Two vaccine toxicity-related genes Agp and Hpx could prove useful for pertussis vaccine safety control

Conventional animal tests such as leukocytosis promoting tests have been used for decades to evaluate toxicity of pertussis vaccine. Here, we examined gene expression in relation to the vaccine toxicity using a DNA microarray. Comparison of conventional animal test data with the DNA microarray-based gene expression data revealed a gene expression pattern highly correlated with leukocytosis in animals. Of 10,490 rat genes analyzed, two genes, alpha1-acid-glycoprotein (Agp) and hemopexin (Hpx), were found up-regulated by the toxin administration in a dose-dependent manner (assayed by a quantitative PCR based on the microarray). Variation of the gene expression was very small amongst the test animals, and the results were highly reproducible. These findings suggest that gene expression analysis of vaccine-treated animals can be used as an accurate and simple method of pertussis vaccine safety assessment.     

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.     

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.     

Application of DNA microarray technology to influenza A/Vietnam/1194/2004 (H5N1) vaccine safety evaluation

We propose that DNA microarray analysis can be used in the quality control of pandemic and endemic influenza vaccine. Based on the expression profiles of 76 genes in the rat lung one day after inoculation of influenza vaccine, we can distinguish whole-virion influenza vaccine (PDv: pandemic influenza vaccine and WPv: whole virion-particle vaccine) and sub-virion vaccine (HA vaccine) from saline. Among these 76 genes, we found genes up-regulated by influenza infection, as well as genes involved in the immune response, and interferon. Hierarchical clustering of each influenza vaccine by the expression profiles of these 76 genes matched data from current quality control tests in Japan, such as the abnormal toxicity test (ATT) and the leukopenic toxicity test (LTT). Thus, it can be concluded that DNA microarray technology is an informative, rapid and highly sensitive method with which to evaluate the quality of influenza vaccines. Using DNA microarray system, consistent with the results of the ATT and LTT, it was clarified that there was no difference in vaccine quality between PDv and WPv.     

Comparative analysis of a Bordetella pertussis patient isolated strain and classical strains used in the pertussis vaccine

A new Bordetella pertussis strain isolated from a whooping cough Brazilian patient was characterized under molecular and immunological aspects and compared with strains used in the production of whole-cell pertussis vaccine. The isolate, named 21A1, exhibited the bacteriological characteristics classically described for B. pertussis. RAPD and SDS-PAGE analysis showed similar DNA and proteic profiles between classical vaccinal strains and the new isolate. Comparative analysis about the efficacy of vaccines in protecting mice against the intracerebral challenge showed that the 21A1 vaccine was able to induce the highest mouse protection when compared with other B. pertussis vaccines. The results presented here indicate that the inclusion of selected new strain isolates in the composition of the B. pertussis vaccines can be an alternative to maintain or increase vaccine potency.     

Pertussis vaccine: present status and future prospects

Use of killed whole-cell Bordetella pertussis vaccines has been a major factor in control of symptomatic whooping cough (pertussis). In the UK, diminished public confidence in the safety of this vaccine led to a reduction in vaccine acceptance which correlated with an increase in the incidence of pertussis. There is a need for acellular pertussis vaccines of low toxicity which, ideally, will prevent colonization and also protect against the disease symptoms. Vaccine design can rely increasingly on knowledge of the roles of individual bacterial components in the pathogenesis of pertussis. Serotype-specific agglutinogens 2 and 3 (fimbriae) and filamentous haemagglutinin are among surface components of B. pertussis which probably mediate adhesion to the respiratory mucosa. Systemic effects of pertussis can largely be attributed to the lymphocytosis promoting factor (pertussis toxin). Vaccines containing detoxified toxin plus one or more purified adhesins are envisaged at present.     

Analysis of Bordetella pertussis isolates collected in Japan before and after introduction of acellular pertussis vaccines

Because of recent concern that whole-cell pertussis vaccination can drive antigenic divergence of circulating isolates of Bordetella pertussis, we compared 12 clinical isolates of B. pertussis collected in Japan, the first country to introduce acellular pertussis vaccines, with the vaccine strain. We used pulsed-field gel electrophoresis, sequencing of ptx and prn genes and expression of fimbriae. Most of the isolates collected before or after introduction of acellular vaccine possess similar restriction patterns. They contain ptx genes and prn alleles similar to the vaccine strain and to European isolates collected before the introduction of vaccination. Two recently collected isolates exhibiting a different pulsed-field gel electrophoresis pattern possess ptxS1 and prn alleles similar to the alleles harbored by European isolates circulating currently. Our preliminary results suggest that, if acellular pertussis vaccine-induced antigenic divergence exists, it is likely to be a slow or rare process.     

