Immunoprotective monoclonals to B anthracis spores

描述（由申请人提供）：炭疽芽孢杆菌孢子最近被用作生物恐怖主义制剂。 在这些故意破坏微生物的事件的许多负面后果中，有一个积极的结果：11名典型的致命吸入型炭疽病患者并没有全部死亡。 事实上，喹诺酮类和其他抗生素的积极使用加上对疾病的早期识别，导致11名患者中有6名存活。 不幸的是，数百名可能接触炭疽孢子的其他人需要延长抗生素治疗疗程。 另一个健康问题是，接受抗生素预防的人在治疗结束后，由于休眠的活孢子发芽，仍可能出现吸入性炭疽。 增加患病患者存活的可能性以及那些暴露于疾病的患者保持健康的可能性的一种方法是防止感染性休眠孢子萌发并随后转化为营养细胞。 最近的证据表明，针对PA（B的两个A亚单位毒素的共享B亚单位）的抗体。炭疽杆菌、水肿因子和致死因子）实际上与孢子表面结合并降低孢子萌发的水平，再加上甲醛灭活的孢子可以在豚鼠中用作对抗炭疽的保护性疫苗，这使我们得出以下假设：抗PA和/或其他孢子表面的mAb-表达的抗原可阻断孢子萌发或使孢子更易被巨噬细胞吞噬并最终杀死。 基于这一理论，我们的目标是开发针对B的免疫保护性mAb。炭疽孢子，在动物模型中提供对炭疽的保护。 最终，我们打算将这些mAb人源化，用作短期预防剂或治疗方式。 具体目标如下。 具体目标1是诱导抗辐照B的小鼠抗血清。炭疽菌休眠孢子、辐射活化孢子、活化孢子表面蛋白提取物和重组PA（rPA）。 我们将在酶联免疫吸附试验（ELISA）、体外发芽试验和巨噬细胞试验中检测这些抗血清，以评估吞噬作用、发芽和杀孢子活性。 然后，我们将从血清显示一种或多种这些抗孢子相关活性的动物制备mAb。 具体目标2是评估这些B的能力。炭疽单克隆抗体预防炭疽病首先在小鼠肠胃外孢子攻击模型，如果保护，在吸入性炭疽的兔模型。 具体目标3是通过N-末端测序鉴定每个mAb识别的孢子表面蛋白。 具体目标4是与苏诺尔Molecular Corporation一起启动赋予保护的那些mAb的人源化版本的工程化。

DESCRIPTION (provided by applicant): Bacillus anthracis spores were recently used as agents of bioterrorism. Among the many negative consequences of these deliberate instances of microbiological sabotage was one positive outcome: not all of the 11 victims with the typically lethal inhalational form of anthrax died. Indeed, the aggressive use of quinolones and other antibiotics coupled with the early recognition of disease resulted in the survival of 6 of the 11 patients. Unfortunately, hundreds of other individuals potentially exposed to the anthrax spores required an extended course of antibiotic therapy. A remaining health concern is that the people who received antibiotic prophylaxis may still present with inhalation anthrax after conclusion of their therapy as dormant viable spores germinate. One way to increase the likelihood that patients with disease will survive and that those exposed will have a higher probability of remaining healthy is to prevent the infectious dormant spores from germinating and subsequently transforming to vegetative cells. Recent evidence that antibodies against the PA (the shared B subunit for the two A subunit toxins of B. anthracis, edema factor and lethal factor) actually bind to the surface of spores and decrease the level of spore germination, taken with the fact that formaldehyde-inactivated spores can serve as a protective vaccine against anthrax in guinea pigs, led us to the following hypothesis: mAbs against PA and/or other spore-surface-expressed antigens can block spore germination or render spores more susceptible to phagocytosis and ultimately killing by macrophages. Based on this theory, our goals are to develop immunoprotective mAbs against B. anthracis spores that confer protection against anthrax in animal models. Ultimately, we intend to humanize those mAbs for use as short-term preventative agents or therapeutic modalities. The Specific Aims are as follows. Specific Aim 1 is to elicit mouse antisera against irradiated B. anthracis dormant spores, irradiated activated spores, activated spore-surface protein extracts, and recombinant PA (rPA). We will test those antisera in enzyme-linked immunosorbent assays (ELISAs), in vitro germination assays, and macrophage assays to evaluate phagocytosis, germination, and sporicidal activity. We will then prepare mAbs from those animals whose sera demonstrate one or more of these anti-spore related activities. Specific Aim 2 is to assess the capacity of these B. anthracis mAbs to prevent anthrax disease first in a mouse parenteral spore challenge model and, if protective, in a rabbit model of inhalational anthrax. Specific Aim 3 is to identify the spore-surface protein(s) recognized by each mAb through N-terminal sequencing. Specific Aim 4 is to initiate, with SUNOL Molecular Corporation, the engineering of humanized versions of those mAbs that confer protection.