Targeting dengue in the mosquito with small molecules

描述（申请人提供）：近年来，登革热已成为最严重的病媒传播病毒性疾病之一。登革热病毒在蚊子载体中传播的分子生物学在很大程度上是未知的，但是基因组序列、反向遗传和功能基因组学工具以及现在可用的高通量筛选方法提供了更详细地剖析蚊子与病毒的相互作用的手段。在这项提案中，我们将采用高度创新的跨学科方法来发现和开发小分子（化合物），可以使埃及伊蚊载体蚊子对病毒产生抵抗力，从而无法传播疾病。Garcia-Blanco博士和Dimopoulos博士之前的合作通过在果蝇细胞系中进行高通量RNAi筛选，确定了300多种潜在的登革热病毒宿主和限制因子。我们现在将扩大这些发现，并确定在特定目标1中的伊蚊中这些因子的同源物。在目标2中，我们将评估伊蚊细胞系和蚊子中选定因子的抗登革热潜力，以选择最适合小分子发现的那些因子。然后，我们将使用强大的已建立的基于酵母的平台来鉴定可以抑制登革热病毒宿主因子的化合物，并使用已建立的基于昆虫细胞系的系统来鉴定可以激活Aim 3中登革热病毒限制因子表达的化合物。在目标4中，我们将通过不同的暴露途径，在伊蚊细胞系和成年雌性蚊子中测试一组选定的这些化合物的抗登革热活性。我们最终将研究所选化合物对蚊子适应性参数的潜在影响，并在目标5中评估它们在野外来源的蚊子品系中的抗登革热活性。这项高度创新的工作的完成将导致过渡到转化研究，将有效的抗登革热化合物递送到天然登革热媒介群体，作为开发新的登革热控制策略的前奏。

DESCRIPTION (provided by applicant): In recent years, dengue fever has emerged as one of the most serious vector-borne viral diseases. The molecular biology of dengue virus propagation in the mosquito vector is largely unknown, but the genome sequence, reverse genetic and functional genomics tools as well as high throughput screening methodologies that are now available provide a means of dissecting the mosquito's interactions with the virus in greater detail. In this proposal, we will apply a highly innovative interdisciplinary approach to discover and develop small molecules (compounds) that can render the Aedes aegypti vector mosquito resistant to the virus and thereby incapable of transmitting disease. A previous collaboration between the groups of Drs. Garcia-Blanco and Dimopoulos identified over 300 potential dengue virus host and restriction factors through a high throughput RNAi screen in a Drosophila cell line. We will now expand on these findings and identify homologues of these factors in the Aedes mosquito in specific Aim 1. In Aim 2 we will assess the anti-dengue potential of selected factors in the Aedes cell line and mosquito to select those that are most suitable for small molecule discovery. We will then use a powerful established yeast-based platform to identify compounds that can repress dengue virus host factors, and an established insect cell line -based system to identify compounds that can activate expression of dengue virus restriction factors in Aim 3. In Aim 4 we will test a selected set of these compounds for anti-dengue activity in Aedes cell lines and adult female mosquitoes through different routes of exposure. We will finally investigate the potential influence of selected compounds on mosquito fitness parameters and assess their anti-dengue activity in field -derived mosquito strains in Aim 5. The completion of this highly innovative work will lead to a transition into translational studis on the delivery of potent anti-dengue compounds to natural dengue vector populations, as a prelude towards the development of a novel dengue control strategy.