Immune Response to Cryptococcal Infections

 描述（由申请人提供）：隐球菌性脑膜炎，由于包囊酵母C。据估计，新生儿（Cn）每年造成60多万人死亡。大多数受害者的CD 4 + T细胞功能受损。近缘种C. gattii（Cg）在通常具有轻度或无免疫功能低下的人中引起地方病。开发一种预防隐球菌病的疫苗是全球卫生的优先事项。该项目的目标是候选疫苗的临床前开发，以保护隐球菌病的风险人群。我们假设这样的疫苗将具有亚单位制剂，并且由隐球菌抗原（Ag）和佐剂一起递送组成，所述佐剂刺激强烈且持久的T细胞和抗体（Ab）应答。目的1合成新型隐球菌疫苗候选抗原Ags并检测其免疫反应性。在初步研究中，我们已经获得了100%的保护，对致命的隐球菌病接种小鼠与葡聚糖颗粒（GP）含有碱性提取物来自Cn。保护性提取物的蛋白质组已被定义;然而，尚不知道哪些Ag负责保护。在目标1A中，将重组表达提取物中的Ag，并使用来自接种疫苗的小鼠和隐球菌病患者的细胞测试其刺激离体CD 4偏向性T细胞应答的能力。认识到疫苗介导的保护可涉及在自然感染中不具有主要作用的免疫系统的臂。在目标1B中，Cn的主要荚膜组分GXM将与白喉类毒素缀合。据预测，这将聚糖转化为能够引发保护性Ab应答的有效免疫原。目的2是制定候选隐球菌疫苗和进行临床前试验。到目标1结束时，我们预期我们将已经鉴定出刺激T细胞应答的~10个Cn Ag，并且我们将已经制备出刺激针对GXM的Ab应答的缀合物疫苗。在目标2A中，我们将确定这些候选抗原中的哪一种在隐球菌病模型中刺激保护性反应。在目标2B中，我们将优化Ag组合和疫苗配方，包括单独和组合测试GP和CpG。将探讨保护的持久性。在目标2C中，我们将探索疫苗诱导保护的免疫机制。在目标2D中，认识到艾滋病是隐球菌病的主要风险因素，将定义CD 4耗竭对CD 8+和Ab疫苗应答的影响。最后，在目标2 E中，我们将测试疫苗介导的保护是否可以针对Cg感染实现。我们预计，在资助期结束时，我们将创造出能够引发强大和持久的Ag特异性Ab和Th 1偏向反应的候选疫苗，这些反应可以保护小鼠免受Cn和Cg的攻击。这些研究解决了NIH确定的开发隐球菌疫苗的需求，并应建立适用于其他疫苗可预防疾病的原则证据，特别是那些T细胞防御至关重要的疾病。

 DESCRIPTION (provided by applicant): Cryptococcal meningitis, due to the encapsulated yeast C. neoformans (Cn), is estimated to kill over 600,000 people annually. Most victims have compromised CD4+ T cell function. A closely related species, C. gattii (Cg), causes endemic disease in persons who generally have mild or no immunocompromise. Development of a vaccine to protect against cryptococcosis is a global health priority. The goal of this project is the preclinical development of candidate vaccines to protect at risk populations from cryptococcosis. We hypothesize such a vaccine will have a subunit formulation and consist of cryptococcal antigens (Ags) delivered with adjuvant(s) that stimulate strong and durable T cell and antibody (Ab) responses. Aim 1 is to synthesize candidate cryptococcal vaccine Ags and test the Ags for immunoreactivity. In preliminary studies, we have obtained 100% protection against lethal cryptococcosis by vaccinating mice with glucan particles (GPs) containing alkaline extracts derived from Cn. The proteome of the protective extract has been defined; however, it is not known which Ags are responsible for protection. In Aim 1A, Ags in the extracts will be recombinantly expressed and tested for their capacity to stimulate ex vivo CD4-biased T cell responses using cells from vaccinated mice and cryptococcosis patients. It is recognized that vaccine-mediated protection can involve arm(s) of the immune system that do not have a predominant role in natural infection. In Aim 1B, GXM, the major capsular component of Cn, will be conjugated to diphtheria toxoid. This is predicted to convert the glycan into a potent immunogen capable of eliciting protective Ab responses. Aim 2 is to formulate candidate cryptococcal vaccines and perform preclinical testing. By the end of Aim 1, we anticipate we will have identified ~10 Cn Ags that stimulate T cell responses and we will have made conjugate vaccines that stimulate Ab responses against GXM. In Aim 2A, we will determine which of these candidate Ags stimulate protective responses in models of cryptococcosis. In Aim 2B, we will optimize Ag combinations and vaccine formulations, including testing GPs and CpG alone and in combination. The durability of protection will be explored. In Aim 2C, we will explore the immune mechanisms of vaccine-induced protection. In Aim 2D, recognizing that AIDS is the major risk factor for cryptococcosis, the effect of CD4-depletion on CD8+ and Ab vaccine responses will be defined. Finally, in Aim 2E, we will test whether vaccine-mediated protection can be achieved against Cg infections. We anticipate that at the end of the funding period, we will have created candidate vaccines capable of eliciting robust and durable Ag-specific Ab and Th1-biased responses that protect mice against challenge with Cn and Cg. The studies address an NIH-identified need for development of cryptococcal vaccines and should establish proofs of principle applicable to other vaccine-preventable diseases, particularly those for which T cell defenses are paramount.