Quantitative Analysis of Cancer Cell Motility using Microfluidic Devices

描述(申请人提供)：一种可以阻止癌细胞离开原发肿瘤的治疗方法可以防止转移并延长癌症患者的生命。然而，这样一种阻止癌细胞迁移的治疗方法还不存在。尽管在过去的50年里进行了大量的研究，但目前与癌症相关的死亡中仍有90%是由转移引起的。预防癌症转移的系统性尝试主要针对癌细胞迁移的背景，通过改变细胞-细胞相互作用，抑制降解基质所需的酶的活性，或改变癌细胞的增殖。尽管这些治疗方法在模型系统中测试时显示出了巨大的潜力，但它们在阻断转移方面的实际成功有限。在这方面，目前研究细胞迁移的模型系统有两个明显的局限性。首先，大多数检测报告了大量细胞的平均迁移，而不能捕捉离群值的行为，即最终导致转移的单个细胞。其次，许多分析改变了细胞过程，进而间接调节细胞的运动；然而，癌细胞的内在运动潜力保持不变，并在逃脱或适应新条件的细胞中变得明显。在这种背景下，需要新的实验模型来研究癌细胞的迁移，能够定量地测量细胞的内在运动性，以及细胞-细胞相互作用、基质降解或底物对癌细胞迁移的调控。我们最近做了一项令人惊讶的观察，当机械地约束在尺寸小于细胞的微型制造的毛细血管中时，来自各种细胞系的癌细胞能够出人意料地快速并持续向一个方向移动几个小时。这种特殊的行为可能与体内的几种情况有关，在这些情况下，癌细胞在组织中的迁移倾向于沿着血管或淋巴管、胶原纤维或白质束等预先存在的路径进行。我们相信，我们的新实验系统可以揭示癌细胞迁移的关键特征，并使我们能够以定量的方式探索许多已知的调控癌细胞迁移的相互作用。为了验证新工具的潜力并展示它们对癌细胞迁移的定量洞察的使用，我们建议：1)定量测量细胞-细胞、细胞-底物和细胞-基质相互作用对微血管内单个癌细胞的矢量运动的影响；2)定量测量异质细胞-细胞相互作用对癌细胞矢量运动的调节。这项拟议的工作将为未来的研究奠定基础，这些研究旨在表征患者癌细胞的运动性，作为实现评估癌症患者预后的新方法、筛选治疗癌症患者转移的新药或个性化癌症治疗的先决条件。 公共卫生相关性：超过90%的癌症相关死亡是由转移引起的。转移瘤是由离开原发肿瘤的细胞形成的，扩散到全身，并在远处的器官如肝、肺、脑或骨骼上定居。了解癌细胞迁移的复杂性的一个主要挑战是准确测量细胞的运动性并量化其受其他细胞过程的调节：例如运动性、侵袭性、粘附性、细胞间通讯。在这项提案中，我们将验证用于研究癌细胞运动的新的微流控工具。了解癌细胞的迁移能力有助于设计新的方法来阻止它们从原发肿瘤转移到组织中。这可以在癌症发生之前防止转移，将癌症转变为一种慢性病，患者可以与之共同生活更长时间。

DESCRIPTION (provided by applicant): A treatment that could stop cancer cells from moving away from the primary tumor could prevent metastases and extend the life of cancer patients. However, such a treatment that stops cancer cell migration does not yet exist. Despite intense research efforts in the past five decades, 90% of the current cancer related deaths are still due to metastases. Systematic attempts to prevent cancer metastases have targeted mainly the context of cancer cell migration, by altering cell-cell interactions, inhibiting the activity of enzymes required to degrade matrix, or altering cancer cell proliferation. Despite the fact that these treatments have shown great potential when tested in model systems, they had only limited practical success on blocking metastases. Two limitations of current model systems for studying cell migration are significant in this respect. First, most assays report the average migration of a large population of cells, and cannot capture the behavior of outliers, single cells that are ultimately responsible for metastases. Second, many assays alter cellular processes which in turn indirectly modulate cell motility; however, the intrinsic motility potential of cancer cells remains the same and becomes manifest in cells that escape or adapt to the new conditions. In this context, new experimental models for studying cancer cell migration are needed that can quantitatively measure the intrinsic motility of cells and the modulation of cancer cell migration by cell-cell interactions, matrix degradation, or substrate. We have recently made the surprising observation that cancer cells from various cell lines are able to move unexpectedly fast and persistent for several hours in one direction, when mechanically constrained in microfabricated capillaries with dimensions smaller than the cells. This particular behavior may be relevant to several in vivo situations where the migration of the cancer cells in tissues is favored along preexisting paths in the form of blood or lymphatic vessels, collagen fibers, or white matter tracts. We believe that our new experimental system could reveal critical characteristics for cancer cell migration and could enable us to probe in quantitative ways many of the interactions known to modulate cancer cells migration. To validate the potential of the new tools and demonstrate their use for quantitative insights into cancer cell migration we propose: 1) to quantitatively measure the effects of cell-cell, cell-substrate, and cell-matrix interaction on the vectorial motility of individual cancer cells inside microcapillaries; 2) to quantitatively measure the modulation of cancer cell vectorial motility by heterogeneous cell-cell interactions. This proposed work will form the basis for future studies aiming at characterizing motility in cancer cells from patients, as a prerequisite for enabling new approaches for evaluating the prognosis of cancer patients, screening new drugs for the therapy of metastases in cancer patients, or personalized cancer treatment. PUBLIC HEALTH RELEVANCE: More than 90% of the cancer related deaths are due to metastases. Metastases are formed by cells that leave the primary tumor, spread in the entire body and colonize distant organs like liver, lungs, brain, or bones. One major challenge in understanding the complexity of cancer cell migration is the accurate measuring of cell motility and quantification of its modulation by other cellular processes: e.g. motility, invasion, adhesion, cell- cell communication. In this proposal we will validate new microfluidic tools for the study of cancer cell motility. Understanding the migration abilities of cancer cells could help designing novel approaches for stopping their migration from the primary tumor and into tissues. This could prevent metastases before they occur, transforming cancer into a chronic disease with which the patient can live a longer period of time.