Production of a plant-derived immunogenic protein targeting ApoB100 and CETP: toward a plant-based atherosclerosis vaccine.

In an effort to initiate the development of a plant-based vaccination model against atherosclerosis, a cholera toxin B subunit (CTB)-based chimeric protein was designed to target both ApoB100 and CETP epitopes associated with immunotherapeutic effects in atherosclerosis. Epitopes were fused at the C-terminus of CTB to yield a protein called CTB:p210:CETPe. A synthetic gene coding for CTB:p210:CETPe was successfully transferred to tobacco plants with no phenotypic alterations. Plant-derived CTB:p210:CETPe was expressed and assembled in the pentameric form. This protein retained the target antigenic determinants, as revealed by GM1-ELISA and Western blot analyses. Higher expresser lines reached recombinant protein accumulation levels up to 10 µg/g fresh weight in leaf tissues and these lines carry a single insertion of the transgene as determined by qPCR. Moreover, when subcutaneously administered, the biomass from these CTB:p210:CETPe-producing plants was able to elicit humoral responses in mice against both ApoB100 and CETP epitopes and human serum proteins. These findings evidenced for the first time that atherosclerosis-related epitopes can be expressed in plants retaining immunogenicity, which opens a new path in the molecular farming field for the development of vaccines against atherosclerosis.