Intranasal administration of a DNA vaccine complexed with sugar-functionalized chitosan induces protective immunity...

The emergence of Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2) and its devastating global impact emphasize the urgent need for effective, non-invasive vaccines. While current mRNA and adenoviral vaccines have mitigated severe disease, they face challenges in preventing mucosal transmission and addressing the emergence of new viral variants. This study investigated an intranasal DNA vaccine encoding the SARS-CoV-2 spike protein, delivered using four sugar-functionalized chitosan-based formulations designed to target antigen-presenting cells (APCs). Two formulations were based on lactobionic acid-modified chitosan, including one combined with laminarin (7.5:1 and LAM25), while the remaining formulations incorporated mannose and/or gluconic acid-modified chitosan (C6M9 and AG40), to modulate APC targeting and intracellular DNA release behavior. All four formulations were first evaluated in human monocyte-derived APCs to assess uptake and immunostimulatory activity, including maturation and pro-inflammatory responses. LAM25 and AG40 emerged as the most active formulations and were selected for further immunological assessment. These were subsequently tested in an autologous mixed lymphocyte reaction, where both promoted strong T cell activation with a Th1-skewed profile. Finally, LAM25 and AG40 were assessed in in vivo intranasal immunization studies in mice, inducing robust systemic and mucosal antibody responses, as well as antigen-specific cellular immunity in spleen and lung tissues and a balanced Th1/Th2/Th17 cytokine profile. These findings underscore the immunostimulatory potential of functionalized chitosan as a mucosal adjuvant, highlighting its promise for enhancing vaccine efficacy and guiding the development of next-generation SARS-CoV-2 vaccines.