The effect of a protein-lipid complex from human milk against pneumococcal otitis

描述（由申请人提供）：急性中耳炎（AOM）是美国最常见的儿童感染，影响超过60%的婴儿在他们的第一年生活，造成严重的疼痛和短暂的听力损失。如果不及时治疗，长期感染可能导致严重的并发症，包括言语和语言发育迟缓，甚至永久性听力障碍。肺炎链球菌（肺炎球菌）是AOM最常见的分离病原体，也是导致大多数感染后并发症的物种。肺炎球菌荚膜结合疫苗在预防侵袭性疾病方面是有效的，但由于非疫苗血清型的感染率增加，对AOM总发作的影响很小。这一事实，加上全球出现的耐抗生素肺炎球菌菌株，强调迫切需要继续努力开发有效的肺炎球菌疾病预防和治疗药物。以前的工作已经从人乳中鉴定出一种蛋白质-脂质复合物（HAMLET），它使用一种不同于普通抗生素的杀菌机制有效地杀死肺炎链球菌，并且不容易受到细菌耐药性的影响。这种机制是特异性的，在小鼠模型中，使用HAMLET治疗已被证明可以有效地显著减少体内鼻咽定植，这显示了未来用于治疗儿童中耳炎的巨大希望。本研究的目的是进一步了解哈姆雷特诱导肺炎链球菌死亡的机制，并评估哈姆雷特作为治疗药物的效用。假设HAMLET靶向肺炎球菌成分和专门用于细胞自杀的遗传机制，并且可以利用这种活性来预防和治疗肺炎球菌定植和中耳炎。这一提议的具体目的是为了直接检验这些假设。Aim 1中提出的研究将描述哈姆雷特诱导肺炎球菌死亡的机制。首先，我们将通过测试各种含胆碱的肺炎球菌表面组分的制备来阐明哈姆雷特的受体结构，以确定它们抑制哈姆雷特诱导死亡的能力，并在固相和体外结合试验中与哈姆雷特相互作用。其次，筛选标记转座子突变文库，以鉴定离子转运体或其他对HAMLET活性至关重要的肺炎球菌成分。在Aim 2中，拟建的研究将使用青鼠（AOM的金标准动物模型）来评估HAMLET在体内预防和根除鼻咽定植和预防中耳炎的能力。这些研究的结果将确定HAMLET在未来儿童治疗研究中的效用，并且具有很大的潜力，可以开发出更好的预防和治疗肺炎球菌定植和AOM的药物，减少耐药性的风险。

DESCRIPTION (provided by applicant): Acute otitis media (AOM) is the most common childhood infection in the United States, affecting over 60% of infants during their first year of life, causing significant pain and transient hearing loss. If left untreated, a prolonged infection can lead to serious complications, including delays in speech and language development and even permanent hearing impairment. Streptococcus pneumoniae (pneumococcal) is the most commonly isolated etiologic agent of AOM, and is also the species responsible for most post-infectious complications. The pneumococcal capsular conjugate vaccine is effective in preventing invasive disease, but has had little impact on total episodes of AOM due to increased rates of infection with non-vaccine serotypes. This fact, coupled with the global emergence of antibiotic-resistant pneumococcal strains, emphasizes the urgent need for continued efforts in developing effective preventive and therapeutic agents against pneumococcal disease. Previous work has identified a protein-lipid complex from human milk (HAMLET) that effectively kills S. pneumoniae, using a bactericidal mechanism that is separate from common antibiotics and is not susceptible to bacterial resistance development. This mechanism is specific, and treatment with HAMLET has been shown to be effective in significantly reducing in vivo nasopharyngeal colonization in a mouse model, revealing great promise for future use in treating otitis media in children. The objectives of this proposal are to develop a greater understanding of the mechanism of HAMLET-induced death in S. pneumoniae, and to assess the utility of HAMLET as a therapeutic agent. It is hypothesized that HAMLET targets pneumococcal components and genetic machinery specifically used for cell suicide, and that this activity can be exploited to prevent and treat pneumococcal colonization and otitis media. The specific aims of this proposal have been designed to directly test these hypotheses. Studies proposed in Aim 1 will characterize the mechanism of HAMLET-induced death in the pneumococcus. First, the receptor structure for HAMLET will be elucidated by testing various preparations of choline-containing pneumococcal surface components for their ability to inhibit HAMLET-induced death and to interact with HAMLET in solid-phase and in vitro binding assays. Secondly, a signature-tagged transposon mutation library will be screened to identify ion transporters or other pneumococcal components critical for HAMLET's activity. In Aim 2, the proposed studies will assess the ability of HAMLET to prevent and eradicate nasopharyngeal colonization and prevent otitis media in vivo using the chinchilla, the gold standard animal model of AOM. Results from these studies will determine the utility of HAMLET for future treatment studies in children and also have great potential to lead to the development of better preventive and therapeutic agents against pneumococcal colonization and AOM, with less risk for resistance development. PUBLIC HEALTH RELEVANCE: Acute otitis media is the most commonly diagnosed childhood infection in the United States. This proposal addresses the anti-bacterial mechanism and potential therapeutic effect of a protein-lipid complex from human milk against Streptococcus pneumoniae, the bacterial pathogen most highly associated with acute otitis media.