[Durable remission of T-cell prolymphocytic leukemia with CLEC16A::IL2 after allogeneic hematopoietic stem cell transplantation].

A 64-year-old woman presented with fine motor impairment in both hands. MRI revealed a contrast-enhanced lesion in the medulla oblongata. Lymphoid cells with abnormal blebs were observed and a CD4+/CD8+ double positive (DP) T cell population was detected by flow cytometry (FCM) in the bone marrow (BM) and the peripheral blood (PB). CLEC16A::IL2 fusion gene was identified by whole exome sequencing with DNA prepared from DP T cells. Clonal rearrangement of the T-cell receptor gene and expression of TCL1A protein were detected. This led to a diagnosis of T-cell prolymphocytic leukemia (T-PLL) with central nervous system (CNS) infiltration. Abnormal cells in BM and PB became undetectable on microscopy and FCM, and the CNS lesion disappeared on MRI after second-line therapy with alemtuzumab. Meanwhile, the CLEC16A::IL2 fusion mRNA remained detectable in PB. Allogeneic hematopoietic stem-cell transplantation was performed, and the fusion mRNA has now been undetectable for more than 5 years since transplantation. This is the first report of a T-PLL case with a CLEC16A::IL2 fusion gene.

Performance evaluation of Espline HTLV-I/II, a newly developed rapid immunochromatographic antibody test for different diagnostic situations.

IMPORTANCE: The World Health Organization estimated that 5-10 million people are infected with human T-cell leukemia virus type 1 (HTLV-1). This number is likely to be underestimated because reliable endemic data are available for only approximately 1.5 billion people worldwide. The point-of-care test is a powerful tool for the easy and quick detection of infections without the requirement for expensive instruments and laboratory equipment. Espline HTLV-I/II, a newly developed rapid immunochromatographic antibody test that was evaluated in this study, might significantly advance our understanding of the global epidemiology of HTLV-1 infection.

Identification of a Self-Assembling Small-Molecule Cancer Vaccine Adjuvant with an Improved Toxicity Profile.

Protein or peptide cancer vaccines usually include immune potentiators, so-called adjuvants. However, it remains challenging to identify structurally simple, chemically accessible synthetic molecules that are effective and safe as vaccine adjuvant. Here, we present cholicamideß (6), a self-assembling small-molecule vaccine adjuvant with an improved toxicity profile and proven efficacy in vivo. We demonstrate that cholicamideß (6), which is less cytotoxic than its parent compound, forms virus-like particles to potently activate dendritic cells with the concomitant secretion of cytokines. When combined with a peptide antigen, cholicamideß (6) potentiated the antigen presentation on dendritic cells to induce antigen-specific T cells. As a therapeutic cancer vaccine adjuvant in mice, a mixture of cholicamideß (6) and a peptide antigen protected mice from the challenges of malignant cancer cells without overt toxicity. Cholicamideß (6) may offer a translational opportunity as an unprecedented class of small-molecule cancer vaccine adjuvants.

Performance evaluation of in vitro diagnostic kits for hepatitis B virus infection using the regional reference panel of Japan.

BACKGROUND: Hepatitis B virus (HBV) infection is a global public health concern. Precise and sensitive detection of viral markers, including HBV DNA and HBs antigen (Ag), is essential to determine HBV infection. METHODS: The sensitivities and specificities of 5 HBV DNA and 14 HBsAg kits were evaluated using World Health Organization International Standards (WHO IS) and the Regional Reference Panel (RRP) consisting of 64 HBsAg-negative and 80 HBsAg-positive specimens. RESULTS: All 5 HBV DNA kits detected HBV DNA in the WHO IS at a concentration of 10 IU/mL. The sensitivity and specificity to the RRP were 98.8-100% and 96.9-100%, respectively. HBV DNA titers were well correlated among the 5 kits regardless of HBV genotype. However, discordance of the HBV DNA titer was found in 5 specimens measured by CAP/CTM HBV v2.0. Among 12 automated HBsAg kits, the minimum detectable concentrations in the WHO IS varied from 0.01 to 0.1 IU/mL. Two lateral flow assays were positive for WHO IS concentrations greater than or equal to 1.0 and 0.1 IU/mL, respectively. When analyzed by the RRP, 12 automated kits exhibited a sensitivity of 98.8-100%, and 2 lateral flow assays showed sensitivities of 93.8% and 100%. The specificities of HBsAg kits were 100%. In the quantification of HBsAg, some kits showed a poor correlation of measurements with each other and showed up to a 1.7-fold difference in the regression coefficient of HBsAg titers. There were variations in the correlations of measurements among HBsAg kits when analyzed by genotype. CONCLUSIONS: Five HBV DNA kits showed sufficient sensitivity and specificity to determine HBV infection. HBV DNA titers were compatible with each other irrespective of HBV genotypes. HBsAg kits had enough sensitivity and specificity to screen for HBV infection. One of the lateral flow assays had a nearly equivalent sensitivity to that of the automated HBsAg kit. HBsAg titers quantified by the evaluated kits were not compatible across the kits. Genotype-dependent amino acid variations might affect the quantification of HBsAg titers.

