Self-assembled ferritin tuberculosis nanovaccines targeting ESAT-6 and CFP-10 elicit potent immunogenicity in mice

Tuberculosis (TB) is a chronic and persistent infectious disease caused by Mycobacterium tuberculosis (Mtb), posing a serious threat to global public health. The only widely available vaccine for TB is Bacillus Calmette-Guérin (BCG), and its limitations have become increasingly apparent in recent years, underscoring the urgent need for new TB vaccines. Moreover, subunit vaccines undergoing clinical trials face challenges such as complex production processes and weak immunogenicity. Efforts must be made to develop a safe, effective, easily mass-produced TB vaccine. Here, early secreted antigenic target of 6 kDa (ESAT6), culture filtrate protein of 10 kDa (CFP10) and ESAT6-CFP10 (EC) fusion epitopes from the Mtb region of difference (RD) were selected to construct traditional subunit antigens and self-assembling ferritin-based nanovaccines, EF, CF, and ECF. The transmission electron microscopy and dynamic light scattering showed that the three antigens coupled with ferritin self-assembled into uniform nanoparticles with even sizes. H&E staining demonstrated that the nanovaccines were safe. EF, CF, and ECF induced higher levels of cell proliferation than BCG and conventional subunit vaccines in immunized mice exposed to antigenic stimulation. TB nanovaccines enhanced the IFN-γ recall response and strongly stimulated the secretion of various TB-related cytokines, particularly Th1-type cytokines. Multi-color flow cytometry revealed that the nanovaccines, especially ECF, induced immune memory involving IFN-γ⁺T_CM/T_EM, CD4⁺IL-2⁺T_EM and CD8⁺IL-2⁺T_CM cells. Ferritin-NPs hold potential as a TB vaccine carrier and compared to BCG and traditional subunit vaccines, Ferritin-based TB nanoparticles produced by prokaryotic expression display superior immunogenicity.