Rationally designed TLR4 ligands for vaccine adjuvant discovery.

Abstract

Abstract Adjuvant properties of bacterial cell wall components like MPLA are well described, and have gained FDA approval for use in vaccines such as Cervarix. MPLA is the product of chemically modified lipooligosaccharide (LOS), altered to diminish toxic proinflammatory effects while retaining adequate immunogenicity. Despite the virtually unlimited number of potential source bacterial strains, useable compounds within this promising class of adjuvants remain unfortunately low. We have developed bacterial enzymatic combinatorial chemistry (BECC) as a method to generate functionally diverse and rationally designed lipid A. BECC removes endogenous, or introduces exogenous, lipid A-modifying enzymes to bacteria, effectively reprogramming the lipid A biosynthetic pathway. In this study, BECC is applied within an avirulent strain of Yersinia pestis (Yp), to develop structurally distinct LOS molecules that elicit differential TLR4 activation. Using reporter cell lines that measure innate immune activated NF-κB transcription, BECC-derived molecules were screened for the ability to induce a lower pro-inflammatory response than E. coli LOS. Structures exhibit varied, dose-dependent TLR4-driven NF-κB activation with both human- and mouse-TLR4 complexes. Additional cytokine secretion screening identified molecules that induce comparable levels of TNF-α and IL-8, as compared to PHAD. Lead candidates demonstrated potent immunostimulation in mouse splenocytes, human primary PBMCs, and human monocyte-derived DC. This newly described BECC system allows for directed programming of lipid A synthesis and has the potential to generate a diverse array of TLR4 agonist candidates comparable if not preferable to PHAD and MPLA.