A High-throughput Route to Synthetic Antibodies for Array-based Cancer Detection

描述（由申请人提供）：能够监测大量蛋白质水平的疾病指标（生物标志物），对包括癌症在内的许多疾病的检测和治疗具有很大的希望。虽然大多数检测人类血清中蛋白质水平的分析技术目前依赖于基于elisa的测定或质谱分析，但蛋白质捕获阵列作为一种未来的蛋白质组分析技术已经获得了相当大的兴趣。本申请中描述的研究旨在开发一种综合的高亲和力合成抗体（合体），可用于检测和分析人类血液和唾液中的癌症和癌症相关蛋白质。免疫球蛋白的高成本和缓慢的生产速度限制了合适的高质量亲和试剂的可用性，与之相反，用低成本的化学试剂快速合成人工抗体为蛋白质配体问题提供了一个可能的解决方案。该应用程序的目标是利用我们称为配体和连接体系统重组（SRLL）的技术，开发针对五种已知癌症生物标志物的合成抗体。该技术将表面等离子体共振（SPR）的多重功能与DNA自组装的纳米级精度相结合，以确定从先前通过筛选或选择方法确定的单价配体生产多价结合剂所需的最佳肽对和肽对分离距离。我们把从这个过程中产生的配体称为“合体”。本建议的具体目标是：

DESCRIPTION (provided by applicant): The ability to monitor the levels of large numbers of proteins for indicators of disease (biomarkers) holds great promise for the detection and treatment of many diseases, including cancer. Although most analytical techniques for detecting protein levels in human serum currently rely on ELISA-based assays or mass spectroscopic analysis, protein capture arrays have gained considerable interest as a future technology for proteome-wide analysis. The studies described in this application seek to develop a comprehensive array of high affinity synthetic antibodies (synbodies) that can be used to detect and profile cancer and cancer-related proteins in human blood and saliva. In contrast to immunoglobins, for which high costs and slow production rates limit the availability of suitable quality affinity reagents, the rapid synthesis of artificial antibodies from low cost chemical reagents offers a possible solution to the protein ligand problem. The goal of this application is to develop synthetic antibodies to five known cancer biomarkers using a technology that we call systematic recombination of ligands and linkers (SRLL). This technology combines the multiplex capability of surface plasmon resonance (SPR) with the nanometer-scale precision of DNA self-assembly to identify optimal peptide pairs and peptide pair separation distances needed to produce multivalent binding agents from monovalent ligands previously identified by screening or selection methods. We have termed the ligands that emerge from this process "synbodies". The Specific Aims of this proposal are to: 1. Determine the minimum number of selection steps needed to produce modest affinity peptide ligands to five well-established cancer biomarkers. 2. Generate an array of high affinity synthetic antibodies to the five cancer biomarkers targeted in Specific Aim 1.The relevance of the proposed research resides in the potential for early cancer detection based on routine proteome-wide analysis using synthetic antibody arrays to detect cancer biomarkers presymptomatically.