The death of neurons that control memory and cognition are responsible for the behavioral symptoms of Alzheimer?s disease (AD). One of the most common pathways for neuronal death in AD is cell cycle re-entry (CCR), which represents the reactivation of neuronal cell cycle machinery. Usually, differentiated neurons never attempt to divide, yet up to 5-10% of the neurons in brain regions affected by AD show signs of CCR. These neurons, which typically have duplicated much of their DNA, fail to undergo cytokinesis. Instead, they eventually die and may account for as much as 90% of the neuronal loss seen in AD. This process initiates with exposure to soluble amyloid-? oligomers (A?Os), which are the building blocks of the insoluble amyloid plaques that accumulate in AD brain. Identifying genes and proteins that mediate A?O-mediated neuronal CCR and defining relevant signaling networks hold promise for early AD diagnosis and for developing new AD therapeutics. We have developed a human neural cell model of A?O-mediated neuronal CCR, which utilizes neural iPS cell lines. We intend to use these iPS lines and derived sublines to develop a high content screening (HCS) assay to identify chemotypes that inhibit A?O-mediated CCR. We believe, through this assay, we can identify small molecules that can block A?O-induced neuronal CCR and function as chemical probes to (1) understand the signaling pathways leading to neuronal death by CCR and (2) serve as parental chemotypes for drugs.
We propose to convert a low throughput assay of A?O-mediated cell cycle reentry to a high content screening assay format. This will enable large-scale screening for small molecule inhibitors of A?O-mediated cell cycle reentry, which is a leading cause of neuronal death in Alzheimer?s Disease. Along with the primary screening assay we are developing, we are proposing to develop several medium throughput secondary confirmation assays that will be used to assist in the prioritization of hit chemotypes. Ultimately, this HCS assay (and supporting secondary confirmation assays) will be used to screen large compound libraries for small molecule inhibitors that will be used as chemical probes and, potentially, as a basis for long term chemotherapeutics for A?O-mediated cell cycle reentry in Alzheimer?s Disease patients.
