Molecular Characterization of C. neoformans Antibodies

描述（由申请方提供）：新型隐球菌是免疫力受损个体的主要真菌病原体，包括晚期HIV感染、器官移植和免疫抑制方案患者。此外，越来越多的证据表明，这种真菌可以在人类中建立潜伏感染，这可能对其他慢性疾病的发展产生深远的影响，包括可能的哮喘。C.新型原虫具有几种特征明确的毒力因子，其中多糖荚膜被认为是最重要的。胶囊由至少三种成分组成，称为葡糖醛酸甘露聚糖（GXM），半乳糖甘露聚糖（GalXM）和称为甘露糖蛋白的高度甘露糖基化蛋白质。对荚膜GXM的抗体应答引发保护性和非保护性抗体。由于隐球菌感染的严重性，人们对利用体液免疫治疗和预防疾病产生了极大的兴趣。单克隆抗体（mAb）已完成初步临床研究，并继续开发。用与蛋白质载体缀合的GXM免疫产生保护性抗体。抗体作用的机制是多因素的，包括调理作用、炎症反应的调节和GXM从酵母细胞释放的消除。值得注意的是，保护性和非保护性mAb可以通过它们在体外影响GXM释放和阻断生物膜形成的能力来区分。虽然现在对抗体在体内和体外的作用机制已经有了很多了解，但保护性和非保护性mAb识别的GXM表位的分子性质仍然未知。此外，抗体在酵母细胞水平消除多糖释放的作用机制尚不清楚。本申请建议继续研究抗体与C.新人类与之前的资助周期相比，当努力集中在抗体分子的分子分析上时，该提案将研究计划重新集中在多糖抗原和胶囊上。除了继续研究针对GXM的抗体之外，本申请还提出探索GalXM在胶囊结构中的作用。提出了三个具体目的：1）确定结合保护性和非保护性mAb的多糖分子结构; 2）确定保护性mAb阻止C. 3）确定GalXM对C.新生儿荚膜结构组织和评估抗体的这种多糖的效力。

DESCRIPTION (provided by the applicant): Cryptococcus neoformans is a major fungal pathogen for individuals with impaired immunity, including those with advanced HIV infection, organ transplants, and on immunosuppressive regimens. Furthermore, there is increasing evidence that this fungus can establish latent infection in humans that could have profound consequences for the development of other chronic diseases, including possibly asthma. C. neoformans has several well-characterized virulence factors, among which a polysaccharide capsule is considered to be the most important. The capsule is composed of at least three components known as Glucuronoxylomannan (GXM), galactoxylomannan (GalXM) and highly mannosylated proteins known as mannoproteins. Antibody responses to the capsular GXM elicit protective and non-protective antibodies. Given the seriousness of cryptococcal infections there has been great interest in harnessing humoral immunity for therapy and prevention of disease. A monoclonal antibody (mAb) has completed preliminary clinical studies and continues in development. Immunization with GXM conjugated to protein carriers elicits protective antibodies. The mechanism of antibody action is multifactorial and includes opsonization, modulation of the inflammatory response, and abrogation of GXM release from yeast cells. Remarkably, protective and non- protective mAbs can be distinguished by their ability to affect GXM release and block biofilm formation in vitro. Although much is now known about the mechanism of antibody action in vivo, and in vitro, the molecular nature of the GXM epitopes recognized by protective and non-protective mAbs is unknown. Furthermore, the mechanisms of action of antibodies at the level of the yeast cell that abrogate polysaccharide release are not understood. This application proposes to continue the study of the interaction of antibodies with the capsule of C. neoformans. In contrast to the prior funding cycles when the effort was focused on molecular analysis of the antibody molecule, this proposal refocuses the research program on the polysaccharide antigen and the capsule. In addition to continuing to study antibodies to GXM this application proposes to explore the role of GalXM in capsule structure. Three specific Aims are proposed: 1) To define the polysaccharide molecular structure(s) that bind protective and non-protective mAbs; 2) To determine the mechanism by which protective mAbs prevent GXM release from C. neoformans; 3) To determine how GalXM contributes to C. neoformans capsular structure organization and evaluate the efficacy of antibodies to this polysaccharide.