Biodefense Proteomics Collaboratory

描述(申请人提供)：体外酶组合选择和裂解合成化学组合方法将被用来开发用于蛋白质组学的“硫代适体芯片”(TACH)--一种诊断工具，用于识别和量化关键蛋白质的差异表达，以响应生物恐怖主义威胁(BT)的病原体。这项新的蛋白质组学技术将利用我们专有的硫代选择和硫代修饰的寡核苷酸“硫代适配子”，与我们的合作伙伴Ciphergen的表面增强激光解吸/电离(SELDI)质谱学技术相结合，针对啮齿动物和人类蛋白质组。特别是，我们将研究Bt制剂挑战的细胞因子和关键转录因子(例如，NF-kappaB)的炎症反应。这5个NF-B/REL家族蛋白可以结合形成15个同源和异源二聚体，每一个都在跨细胞核膜转位并与基因的启动子区域结合时执行特定的信号功能。与赛弗吉公司合作，我们还将开发新的、大规模平行的硫代适配子珠基蛋白质组筛选和SELDI质谱学方法，以识别参与Bt病毒免疫反应的未知蛋白质。我们来自TACH/SELDI方法的结果将通过2D凝胶质谱学蛋白质组学方法进行验证。我们还将应用生物信息学分析，将蛋白质表达的变化与现有的基因组数据相关联，这些数据是病毒感染或休克后炎症导致的基因表达变化的结果。阐明这些蛋白表达的变化将有助于病毒疾病的早期诊断和改善预后，以及随后开发有效的药物和免疫干预措施。最初的具体病毒目标包括ArenaVirus、Pichinde和Lassa(后者都在NIH和CDC的A类名单上)。

DESCRIPTION (provided by applicant): In vitro enzymatic combinatorial selection and split-synthesis chemical combinatorial methods will be used to develop a "ThioAptamer Chip" (TACh) for proteomics - a diagnostic tool to identify and quantify the differential expression of key proteins in response to pathogens of concern for bioterrorism threat (BT). This new proteomics technology will utilize our proprietary thioselection and phosphorothioate-modified oligonucleotide "thioaptamers," combined with the surface enhanced laser desorption/ionization (SELDI) mass spectroscopy technology of our collaborating partner, Ciphergen, to target both rodent and human proteomes. In particular we will study the inflammatory response of cytokines and key transcription factors (e.g., NF-kappaB) challenged with BT agents. The five NF-?B/Rel family proteins can combine to form 15 homo- and heterodimers, each performing a specific signaling function upon translocation across the cell nuclear membrane and binding to a gene's promoter region. In partnership with Ciphergen, we will also develop new, massively parallel, thioaptamer bead-based screening of the proteome with SELDI mass-spectrometric methods to identify uncharacterized proteins involved in the immune response to BT viruses. Our results from the TACh/SELDI approaches will be validated by 2D gel mass spectrometric proteomic methods. We will also apply bioinformatic analyses to correlate changes in protein expression with available genomic data on changes in gene expression as a result of inflammation after viral infection or shock. Elucidating these protein expression changes will allow early diagnosis and enhanced prognosis of viral disease, and subsequent development of effective pharmacological and immunological interventions. Specific initial viral targets include arenaviruses, Pichinde and Lassa (the latter on both the NIH and CDC class A lists).