Systemically inoculated adjuvants stimulate pDC-dependent IgA response in local site.

The stimulation of local immunity by vaccination is desirable for controlling virus replication in the respiratory tract. However, the local immune stimulatory effects of adjuvanted vaccines administered through the non-mucosal route are poorly understood. Here, we clarify the mechanisms by which non-mucosal inoculation of adjuvants stimulates the plasmacytoid dendritic cell (pDC)-dependent immunoglobulin (Ig)A response in the lungs. After systemic inoculation with type 1 interferon (IFN)-inducing adjuvants, type 1 IFN promotes CXCL9/10/11 release from alveolar endothelial and epithelial cells and recruits CXCR3-expressing pDCs into the lungs. Because adjuvant-activated pulmonary pDCs highly express major histocompatibility complex II, cluster of differentiation 80, and cluster of differentiation 86, transplantation of such cells into the lungs successfully enhances antigen-specific IgA production by the intranasally sensitized vaccine. In contrast, pDC accumulation in the lungs and subsequent IgA production are impaired in pDC-depleted mice and Ifnar1-/- mice. Notably, the combination of systemic inoculation with type 1 IFN-inducing adjuvants and intranasal antigen sensitization protects mice against influenza virus infection due to the pDC-dependent IgA response and type I IFN response. Our results provide insights into the novel mucosal vaccine strategies using non-mucosal inoculated adjuvants.

[Lymphoplasmacytic lymphoma accompanied by severe myelofibrosis].

A 61-year-old female was referred to our hospital because of pancytopenia and febrile neutropenia. On admission, computed tomography showed mild hepatosplenomegaly and intra-abdominal abscess formation in the right pelvic region; however, no lymphadenopathy was found. Bone marrow (BM) examination showed severe fibrosis by silver staining. Several small- to medium-sized lymphocytes with a constriction in the nuclei were observed, exhibiting CD3 (-), CD10 (-), CD20 (+), BCL-2 (+-), and CD138 (+-). Genetic testing revealed that BM cells were positive for MYD88 mutation and positive for IgH rearrangement, whereas neither JAK2 nor CALR mutation was positive. A diagnosis of BM infiltration of lymphoplasmacytic lymphoma (LPL) was made. Rituximab monotherapy was administered once a week for four times. BM examination 4 weeks after the end of treatment showed that lymphoma cells had disappeared and that myelofibrosis had been almost gone. The MYD88 mutation of BM turned out to be negative at that moment.

Safety and immunogenicity of the Pfizer/BioNTech SARS-CoV-2 mRNA third booster vaccine dose against the BA.1 and BA.2 Omicron variants.

BACKGROUND: The Omicron variant of severe acute respiratory syndrome-coronavirus-2 (SARS-CoV-2) was identified in Japan in November 2021. This variant contains up to 36 mutations in the spike protein, the target of neutralizing antibodies, and can escape vaccine-induced immunity. A booster vaccination campaign began with healthcare workers and high-risk groups. The safety and immunogenicity of the three-dose vaccination against Omicron remain unknown. METHODS: A total of 272 healthcare workers were initially evaluated for long-term vaccine safety and immunogenicity. We further established a vaccinee panel to evaluate the safety and immunogenicity against variants of concern (VOCs), including the Omicron variants, using a live virus microneutralization assay. FINDINGS: Two-dose vaccination induced robust anti-spike antibodies and neutralization titers (NTs) against the ancestral strain WK-521, whereas NTs against VOCs were significantly lower. Within 93-247 days of the second vaccine dose, NTs against Omicron were completely abolished in up to 80% of individuals in the vaccinee panel. Booster dose induced a robust increase in anti-spike antibodies and NTs against the WK-521, Delta, and Omicron variants. There were no significant differences in the neutralization ability of sera from boosted individuals among the Omicron subvariants BA.1, BA.1.1, and BA.2. Boosting increased the breadth of humoral immunity and cross-reactivity with Omicron without changes in cytokine signatures and adverse event rate. CONCLUSIONS: The third vaccination dose is safe and increases neutralization against Omicron variants. FUNDING: This study was supported by grants from AMED (grants JP21fk0108104 and JP21mk0102146).

