Response To Polysaccharide/Oligosaccharide/Peptide-Prote

对多糖/低聚糖/肽蛋白的反应

• 批准号: 7208898
• 负责人: RACHEL SCHNEERSON
• 金额: --
• 依托单位: EUNICE KENNEDY SHRIVER NATIONAL INSTITUTE OF CHILD HEALTH & HUMAN DEVELOPMENT
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/7208898
• 关键词: Bordetella pertussis; Borrelia; Haemophilus ducreyi; Neisseria meningitidis; Shigella; active immunization; bacterial polysaccharides; bacterial proteins; bacterial vaccines; bactericidal immunity; biotechnology; clinical trial phase III; cross immunity; human subject; human therapy evaluation; immune response; immunoconjugates; laboratory mouse; oligosaccharides; patient oriented research; peptides; preschool child (1-5); synthetic vaccines; vaccine development; vaccine evaluation

SHIGELLAE: Surface polysaccharides of pathogenic bacteria, including capsular polysaccharides (CPS) or the O-specific polysaccharide (O-SP) of lipopolysaccharides (LPS), serve both as essential virulence factors and as protective antigens. Covalent binding of CPS or of O-SP to medically-useful proteins to form conjugates both increases their immunogenicity and confers T-cell dependence to these saccharides making them suitable vaccines for infants and children. The O-SP of Shigella sonnei bound to recombinant non toxic P. aeruginosa exoprotein A (rEPA) and of S. flexneri 2a bound to the succinylated, exoprotein A (rEPA-succ) were safe and induced IgG antibodies to the homologous LPS in 1-4 year-olds. A randomized, blinded, phase 3 study of these conjugates in 1-4 year-olds with each conjugate serving as a control for the other is in progress. CROSS REACTING POLYSACCHARIDES: H. influenzae type b (Hib), Bacillus pumilis: A study of 10 year old children injected in infancy with Hib-TT showed a GM serum level of 4.16 mcg of IgG anti-Hib/mL with no change in this level after a DT booster but with an increase of IgG anti-TT from 0.09 to 4.58 IU/mL. To investigate if concurrent administration of a cross-reacting polysaccharide along with a homologous CPS has an advantage over the use of the homologous CPS alone, the cell wall polysaccharide (PS) of Bacillus pumilis Sh18, cross reactive with Hib CPS, was isolated and its structure investigated using GC-MS, NMR, fast atom bombardment and several sugar degrading techniques. It was shown to be composed of polyribitolphosphate, polyribitolphosphate substituted at C2 with N-acetyl glucosamine and polyglycerolphosphate. In addition to the anti-Hib it reacted with anti-Staphylococcus epidermidis known to contain polyribitol phosphate (poly Rib-P) in its teichoic acid. The Sh18 PS was conjugated to carrier proteins and its immunogenicity evaluated in general purpose mice. Conjugate-induced antibodies reacted with the homologous and with several cross-reacting polysaccharides. Poly (Rib-P) was synthesized to investigate its cross reactivity with poly (Rib-P) containing polysaccharides. NEISSERIA MENINGITIDIS: N. Meningitidis group A (Men. A): causes endemic and epidemic meningitis, notably in the meningitis belt of Africa. A CPS vaccine, effective and available, is underutilized. To improve its immunogenicity, Men. A CPS was conjugated to BSA using ADH as a linker. Contrary to the CPS alone the conjugates were immunogenic in mice, with booster responses after the 2nd and 3rd injections and bactericidal activity related to IgG anti-CPS levels. Conjugates of the cross reactive polysaccharides, E. coli K93 and B. pumilus Sh17, did not induce anti-Men. A CPS. Outbreaks of meningitis due to N. meningitides group W135 were reported recently in West Africa, Saudi Arabia, Europe, and Australia. Similar to other polysaccharides W135 CPS is not immunogenic in young children. We prepared conjugates of this CPS bound to a recombinant Staphylococcus aureus enterotoxin C1 (rSEC) using ADH as a linker. Opposed to the CPS alone the conjugates induced IgG anti-CPS in mice, with booster responses upon reinjection. Neutralizing SEC antibodies were induced as well. Injection of Group A and W135 conjugates combined induced antibody levels comparable to those