An aerosol challenge model of Bordetella pertussis infection as a potential bioassay for acellular pertussis vaccines

Whole cell and five different types of acellular pertussis vaccine were assayed using a mouse aerosol challenge model which permitted delivery of a controlled, consistent dose of Bordetella pertussis to the lower respiratory tract. Using this system, the viable counts in the lungs of vaccinated mice were immunisation dose-dependent and allowed the protective capacity of different vaccine preparations to be distinguished. This model may thus provide the basis for a protection assay for pertussis vaccines. Comparison of acellular vaccines with a whole cell pertussis vaccine showed that the latter gave better active protection in mice but with a different dose-response relationship. Thus the two types of vaccine are not directly comparable in the same assay and require different reference standards. A pentavalent type acellular vaccine is suggested as a possible candidate standard for the acellular vaccine potency test. The results suggest that this aerosol challenge model has potential for use as a potency test for acellular pertussis vaccines.     

WHO working group on standardisation and control of acellular pertussis vaccines--report of a meeting held on 16-17 March 2006, St. Albans, United Kingdom

This report reflects the discussion and conclusions of a WHO group of experts from national regulatory authorities, national control laboratories, vaccine industry and other relevant institutions involved in standardisation and control of acellular pertussis vaccines, held on 16-17 March 2006, in St. Albans, UK. Following previous discussions (Bethesda, 2000; Ferney-Voltaire, 2003; Geneva, 2005) and collection of relevant data for quality control, on the one hand, and clinical evaluation of acellular pertussis vaccines, on the other, this meeting was intended to review the scientific basis for the revision of WHO guidelines adopted in 1996 [Guidelines for the production and control of the acellular pertussis component of monovalent or combined vaccines. In: WHO Expert Committee on Biological Standardisation. Forty-seventh report. Geneva, World Health Organisation, 1998 (WHO Technical Report Series, No. 878), Annex 2]. The discussion on animal protection models, immunogenicity and toxicity testing was focused on three main aspects: value of the assay for the purpose of licensing and/or lot release; validity criteria and potential optimisation of the assays. The group agreed that establishment of JNIH-3 as a potential International Standard (IS) for modified intra-cerebral challenge assay should be under consideration. It was suggested that the inclusion of a reference vaccine, such as JNIH-3 in the intra-nasal challenge model could improve the standardisation of this assay. It was proposed that the development of stable reference vaccines for immunogenicity testing should be encouraged. Further collection of the data from the countries with established lot release of acellular pertussis vaccines will be undertaken to prepare a solid basis for recommendations on toxicity tests. In the context of recommendations for clinical assessment of new vaccines, the group emphasised the importance of comparability studies with antigens that have already undergone efficacy trials in the past. The outline for the section on clinical evaluation of acellular pertussis vaccines was presented and after the consultation further additions were made. Post-marketing surveillance was recognised as an important part of overall vaccine evaluation and a unique opportunity to understand vaccine performance in the population and to establish a link with quality control.     

Comparison of acellular pertussis-based combination vaccines by Japanese control tests for toxicities and laboratory models for local reaction

Two batches each of diphtheria–tetanus–acellular pertussis vaccine (DTaP) and that combined with inactivated polio vaccine purchased from the U.S.A., European and Asian markets were compared with Japanese DTaPs by Japanese control tests for DTaP and laboratory models for local reaction. All the imported vaccines met Japanese criteria for toxicities of acellular pertussis vaccine except for the toxicity to mouse weight gain (body weight decreasing (BWD) toxicity). When injecting into mouse footpad, rabbit back skin and mouse quadriceps muscle, the imported vaccines induced much severer inflammation and tissue injury comparing to Japanese DTaPs irrespective of animal species, injection site and injection volume suggesting that these vaccines may induce stronger local reactogenicity.     

WHO Working Group meeting on standardization of acellular pertussis vaccines: Potency assay Beijing, China, 7-9 November 2007

New acellular pertussis vaccines have recently been developed in China and India. In this context, potency testing and potential improvements of the protective animal models with inclusion of a reference material were recognized as critical issues in the quality assessment of acellular pertussis vaccines. One of these models, namely Modified Intracerebral Challenge Assay (MICA), is currently used as a potency assay in Japan, China and Korea. A collaborative study comparing whole cell references, a candidate acellular pertussis vaccine reference (JNIH-3) and various acellular pertussis products was undertaken in 2006. The results of the collaborative study showed that MICA worked reliably and gave consistent results between laboratories. JNIH-3 was found to give similar dose-response lines to a variety of acellular pertussis vaccines and DTaP formulations, irrespective of the differences in acellular pertussis components. The WHO Working Group agreed that proposal for establishing JNIH-3 as the First International Standard for acellular pertussis vaccine in MICA should be submitted to the Expert Committee on Biological Standardization at its meeting in October 2008.     