RAISING is a high-performance method for identifying random transgene integration sites.

Both natural viral infections and therapeutic interventions using viral vectors pose significant risks of malignant transformation. Monitoring for clonal expansion of infected cells is important for detecting cancer. Here we developed a novel method of tracking clonality via the detection of transgene integration sites. RAISING (Rapid Amplification of Integration Sites without Interference by Genomic DNA contamination) is a sensitive, inexpensive alternative to established methods. Its compatibility with Sanger sequencing combined with our CLOVA (Clonality Value) software is critical for those without access to expensive high throughput sequencing. We analyzed samples from 688 individuals infected with the retrovirus HTLV-1, which causes adult T-cell leukemia/lymphoma (ATL) to model our method. We defined a clonality value identifying ATL patients with 100% sensitivity and 94.8% specificity, and our longitudinal analysis also demonstrates the usefulness of ATL risk assessment. Future studies will confirm the broad applicability of our technology, especially in the emerging gene therapy sector.

Salvage Cord Blood Transplantation Using a Short-term Reduced-intensity Conditioning Regimen for Graft Failure.

Objective Graft failure (GF) is a life-threatening complication of hematopoietic stem cell transplantation (HSCT). A standardized conditioning regimen and an appropriate graft source of salvage HSCT for GF have not yet been established. Some case series have shown good hematopoietic recoveries after salvage HSCT using a short-term reduced-intensity preparative regimen consisting of fludarabine (30-90 mg/m2), cyclophosphamide (2 g/m2), and total-body irradiation (2 Gy). However, the dose of fludarabine has varied in these reports based on the clinical condition of the patients, resulting in very limited experiences with each dose of fludarabine. Methods We retrospectively analyzed 10 patients who developed GF after allogeneic HSCT and underwent salvage cord blood transplantation (CBT) using the above-mentioned conditioning regimen with a fixed dose (90 mg/m2) of fludarabine. Results Eight patients (80.0%) achieved neutrophil engraftment within 30 days from salvage HSCT with a median of 21 (range, 17-23) days. The 1-year overall survival (OS) rate after the salvage HSCT was 50.0%, and the median OS was 281 (range, 23-1,638) days. Cumulative incidences of non-relapse mortality and relapse at 1 year were 50.0% and 10.0%, respectively. Conclusion CBT using this short-term reduced-intensity conditioning regimen may be a promising salvage therapy for GF.

Impact of injection buffer volume to perform bronchoalveolar lavage fluid collection for isolating alveolar macrophages to investigate fine particle-induced IL-1α secretion.

The importance of alveolar macrophages has been reported in many toxicology/immunology studies. Alveolar macrophages release interleukin (IL)-1α as a damage-associated molecular pattern (DAMP) when stimulated by fine particles. However, it is unclear whether cell isolation procedures affect ex vivo particle-induced responses in primary mouse alveolar macrophages (mAM). In this study, effects of injection buffer volume used to perform bronchoalveolar lavage fluid (BALF) collection to isolate mAM for use in ex vivo particle-induced responses were assessed. Among the mAM obtained from BALF collected using a 0.55 or 0.75 ml, but not a 1.0 ml buffer injection volume, decreased cell viability and IL-1α release were observed when cells were stimulated ex vivo with silica crystal or aluminum salt. Injected buffer composition did not affect the IL-1α release. On the other hand, IL-6 secretion induced by lipopolysaccharide (LPS) did not differ among mAM obtained from BALF collected using the different volumes. Expression levels of cell surface markers like CD11c, SiglecF, and CD64 did not differ among mAM obtained from BALF collected using the different injection buffer volumes. IL-1α release (and also necroptosis) induced by ex vivoparticle stimulation was suppressed by RIPK3 inhibitor or cytochalasin D co-treatment. Decreases in RIPK3 phosphorylation were noted in mAM obtained in BALF collected using the 1.0 ml injection volume compared with mAM obtained in BALF using 0.55 or 0.75 ml buffer. These observations illustrate that larger volumes of buffer used to collect BALF from mice can affect sensitivity of the isolated mAM to ex vivo particle-induced responses by inhibiting their functions.