of each alone. Group B N. meningitidis (GBM) and Escherichia coli k1 continue to cause serious and difficult to treat diseases and there is no licensed vaccine for their prevention. The polysaccharide, alpha (2-8) N-acetylneuraminic acid (PSA), is the capsular polysaccharide (CPS) of GBM and of E. coli K1 and a component of mammalian glycopeptides. Because it is a "self antigen", vaccines designed to induce PSA immune responses have been considered as a potential cause of immunopathology. Antibodies to PSA bind to human tissues in-vitro but have not been shown to induce pathology in-vivo. The incidence, severity, and nature of systemic infection caused by GBM are similar to those caused by the closely related group C meningococci (GCM). As shown for other polysaccharides, there is an age-related acquisition of serum PSA antibodies in humans: most human neonates and adults have IgG anti-PSA. Purified PSA is not immunogenic but as a component of bacteria or conjugated to proteins this CPS induces antibodies of the 3 major isotypes with its IgM and IgG components being protective in in-vitro and in-vivo models. Despite its similarity to mammalian glycoproteins, there are no data indicating an association of IgG anti-PSA with immunopathology in experimental animals or in humans. In collaboration with Mark Miller, FIC, the Lanspitali University Hospital, Iceland, and the Statens Serum Institut, Denmark, we are conducting long term examination of survivors of groups B and C meningococcal meningitis to unearth any unusual diseases considered to be "autoimmune". We propose that clinical trials of PSA conjugate vaccines, shown to be immunogenic and protective in animals, be considered. BORRELIA BURGDORFERI, a spirochete transmitted through the bite of infected Ixodes ticks, is the etiologic agent of Lyme disease. A protein vaccine against it is not effective below the age of 12 years, and has been taken off the market recently. LPS has been described in other spirochetes but its presence in B. burgdorferi has been debated. We have not been able to confirm its presence. The search for LPS revealed 2 unique glycolipids: cholesteryl 6-O-acyl-beta-D-galactopyranoside (BbGL I) and 1,2-di-O-acyl-3-O-alpha-D-galactopyranosyl-sn-glycerol (BbGL II) We found evidence that they are surface exposed. Injected in various formulations into mice BbGL I induced specific antibodies, in order of induced levels: CFA, PBS, DMSO, and squalene. Investigation of the effect of BbGL-I and II upon human PMNs and monocytes showed increased levels of IFN-gamma, IL-4 and TNF-alpha secreted in their presence. BORDETELLAE: B. parapertussis causes pertussis in humans, though of a milder form and lower frequency than B. pertussis. B. bronchiseptica causes respiratory infection in animals, rarely in humans. B. parapertussis and B. bronchiseptica do not produce pertussis toxin. They reportedly share their LPS structure, which is different from that of B. pertussis, but variants in the LPS of B. bronchiseptica were found. The LPS of B. parapertussis and of 2 strains of B. bronchiseptica were purified and their O-SP isolated. Their structural and immunological properties are being studied. HAEMOPHILUS DUCREYI: is the cause of chancroid, a sexually transmitted disease characterized by painful genital ulceration. Importantly, chancroid also enhances the spread of HIV infection. A major virulence factor and a potentially protective antigen of H. ducreyi is its lipooligosaccharide (LOS). The LOS was isolated, its lipid A hydrolyzed and the O-Specific oligosaccharide (O-SP) bound to carrier proteins by two new methods: 1. binding the carbonyl group at the KDO of the reducing end of the O-SP to an aminooxy group of a bifunctional linker, prepared in our laboratory, bound to carrier proteins; 2. using adipic acid dihydrazide to bind to the carbonyl group of the O-SP and to benzaldehyde derivatized carrier proteins. These conjugates preserved the external, nonreducing end of the O-SP and precipitated with H. ducreyi antisera. Their immunogenicity in mice is being investigated.

