MRC/FAPESP - bilateral agreement: Multiple-epitope vaccine to confer serotype-independent protection against pneumonia

肺炎球菌感染占5岁以下儿童死亡总数的11%，而在老年人中，肺炎链球菌是社区获得性肺炎的主要原因。儿童和成人的肺炎球菌感染都有可能通过接种疫苗来预防，但目前的疫苗不能提供所需的保护水平。此外，多糖结合疫苗的使用与非疫苗类型引起的疾病有关，这种现象称为血清型替代。bbbb90不同肺炎球菌类型的存在意味着血清型替代将是抗荚膜疫苗的一个持续问题。基于所有菌株共有的肺炎球菌蛋白的疫苗的开发，提供了不依赖血清型的保护和完全根除携带的潜力。肺炎球菌表面蛋白A （PspA）和肺炎球菌表面蛋白C （PspC）是基于蛋白质的疫苗最有希望的候选物。尽管这些蛋白在不同的临床分离株中是可变的，但我们发现一些变体能够诱导具有广泛反应性的抗体，识别不同的PspA和PspC变体。我们还表明，PspA蛋白片段的融合是一种有效的策略，以延长蛋白质提供的保护，但理想的表位（蛋白质的一部分）包括在疫苗中尚未确定。选择能够诱导具有广泛交叉反应性的抗体的表位（能够识别PspA和PspC的所有变体）对于使用这种方法成功开发疫苗具有临床重要性。我们将使用肽阵列技术来描述已知具有交叉反应性的血清（小鼠免疫血清和人血清）靶向的整个交叉反应性表位范围，并确定疫苗中包含的最佳表位。使用肽阵列的方法已成功地用于鉴定候选疟原虫和钩端螺旋体疫苗的免疫原性表位，但尚未用于肺炎链球菌。我们在LSTM的小组已经成功建立了实验性人类肺炎球菌携带（EHPC）模型，并建立了肺炎患者队列。我们将使用一组独特的肺炎球菌鼻内接种前和后样本来解剖对已知携带事件的PspA和PspC表位的反应。我们将使用肺炎患者的样本来比较健康和易感的抗体功能。在先前的人类载体实验研究中获得的结果表明，血清中抗PspA抗体的存在与无法建立载体相关。我们观察到血清、鼻洗和支气管肺泡灌洗对PspA和PspC的抗体反应增加。免疫球蛋白对暴露、携带和疾病的反应是不同的，并表现出区隔性。目前迫切需要一种对儿童和老年都有效的肺炎疫苗。我们建议一种最佳疫苗将保护两个年龄组免受所有血清型肺炎球菌引起的肺炎。我们将通过鉴定PspA和PspC的表位来生产多表位疫苗，在侵袭性疾病的小鼠模型中测试该疫苗，并使用广泛的肺炎球菌临床分离物小组确认抗体的交叉反应性，从而实现这一目标。此外，我们将利用保护健康成人免受携带和肺炎患者缺陷反应的有效获得性免疫来确定该候选疫苗抗体的有效水平和功能。最后，我们将证实小鼠免疫诱导的细胞反应，以及健康成人和肺炎患者中存在的多表位抗原特异性CD4 T细胞。

Pneumococcal infections account for 11% of all deaths in children under 5 years of age and in older adults, Streptococcus pneumoniae is the leading cause of community-acquired pneumonia. Pneumococcal infections in both children and adults are potentially preventable by vaccination but current vaccines do not offer the level of protection needed. Furthermore, the use of polysaccharide conjugate vaccine is associated with disease caused by non-vaccine types, a phenomenon known as serotype replacement. The existence of >90 different pneumococcal types implies that serotype replacement will be a persistent problem for anti-capsular vaccines. The development of a vaccine based on pneumococcal proteins common to all strains offers the potential for serotype-independent protection and the complete eradication of carriage. Pneumococcal surface proteins A (PspA) and Pneumococcal surface protein C (PspC) among the most promising candidates for a protein-based vaccine. Although these proteins are variable among different clinical isolates, we have found some variants that are able to induce antibodies with broad reactivity, recognizing different PspA and PspC variants. We have also shown that fusion of PspA protein fragments is an efficient strategy to extend protection provided by a protein, but the ideal epitopes (parts of the protein) to be included in the vaccine has yet to be defined. The selection of epitopes capable of inducing antibodies with broad cross-reactivity (able to recognize all variants of PspA and PspC) is of clinical importance to the successful development of a vaccine using this approach. We will use peptide array techniques to describe the entire range of cross-reactive epitopes target by sera that are known to be cross-reactive (murine immunized and human sera) and determine the optimal epitopes to be included in the vaccine. The approach of using peptide arrays has been successful used to identify immunogenic epitopes of candidates for Plasmodium and Leptospira vaccine but not yet for S. pneumoniae. Our group at LSTM has successful established an Experimental Human Pneumococcal Carriage (EHPC) model and has established pneumonia patient cohorts. We will use a unique set of pre- and post-pneumococcal intranasal inoculation samples to dissect responses to PspA and PspC epitopes to a known carriage event. We will use samples from pneumonia patients to compare healthy and susceptible antibody function. Results obtained in a previous experimental human carriage study showed that presence of serum antibodies against PspA correlated with failure to establish carriage. We have observed increased antibody responses to PspA and PspC in serum, nasal wash and bronchoalveolar lavage following carriage. Immunoglobulin reponses to exposure, carriage and disease are different and show compartmentalization. There is an urgent need for a pneumonia vaccine that is effective in childhood and in old age. We propose that an optimal vaccine will protect both age groups against pneumonia caused by all serotypes of pneumococci. We will achieve this by identifying epitopes of PspA and PspC to produce a multiple-epitope vaccine, test this vaccine in murine models of invasive disease and confirm cross-reactivity of antibodies using a wide panel of pneumococcal clinical isolates. Further we will use the effective acquired immunity that protects healthy adults against carriage and the deficient response of pneumonia patients to determine the effective levels and function of antibodies to this vaccine candidate. Finally, we will confirm induction of cellular responses elicited by immunisation in mice and the presence of multiple-epitope antigen specific CD4 T cells in healthy adults and pneumonia patients.