Oral Delivery of mRNA Vaccine by Plant-Derived Extracellular Vesicle Carriers

Abstract

mRNA-based vaccines were effective in contrasting SARS-CoV-2 infection. However, they presented several limitations of storage and supply chain, and their parenteral administration elicited a limited mucosal IgA immune response. Extracellular vesicles (EVs) have been recognized as a mechanism of cell-to-cell communication well-preserved in all life kingdoms, including plants. Their membrane confers protection from enzyme degradation to encapsulated nucleic acids favoring their transfer between cells. In the present study, EVs derived from the juice of an edible plant (Citrus sinensis) (oEVs) were investigated as carriers of an orally administered mRNA vaccine coding for the S1 protein subunit of SARS-CoV-2 with gastro-resistant oral capsule formulation. The mRNA loaded into oEVs was protected and was stable at room temperature for one year after lyophilization and encapsulation. Rats immunized via gavage administration developed a humoral immune response with the production of specific IgM, IgG, and IgA, which represent the first mucosal barrier in the adaptive immune response. The vaccination also triggered the generation of blocking antibodies and specific lymphocyte activation. In conclusion, the formulation of lyophilized mRNA-containing oEVs represents an efficient delivery strategy for oral vaccines due to their stability at room temperature, optimal mucosal absorption, and the ability to trigger an immune response.

Edible Plant-Derived Extracellular Vesicles for Oral mRNA Vaccine Delivery

“…Using a proprietary technique [ 115 ], we engineered EVs purified from orange juice (Citrus sinensis) (oEVs) with SARS-CoV-2 mRNA coding for Spike S1 subunit (S), Full Spike (FS), and nucleocapsid (N) proteins using cation-based interaction combined with controlled osmotic shock ( Figure 2 ) [ 110 ].…”

“…Protection was due to encapsulation into oEVs, since Triton X-100 permeabilization of oEV lipid membrane abrogated the resistance to RNase. In vitro experiments demonstrated that the oEV-mediated delivery of viral mRNAs to macrophages was followed by translation into N, S1, and FS proteins and lymphocyte activation [ 110 ]. Moreover, oEV incorporation of mRNA conferred resistance at room temperature up to one year after lyophilization [ 110 ].…”

“…The vaccination also triggered a Th1 immune response [ 110 ]. Moreover, the stability of the vaccine formulation was evaluated after one year and showed intact and functional mRNA inside lyophilized oEVs [ 110 ].…”

“…Mucosal vaccines currently undergoing clinical trials are mainly based on protein antigens and live attenuated viruses and are preferentially delivered via viral vectors [ 117 ]. The formulation of lyophilized mRNA-EVs could be an efficient strategy for oral vaccine development due to the fact that they are stable at room temperature, optimally absorbed at the mucosal level, and able to induce an immune response [ 110 , 120 ].…”

The application of plant-exosome-like nanovesicles as improved drug delivery systems for cancer vaccines

“…Plant-derived EVs or PELNVs are alternatives to human-cell-derived EVs for mRNA vaccine delivery. PELNVs are promising for delivering gene-based vaccines since their production is frugal and with a higher yield, they do not cause toxicity, they prevent damage to nucleic acid cargoes from enzymes or stress, and they are modifiable [ 80 , 164 – 166 ]. PELNVs isolated from plants are resistant to the gastrointestinal tract, have stability, and are without immunogenicity [ 167 ].…”