Optimizing mucosal vaccination: Exploiting Lactobionic acid-modified chitosan for superior gene delivery systems

The increasing prevalence of respiratory disorders highlights the urgent need for effective mucosal vaccines that elicit targeted immune responses at pathogen entry sites. However, the advancement of mucosal vaccines is limited by challenges in antigen delivery and overcoming mucosal immune tolerance. In this study, we developed a gene delivery platform using chitosan functionalized with lactobionic acid (LA) to enhance targeting of antigen-presenting cells and to form stable DNA polyplexes with high transfection efficiency. ATR-FTIR, NMR, and ninhydrin assays confirmed successful functionalization and structural changes in chitosan after LA incorporation. The mixture of the modified polymer with luciferase plasmid at a polymer-to-DNA ratio of 7.5:1, along with the addition of adjuvants including laminarin (LAM), yielded polyplexes that significantly enhanced transfection efficiency in A549 cells. Moreover, the polyplexes induced ROS production via ASGPR-mediated pathways in HepG2 cells, confirming receptor-specific uptake. The 7.5:1 LAM25 formulation demonstrated the strongest immunostimulatory effect in human dendritic cells, significantly increasing the expression of maturation markers and the secretion of the pro-inflammatory cytokines TNF-α and IL-12, likely through dual interactions with macrophage galactose-type lectin (MGL) and dectin-1 receptors. These findings highlight LA-modified chitosan as a versatile and promising platform for mucosal vaccine development. • Lactobionic Acid-modified chitosan boosts DNA delivery and APC targeting. • Polyplexes show potential for Th1-like responses, relevant for respiratory immunity. • Formulations trigger ROS via ASGPR/clathrin, enhancing immune responses. • Dual receptor binding augments APC activation and cytokine secretion. • An affordable, adaptable platform for respiratory vaccine design.