Biomolecular Analysis using Liquid Crystals

描述（由申请人提供）：威斯康星大学生物工程研究合作伙伴关系（UW-BRP）将专注于开发新的分子分析工具，这些工具具有用于识别和验证生物终点的潜力，从而可以更准确和快速地评估新型抗癌药物的活性，以确定其分子机制和临床相关性。鉴于表皮生长因子受体（EGFR）的过度表达和突变与一些最无法治愈的癌症密切相关，这项工作最初将集中在表皮生长因子受体（EGFR）的分析上。然而，预计未来所开发的原理将具有足够的通用性，可以应用于其他关键信号分子。鉴于现有的高通量筛选的基础很大程度上限制了它们在体外对受体或其他信号分子富集制剂的分子分析的应用，UW-BRP寻求建立工具的原则，这些工具也可以应用于对培养细胞、异种活检和自发肿瘤组织样本的分析。这种能力最终将使一种跨越分子、细胞和组织水平的基本方法成为可能，并将用于基础研究以及动物和人体临床试验。在本提案中，一个由化学和生物工程、化学和生物化学以及生物分子和生物医学科学等不同专业知识的多学科研究人员组成的团队提出了一种广泛适用的生物分析方法，该方法整合了以下领域的进展：A)表面的纳米制造，b)蛋白质和肽表面共价定向固定的合成和生化策略的发展，c)液晶作为表面捕获蛋白质存在的高灵敏度报告的实现，d)参与与致癌相关过程的关键细胞信号蛋白的研究。具体来说，将在一项研究中将用于报告公认的抗癌靶点（即EGF受体）行为的液晶分析特性与传统分析方法进行比较，该研究将a)快速敏感地评估生物样品中野生型和突变型人类EGF受体的水平和活性，b)验证野生型和致癌形式的EGF受体表现出不同抑制剂特异性的假设。c)评估在体外有效抑制EGF介导事件的药物是否也会在细胞培养中表现出拮抗EGF受体表达和/或活性的能力。这些研究将使用EGF受体系统作为原型，预计该技术将很容易适应于广泛的其他分子靶标。从长远来看，这些新工具应该有助于评估抗癌药物的分子机制和后果，从而促进其从基础生物学研究到临床疗效评估。

DESCRIPTION (provided by applicant): The University of Wisconsin Bioengineering Research Partnership (UW-BRP) will focus on the development of new molecular analysis tools that possess the potential to be used to identify and validate biological endpoints whereby the activity of novel anti-cancer agents can be more accurately and rapidly evaluated as to their molecular mechanism(s) and clinical relevance. The work will initially be focused on the analysis of the epidermal growth factor receptor (EGFR), given that its over expression and mutation has been well-associated with some of the most incurable cancers. However, it is anticipated that the principles to be developed will be sufficiently versatile to be applied to other key signaling molecules in the future. Whereas the basis of existing high throughput screens largely restricts their application to in vitro molecular analyses of enriched preparations of receptors or other signaling molecules, the UW-BRP seeks to establish principles for tools that can also be applied to the analysis of samples from cultured cells and from biopsies of xenographs and spontaneous tumor tissues. This capability will ultimately enable a fundamental approach that will span the molecular, cellular and tissue levels and will be used in both basic research and in animal and human clinical trials. In the present proposal, a multidisciplinary team of researchers with diverse expertise in chemical and biological engineering, chemistry and biochemistry, and the biomolecular and biomedical sciences proposes to develop a broadly-applicable bioanalytical approach that integrates advances in the following areas: a) the nano-fabrication of surfaces, b) the development of synthetic and biochemical strategies for the covalent and oriented immobilization of proteins and peptides on surfaces, c) the implementation of liquid crystals as highly sensitive reporters of the presence of proteins captured on surfaces, and d) the investigation of key cell signaling proteins that participate in processes associated with carcinogenesis. Specifically, the analytical characteristics of liquid crystals for reporting the behavior of the well-recognized anti-cancer target, i.e. the EGF receptor, will be compared to conventional analytical methods in a study that will a) rapidly and sensitively assess the levels and activity of wild-type and mutant human EGF receptor in biological samples, b) test the hypothesis that wild-type and oncogenic forms of the EGF receptor exhibit differential inhibitor specificity, and c) assess if agents that potently inhibit EGF-mediated events in vitro will also exhibit a capacity to antagonize EGF receptor expression and/or activity in cell culture. These studies will use the EGF receptor system as a prototype and it is anticipated that the technology will be readily adaptable to a wide range of other molecular targets. In the long term, these new tools should be useful for the assessment of the molecular mechanisms and consequences of anti-cancer agents, thereby facilitating their research from basic biology through to clinical assessment of efficacy.