志贺氏菌：病原菌的表面多糖，包括荚膜多糖 (CPS) 或脂多糖 (LPS) 的 O 特异性多糖 (O-SP)，既可作为必需毒力因子，又可作为保护性抗原。 CPS 或 O-SP 与医学上有用的蛋白质共价结合形成缀合物，既增加了它们的免疫原性，又赋予了 T 细胞对这些糖的依赖性，使其成为适合婴儿和儿童的疫苗。宋内志贺氏菌与重组无毒铜绿假单胞菌外蛋白 A (rEPA) 结合的 O-SP 和与琥珀酰化外蛋白 A (rEPA-succ) 结合的弗氏志贺氏菌 2a 的 O-SP 是安全的，可在 1-4 岁儿童中诱导针对同源 LPS 的 IgG 抗体。对 1-4 岁儿童进行的一项随机、盲法 3 期研究正在进行中，其中每种缀合物作为另一种缀合物的对照。 交叉反应多糖：b 型流感嗜血杆菌 (Hib)、短小芽孢杆菌：一项针对婴儿期注射 Hib-TT 的 10 岁儿童的研究显示，GM 血清水平为 4.16 mcg IgG 抗 Hib/mL，DT 加强后该水平没有变化，但 IgG 抗 TT 从 0.09 IU/mL 增加至 4.58 IU/mL。为了研究同时施用交叉反应多糖和同源 CPS 是否优于单独使用同源 CPS，分离了与 Hib CPS 交叉反应的短小芽孢杆菌 Sh18 的细胞壁多糖 (PS)，并使用 GC-MS、NMR、快原子轰击和几种糖降解技术研究了其结构。它显示出由聚核糖醇磷酸酯、在C2处被N-乙酰葡糖胺取代的聚核糖醇磷酸酯和聚甘油磷酸酯组成。除了抗 Hib 抗体外，它还与抗表皮葡萄球菌发生反应，已知其磷壁酸中含有聚核糖醇磷酸盐 (poly Rib-P)。 Sh18 PS 与载体蛋白缀合，并在通用小鼠中评估其免疫原性。缀合物诱导的抗体与同源物和几种交叉反应的多糖发生反应。合成聚（Rib-P）以研究其与含有多糖的聚（Rib-P）的交叉反应性。 脑膜炎奈瑟菌：A 组脑膜炎奈瑟菌（Men. A）：引起地方性和流行性脑膜炎，特别是在非洲的脑膜炎带。有效且可用的 CPS 疫苗尚未得到充分利用。为提高其免疫原性，Men．使用 ADH 作为连接体将 CPS 与 BSA 缀合。与单独的 CPS 相反，缀合物在小鼠中具有免疫原性，在第 2 次和第 3 次注射后具有增强反应，并且杀菌活性与 IgG 抗 CPS 水平相关。交叉反应性多糖、大肠杆菌 K93 和短小芽孢杆菌 Sh17 的缀合物不会诱导抗 Men。一个 CPS。 最近在西非、沙特阿拉伯、欧洲和澳大利亚报告了由 W135 脑膜炎奈瑟菌群引起的脑膜炎暴发。与其他多糖类似，W135 CPS 对幼儿不具有免疫原性。我们使用 ADH 作为连接子，制备了该 CPS 与重组金黄色葡萄球菌肠毒素 C1 (rSEC) 结合的缀合物。与单独的 CPS 不同，缀合物在小鼠体内诱导 IgG 抗 CPS，并在重新注射后产生增强反应。还诱导了中和 SEC 抗体。注射 A 组和 W135 缀合物组合诱导的抗体水平与单独注射的抗体水平相当。 B 组脑膜炎奈瑟菌 (GBM) 和大肠杆菌 k1 继续引起严重且难以治疗的疾病，并且没有获得许可的疫苗可用于预防。多糖 α (2-8) N-乙酰神经氨酸 (PSA) 是 GBM 和大肠杆菌 K1 的荚膜多糖 (CPS)，也是哺乳动物糖肽的组成部分。