Development of a Flexible Micro Spring Array device for viable circulating tumor

描述（由申请人提供）：用于活循环肿瘤细胞富集、分析、原代培养和离体治疗反应测试的柔性微弹簧阵列装置的开发转移性癌症占癌症相关死亡的90%以上。癌症转移是恶性循环肿瘤细胞（CTC）从原发性肿瘤分离、通过外周血扩散、然后在远处器官中侵入和增殖的过程。检测血液样本中CTC的根本挑战是它们非常罕见，需要数十亿血细胞中仅少数肿瘤细胞的检测灵敏度。由于大多数实体瘤细胞是上皮来源的，并且几乎总是比正常血细胞显著更大和更刚性，因此基于尺寸的分离已被证明是用于CTC捕获的有效方法。该项目的第一个目标是表征和优化一种新的柔性微弹簧阵列（FMSA）装置，用于从外周血样品中富集CTC。结合到装置设计中的新型弹簧结构最小化CTC所经历的集中应力，并促进其健康、存活和增殖。我们已经优化了装置的差距尺寸，以可靠地实现超过85%的回收率、针对白细胞的104富集、80%的存活率以及EDTA管中5mL新鲜血液在5分钟内的通量。该技术是便携式的，便宜的，快速的，比CellSearchTM更有效，这是目前FDA批准的唯一一种用于有限种类癌症的CTC计数系统。在我们的初步研究中，我们还发现，驱动压差是至关重要的细胞活力和增殖能力。安全操作压力比通常用于常规细胞过滤的压力低几个数量级。因此，我们的第二个目标是开发一个MEMS启用自动低压控制系统，以促进FMSA设备。与FMSA装置一起，整个系统可以解决40多年来广泛使用的常规过滤因细胞损伤和装置堵塞而无法富集活的哺乳动物细胞的问题。更重要的是，FMSA系统能够捕获可用于原代培养和离体治疗药物疗效测试的活肿瘤细胞，这是拟议研究的第三个目标。成功捕获有活力的CTC将促进建立用于其生长成原代培养物的系统和方案。这将为体外测试肿瘤细胞对多种抗癌药物的反应提供基础。如果证明 为了有效，可以开发更具体和个性化的治疗计划，而不必使患者不必要地暴露于化疗的成本和毒性作用。随着对装置灵活性和原代细胞培养的进一步优化，以及自动低压控制原型的建立，FMSA富集系统可以容易地适应临床使用。从我们的临床合作者那里获得的临床样本的初步研究将 使用FMSA系统评估CTC检测、原代培养和离体药物效力测试。 公共卫生相关性：最致命的癌症形式可以释放出攻击性细胞，这些细胞通过血液传播并扩散到全身。该项目将探索使用一种新的微制造设备来捕获这些入侵细胞并将其从患者血液样本中分离出来。通过培养和分析这些细胞获得的信息可用于改进检测、诊断和制定个性化治疗计划。

DESCRIPTION (provided by applicant): Development of a flexible micro spring array device for viable circulating tumor cell enrichment, analysis, primary culture and ex vivo therapeutic response test Metastatic cancer accounts for over 90% of cancer related deaths. Cancer metastasis is the process by which malignant circulating tumor cells (CTCs) detach from a primary tumor, spread through peripheral blood, then invade and proliferate in distant organs. The fundamental challenge of detecting CTCs in blood samples is the fact that they are so rare, requiring detection sensitivity of only a few tumors cells in billions of blood cells. Since the majority of solid tumor cells are of epithelial origin, and are almost always significantly larger and more rigid than normal blood cells, size based separation has been demonstrated as an effective method for CTC capture. The first goal of this project is to characterize and optimize a new flexible micro spring array (FMSA) device for the enrichment of CTCs from peripheral blood samples. The novel spring structures incorporated into the device design minimize concentrated stresses experienced by the CTCs and encourage their health, survival and proliferation. We have optimized the gap size of the device to reliably achieve over 85% recovery, 104 enrichment against leukocytes, 80% viability, and throughput of 5mL fresh blood in EDTA tube in 5 minutes. This technology is portable, cheap, fast, and more effective than CellSearchTM, which is the only current FDA approved system for CTC enumeration in a limited variety of cancers. In our preliminary study, we also found the driving differential pressure is critical for ell viability and proliferability. The safe operation pressure is orders of magnitude lower than the pressure normally used for conventional cell filtration. Thus our second goal is to develop a MEMS enabled automatic low pressure control system to facilitate the FMSA device. Together with the FMSA device, the whole system can solve the over 40 years' problem that widely used conventional filtration can't enrich viable mammalian cells because of cell damage and device clogging. More importantly, the FMSA system is capable of the capture of viable tumor cells that are available for primary culture and ex vivo therapeutic drug efficacy test, which is the third goal of the proposed study. The successful capture of viable CTCs will prompt the establishment of systems and protocols for their growth into a primary culture. This will provide the basis for testing the tumor cells' response to a variety of anticancer drugs ex vivo. If proven to be effective, a more specific and personalized treatment plan may be developed without having to unnecessarily expose a patient to the cost and toxic effects of chemotherapy. With further optimization for device flexibility and primary cell culture, and establishment of an automated low pressure control prototype, the FMSA enrichment system can be readily adapted for clinical usage. A pilot study of clinical samples obtained from our clinical collaborators will evaluate CTC detection, primary culture, and ex vivo drug efficacy tests with the FMSA system. PUBLIC HEALTH RELEVANCE: The most deadly forms of cancer can release aggressive cells that travel through the bloodstream and spread throughout the body. This project will explore the use of a new microfabricated device to capture these invading cells and isolate them from a patient blood sample. The information obtained by growing and analyzing these cells can be used for improved detection, diagnosis, and development of personalized treatment plans.