Target identification of small molecule LDN/OSU-215111

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Glutamate transporter EAAT2 plays a critical role in the homeostatic regulation of extracellular glutamate levels. EAAT2 also plays an essential role in cognitive memory functions. However, loss of EAAT2 protein and function is commonly found in Alzheimer’s Disease (AD) patients. We have discovered and developed a novel series of small molecules that can increase EAAT2 expression via a novel translational activation mechanism. These molecules have been proved to be capable of normalizing glutamate dyshomeostasis and providing significant benefits in two mouse models of AD. This project is currently at the pre-clinical development stage. However, our original lead compound was identified with a phenotypic approach. At this point, we still don't know the compound target. The focus of this study is to identify the target protein of LDN/OSU-215111, which is our pre-clinical candidate. In Aim 1, we will utilize, in tandem, an affinity probe pull-down approach and a protein microarray approach to identify putative compound targets. In Aim 2, We will investigate the identified candidates and determine the target protein that is responsible for LDN/OSU-215111-mediated EAAT2 induction. We will first validate the interaction of LDN/OSU-215111 with the identified proteins in vitro. Once the interaction is confirmed, we will investigate if knockdown of the candidate protein results in loss of LDN/OSU-215111-mediated EAAT2 induction. The goal is to determine the compound target.