Increasing FIM2/3 antigen-content improves efficacy of Bordetella pertussis vaccines in mice in vivo without altering vaccine-induced human reactogenicity biomarkers in vitro

Current acellular-pertussis (aP) vaccines appear inadequate for long-term pertussis control because of short-lived efficacy and the increasing prevalence of pertactin-negative isolates which may negatively impact vaccine efficacy. In this study, we added fimbriae (FIM)2 and FIM3 protein to licensed 2-, 3- or 5-component aP vaccines (Pentavac®, Boostrix®, Adacel®, respectively) to assess whether an aP vaccine with enhanced FIM content demonstrates enhanced efficacy. Vaccine-induced protection was assessed in an intranasal mouse challenge model. In addition, potential reactogenicity was measured by biomarkers in a human whole blood assay (WBA) in vitro and benchmarked the responses against licensed whole cell pertussis (wP) and aP vaccines including Easyfive®, Pentavac® and Pentacel®. The results show that commercial vaccines demonstrated reduced efficacy against pertactin-negative versus pertactin-positive strains. However, addition of higher amounts of FIM2/3 to aP vaccines reduced lung colonization and increased vaccine efficacy against a pertactin-negative strain in a dose-dependent manner. Improvements in efficacy were similar for FIM2 and FIM3-expressing strains. Increasing the amount of FIM2/3 proteins in aP formulations did not alter vaccine-induced biomarkers of potential reactogenicity including prostaglandin E₂, cytokines and chemokines in human newborn cord and adult peripheral blood tested in vitro. These results suggest that increasing the quantity of FIM proteins in current pertussis vaccine formulations may further enhance vaccine efficacy against B. pertussis infection without increasing the reactogenicity of the vaccine.     

The efficacy of whole cell pertussis immunisation: collected data on a vaccine produced in France

The place of whole cell pertussis vaccines in paediatric immunisation schedules is under re-evaluation by public health authorities in many countries, with the expectation that the newly licensed acellular Bordetella pertussis vaccines will induce fewer adverse events while providing equivalent efficacy. In France, for instance, the CSHPF (Conseil supérieur d'hygiène public de France) recently modified its long-standing recommendation that French children only receive whole cell pertussis vaccine. Consequently, an acellular pertussis vaccine may be used for the first booster, at 16-18 months of age, and should be used for the reinforcing dose at 11-13 y of age. French children, nonetheless, continue to receive whole cell pertussis vaccine for the primary series immunisations at 2, 3, and 4 months, as the only whole cell pertussis vaccine available in France (licensed by Aventis Pasteur) has a long-established record of safety and protective efficacy. A review of its unpublished and published clinical results, obtained from studies throughout the world, demonstrates an efficacy of from 84-100% in six different retrospective analyses or outbreak investigations and a protective efficacy of 92% by clinical trial.     

Systematic review and meta-analysis of the effect of pertussis vaccine in pregnancy on the risk of chorioamnionitis,non-pertussis infectious diseases and other adverse pregnancy outcomes

BackgroundSeveral countries have introduced maternal immunisation with pertussis vaccine to provide protection against pertussis in early infancy. There is increasing interest in non-specific effects of vaccines including that non-live vaccines may enhance susceptibility to non-targeted infections in females. Some studies have shown increased risk of chorioamnionitis among women receiving pertussis vaccine during pregnancy. We aimed to conduct a systematic review and meta-analysis of the effect of maternal pertussis immunisation on the risk of chorioamnionitis, as well as the secondary outcomes of non-pertussis infections in women, non-pertussis infections in infants, spontaneous abortion or stillbirth, maternal death and infant death.MethodsWe searched PubMed and Embase for articles published until January 14, 2021. We screened articles for eligibility and extracted data using Covidence. Quality was assessed using Cochrane RoB tool and Newcastle-Ottawa Scale. Data were imported into RevMan for pooling and conduction of a meta-analysis stratified by study type. Outcomes are presented as risk ratios.ResultsWe identified 13 observational studies and six randomized controlled trials eligible for inclusion. We pooled data on chorioamnionitis from six observational studies and found maternal pertussis vaccine (mostly compared with other maternal immunizations with non-live vaccines) to be associated with an increased risk among the pertussis vaccinated women, RR = 1.27 [CI 95%: 1.14–1.42]. We found no difference in the analysis of our secondary outcomes of non-pertussis infections, spontaneous abortion or stillbirth and death.ConclusionWe found an increased risk of chorioamnionitis among women who received pertussis vaccine in pregnancy. The large number of women receiving pertussis vaccine during pregnancy, as well as the growing evidence of non-live vaccines causing increased susceptibility to infections, indicates a need for further randomised trials to assess potential adverse effects of maternal immunisation with pertussis-containing vaccines.     

