Detecting Cancer at the Single-Cell Level Using Endogenous Signal Biomolecular Sensors

癌症是一类以不受控制的生长、侵袭和转移为特征的细胞疾病。尽管癌症是多方面的，具有复杂的潜在遗传决定因素，但新出现的证据表明，在不同组织和癌症类型的细胞内信号水平上存在共同的特征。能够检测细胞内这些细胞内分子信号组合的系统在单细胞水平上对疾病的诊断、预防、治疗和监测具有重要的潜力。最近，科学家们开发了第一代基因电路和传感器，它们在细胞中作为信息处理系统运行。拟议的研究将开发和研究用于检测和监测内源性microrna的分子生物传感器。microrna是一类小的非编码RNA，通过靶向RNA转录物来调节基因表达。拟议研究的目标是：(a)设计和优化对内源性microrna组合具有敏感性和特异性反应的分子生物传感器，(b)基于NCI-60小组开发具有集成生物传感器的单克隆肿瘤细胞系，以及(c)通过监测microrna在细胞周期中对药物治疗的表达来研究microrna及其与疾病的关系。知识价值：microrna已经成为发育和疾病（包括癌症）的重要调节因子。实验证据暗示在大多数人类恶性肿瘤中异常的microRNA表达谱。可靠地（以单细胞分辨率）处理microRNA信息的能力将为基础癌症研究产生新的工具，并导致新的癌症治疗方式。具体来说，将microRNA生物传感器应用于肿瘤细胞系将极大地促进我们对活细胞中microRNA表达和动力学的理解。单克隆细胞系与选定的microRNA传感器的发展将铺平道路，为microRNA的研究一类新的工具。这将是一个重要的新方向，将在其他细胞系和其他microrna中引发一系列类似的研究。更广泛的影响：实验将在NCI-60结肠、血液、肺和胰腺肿瘤细胞系中进行。选择特定细胞系的标准是：基于肺癌和结肠癌在美国的发病率（分别是男性和女性癌症死亡的主要原因和男性和女性癌症死亡的第二大原因）；由于胰腺癌和血液癌的5年生存率非常低（尤其是胰腺癌），因此迫切需要新的诊断和治疗工具。包含生物分子网络的单克隆细胞系的收集将提供给科学界。拟议的研究也将形成一个教育平台，学生将接触到一个综合的研究环境，结合生物学和工程学交叉的理论和实验。

Cancer is a class of cellular diseases characterized by uncontrolled growth, invasion, and metastasis. Although cancer is multifaceted with complex underlying genetic determinants, emerging evidence points to common signatures at the level of intracellular signaling across different tissue and cancer types. A system capable of detecting combinations of these intracellular molecular signals in cells has significant potential for the diagnosis, prevention, treatment, and monitoring of the disease at a single-cell level. Recently, scientists have developed first generation genetic circuits and sensors that operate as information processing systems in cells. The proposed research will develop and study molecular biosensors engineered to detect and monitor endogenous microRNAs, a class of small, noncoding RNAs that regulate gene expression by targeting RNA transcripts. The objectives of the proposed research are to: (a) design and optimize molecular biosensors that respond with sensitivity and specificity to combinations of endogenous microRNAs, (b) develop monoclonal tumor cell lines with integrated biosensors based on the NCI-60 panel, and (c) study microRNAs and their relationship to disease by monitoring their expression through the cell cycle in response to drug treatment.Intellectual merit: MicroRNAs have emerged as important modulators in both development and disease, including cancer. Experimental evidence implicates aberrant microRNA expression profiles in most human malignancies. The ability to process microRNA information reliably (with single-cell resolution) will generate new tools for basic cancer research and lead to new cancer treatment modalities. Specifically, the application of the microRNA biosensors to tumor cell lines will dramatically advance our understanding of microRNA expression and dynamics in living cells. The development of monoclonal cell lines with the selected microRNA sensors will pave the way for a new class of tools for the study of microRNAs. This will be a major new direction that will spark a range of equivalent studies in other cell lines and with other microRNAs. Broader impact: The experiments will be performed in NCI-60 colon, hematologic, lung, and pancreatic tumor cell lines. The criteria for the selection of the particular cell lines was based: on the prevalence of lung cancer and colon cancer in the United States (the leading cause of cancer death for both men and women and the second leading cause of cancer deaths for both men and women combined, respectively); because pancreatic and hematologic cancers have very poor 5-year survival (pancreatic cancer in particular) underscoring the urgent need for new diagnostic and treatment tools. The collection of monoclonal cell lines harboring the biomolecular networks will be made available to the scientific community. The proposed research will also form an educational platform where students will be exposed to an integrated research environment, combining theory and experiments at the intersection of biology and engineering.