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Nuclease-Resistant Aptamers for Anthrax Opsonization

用于炭疽调理作用的核酸酶抗性适体

基本信息

批准号：
6735824
负责人：
John G Bruno
金额：
$ 9.99万
依托单位：
OPERATIONAL TECHNOLOGIES CORPORATION
依托单位国家：
美国
项目类别：
财政年份：
2004
资助国家：
美国
起止时间：
2004-04-01 至 2004-10-31
项目状态：
已结题

项目摘要

DESCRIPTION (provided by investigator): The anti-phagocytic poly-D-glutamic acid (PDGA) capsule of Bacillus anthracis is a major virulence factor of inhalation anthrax. Although generation of antibodies to PDGA might be of value in opsonizing vegetative anthrax, such antibodies would require "humanization". A simpler and less expensive approach would be the development of nuclease-resistant ('shielded') DNA aptamers against PDGA with an Fc-TR or C3bR binding domain at the other end of a 'hybrid' aptamer. In Phase I, Operational Technologies (OpTech) in conjunction with the Biochemistry Department of the University of Texas at San Antonio (UTSA), proposes to develop 2'- amino pyrimidine modified shielded DNA aptamers against PDGA, murine Fc-yR, and C3bR by the SELEX process. OpTech will link the aptamers and compare phagocytosis of PDGA-conjugated microbeads by the RAW 264.7 macrophage-like cell line in the presence and absence of shielded aptamers and versus serum opsonized PDGA-conjugated microbeads. Since polyanionic PDGA appears to inhibit phagolysosome formation and killing of ingested bacteria, targeting PDGA with 2'-amino-modified aptamers may serve to partially neutralize the polyanionic charge and enable bacterial killing by the respiratory burst. In Phase II, OpTech will clone and sequence all aptamers, then assess aptamer-opsonization and bacterial killing in RAW 264.7 cells with virulent anthrax at the Southwest Research Foundation. Since anthrax and some other infectious bacteria utilize capsules to evade phagocytosis, any inexpensive reagent that acts as an effective opsonin would be of tremendous value in the medical, veterinary or agricultural communities. If successful, OpTech will possess a pharmaceutical substance of great commercial value in the post-exposure treatment of anthrax. Further development of the concept for shielded aptamers against hyaluronic acid or other capsule materials, for example, would also lead to better treatments for many Streptococcus-related infections. Such shielded aptamers could even be used in inhalers to enhance alveolar macrophage phagocytosis.

描述（由研究者提供）：炭疽芽孢杆菌的抗吞噬聚-D-谷氨酸（PDGA）胶囊是吸入性炭疽的主要毒力因子。虽然PDGA抗体的产生可能在调理植物炭疽中有价值，但这种抗体需要“人源化”。一种更简单且更便宜的方法是开发针对PDGA的核酸酶抗性（“屏蔽”）DNA适体，其在“杂合”适体的另一端具有Fc-TR或C3 bR结合结构域。在第I阶段，Operational Technologies（OpTech）与德克萨斯大学圣安东尼奥分校（UTSA）的生物化学系合作，提出通过SELEX方法开发针对PDGA、鼠Fc-γ R和C3 bR的2 '-氨基嘧啶修饰的屏蔽DNA适体。OpTech将连接适体，并比较在存在和不存在屏蔽适体的情况下RAW 264.7巨噬细胞样细胞系对PDGA缀合微珠的吞噬作用，以及与血清调理的PDGA缀合微珠的吞噬作用。由于聚阴离子PDGA似乎抑制吞噬溶酶体形成和杀死摄入的细菌，因此用2 '-氨基修饰的适体靶向PDGA可用于部分中和聚阴离子电荷并通过呼吸爆发实现细菌杀死。在第二阶段，OpTech将对所有适体进行克隆和测序，然后在西南研究基金会评估具有毒性炭疽的RAW 264.7细胞中的适体调理作用和细菌杀伤作用。由于炭疽和其他一些传染性细菌利用荚膜来逃避吞噬作用，因此任何充当有效调理素的廉价试剂在医学、兽医或农业界都具有巨大价值。如果成功，OpTech将拥有一种在炭疽病暴露后治疗方面具有巨大商业价值的药物。例如，针对透明质酸或其他胶囊材料的屏蔽适体概念的进一步发展也将导致对许多链球菌相关感染的更好治疗。这种屏蔽的适体甚至可以用于吸入器中以增强肺泡巨噬细胞的吞噬作用。