Immunogenicity and Toxicity of Different Adjuvants Can Be Characterized by Profiling Lung Biomarker Genes After Nasal Immunization.

The efficacy of vaccine adjuvants depends on their ability to appropriately enhance the immunogenicity of vaccine antigens, which is often insufficient in non-adjuvanted vaccines. Genomic analyses of immune responses elicited by vaccine adjuvants provide information that is critical for the rational design of adjuvant vaccination strategies. In this study, biomarker genes from the genomic analyses of lungs after priming were used to predict the efficacy and toxicity of vaccine adjuvants. Based on the results, it was verified whether the efficacy and toxicity of the tested adjuvants could be predicted based on the biomarker gene profiles after priming. Various commercially available adjuvants were assessed by combining them with the split influenza vaccine and were subsequently administered in mice through nasal inoculation. The expression levels of lung biomarker genes within 24 h after priming were analyzed. Furthermore, we analyzed the antibody titer, cytotoxic T lymphocyte (CTL) induction, IgG1/IgG2a ratio, leukopenic toxicity, and cytotoxicity in mice vaccinated at similar doses. The association between the phenotypes and the changes in the expression levels of biomarker genes were analyzed. The ability of the adjuvants to induce the production of antigen-specific IgA could be assessed based on the levels of Timp1 expression. Furthermore, the expression of this gene partially correlated with the levels of other damage-associated molecular patterns in bronchoalveolar lavage fluid. Additionally, the changes in the expression of proteasome- and transporter-related genes involved in major histocompatibility complex class 1 antigen presentation could be monitored to effectively assess the expansion of CTL by adjuvants. The monitoring of certain genes is necessary for the assessment of leukopenic toxicity and cytotoxicity of the tested adjuvant. These results indicate that the efficacy and toxicity of various adjuvants can be characterized by profiling lung biomarker genes after the first instance of immunization. This approach could make a significant contribution to the development of optimal selection and exploratory screening strategies for novel adjuvants.

Development of screening method for intranasal influenza vaccine and adjuvant safety in preclinical study.

Recently, many vaccine adjuvants have been developed; however, most of the newly developed adjuvants have been dropped out of preclinical and clinical trials owing to their unexpected toxicity. Thus, the development of highly quantitative and comparable screening methods for evaluating adjuvant safety is needed. In a previous study, we identified specific biomarkers for evaluating the safety of an intranasal influenza vaccine with CpG K3 adjuvant by comparing biomarker expression. We hypothesized that these biomarkers might be useful for screening newly developed adjuvant safety. We compared the expression of biomarkers in mouse lungs by the intranasal administration of 4 types of adjuvants: Alum, Pam3CSK4, NanoSiO2, and DMXAA with subvirion influenza vaccine. The control adjuvant alum did not show any significant increase in biomarker expression or preclinical parameters; however, NanoSiO2 and Pam3CSK4 increased the expression of biomarkers, such as Timp1 and Csf1. DMXAA at 300 µg induced the expression of over 80% of biomarkers. Hierarchical clustering analysis showed that 300 µg DMXAA was classified in the toxicity reference whole-particle influenza vaccine cluster. FACS analysis to confirm specific phenotypes that the number of T cells decreased in DMXAA-treated mouse lungs. Thus, our biomarkers are useful for initial adjuvant safety and toxicity screening.

A novel vaccinological evaluation of intranasal vaccine and adjuvant safety for preclinical tests.

Vaccines are administered to healthy humans, including infants, so the safety and efficacy must be very high. Therefore, evaluating vaccine safety in preclinical and clinical studies, according to World Health Organization guidelines, is crucial for vaccine development and clinical use. A change in the route of administration is considered to alter a vaccine's immunogenicity. Several adjuvants have also been developed and approved for use in vaccines. However, the addition of adjuvants to vaccines may cause unwanted immune responses, including facial nerve paralysis and narcolepsy. Therefore, a more accurate and comprehensive strategy must be used to develope next-generation vaccines for ensuring vaccine safety. Previously, we have developed a system with which to evaluate vaccine safety in rats using a systematic vaccinological approach and 20 marker genes. In this study, we developed a safety evaluation system for nasally administered influenza vaccines and adjuvanted influenza vaccines using these marker genes. Expression of these genes increased dose-dependent manner when mice were intranasally administered the toxicity reference vaccine. When the adjuvant CpG K3 or a CpG-K3-combined influenza vaccine was administered intranasally, marker gene expression increased in a CpG-K3-dose-dependent way. A histopathological analysis indicated that marker gene expression correlated with vaccine- or adjuvant-induced phenotypic changes in the lung and nasal mucosa. We believe that the marker genes expression analyses will be useful in preclinical testing, adjuvant development, and selecting the appropriate dose of adjuvant in nasal administration vaccines.

A new method for the evaluation of vaccine safety based on comprehensive gene expression analysis.

For the past 50 years, quality control and safety tests have been used to evaluate vaccine safety. However, conventional animal safety tests need to be improved in several aspects. For example, the number of test animals used needs to be reduced and the test period shortened. It is, therefore, necessary to develop a new vaccine evaluation system. In this review, we show that gene expression patterns are well correlated to biological responses in vaccinated rats. Our findings and methods using experimental biology and genome science provide an important means of assessment for vaccine toxicity.