HBcAg produced in transgenic tobacco triggers Th1 and Th2 response when intramuscularly delivered.

Hepatitis B core Antigen (HBcAg) assembled into Capsid-Like Particles (CLPs) is investigated as a therapeutic vaccine in treatment of chronic hepatitis B (CHB) and in diagnostic tests or as a carrier for various epitopes. While the expression of HBcAg has been thoroughly clarified in E. coli and yeast, it has also been investigated in other expression systems. Stably transformed tobacco expressed HBcAg at a level of 110-250µg/g fresh weight, therefore in view of its large leaf biomass it offers a production platform comparable with transient expression systems regarding the final yield of HBcAg. Several extraction and purification methods were tested and finally the antigen was purified up to 43% using sucrose density gradient centrifugation. The purified HBcAg retained its antigenicity, as confirmed by ELISA and western blot, while maintaining its CLP-structure as observed in TEM. In mice HBcAg intramuscularly delivered at 2×10µg triggered a significant response (serum anti-HBc titre around 150,000), being statistically equivalent to that induced by the reference antigen. Among anti-HBc IgG isotypes, IgG2a and then IgG1 were increasing during immune response. However IgG2b and IgG3 were also induced, especially in mice immunised with the plant-derived antigen. Analysis of the isotype profile indicates mainly Th1 polarisation, but completed with Th2 response. Obtained results indicate a considerable potential of plant-derived HBcAg as a therapeutic vaccine, since a mixed immune response with a stronger Th1 component is particularly required for treatment of CHB.