Magnetorotation: a Rapid Assay for Single Cell Drug Sensitivity of Cancer Cells

描述(申请人提供)：目前许多用于癌症研究、药物开发和精确治疗的高通量细胞检测技术，通常需要在2D表面生长，并需要耗时和劳动密集型的培养步骤。此外，目前的检测方法使用的是大量细胞，这在处理至关重要但数量较少的癌症干细胞(CSCs)时可能会构成挑战。在研究CSCs时，分化可以发生在1-2个细胞分裂中。目前，还没有高通量的方法来测量悬浮的肿瘤干细胞的细胞毒效应。因此，需要一种对单个悬浮细胞和小球体的体积和形状变化敏感的高通量检测方法。为了应对当前方案的局限性，我们正在开发一种基于异步磁珠旋转(AMBR)的细胞磁旋转方法，该方法对单个细胞的变化很敏感，并将允许对单个悬浮细胞和球体进行快速生长和细胞毒性分析。由于其亚显微分辨率、高通量和观察时间短，我们预计这种AMBR检测将显著减少培养步骤时间、结果的总体时间和这种检测所需的细胞数量。为了初步验证这一新方法，我们选择了与前列腺癌相关的单细胞、单干细胞和单球体进行研究。我们利用科佩尔曼实验室的纳米技术和纳米医学专业知识，以及皮恩塔实验室的前列腺癌和药物敏感性专业知识，建立了一个合作关系。基于单个癌细胞的初步结果，我们假设癌细胞和球体对化疗药物的反应所引起的体积变化可以被准确和灵敏地监测。此外，由于所提出的方法的敏感性、速度和简单性，通常数量很少的干细胞可以在转化为异质群体之前进行快速测试。这将使对癌症干细胞和含有癌症干细胞的球体进行快速药物敏感性测试成为可能，从而为特定患者量身定做一种快速而智能的治疗剂选择方案。我们计划在单个前列腺癌和癌症干细胞上实现生长和细胞毒性的示范测量。我们还计划证明AMBR方法可以用来测量前列腺癌球体的生长以及它们对细胞毒剂多西他赛的反应。我们的长期目标是开发一种检测设备，它将直接在液体临床样本(患者血液中的循环肿瘤细胞)中检测靶细胞，并提供自动化结果，无需手动步骤，用于任何形式的癌症的个性化识别和药物敏感性测试。

DESCRIPTION (provided by applicant): The current state of the art of many high throughput cell-based assays, used for cancer research, drug development and determination of precision therapy, often requires growth on 2D surfaces and requires the time-consuming and labor-intensive culturing step. Furthermore, current assays use large cell populations, which can pose a challenge when working with the critically important, but small in number, cancer stem cells (CSCs). When investigating CSCs, differentiation can occur in as little as 1-2 cell divisions. Currently, no high throughput method exists to measure cytotoxic effects on suspended cancer stem cells. So it is desirable to have a high throughput assay that is sensitive to volume and shape changes in single suspended cells and small spheroids. In response to the limitation of current protocols, we are developing a cell magnetorotation method, based on asynchronous magnetic bead rotation (AMBR), that is sensitive to single-cell changes, and that will allow for rapid growth and cytotoxicity analysis of individual suspended cells and spheroids. Due to its submicroscopic resolution, high throughput, and short observation time, we anticipate that this AMBR assay will drastically reduce the culture step time, the overall time to results, and the number of cells needed for such an assay. For the preliminary validation of this new approach we selected to investigate single cells, single stem cells and single spheroids relating to prostate cancer. We set up a collaboration using the nanotechnology and nanomedicine expertise of the Kopelman Lab and the prostate cancer and drug sensitivity expertise of the Pienta Lab. Based on preliminary single cancer cell results, we hypothesize that the volumetric changes, resulting from the response to chemotherapy agents by cancer cells and spheroids, can be accurately and sensitively monitored. Furthermore, due to the combined sensitivity, speed and simplicity of the proposed approach, the typically very small number of stem cells can be tested rapidly, before their turning into a heterogeneous population. This will enable rapid drug sensitivity tests on cancer stem cells and on spheroids containing cancer stem cells, leading to a fast and smart selection protocol for therapeutic agents that can be tailored to a specific patient. We plan to achieve growth and cytotoxicity demonstration measurements on individual prostate cancer and cancer stem cells. We also plan to demonstrate that the AMBR method can be used to measure the growth of prostate cancer spheroids and their response to the cytotoxic agent docetaxel. Our long-term objective is to develop an assay device that will detect target cells directly in liquid clinical specimens (circulating tumor cells from patient blood) and provide automated results, with no manual steps, for personalized identification and drug sensitivity testing, for any form of cancer.