Abstract Our early-stage ADDP proposal aims to develop a novel genetically engineered bacterial biologic to treat the most common early symptoms of Alzheimer's disease (AD), including cognitive impairment and other neuropsychological symptoms, such as anxiety and depression. This debilitating disease imposes a huge emotional, social and financial burden on society. No effective disease-modifying AD drug exists to dampen the Aβ and tau proteinopathies associated with disease progression. Current FDA-approved cholinergic and glutamatergic neurotherapeutics are very modest at best in rescuing memory in mild cognitive impairment (MCI) and prodromal or early stages of AD cases, and often worsen anxiety, apathy, depression, agitation, and other neurobehavioral symptoms, GI irritations, and even mortality. Recent biological evidence indicates that AD is a neural circuit disorder. The onset and progression of cognitive and behavioral symptoms involve a deficiency in monoamine neurotransmitter signaling networks, including norepinephrine (NE) and dopamine (DA). Thus, we propose that restoring brain DA/NE inputs holds the excellent potential to be an effective approach to alleviating cognitive and behavioral deficits in AD and could even delay disease onset. Currently, oral tablet dosing of L- DOPA/carbidopa 3-4 times/day remains the most effective therapy at restoring brain DA/NE levels in humans. However, this repeated chronic pulsatile delivery causes severe side effects. Thus, our therapeutic hypothesis to address this unmet clinical problem is that systemic delivery of genetically engineered L-DOPA bacterial live- therapeutics (LDBL) will avoid large fluctuations in plasma L-DOPA and provide more consistent delivery of L- DOPA to the brain for restoring DA/NE to stable levels that better relieve AD symptoms without additional side effects. Our proof-of-concept data support that 1) the genetically engineered probiotic E. coli Nissle 1917 strains (EcNL-DOPA) efficiently produce L-DOPA both in vitro and in vivo, 2) oral dosing of EcNL-DOPA readily colonizes the mouse gut, achieves a steady-state plasma L-DOPA level that corresponds to the clinically effective plasma level in humans, and increases L-DOPA and DA/NE levels in the brain of rodents and canines, and 3) EcNL-DOPA treatment leads to improved neurobehavioral outcomes and reduces Aβ levels in AD animal models including canines. The overarching goal of our patent-pending ADDP strategy is to optimize the lead LDBL and test its preclinical efficacy in alleviating the cognitive and behavioral deficits, such as apathy, of early AD. To achieve this goal, we will pursue the following specific aims: (i) Optimize the lead LDBL for animal testing, (ii) Evaluate the chronic pharmacokinetic (PK), and safety profile of the lead LDBLs for preclinical efficacy studies, (iii) Determine in vivo pharmacodynamic (PD) efficacy of two lead LDBLs in transgenic (Tg) AD rodent models, and (iv) Assess the efficacy of the ...