Neuroprotective Engineered Bacteria for Alzheimer's Disease Treatment

ABSTRACT Alzheimer’s disease is the most common neurodegenerative disease, and is characterized by the severe deterioration of cognition and memory due to the production of amyloid-β plaques, and neurofibrillary tau tangles in the brain. Current drugs on the market reduce symptoms of AD, but none cure the disease. There is a need for innovative new therapeutic approaches. Increasing evidence suggests that alterations in the gut microbiota can modulate disease-associated pathology in the brain, such as inflammation and toxic metabolite accumulation. Recent studies show that microbiome imbalances are involved in the early stages of Alzheimer’s disease and increase neuroinflammation. Therefore, by reducing systemic inflammation and gut-derived toxic metabolites, the brain can be protected from neuroinflammation, curtailing the progression of neurodegenerative disease. Akkermansia muciniphila is a mucin-degrading bacterium found in the human gut. It plays a key role in regulating gut barrier integrity, intestinal inflammation, and toxic metabolites. By enhancing intestinal barrier integrity, A. muciniphila reduces the translocation of LPS into circulation, and subsequently decreases pro- inflammatory cytokines. Additionally, A. muciniphila was found to downregulate systemic levels of trimethylamine N-oxide (TMAO), a metabolite associated with neurodegenerative pathology23,24, and that was found elevated in cerebrospinal fluid of patients with mild cognitive impairment and AD. Overall, the data suggest that A. muciniphila offers an innovative multifaceted therapeutic approach to inflammatory and neurodegenerative diseases. As such, we propose to leverage our proprietary isolate collection, to select the most suitable A. muciniphila candidate and enhance its neuroprotective properties by engineering beneficial metabolites into its genome. We have identified three anti-inflammatory metabolites to engineer into A. muciniphila: the short chain fatty acid butyrate, the ketone body beta-hydroxybutyrate, and the bile acid UDCA (converted by the liver in TUDCA), all three of which have been shown to reduce inflammation in the CNS. Therefore, our central hypothesis is that engineering these anti-inflammatory compounds into the well-established probiotic A. muciniphila will create a highly effective, targeted and safe therapeutic for Alzheimer’s disease. To test this hypothesis, we propose three Specific Aims. Specific Aim 1 will validate the efficacy of a cocktail of BHB, butyrate and UDCA in a mouse models of AD. This will act as a proof-of-concept study to test the effects of the metabolites in a model of AD pathology, and justify the following aims. Specific Aim 2 will build a single strain of E. coli Nissle to produce butyrate, UDCA and BHB, to precisely identify the best combination of genes and regulatory elements to achieve the target production rates. Specific Aim 3 will involve the rational selection and engineering of Akkermansia muciniphila strain. This will include first screening Bloom’s library of Akkermansia isolates to identify top candidates for AD therapeutics, and secondly engineering the metabolites into the genome of Akkermansia, and progress to a phase II project and begin pre-clinical characterization.