清野 宏

We previously established the selection-marker-free rice-based oral cholera vaccine (MucoRice-CTB) line 51A for human use by Agrobacterium-mediated co-transformation and conducted a double-blind, randomized, placebo-controlled phase I trial in Japan and the United States. Although MucoRice-CTB 51A was acceptably safe and well tolerated by healthy Japanese and U.S. subjects and induced CTB-specific antibodies neutralizing cholera toxin secreted by Vibrio cholerae, we were limited to a 6-g cohort in the U.S. trial because of insufficient production of MucoRice-CTB. Since MucoRice-CTB 51A did not grow in sunlight, we re-examined the previously established marker-free lines and selected MucoRice-CTB line 19A. Southern blot analysis of line 19A showed a single copy of the CTB gene. We resequenced the whole genome and detected the transgene in an intergenic region in chromosome 1. After establishing a master seed bank of MucoRice-CTB line 19A, we established a hydroponic production facility with LED lighting to reduce electricity consumption and to increase production capacity for clinical trials. Shotgun MS/MS proteomics analysis of MucoRice-CTB 19A showed low levels of α-amylase/trypsin inhibitor-like proteins (major rice allergens), which was consistent with the data for line 51A. We also demonstrated that MucoRice-CTB 19A had high oral immunogenicity and induced protective immunity against cholera toxin challenge in mice. These results indicate that MucoRice-CTB 19A is a suitable oral cholera vaccine candidate for Phase I and II clinical trials in humans, including a V. cholerae challenge study.

Glycoprotein 2 (GP2) is a widely distributed protein in the digestive tract, contributing to mucosal barrier maintenance, immune homeostasis, and antigen-specific immune response, while also being linked to inflammatory bowel disease (IBD) pathogenesis. This review sheds light on the extensive distribution of GP2 within the gastrointestinal tract and its intricate interplay with the immune system. Furthermore, the significance of GP2 autoantibodies in diagnosing and categorizing IBD is underscored, alongside the promising therapeutic avenues for modulating GP2 to regulate immunity and maintain mucosal balance.

Abstract We previously demonstrated that Alcaligenes-derived lipid A (ALA), which is produced from an intestinal lymphoid tissue-resident commensal bacterium, is an effective adjuvant for inducing antigen-specific immune responses. To understand the immunologic characteristics of ALA as a vaccine adjuvant, we here compared the adjuvant activity of ALA with that of a licensed adjuvant (monophosphoryl lipid A, MPLA) in mice. Although the adjuvant activity of ALA was only slightly greater than that of MPLA for subcutaneous immunization, ALA induced significantly greater IgA antibody production than did MPLA during nasal immunization. Regarding the underlying mechanism, ALA increased and activated CD11b+ CD103− CD11c+ dendritic cells in the nasal tissue by stimulating chemokine responses. These findings revealed the superiority of ALA as a mucosal adjuvant due to the unique immunologic functions of ALA in nasal tissue.

Sapoviruses, like noroviruses, are single-stranded positive-sense RNA viruses classified in the family Caliciviridae and are recognized as a causative pathogen of diarrhea in infants and the elderly. Like human norovirus, human sapovirus (HuSaV) has long been difficult to replicate in vitro. Recently, it has been reported that HuSaV can be replicated in vitro by using intestinal epithelial cells (IECs) derived from human tissues and cell lines derived from testicular and duodenal cancers. In this study, we report that multiple genotypes of HuSaV can sufficiently infect and replicate in human-induced pluripotent stem cell-derived IECs. We also show that this HuSaV replication system can be used to investigate the conditions for inactivation of HuSaV by heat and alcohol, and the effects of virus neutralization of antisera obtained by immunization with vaccine antigens, under conditions closer to the living environment. The results of this study confirm that HuSaV can also infect and replicate in human normal IECs regardless of their origin and are expected to contribute to future virological studies.

Cationic cholesteryl-group-bearing pullulan nanogel (cCHP-nanogel) is an effective drug-delivery system for nasal vaccines. However, cCHP-nanogel-based nasal vaccines might access the central nervous system due to its close proximity via the olfactory bulb in the nasal cavity. Using real-time quantitative tracking of the nanogel-based nasal botulinum neurotoxin and pneumococcal vaccines, we previously confirmed the lack of deposition of vaccine antigen in the cerebrum or olfactory bulbs of mice and non-human primates (NHPs), rhesus macaques. Here, we used positron emission tomography to investigate the biodistribution of the drug-delivery system itself, cCHP-nanogel after mice and NHPs were nasally administered with 18F-labeled cCHP nanogel. The results generated by the PET analysis of rhesus macaques were consistent with the direct counting of radioactivity due to 18F or 111In in dissected mouse tissues. Thus, no depositions of cCHP-nanogel were noted in the cerebrum, olfactory bulbs, or eyes of both species after nasal administration of the radiolabeled cCHP-nanogel compound. Our findings confirm the safe biodistribution of the cCHP-nanogel-based nasal vaccine delivery system in mice and NHPs.