New acellular pertussis-containing paediatric combined vaccines

Combined pediatric vaccines have the advantages of conferring protection against multiple common infectious diseases with a reduced number of injections. Their use should lead to better compliance to recommended vaccination schedules. Diphtheria (D), tetanus (T) and whole-cell pertussis vaccine (P) have been successfully combined, with or without inactivated poliovirus vaccine (IPV) in the same syringe for many years. Recently developed acellular pertussis (aP) Haemophilus influenzae type B (Hib), inactivated poliomyelitis virus and hepatitis B vaccines are ideal candidates for inclusion in current combined vaccines. Nevertheless, the development of new combinations has to face preclinical and clinical issues: the appropriate formulation of the new antigen(s) and other vaccine components needs to be determined to ensure compatibility and guard against potential additive or unexpected adverse reactions; potential immunological interference between antigens and the negative impact of other vaccine components on immunogenicity may occur, and these have to be examined also. Whole-cell pertussis vaccines are highly protective against whooping cough, but the severe adverse reactions that these vaccines sometimes produce have led to hesitation over their use, including the decision of some countries to stop pertussis immunization. To increase the acceptability of pertussis vaccination, Pasteur Mérieux Connaught has developed a combined D, T and a two-component acellular pertussis vaccine (DTaP), composed of purified pertussis toxoid (PT) and filamentous haemagglutinin (FHA), which has been shown to be effective in an efficacy trial conducted in Senegal. Acellular DTaP vaccines are immunogenic and have a better safety profile than DTP vaccines, when given either for the primary series, for the booster vaccination or for both. In order to meet worldwide demands, the combined DTaP-IPV or DTP-IPV has been developed for countries where IPV is recommended. Following the encouragement of the WHO, an H. influenzae type B tetanus-conjugated (Act-HIB) vaccine, has been combined in a full liquid formulation with the whole-cell DTP. This vaccine showed a good safety and immunogenicity profile in infants and in toddlers. A combined DTaP-IPV-PRP-T vaccine (where the Act-HIB vaccine is reconstituted by the full-liquid DTaP-IPV) also has been successfully developed both for the primary series and for booster vaccination; although, a reduced immunogenicity against PRP observed after the primary series, this did not affect vaccine priming. Hepatitis B immunization campaigns targeting high-risk groups have failed to control the disease in areas of low endemicity. In 1992, the WHO recommended that hepatitis B vaccination should be integrated into the EPI in all countries by 1997-1999. For that purpose, hepatitis B vaccine is currently evaluated in pediatric combined vaccines. Developing new combination vaccines is a difficult but essential process for maintaining high immunization rates worldwide against infectious diseases, provided that the costs are acceptable. New combined vaccines including pneumococcal and meningococcal component are under wide-scale development.     

Development and clinical testing of multivalent vaccines based on a diphtheria-tetanus-acellular pertussis vaccine: difficulties encountered and lessons learned

The widespread use of whole-cell pertussis vaccines in the second half of the 20th century have reduced the incidence of the disease significantly. However, in some countries, concerns about the reactogenicity and potential neurological damage associated with whole-cell vaccines led to a decrease in vaccine acceptance and an increase in morbidity and mortality of pertussis in several countries. This prompted the development of less reactogenic acellular pertussis vaccines combined with diphtheria and tetanus toxoids, initially in Japan and later in other countries. In Europe, the improved diphtheria, tetanus and acellular pertussis (DTPa) vaccine was first introduced in March 1994.The pertussis component of this DTPa vaccine, developed by Glaxo SmithKline, consists of pertussis toxoid, filamentous haemagglutinin and pertactin. The vaccine is well tolerated, with a lower incidence of adverse reactions than after administration of whole-cell vaccines.The long-lasting efficacy and effectiveness of DTPa vaccines have been extensively documented and these are now the cornerstone of a large range of combined vaccines including DTPa-hepatitis B (HBV), DTPa-inactivated polio (IPV) and DTPa-HBV-IPV. A lyophilised Haemophilus influenzae type b (Hib) vaccine can be reconstituted with all of these liquid combinations. The introduction of well-tolerated and efficacious DTPa vaccines and their more polyvalent combinations has improved the acceptance and simplified the implementation of childhood immunisation. This paper is a review of the technical and scientific difficulties encountered and the lessons learned over the 10-year period that it took to develop and introduce six multivalent vaccines using the Glaxo SmithKline DTPa as a building block.     