[Acquired platelet dysfunction with severe bleeding tendency in triple-negative myelofibrosis].

A 50-year-old man demonstrated markedly increased number of white blood cells, anemia, severe splenomegaly, and bleeding tendency. Bone marrow analysis revealed remarkable hypercellularity; dysplasia in multilineage cells, including megakaryocytes; and fibrosis. He was eventually diagnosed with triple-negative myelofibrosis. A massive hematoma developed at the bone marrow biopsy site. A similar episode recurred after the second bone marrow biopsy. The von Willebrand factor and other coagulation factor activities were within normal ranges. Platelet aggregation analyses demonstrated highly impaired aggregation induced by ADP, collagen, and epinephrine. Treatment with hydroxyurea and ruxolitinib, a JAK inhibitor, was ineffective, and he eventually died on day 144 after hospitalization. Acquired platelet dysfunction uncommonly occurs in patients with myelodysplastic syndromes (MDS) and myeloproliferative neoplasms (MPN), without precise elucidation of the frequency and underlying mechanism. The onset of bleeding tendency in the current patient suggested that platelet dysfunction may be caused by somatic genetic events. Here, we discuss the mechanisms of acquired platelet dysfunction in MDS or MPN with a literature review.

Nasal alum-adjuvanted vaccine promotes IL-33 release from alveolar epithelial cells that elicits IgA production via type 2 immune responses.

Aluminum hydroxide salts (alum) have been added to inactivated vaccines as safe and effective adjuvants to increase the effectiveness of vaccination. However, the exact cell types and immunological factors that initiate mucosal immune responses to alum adjuvants are unclear. In this study, the mechanism of action of alum adjuvant in nasal vaccination was investigated. Alum has been shown to act as a powerful and unique adjuvant when added to a nasal influenza split vaccine in mice. Alum is cytotoxic in the alveoli and stimulates the release of damage-associated molecular patterns, such as dsDNA, interleukin (IL)-1α, and IL-33. We found that Ag-specific IgA antibody (Ab) production was markedly reduced in IL-33-deficient mice. However, no decrease was observed in Ag-specific IgA Ab production with DNase I treatment, and no decrease was observed in IL-1α/ß or IL-6 production in IL-33-deficient mice. From the experimental results of primary cultured cells and immunofluorescence staining, although IL-1α was secreted by alveolar macrophage necroptosis, IL-33 release was observed in alveolar epithelial cell necroptosis but not in alveolar macrophages. Alum- or IL-33-dependent Ag uptake enhancement and elevation of OX40L expression were not observed. By stimulating the release of IL-33, alum induced Th2 immunity via IL-5 and IL-13 production in group 2 innate lymphoid cells (ILC2s) and increased MHC class II expression in antigen-presenting cells (APCs) in the lung. Our results suggest that IL-33 secretion by epithelial cell necroptosis initiates APC- and ILC2-mediated T cell activation, which is important for the enhancement of Ag-specific IgA Ab production by alum.

Performance Evaluation of In Vitro Screening and Diagnostic Kits for Hepatitis C Virus Infection.

