Ricin toxin: Neutralizing antibodies and vaccine design

描述(由申请人提供)：国家过敏和传染病研究所(NIAID)正在支持开发针对B类病原体蓖麻毒素的重组亚单位疫苗的努力。目前的焦点是毒素的267个氨基酸的酶A亚单位(RTA)。到目前为止，疫苗设计的目标是在RTA中进行点突变和定点删除，以减弱其酶活性，使其在人类身上使用是安全的。 很少有人注意到这些突变如何影响RTA上的B细胞表位，而RTA上的B细胞表位对于诱导保护性免疫至关重要。众所周知，中和抗体只占RTA免疫所产生的总抗体的一小部分。压倒性的抗体反应由非中和抗体和毒素增强抗体组成。这一事实可能解释了目前的主要候选疫苗在诱导可检测到的血清中和活性方面相对无效，尽管抗毒素抗体效价很高。因为血清中和抗体效价是对蓖麻毒素免疫的唯一相关因素，因此基于RTA的亚单位疫苗激发可检测的血清中和活性是必不可少的。不幸的是，在缺乏全面的RTA B细胞表位图的情况下，不可能设计出RTA的衍生物，在RTA中保留关键的保护性B细胞表位，同时消除产生非中和(甚至有害的)抗体的表位。这项应用建议生成一个全面的和高分辨率的RTA B细胞表位图。这将使用一组独特的资源来实现，包括一大组RTA特异性单抗，以及一组在蛋白质的所有已知免疫优势区域发生突变的RTA衍生物的集合。这项拟议的研究项目具有重要意义，因为所得到的B细胞表位图将与RTA的现有结构信息结合起来设计新的抗原，其中中和抗体的靶标被保存下来，并在最有效地诱导保护性免疫的背景下呈现。

DESCRIPTION (provided by applicant): The National Institute of Allergy and Infectious Diseases (NIAID) is supporting efforts to develop a recombinant subunit vaccine against the Category B agent ricin toxin. The current focus is on the toxin's 267- amino acid enzymatic A subunit (RTA). To date, vaccine design has been aimed at making point mutations and site-specific deletions in RTA to attenuate its enzymatic activity so that it is safe for use in humans. Very little attention has been paid to how these mutations may affect B-cell epitopes on RTA that are critical for eliciting protective immunity. It is known that neutralizing antibodies consttute only a very small fraction of the total antibody pool elicited by RTA immunization. The overwhelming antibody response is made up of non- neutralizing and toxin-enhancing antibodies. This fact may explain the relative ineffectiveness of the current lead vaccine candidate to elicit detectable serum neutralizing activity despite high titer anti-toxin antibodies Because serum neutralizing antibody titers are the singular correlate of immunity to ricin, it is essential that RTA-based subunit vaccine stimulate measurable serum neutralizing activities. Unfortunately, in the absence of a comprehensive B-cell epitope map of RTA it is not possible to engineer derivatives RTA in which key protective B-cell epitopes are preserved, while epitopes that give rise to non-neutralizing (or even deleterious) antibodies are eliminated. This applicatio proposes to generate a comprehensive and high-resolution B-cell epitope map of RTA. This will be accomplished using a unique collection of resources, including a large panel of RTA-specific mAbs, as well as a collection of RTA derivatives with mutations in all of the known immunodominant regions of the protein. The proposed research project is significant in that the resulting B-cell epitope map will be used in conjunction with the available structural information about RTA to design novel antigens in which targets of neutralizing antibodies are preserved and presented in a context designed to most effectively elicit protective immunity.