Laboratory assessment problems of the efficacy of acellular pertussis vaccines

Although composition of acellular pertussis vaccines is better defined than whole-cell vaccines, differences in the formulation, content, and detoxification of pertussis vaccine antigens led to a unique character of each of differently produced acellular vaccine. Currently used methods for laboratory evaluation of whole-cell pertussis vaccine efficacy were found not suitable for acellular vaccines. There is a strong need to perform analysis and evaluation of the safety and efficacy profiles of acellular pertussis vaccines combined with other vaccine antigens (e.g. Hib) both before and after conjugation. Mechanisms of interactions seen after conjugation inducing weaker immunogenicity or efficacy are still poorly recognized.     

Pertussis vaccine effectiveness and duration of protection – A systematic review and meta-analysis

A comprehensive review of observational pertussis vaccine effectiveness (VE) studies is needed to update gaps from previous reviews. We conducted a systematic review of VE and duration of protection studies for the whole-cell (wP) and acellular (aP) pertussis vaccines and conducted a formal meta-analysis using random effects models. Evidence continues to suggest that receipt of any pertussis vaccine confers protection in the short-term against disease although this protection wanes rapidly for aP vaccine. We detected significant heterogeneity in pooled estimates due, in part, to factors such as bias and confounding which may be mitigated by study design. Our review of possible sources of heterogeneity may help interpretation of other VE studies and aid design decisions in future pertussis VE research.     

Lung pathology and immediate hypersensitivity in a mouse model after vaccination with pertussis vaccines and challenge with Bordetella pertussis

While evaluating vaccine efficacy against clinical Bordetella pertussis isolates in mice, after challenge vaccinated mice showed increased lung pathology with eosinophilia, compared to challenged, non-vaccinated animals. This led us to study bacterial clearance, lung pathology, lung TNF-alpha expression, and parameters of immediate hypersensitivity (IH), being serum IgE levels, eosinophil numbers in the bronchoalveolar lavage fluid, and ex vivo IL-4, IL-5, IL-10, IL-13, and IFN-gamma production by the bronchial lymph node cells. BALB/c mice received a combined Diphtheria (D), Tetanus (T), Poliomyelitis, and whole-cell Pertussis vaccine (WCV), a combined D, T, and three-component acellular Pertussis vaccine (ACV), aluminium hydroxide adjuvant, or PBS, 28 and 14 days before B. pertussis infection. Similarly treated non-infected mice were taken as a control. Infection induced pathology; this induction was stronger after (especially WCV) vaccination. WCV but not ACV vaccination induced TNF-alpha expression after challenge. After challenge, IH parameters were strongly increased by (especially ACV) vaccination. Vaccinated IL-4 KO mice showed similar clearance and pathology, in the absence of IgE and with reduced numbers of eosinophils. Vaccinated (Th1-deficient) T-bet KO mice showed reduced clearance and similar pathology. In summary, after challenge vaccination increased lung pathology, TNF-alpha expression (only WCV), and IH parameters. Th1 cells were critical for clearance.     

Comparison of acellular pertussis vaccines-induced immunity against infection due to Bordetella pertussis variant isolates in a mouse model

A significant increase in the incidence of pertussis in adolescents and adults has been observed in vaccinated populations. Concomitantly, emergence of novel pertussis toxin and pertactin types in circulating Bordetella pertussis isolates was noticed. In this study, immunity induced by acellular vaccines against infection due to isolates expressing different pertactin types and fimbriae was monitored in a mouse model. In accordance with previous studies, the effect of a bicomponent DTPa vaccine on bacterial clearance was lower when compared with tri- or pentavalent DTPa vaccines. Whatever the isolates used to infect mice, the tri- or pentavalent DTPa vaccines were both efficacious in inducing immunity that resulted in clearance of infection. These findings suggest that re-emergence of pertussis might not be related to emergence of isolates escaping vaccine protection. The present study reduces potential concerns about acellular vaccine efficacy, but frequent monitoring of protection and surveillance of the evolution of the B. pertussis population remains of particular importance.