy, persistently alters both the composition of the stroma in mice/3 and cell-cell and ceIl-ECM interaction in human breast epithelial cells in three-dimensional cultures (3D)/4. As a result of fractionated therapy, treated tumor tissue is the norm at the star of therapy. It is thus important to be mindful of the fact that the phenotype of the tumors is constantly changing as a function of the stage and duration, as well as modality of therapy. The objective of this proposal is to critically examine the influence of microenvironmental interactions (i.e. cell-ECM, cell-cell- myoepithelial and stromal- interactions) on therapeutic responses and to use this information to increase the accuracy with which we can model the signaling pathways both experimentally and computationally. We will accomplish this by studying representative subclasses of the 60 breast cancer cell lines that comprise the "system" that is the overall focus of this Integrative Cancer Biology Project. Specifically, we will: Aim 1. Develop 3D cultures of interactions between breast cancer ceils and fibroblasts and/or myoepithelial cells that originate from either normal or tumor tissue and define the breast cancer cell Pathway Logic starting state as a function of co-cultures. Our goal in this aim is to redefine the Pathway Logic model initial conditions so they match those in co-cultures. The revised model can then be used to predict responses to Raf-MEK-ERK pathway manipulations in co-cultures that may be different from those observed in analyses of 2D and 3D cultures in Project 2. Aim 2. Determine the response of breast cancer ceils to Raf-MEK-ERK module inhibitors in the context of specific microenvironments developed in Aim 1, by measuring growth response, gene expression, and phenotype in vitro. These studies will test the Pathway Logic model predictions made based on information from Aim 1. Response information will be used to further refine the model. Aim 3. Develop in vivo models of humanized stroma to test agents that have microenvironment? based signatures of response as defined in Aim 2. These studies will provide Pathway Logic model initial conditions that are consistent with those found in vivo. Pathway Logic model predictions of in vivo responses to Raf-MEK-ERK perturbations will then be tested experimentally.