Efficacy of Pam3CSK4 as a cross-species adjuvant for polysaccharide vaccines in humanized mouse and non-human primate models.

Polysaccharide-based vaccines are essential for preventing bacterial infections, but their effectiveness is limited by weak antibody responses and lack of suitable adjuvants. TLR4 agonists enhance polysaccharide-specific antibody responses through B cell-intrinsic TLR4-MyD88 signaling in mice, but this mechanism is not conserved in primates, prompting the search for alternative MyD88-activating agonists. In vitro, the TLR1/2 agonist Pam3CSK4 synergizes with strong BCR crosslinking to enhance activation and antibody secretion by mouse and human B cells. In vivo, Pam3CSK4 in squalene emulsion increases protective pneumococcal polysaccharide-specific antibody responses in both immunocompetent and humanized mice. Although a dual TLR2/7 agonist shows strong in vitro activity, it fails to enhance polysaccharide-specific IgG responses in vivo, consistent with antagonism observed when Pam3CSK4 and TLR7 agonists are combined. In contrast, incorporating Pam3CSK4 into an adjuvant containing a TLR4 agonist, synthetic cord factor, and squalene emulsion further enhances memory B cell generation and protective antibody responses in mice and restores adjuvant activity in non-human primates, supporting Pam3CSK4-based formulations as promising adjuvants for polysaccharide vaccines.