The mechanical role of the glycocalyx in cancer cell adhesion

项目摘要 许多类型的癌症--如复发性多形性胶质母细胞瘤，总是致命的--过度表达 粗大的糖蛋白形成一层厚厚的糖帽层。糖萼在物理上将细胞与其 但最近的研究表明，糖基化会矛盾地增加软组织的粘附性。 从而促进癌细胞的转移。这种令人惊讶的现象之所以会发生，是因为 糖萼迫使细胞表面的黏附分子(称为整合素)形成簇状。这些整合素簇 具有协同作用，使它们与周围组织形成更强的粘附力 有可能与同等数量的非聚集整合素。这些合作机制一直在激烈地 近几年的仔细研究；对糖萼的生物物理基础有了更细微的了解- 介导性黏附可以发现治疗靶点，加深我们对癌症转移的一般理解， 并阐明远远超出癌症研究领域的一般生物物理过程。 在这里，我提出了一个假设，即糖萼具有增加机械张力的额外效果 簇状整合素所经历的。整合素作为机械传感器，经历结构转换为 在机械张力作用下的“主动”构象。因此，如果我的假设是真的，我的假设将表明 糖萼在粘连中比以前认识到的更直接的调节作用。这一假说(我 称为本地组织假说)通过研究文献中的间接观察得到了很好的支持 但由于与测量个人机械张力相关的挑战，尚未进行直接测试 活细胞中的生物分子。然而，我职业生涯的大部分时间都致力于开发基于DNA的 机械传感器工具，可用于直接测量活细胞中微牛顿规模的整合素张力。这里 我建议利用这些工具来检验当地组织假说，并澄清这一快速增长的假设 研究领域。我计划在细胞黏附的早期阶段成像整合素的力量，并测试一组 支持或反驳当地组织假说的具体观察结果。同时，我计划利用 我的计算技能，用一种复杂的化学机械模拟方法来检验这个假说。 对于我的博士后(K00)工作，我将过渡到翻译工作，并开发利用 用于癌症诊断的糖萼介导的黏附原理。因为呈现糖基化的是 癌细胞更容易附着在软基质上，我将开发一种基于流动的方法来检测循环 肿瘤细胞(CTCs)使用柔软的底物和不同硬度的底物，便于机械选择 呈现糖基化反应的癌细胞。这种基于机械选择的方法将使机械分析能够 CTC以确定哪些类型的组织最容易转移，并将允许常规 并行的生化图谱分析。这项工作将大大加强对这些机制的理解 以及可以将这一改进的理解用于临床诊断的工具。

Project Summary Many types of cancer – such as recurrent glioblastoma multiform, which is invariably lethal – overexpress bulky glycoproteins to form a thick glycocalyx layer. The glycocalyx physically separates the cell from its surroundings, but recent work has shown that the glycocalyx can paradoxically increase adhesion to soft tissues and therefore promote the metastasis of cancer cells. This surprising phenomenon occurs because the glycocalyx forces adhesion molecules (called integrins) on the cell's surface into clusters. These integrin clusters have cooperative effects that allow them to form stronger adhesions to surrounding tissues than would be possible with equivalent numbers of un-clustered integrins. These cooperative mechanisms have been intensely scrutinized in recent years; a more nuanced understanding of the biophysical underpinnings of glycocalyx- mediated adhesion could uncover therapeutic targets, deepen our general understanding of cancer metastasis, and elucidate general biophysical processes that extend far beyond the realm of cancer research. Here I present the hypothesis that the glycocalyx has the additional effect of increasing mechanical tension experienced by clustered integrins. Integrins function as mechanosensors that undergo structure-switching into “active” conformations when subjected to mechanical tension. As such, my hypothesis would, if true, suggest a more immediate regulatory role of the glycocalyx in adhesion than previously realized. This hypothesis (which I call the local organization hypothesis) is well-supported through indirect observation in the research literature but has not been directly tested due to challenges associated with measuring mechanical tension on individual biomolecules in live cells. However, I have devoted much of my career to developing DNA-based mechanosensor tools that can be used to directly measure piconewton-scale integrin tension in live cells. Here I propose to utilize these tools to test the local organization hypothesis and bring clarity to this rapidly-growing research field. I plan to image integrin forces during the early stages of cellular adhesion and test for a set of specific observations that would support or refute the local organization hypothesis. In parallel, I plan to leverage my computational skills to test this hypothesis using a sophisticated chemomechanical simulation method. For my postdoctoral (K00) work, I will transition into translational work and develop tools that leverage the principles of glycocalyx-mediated adhesion for cancer diagnostic purposes. Because glycocalyx-presenting cancer cells adhere more readily to soft substrates, I will develop a flow-based method for detecting circulating tumor cells (CTCs) using soft substrates and substrates of varying stiffness that facilitate mechanoselection of glycocalyx-presenting cancer cells. This mechanoselection-based method will enable mechanical profiling of CTCs to determine what types of tissues are most vulnerable to metastasis and will also allow for conventional biochemical profiling in parallel. This work will deliver a substantially enhanced understanding of the mechanisms of metastasis, as well as tools that can put this improved understanding to diagnostic use in clinical settings.