Protective anti-LPS antibodies: what are they?

描述(由申请人提供)：基于霍乱弧菌脂多糖(脂多糖)的霍乱疫苗的开发是合理的，因为杀弧性抗脂多糖反应与免疫相关。两个O1(血清组)内毒素血清型Inaba和Ogawa与地方性和大流行性霍乱有关。这两种内毒素血清型在感染或接种后都会产生保护性抗体。在结构上，除了末端糖(过氧胺)在Ogawa的第2位有甲氧基，在稻叶有羟基外，其他血清型是相同的。小川脂多糖的末端糖是一种保护性B细胞表位。合成霍乱弧菌血清Ogawa偶联BSA的过氧胺六糖(CHO-BSA)诱导保护性抗体。我们推测稻叶内毒素的末端糖也能诱导保护性抗体。稻叶新糖结合物确实诱导抗体，但令人惊讶的是，它们没有保护性。相比之下，纯化的稻叶内毒素免疫后可产生高度保护性抗体。初步数据表明，由Inaba Cho-BSA结合物诱导的Inaba抗体具有足够的亲和力，可以在ELISA法中得分，但不足以在体内具有保护性。用原生内毒素预置，然后提供稻叶CHO-BSA，在10天内可产生高效价的ELISA和保护性抗体。我们推测稻叶内毒素和稻叶Cho-BSA选择不同的B细胞，并且稻叶Cho-BSA可以与稻叶内毒素选择的B细胞交叉反应扩增它们，从而可能为那些生活在流行区具有记忆性B细胞但体液免疫减弱的人提供一种有效的疫苗免疫原。将产生针对这两种稻叶免疫原的单抗，以确定在免疫期间是否选择了不同的B细胞。R21拨款非常适合这些需要开发试剂以解决霍乱生物学中的一个基本问题的研究。一种新的技术，表面等离子体共振(SPR)将被用来表征mAbs的亲和力。这些实验是有风险的，因为它们需要产生多种试剂来完成抗内毒素抗体的群体分析。这项研究的回报很高，因为无论确切的特异性如何，都会产生试剂，这些试剂将绘制稻叶内毒素末端糖的抗体库。我们将澄清抗V病毒的“类型”。具有保护性的霍乱内毒素抗体，从而证实或不使用稻叶新结合物作为疫苗成分。

DESCRIPTION (provided by applicant): Development of a V. cholerae LPS (lipopolysaccharide)- based cholera vaccine is justified by the correlation of vibriocidal anti-LPS responses with immunity. Two O1 (serogroup) LPS serotypes, Inaba and Ogawa are associated with endemic and pandemic cholera. Both LPS serotypes induce protective antibody following infection or vaccination. Structurally, the serotypes are identical except for the terminal sugar (perosamine) which has a methoxyl group at position 2 in Ogawa, but a hydroxyl group in Inaba. The terminal sugar of the Ogawa LPS is a protective B cell epitope. Synthetic perosamine hexasaccharides of V. cholerae, serotype Ogawa coupled to BSA (CHO-BSA) induced protective antibody. We hypothesized that the terminal sugar of Inaba LPS would also induce protective antibody. Inaba neoglycoconjugates did induce antibodies but surprisingly they were not protective. In contrast, immunization with purified native Inaba LPS induced highly protective antibody. Preliminary data indicate that Inaba antibodies induced by the Inaba CHO-BSA conjugates have sufficient affinity to score in an ELISA but not enough to be protective in vivo. Priming with native LPS and then providing the Inaba CHO-BSA resulted in high ELISA titers in 10 days and antibodies that are protective. We hypothesize that Inaba LPS and Inaba CHO-BSA select different B cells, and that Inaba CHO-BSA can cross react with Inaba LPS-selected B cells to expand them and thus may provide an effective vaccine immunogen for those living in endemic areas with memory B cells but waning humoral immunity. Monoclonal antibodies (mAb) will be generated to the two Inaba immunogens to determine if different B cells are selected during immunization. An R21 grant is well suited for these studies that require reagent development to address a fundamental question in cholera biology. A new technology, surface plasmon resonance (SPR) will be used to characterize the affinity of the mAbs. The experiments are risky as they require generation of multiple reagents to complete the population analysis of anti-LPS antibodies. The study is high pay off because reagents, regardless of the exact specificity, will be generated that will map the antibody repertoire for Inaba LPS's terminal sugar. We will clarify the "type" of anti-V. cholerae LPS antibodies that are protective and thus substantiate or not the use of Inaba neoconjugates as a vaccine component.