title: MucoRice-cholera toxin B-subunit, a rice-based oral cholera vaccine, down-regulates the expression of α-amylase/trypsin inhibitor-like protein family as major rice allergens.
abstract: To develop a cold chain- and needle/syringe-free rice-based cholera vaccine (MucoRice-CTB) for human use, we previously advanced the MucoRice system by introducing antisense genes specific for endogenous rice storage proteins and produced a molecularly uniform, human-applicable, high-yield MucoRice-CTB devoid of plant-associated sugar. To maintain the cold chain-free property of this vaccine for clinical application, we wanted to use a polished rice powder preparation of MucoRice-CTB without further purification but wondered whether this might cause an unexpected increase in rice allergen protein expression levels in MucoRice-CTB and prompt safety concerns. Therefore, we used two-dimensional fluorescence difference gel electrophoresis and shotgun MS/MS proteomics to compare rice allergen protein expression levels in MucoRice-CTB and wild-type (WT) rice. Both proteomics analyses showed that the only notable change in the expression levels of rice allergen protein in MucoRice-CTB, compared with those in WT rice, was a decrease in the expression levels of α-amylase/trypsin inhibitor-like protein family such as the seed allergen protein RAG2. Real-time PCR analysis showed mRNA of RAG2 reduced in MucoRice-CTB seed. These results demonstrate that no known rice allergens appear to be up-reregulated by genetic modification of MucoRice-CTB, suggesting that MucoRice-CTB has potential as a safe oral cholera vaccine for clinical application.
authors.affiliation: Division of Mucosal Immunology, Department of Microbiology and Immunology, The Institute of Medical Science, The University of Tokyo 108-8639, Japan.
authors.forename: Shiho
authors.initials: S
authors.lastname: Kurokawa
authors.forename: Rika
authors.initials: R
authors.lastname: Nakamura
authors.forename: Mio
authors.initials: M
authors.lastname: Mejima
authors.forename: Hiroko
authors.initials: H
authors.lastname: Kozuka-Hata
authors.forename: Masaharu
authors.initials: M
authors.lastname: Kuroda
authors.forename: Natsumi
authors.initials: N
authors.lastname: Takeyama
authors.forename: Masaaki
authors.initials: M
authors.lastname: Oyama
authors.forename: Shigeru
authors.initials: S
authors.lastname: Satoh
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Kiyono
authors.forename: Takehiro
authors.initials: T
authors.lastname: Masumura
authors.forename: Reiko
authors.initials: R
authors.lastname: Teshima
authors.forename: Yoshikazu
authors.initials: Y
authors.lastname: Yuki
mesh_terms: D000284:Administration, Oral; D000485:Allergens / Q000235:genetics / Q000302:isolation & purification; D052179:Antigens, Plant / Q000096:biosynthesis / Q000235:genetics*; D002771:Cholera / Q000188:drug therapy / Q000473:pathology / Q000517:prevention & control*; D002772:Cholera Toxin / Q000235:genetics* / Q000627:therapeutic use; D022121:Cholera Vaccines / Q000008:administration & dosage / Q000235:genetics; D015536:Down-Regulation; D018506:Gene Expression Regulation, Plant; D006801:Humans; D012275:Oryza / Q000235:genetics / Q000276:immunology; D010940:Plant Proteins / Q000096:biosynthesis / Q000235:genetics*; D030821:Plants, Genetically Modified / Q000235:genetics; D040901:Proteomics; D012639:Seeds / Q000235:genetics / Q000378:metabolism; D053719:Tandem Mass Spectrometry; D014361:Trypsin Inhibitors / Q000096:biosynthesis; D000516:alpha-Amylases / Q000037:antagonists & inhibitors / Q000096:biosynthesis*
chemical_list: D000485:Allergens; D052179:Antigens, Plant; D022121:Cholera Vaccines; D010940:Plant Proteins; C079480:RAG2 protein, Oryza sativa; D014361:Trypsin Inhibitors; D002772:Cholera Toxin; D000516:alpha-Amylases