Characterizing trained innate immunity mediated by AMMO - Project Summary / Abstract Two million Hospital Acquired Infections (HAIs) occur per year in the US, killing tens of thousands of patients and costing ~$100 billion. Preventing HAIs is a top NIH/DHHS priority, and altering immunity is a novel strategy to achieve this goal. However, the breadth of HAI pathogen coverage required is daunting to accomplish via protein-based, pathogen-specific vaccines. A more innovative approach may be required. Our initial discovery was that a protein-free mixture of aluminum hydroxide [AlOH3] + monophosphoryl lipid [MPL] + whole glucan particles [WGP] protected mice against lethal bacteremia caused by methicillinresistant S. aureus (MRSA) or the Gram negative pathogen, Acinetobacter baumannii. Dual regimens were not protective. Replacement of WGP with fungal mannan enhanced protection against Gram positive (S. aureus) and negative pathogens (A. baumannii, K. pneumoniae, P. aeruginosa), and the mold mucormycosis. Thus, our lead regimen consists of Al(OH)3 + MPL + mannan. Efficacy lasts for up to 21 days at a 3x dose, which is adequate to prevent infections for >95% of hospital admissions (avg duration 5 days). The regimen does not induce antibodies, and T and B cells play no role in protection. In contrast, macrophages are required for protection, and NK cells play a role in protection against S. aureus but not A. baumannii. Finally, DNA acetylation and RNA transcriptional changes occur in macrophages consistent with Trained Immunity. This innate immune-modulatory mixture has potential to prevent HAIs caused by the highest priority, antibiotic-resistant, pathogens. We seek to optimize efficacy, identify immune correlates of protection, and define mechanisms of protection to facilitate clinical development and deployment. AIM 1: Optimize dosing for maximal efficacy and duration against multiple pathogens causing bloodstream infections and pneumonia in mice. We will determine if efficacy can be further improved, and durability prolonged, by altering component doses against a clinical blood isolate of MRSA (LAC), a carbapenem-resistant clinical isolate of A. baumannii (HUMC1), or a clinical isolate of Rhizopus (99-880). AIM 2: Define immune correlates of protection against multiple pathogens in wild type mice. We will establish immune correlates of protection, which will inform mechanistic disruption experiments in Aim 3 and identify biomarkers for study in patients in future clinical trials. Biomarkers will be measured in mice immunized with more or less effective regimens and then infected with each pathogen. AIM 3: Determine receptor/effector mechanisms of protection in specific, immune-compromised mouse models. We will determine efficacy in mice depleted of specific receptors or cytokines, including Nalp3 (Al(OH)3), TLR4 (MPL), mannose receptors, and other biomarkers identified as immune correlates in Aim 2. We will also test efficacy in mice that are diabetic or elderly, or treated with corticosteroids. IMPACT: These results will lay the groundwork for future clinical development and deployment.