AIM: A reliable kit with high sensitivity and specificity is indispensable for diagnosing hepatitis C virus (HCV) infection. Detection kits for anti-HCV antibodies (anti-HCV) are used for screening, and quantification kits for HCV RNA and HCV antigen (Ag) are used for the definite diagnosis of HCV infection or the evaluation of the pathological condition of and therapeutic effects in patients with chronic hepatitis C. Several kits are currently available for these purposes and are provided for clinical use in Japan. In this study, we aimed to evaluate the performance of these kits. METHODS: We used International Standards for HCV RNA and HCV Ag and a regional reference panel to evaluate the performance of thirteen anti-HCV, five HCV RNA, and two HCV Ag kits. RESULTS: All specimens in the regional reference panel were diagnosed correctly by all anti-HCV kits, although the distributions of the quantified values varied, and the ratios of titer classification were not identical across kits. All HCV RNA kits quantified the International Standard with minimum deviation and diagnosed the specimens of the reference panel correctly. The quantified values of the International Standard by two HCV Ag kits were inconsistent. HCV Ag titers of some specimens were underestimated owing to the amino acid polymorphisms in comparison with HCV RNA titers. CONCLUSIONS: The evaluation with International Standards and the regional reference panel was useful for assessing the quality of screening and diagnostic kits for HCV infection, and such quality control is essential for the clinical usage of these kits.

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.

A high-throughput detection method for the clonality of Human T-cell leukemia virus type-1-infected cells in vivo.

Approximately 10-20 million of Human T-cell leukemia virus type-1 (HTLV-1)-infected carriers have been previously reported, and approximately 5% of these carriers develop adult T-cell leukemia/lymphoma (ATL) with a characteristic poor prognosis. In Japan, Southern blotting has long been routinely performed for detection of clonally expanded ATL cells in vivo, and as a confirmatory diagnostic test for ATL. However, alternative methods to Southern blotting, such as sensitive, quantitative, and rapid analytical methods, are currently required in clinical practice. In this study, we developed a high-throughput method called rapid amplification of integration site (RAIS) that could amplify HTLV-1-integrated fragments within 4 h and detect the integration sites in > 0.16% of infected cells. Furthermore, we established a novel quantification method for HTLV-1 clonality using Sanger sequencing with RAIS products, and the validity of the quantification method was confirmed by comparing it with next-generation sequencing in terms of the clonality. Thus, we believe that RAIS has a high potential for use as an alternative routine molecular confirmatory test for the clonality analysis of HTLV-1-infected cells.

Dasatinib Is an Effective Treatment for Angioimmunoblastic T-cell Lymphoma.

Recurrent hotspot (p.Gly17Val) mutations in RHOA encoding a small GTPase, together with loss-of-function mutations in TET2 encoding an epigenetic regulator, are genetic hallmarks of angioimmunoblastic T-cell lymphoma (AITL). Mice expressing the p.Gly17Val RHOA mutant on a Tet2-null background succumbed to AITL-like T-cell lymphomas due to deregulated T-cell receptor (TCR) signaling. Using these mice to investigate therapeutics for AITL, we found that dasatinib, a multikinase inhibitor prolonged their survival through inhibition of hyperactivated TCR signaling. A phase I clinical trial study of dasatinib monotherapy in 5 patients with relapsed/refractory AITL was performed. Dasatinib was started at a dose of 100 mg/body once a day and continued until days 10-78 (median day 58). All the evaluable patients achieved partial responses. Our findings suggest that AITL is highly dependent on TCR signaling and that dasatinib could be a promising candidate drug for AITL treatment. SIGNIFICANCE: Deregulated T-cell receptor signaling is a critical molecular event in angioimmunoblastic T-cell lymphoma and can be targeted with dasatinib.

Changes of urine metabolite profiles are induced by inactivated influenza vaccine inoculations in mice.

The safety evaluation of vaccines is critical to avoid the development of side effects in humans. To increase the sensitivity of detection for toxicity tests, it is important to capture not only pathological changes but also physiological changes. 1H nuclear magnetic resonance (NMR) spectroscopy analysis of biofluids produces profiles that show characteristic responses to changes in physiological status. In this study, mouse urine metabolomics analysis with 1H NMR was performed using different influenza vaccines of varying toxicity to assess the usefulness of 1H NMR in evaluating vaccine toxicity. Two types of influenza vaccines were used as model vaccines: a toxicity reference vaccine (RE) and a hemagglutinin split vaccine. According to the blood biochemical analyses, the plasma alanine transaminase levels were increased in RE-treated mice. Changes in metabolite levels between mice administered different types of influenza vaccines were observed in the 1H NMR spectra of urine, and a tendency toward dosage-dependent responses for some spectra was observed. Hierarchical clustering analyses and principal component analyses showed that the changes in various urine metabolite levels allowed for the classification of different types of vaccines. Among them, two liver-derived metabolites were shown to largely contribute to the formation of the cluster. These results demonstrate the possibility that urine metabolomics analysis could provide information about vaccine-induced toxicity and physiological changes.

