Phosphorylated Hexa-Acyl Disaccharides Augment Host Resistance Against Common Nosocomial Pathogens

Abstract

Objectives: To determine whether synthetic phosphorylated hexa-acyl disaccharides provide antimicrobial protection in clinically relevant models of bacterial infection. Design: Laboratory study. Setting: University laboratory. Subjects: BALB/c, C57BL/10J, and C57BL/10ScNJ mice. Interventions: Mice were treated with lactated Ringer’s (vehicle) solution, monophosphoryl lipid A, or phosphorylated hexa-acyl disaccharides at 48 and 24 hours prior to intraperitoneal Pseudomonas aeruginosa or IV Staphylococcus aureus infection. Leukocyte recruitment, cytokine production, and bacterial clearance were measured 6 hours after P. aeruginosa infection. In the systemic S. aureus infection model, one group of mice was monitored for 14-day survival and another for S. aureus tissue burden at 3 days postinfection. Duration of action for 3-deacyl 6-Acyl phosphorylated hexa-acyl disaccharide was determined at 3, 10, and 14 days using a model of intraperitoneal P. aeruginosa infection. Effect of 3-deacyl 6-Acyl phosphorylated hexa-acyl disaccharide on in vivo leukocyte phagocytosis and respiratory burst was examined. Leukocyte recruitment, cytokine production, and bacterial clearance were measured after P. aeruginosa infection in wild-type and toll-like receptor 4 knockout mice treated with 3-deacyl 6-Acyl phosphorylated hexa-acyl disaccharide or vehicle to assess receptor specificity. Measurements and Main Results: During intraperitoneal P. aeruginosa infection, phosphorylated hexa-acyl disaccharides significantly attenuated infection-induced hypothermia, augmented leukocyte recruitment and bacterial clearance, and decreased cytokine production. At 3 days post S. aureus infection, bacterial burden in lungs, spleen, and kidneys was significantly decreased in mice treated with monophosphoryl lipid A or phosphorylated hexa-acyl disaccharides, which was associated with improved survival. Leukocyte phagocytosis and respiratory burst functions were enhanced after treatment with monophosphoryl lipid A or phosphorylated hexa-acyl disaccharides. A time course study showed that monophosphoryl lipid A– and 3-deacyl 6-Acyl phosphorylated hexa-acyl disaccharide–mediated protection against P. aeruginosa lasts for up to 10 days. Partial loss of augmented innate antimicrobial responses was observed in toll-like receptor 4 knockout mice treated with 3-deacyl 6-Acyl phosphorylated hexa-acyl disaccharide. Conclusions: Phosphorylated hexa-acyl disaccharides significantly augment resistance against clinically relevant Gram-negative and Gram-positive infections via enhanced leukocyte recruitment, phagocytosis, and respiratory burst functions of innate leukocytes. Improved antimicrobial protection persists for up to 10 days and is partially mediated through toll-like receptor 4.

TLR Agonists as Mediators of Trained Immunity: Mechanistic Insight and Immunotherapeutic Potential to Combat Infection

“…PHADs similarly bind TLR4 and activate both MyD88- and TRIF-dependent signaling. Hernandez and colleagues recently showed that treating mice with PHADs confers protection to P. aeruginosa and S. aureus , both of which are of high clinical relevance ( 77 ). They found the survival benefit to be associated with increased bacterial clearance, an effect which was observed up to 10 days after treatment.…”

“…They found the survival benefit to be associated with increased bacterial clearance, an effect which was observed up to 10 days after treatment. Further, treatment with PHADs increased leukocyte recruitment and antimicrobial functions while attenuating systemic and local levels of proinflammatory cytokines ( 77 ).…”

Immune Checkpoints: Novel Therapeutic Targets to Attenuate Sepsis-Induced Immunosuppression

“…Trained immunity refers to a long-term functional reprogramming of innate leukocytes, evoked by a prior exposure to microbial ligands or a non-lethal infectious insult, which leads to an amplified anti-microbial response to a broad range of secondary infections (147). Trained immunity induced by microbial ligands such as Toll-like receptor 4 agonists (TLR4), including Monophosphoryl lipid-A (MPLA) and fungal ligand b-glucan, potently protects against life-threatening pathogens such as Pseudomonas aeruginosa, Staphylococcus aureus, and Mycobacterium tuberculosis (148)(149)(150)(151). TLR4 agonist-induced trained immunity involves metabolic reprogramming of macrophages including mitochondrial biogenesis and augmentation of glycolysis and mitochondrial oxidative phosphorylation (149,152).…”

MyD88-Dependent Signaling Drives Toll-Like Receptor-Induced Trained Immunity in Macrophages

“…Investigation of trained immunity has been an active field of research with major discoveries advancing the field towards therapeutic translation for several applications; however, much remains to be explored before the true immunomodulatory potential of training agents can come to fruition. Toll-like receptor (TLR)-4 agonists have been shown to trigger host resistance to infection that lasts for up to two weeks in several clinically relevant models of infection ( 10, 11, 15 ), which our group has revealed to be driven by long-term changes in innate immune function supported by metabolic rewiring ( 13 ). We sought to define which signaling pathway(s) mediate immune training given that TLR4 uniquely signals through both MyD88- and TRIF-dependent signaling cascades.…”