Alzheimer's disease is predicted to be one of the highest priority global health risks in coming decades, yet there are currently no effective prophylactic or disease modifying therapies. The primary focus in the field has been to target reduction of the synaptotoxic amyloid ?-peptide (A?) however, recent late-stage clinical trials have proven disappointing. Therefore, new strategies and cellular targets are required. Recent work has found that genetic alteration of lipid modifying enzymes can ameliorate behavioral and synaptic defects in mouse models of the disease. Phospholipases, lipid kinases, lipid activated protein kinases and a lipid phosphatase have all effectively been targeted for ameliorating behavioral deficits in mouse models of the disease indicating that phospholipid modulation may be a key factor of the disease phenotype. A specific class of phospholipids, phosphoinositides (PI), known to be critical for cell and neuronal signaling, has been shown historically and in recent work to be altered in AD affected patient brain as well as in mouse models. The activities of enzymes that control phosphoinositide metabolism and phosphorylation are extensively interconnected and represent a family of tractable lipid modifying enzymes which control levels of signaling lipids. Specifically, the critical signaling lipid phosphatidylinositol-4,5-bisphosphae [P(4,5)P2], is altered by treatment of neurons with A? and in synapses of mouse models of the disease. In order to study lipid changes in response to A?, a physiologically relevant neuronal model is required. Mouse embryonic stem cell derived neurons (ESN) are a scalable neuronal model amenable to mid to high throughput platforms and can be used effectively for screening paradigms. Use of ESN overcomes the limitations of other transformed cell lines commonly used for screening by expression of critical neuronal phenotypes as well as the limitation of dissociated neuronal cultures by scalability. ESN form morphologically mature and functional synapses which are sensitive to A? challenge resulting in a loss of synaptic connections. These neurons display a prominent loss of post-synaptic protein 95 (PDS-95) after treatment with A?. We propose to establish and miniaturize an assay to detect synapse/PSD-95 loss and use RNA interference to screen candidate components of the PI metabolic pathway for synapse maintenance. Since previous research has identified several PI modifying enzymes in connection with AD phenotypes, we expect that this system-wide approach will be able to identify both known and novel targets for amelioration of A?-triggered synaptic loss as well as validate the high content screening platform. These studies will support future investigations into the role of PI in cellular pathways underlying A?-triggered deficits as well as pave the way for more extensive screening paradigms using ESN.
Alzheimer's disease is becoming one of the most pervasive health risks in the aging population, yet currently there are no prophylactic or disease modifying therapies and several recent disappointments in late-stage clinical trials have highlighted the need for new disease relevant therapeutic targets. Lipid modifying enzymes are currently being investigated for their ameliorative effects for both in vitro and in vivo models of AD and represent a network of potential tractable therapeutic targets. The use of mouse embryonic stem cell derived neurons for early stage pre-clinical investigations, including relevant screening platforms, augments the likelihood of identification of targets which will withstand validation and testing throughout the drug discovery process.
