Virus-like particle expression and assembly in plants: hepatitis B and Norwalk viruses

• Published in: Vaccine Vol. 23; no. 15; pp. 1851 - 1858
• Main Authors: Huang, Zhong, Elkin, Galina, Maloney, Bryan J., Beuhner, Norene, Arntzen, Charles J., Thanavala, Yasmin, Mason, Hugh S.
• Format: Journal Article Conference Proceeding
• Language: English
• Published: Oxford Elsevier Ltd 07.03.2005; Elsevier; Elsevier Limited
• Subjects: Administration, Oral; Amino acids; Animals; Antigens; Antigens - biosynthesis; Antigens - immunology; Antigens - isolation & purification; Applied microbiology; Biological and medical sciences; Blotting, Western; Centrifugation, Density Gradient; Crop yield; Enzymes; Female; Freeze drying; Fruits; Fundamental and applied biological sciences. Psychology; Glycine max; HBsAg; Hepatitis; Hepatitis B - immunology; Hepatitis B Surface Antigens - genetics; Hepatitis B Surface Antigens - immunology; Human viral diseases; Immunogenicity; Infectious diseases; Ingestion; Insects; Leaves; Lycopersicon esculentum; Medical sciences; Mice; Mice, Inbred BALB C; Microbiology; Miscellaneous; Nicotiana - immunology; Nicotiana - metabolism; Norwalk virus; Norwalk virus - immunology; Oral vaccine; Plant expression; Plant Extracts - chemistry; Plant Leaves - chemistry; Plants, Genetically Modified - immunology; Plasmids; Plasmids - genetics; Proteins; Rhizobium - immunology; Solanum lycopersicum - genetics; Solanum lycopersicum - immunology; Solanum tuberosum; Soybeans; Tobacco; Tomatoes; Transgenic plants; Vaccines; Vaccines, antisera, therapeutical immunoglobulins and monoclonal antibodies (general aspects); Vaccines, Synthetic - administration & dosage; Vaccines, Synthetic - biosynthesis; Vaccines, Synthetic - immunology; Viral diseases; Viral hepatitis; Viral Vaccines - administration & dosage; Viral Vaccines - biosynthesis; Viral Vaccines - immunology; Virology; VLP; Plant expression; HBsAg; Oral vaccine; VLP; Norwalk virus; Hepatitis B surface antigen; Human calicivirus; Virus like particle; Orthohepadnavirus; Hepatic disease; Vaccine; Infection; Virus; Viral hepatitis B; Calicivirus; Hepadnaviridae; Caliciviridae; Viral disease; Digestive diseases; Hepatitis B virus
• ISSN: 0264-410X; 1873-2518
• DOI: 10.1016/j.vaccine.2004.11.017

Expression of vaccine antigens in plants and delivery via ingestion of transgenic plant material has shown promise in numerous pre-clinical animal studies and in a few clinical trials. A number of different viral antigens have been tested, and among the most promising are those that can assemble virus-like particles (VLP), which mimic the form of authentic virions and display neutralizing antibody epitopes. We have extensively studied plant expression, VLP assembly, and immunogenicity of hepatitis B surface antigen (HBsAg) and Norwalk virus capsid protein (NVCP). The HBsAg small protein (S protein) was found by TEM to assemble tubular membrane complexes derived from endoplasmic reticulum in suspension cultured cells of tobacco and soybean, and in potato leaf and tuber tissues. The potato material was immunogenic in mice upon delivery by ingestion. Here we describe the plant expression and immunogenicity of HBsAg middle protein (M protein or pre-S2 + S) which contains additional 55 amino acid pre-S2 region at N-terminus of the S protein. Plant-derived recombinant M protein provoked stronger serum antibody responses against HBsAg than did S protein when injected systemically in mice. We discuss implications for use of fusion proteins for enhanced immunogenicity and mucosal targeting of HBsAg, as well as delivery of heterologous fused antigens. NVCP expressed in plants assembled 38 nm virion-size icosahedral ( T = 3) VLP, similar to those produced in insect cells. The VLP stimulated serum IgG and IgA responses in mice and humans when they were delivered by ingestion of fresh potato tuber. Here we show that freeze-drying of transgenic NVCP tomato fruit yielded stable preparations that stimulated excellent IgG and IgA responses against NVCP when fed to mice. However, the predominant VLP form in tomato fruit was the small 23 nm particle also observed in insect cell-derived NVCP.