由于它是一种“自身抗原”，旨在诱导 PSA 免疫反应的疫苗已被认为是免疫病理学的潜在原因。 PSA 抗体在体外与人体组织结合，但尚未显示出会在体内诱发病理。 GBM 引起的全身感染的发生率、严重程度和性质与密切相关的 C 族脑膜炎球菌 (GCM) 引起的全身感染相似。正如其他多糖所示，人类血清 PSA 抗体的获得与年龄相关：大多数人类新生儿和成人都有 IgG 抗 PSA。纯化的 PSA 不具有免疫原性，但作为细菌的成分或与蛋白质缀合，该 CPS 会诱导 3 种主要同种型的抗体，其 IgM 和 IgG 成分在体外和体内模型中具有保护作用。尽管它与哺乳动物糖蛋白相似，但没有数据表明 IgG 抗 PSA 与实验动物或人类的免疫病理学相关。我们与 Mark Miller、FIC、冰岛 Lanspitali 大学医院和丹麦 Statens 血清研究所合作，对 B 型和 C 型流行性脑膜炎幸存者进行长期检查，以发现任何被认为是“自身免疫”的异常疾病。我们建议考虑对已证明在动物中具有免疫原性和保护性的 PSA 结合疫苗进行临床试验。 伯氏疏螺旋体是一种通过受感染的硬蜱叮咬传播的螺旋体，是莱姆病的病原体。针对该病的蛋白质疫苗对 12 岁以下儿童无效，最近已从市场上撤下。其他螺旋体中也有 LPS 的描述，但伯氏疏螺旋体中是否存在 LPS 仍存在争议。我们无法确认它的存在。 LPS 的搜索揭示了 2 种独特的糖脂：胆固醇 6-O-酰基-β-D-吡喃半乳糖苷 (BbGL I) 和 1,2-二-O-酰基-3-O-α-D-吡喃半乳糖基-sn-甘油 (BbGL II) 我们发现证据表明它们是表面暴露的。以各种制剂注射到小鼠体内，BbGL I 诱导了特异性抗体，按照诱导水平的顺序：CFA、PBS、DMSO 和角鲨烯。 BbGL-I 和 II 对人 PMN 和单核细胞的影响的研究表明，在它们存在的情况下，分泌的 IFN-γ、IL-4 和 TNF-α 水平增加。 博德特氏菌：副百日咳博德特氏菌会引起人类百日咳，但其形式较百日咳博德特氏菌更温和且频率较低。支气管败血杆菌会引起动物呼吸道感染，但很少会引起人类呼吸道感染。副百日咳博德特氏菌和支气管败血博德特氏菌不产生百日咳毒素。据报道，它们具有与百日咳博德特氏菌相同的脂多糖结构，但发现了支气管败血博德特氏菌的脂多糖变异。纯化副百日咳博德特氏菌和2株支气管败血博德特氏菌的LPS并分离其O-SP。它们的结构和免疫学特性正在研究中。 杜克雷嗜血杆菌：是软下疳的病因，软下疳是一种以生殖器疼痛性溃疡为特征的性传播疾病。重要的是，软下疳还会促进艾滋病毒感染的传播。杜克雷嗜血杆菌的主要毒力因子和潜在保护性抗原是其脂寡糖 (LOS)。通过两种新方法分离出LOS，其脂质A被水解，O-特异性寡糖（O-SP）与载体蛋白结合：1.将O-SP还原端KDO处的羰基与我们实验室制备的双功能连接子的氨氧基结合，与载体蛋白结合； 2.使用己二酸二酰肼结合O-SP的羰基和苯甲醛衍生的载体蛋白。这些缀合物保留了 O-SP 的外部非还原端，并用杜克雷螺杆菌抗血清沉淀。正在研究它们在小鼠中的免疫原性。