Evaluation of in vitro screening and diagnostic kits for hepatitis B virus infection.

BACKGROUND: For the diagnosis of hepatitis B virus (HBV) infection, the detection and quantification of hepatitis B surface antigen (HBsAg) and HBV DNA are used. Several kits are available for this purpose, and there is a growing need for the evaluation of these kits because their performance may be affected by HBV genotype- or strain-specific polymorphisms. OBJECTIVES AND STUDY DESIGN: In this study, we used International Standards and the established regional reference panel to evaluate the performance of two HBV DNA quantitative kits, five HBsAg qualitative kits, seven HBsAg quantitative kits and three rapid immune-chromatographic tests for HBsAg. RESULTS: The quantification values of two HBV DNA quantitative kits exhibited excellent correlation. In the evaluation of HBsAg qualitative and quantitative kits, the titers of several specimens in the HBV-positive panel were below the detection limits of a few kits, and the specimens were determined as HBV-negative. Notably, the quantitative kit results exhibited low correlation values. However, when these data were analyzed for each genotype, the correlations improved. These results suggest that the HBsAg quantification data are influenced by HBV genotypes. The novel rapid immune-chromatographic test exhibited the comparable level of sensitivity to the HBsAg quantitative kits. CONCLUSIONS: We evaluated the performance of kits for the detection of HBV infection. The HBV DNA quantification data correlated with an excellent agreement, whereas the HBsAg quantification data were affected by HBV genotype. Such evaluations will be useful for estimating the quality of currently available and new HBV assay kits, and for the quality control of these kits.

Establishment of a novel safety assessment method for vaccine adjuvant development.

Vaccines effectively prevent infectious diseases. Many types of vaccines against various pathogens that threaten humans are currently in widespread use. Recently, adjuvant adaptation has been attempted to activate innate immunity to enhance the effectiveness of vaccines. The effectiveness of adjuvants for vaccinations has been demonstrated in many animal models and clinical trials. Although a highly potent adjuvant tends to have high effectiveness, it also has the potential to increase the risk of side effects such as pain, edema, and fever. Indeed, highly effective adjuvants, such as poly(I:C), have not been clinically applied due to their high risks of toxicity in humans. Therefore, the task in the field of adjuvant development is to clinically apply highly effective and non- or low-toxic adjuvant-containing vaccines. To resolve this issue, it is essential to ensure a low risk of side effects and the high efficacy of an adjuvant in the early developmental phases. This review summarizes the theory and history of the current safety assessment methods for adjuvants, using the inactivated influenza vaccine as a model. Our novel method was developed as a system to judge the safety of a candidate compound using biomarkers identified by genomic technology and statistical tools. A systematic safety assessment tool for adjuvants would be of great use for predicting toxicity during novel adjuvant development, screening, and quality control.

Gene expression profiling toward the next generation safety control of influenza vaccines and adjuvants in Japan.

Influenza becomes epidemic worldwide every year, and many individuals receive vaccination annually. Quality control relating to safety and potency of influenza vaccines is important to maintain public confidence. The safety of influenza vaccines has been assessed by clinical trials, and animal safety tests are performed to monitor the consistent quality between vaccines used for clinical trials and marketing; the biological responses in vaccinated animals are evaluated, including changes in body weight and white blood cell count. Animal safety tests have been contributing to the quality relating to the safety of influenza vaccines for decades, but improvements are needed. Although precise mechanisms involving biological changes in animal safety tests have not been fully elucidated, the application of cDNA microarray technology make it possible to reliably identify genes related to biological responses in vaccinated animals. From analysis of the expression profile of >10,000 genes of lung in animals treated with an inactivated whole virion influenza vaccine, we identified 17 marker genes whose expression patterns correlated well to changes in body weight and leukocyte count in vaccinated animals. In influenza HA vaccine-treated animals exhibiting subtle changes in biological responses, a robust expression pattern of marker genes was found. Furthermore, these marker genes could also be used in the evaluation of adjuvanted influenza vaccines. The expression profile of marker genes is expected to be an alternative indicator for safety control of various influenza vaccines conferring high sensitivity and short turnaround time. Thus, gene expression profiling may be a powerful tool for safety control of vaccines in the future.