Determining unique gene control mechanisms and expression patterns in children wi

描述（由申请人提供）：在疟疾流行的环境中，需要对昏迷儿童采取预防死亡率的干预措施。这些干预措施要求将具有不同疾病状态的患者仔细地划分为精确的亚组，以便提供影响特定死亡率的治疗方法。新的干预措施可能来自对寄生虫体内生理学（由遗传学和表达谱定义）的深入了解，这不仅在这些儿童中是独特的，而且在体外也不可复制。我们的实地研究率先使用眼科检查在临床脑疟疾患者中识别疟疾视网膜病变——这些发现清楚地将患者分为“真正的”脑疟疾（视网膜病变阳性）和其他原因的昏迷（视网膜病变阴性）。此外，我们的数据库和组织库中有超过100例来自脑疟疾前瞻性临床病理相关性研究的儿童尸检，是目前最全面的。脑型疟疾患者体内恶性疟原虫的生物学特性包括毒力变异基因的表达以及在体外从未表达过的一组800个基因。利用一组严格定义的昏迷患者（包括脑型疟疾患者和其他原因昏迷的寄生虫患者）的档案和前瞻性收集，我们将有机会彻底描述恶性疟原虫变异基因和独特体内基因的体内生物学特征，并利用综合系统生物学方法了解它们在人类疾病中的控制和功能。我们将根据我们的分子基因分型工具，对符合上述定义并具有低复杂性感染的患者（包括临床病例和尸检组织）的寄生虫进行全基因组测序。我们将利用我们的新型纳米链方法（通过测量来自微小RNA样本的330个标记基因，就可以实现基因表达的完全估算）将这些全基因组信息与来自同一种寄生虫的表达数据结合起来，并通过RNA表达和数字基因表达进行确认。这两个强大的数据集的整合将使我们能够定义基因组内的表达数量性状位点，既指导整体体内生物学，又明确阐明仅在人类感染期间发现的独特基因的控制和功能。在本研究结束时，我们将定义人类感染过程中独特的体内表达基因网络，表征var基因在个体患者中的特异性表达，并确定其中哪些是干预策略的假定目标。

DESCRIPTION (provided by applicant): Interventions that thwart mortality are needed for comatose children in malaria endemic settings. These interventions require that patients who have variable disease states be carefully stratified into an exact subgroup in order to provide therapies that impact specific mortality. New interventions may come from an in depth understanding of the in vivo physiology (define by genetics and expression profiling) of the parasite which is not only unique in these children but also not reproducible in vitro. Our field study has pioneered the use of eye exams to identify malaria retinopathy in the setting of patients with clinical cerebral malaria - these findings cleanly separate patients into "true" cerebral malaria (retinopathy positive) and coma of other causes (retinopathy negative). Moreover, our database and tissue repository of over 100 pediatric autopsies from a prospective clinicopathological correlation study of cerebral malaria is the most comprehensive available. The biology of Plasmodium falciparum in vivo in cerebral malaria includes the expression of the virulent var genes as well as a set of 800 genes which are never expressed in vitro. Using an archive and prospective collection from a stringently defined cohort of comatose patients including both cerebral malaria patients and parasitemic patients with other causes of coma, we will have the opportunity to thoroughly characterize the in vivo biology of both var and unique in vivo genes of Plasmodium falciparum and understand their control and function within human disease using a comprehensive systems biology approach. We will perform full genome sequencing of parasites from patients (including clinical cases and autopsy tissues) who meet the above definitions and have low complexity infections based on our molecular genotyping tools. We will combine this full genome information with expression data from the same parasites using our novel Nanostring approach (which allows for full imputation of gene expression by measuring only 330 marker genes from minute samples of RNA) with confirmation by RNA expression and digital gene expression. The integration of these two robust data sets will allow us to define expression quantitative trait loci within the genome both directing the overall in vivo biology and specifically elucidating control and function within the unique genes found only during human infection. At the conclusion of this study, we will have defined the network of unique in vivo expressed genes during human infection, characterized the specific expression of var genes within individual patients, and identified which of these are putative targets for intervention strategies.