New Glucosamine-Based TLR4 Agonists: Design, Synthesis, Mechanism of Action, and In Vivo Activity as Vaccine Adjuvants

We disclose here a panel of small-molecule TLR4 agonists (the FP20 series) whose structure is derived from previously developed TLR4 ligands (FP18 series). The new molecules have increased chemical stability and a shorter, more efficient, and scalable synthesis. The FP20 series showed selective activity as TLR4 agonists with a potency similar to FP18. Interestingly, despite the chemical similarity with the FP18 series, FP20 showed a different mechanism of action and immunofluorescence microscopy showed no NF-kappa B nor p-IRF-3 nuclear translocation but rather MAPK and NLRP3-dependent inflammasome activation. The computational studies related a 3D shape of FP20 series with agonist binding properties inside the MD-2 pocket. FP20 displayed a CMC value lower than 5 mu M in water, and small unilamellar vesicle (SUV) formation was observed in the biological activity concentration range. FP20 showed no toxicity in mouse vaccination experiments with OVA antigen and induced IgG production, thus indicating a promising adjuvant activity. [GRAPHICS]

Automated summary

In this study, a panel of small-molecule TLR4 agonists (the FP20 series) was developed based on previously developed TLR4 ligands (the FP18 series). The FP20 series demonstrated improved chemical stability and a more efficient synthesis method. The molecules showed selective TLR4 agonist activity similar to FP18, but with a different mechanism of action involving MAPK and NLRP3-dependent inflammasome activation. Computational studies suggested that the 3D shape of the FP20 series contributes to their agonist binding properties within the MD-2 pocket. FP20 exhibited low toxicity and promising adjuvant activity in mouse vaccination experiments.