Project Summary/Abstract Significant effort has been put toward developing therapies targeted at amyloid-β peptide (Aβ), the hallmark toxic aggregatory protein associated with AD. Unfortunately, Aβ targeted therapies have resulted in several costly Phase III clinical failures. This research focuses on developing novel cognition enhancing agents which do not directly target Aβ but may reverse the effects of Aβ on cognitive function and provide neurorestorative effects by up-regulating neurogenic gene products. Nitric oxide (NO) mimetics activate an intracellular 2nd messenger known as soluble guanylyl cyclase (sGC) leading to increased cyclic GMP (cGMP) production and increased phosphorylation/activation of cAMP response element binding protein- CREB (pCREB). CREB phosphorylation is recognized as being a crucial regulator of synaptic plasticity, resulting in the production of pro-growth gene products, such as brain derived neurotrophic factor (BDNF), and enhanced synaptic transmission. NO/cGMP/CREB signaling is disrupted in AD via Aβ-mediated inhibition of NO-induced CREB phosphorylation and synaptic plasticity. Reversal of Aβ induced memory impairment via agents which activate NO/cGMP signaling results in improved cognitive function. Hence, NO/cGMP activating agents show potential for the treatment of AD. Furoxans are a class of thiol-dependent NO mimetics which may hold potential as novel neurorestorative therapies. Furoxans are distinct because they exhibit `tunable' NO mimetic effects. A unique molecular structure distinguishes furoxans from classical NO mimetic nitrates and makes it possible to engineer molecules with significantly reduced rates of NO mimetic activity. HPLC-MS/MS analysis reveals that furoxan reactivity can be manipulated in a predictable manner to avoid the adverse systemic hypotensive side-effects associated with transient fluxes of NO. Preliminary studies indicate furoxans have good brain penetration, neuroprotective activity, and cognition enhancing effects via NO/cGMP/CREB signaling. This project represents a hit-to-lead optimization campaign for the development of furoxans as novel agents for AD. Our approach includes- 1) synthesis of novel analogs and preliminary screening in PC12 cells for protection against oxidative stress; 2) Counter screening active analogs in a focused in vitro pharmacokinetic battery; 3) validating efficacy to improve synaptic function (ex vivo LTP experiments) and protect primary cortical neurons from Aβ induced toxicity; 4) a focused PK/PD study to define a relationship between orally administered furoxan, unbound furoxan in the hippocampus, and engagement of NO/cGMP signaling. A brief dose escalation study will confirm that furoxans do not affect systemic blood pressure or possess acute toxicity prior to conducting a pilot in vivo efficacy in 3xTg transgenic AD mice. Primary outcomes focus on the ability to improve spatial working and contextual fear memory in 3xTg mice.