Immunogenicity studies performed within clinical trials have shown that there is no clear correlation between the presence of Ab to pertussis antigens and protection from disease (1
). Thus, the Ab response, and in particular the assessment of the Ab binding to antigens, may not be sufficient to assure the potency of aP vaccine preparations in animal models. Moreover, measurement of Ab elicited by test vaccines gives information only on the consistency of the vaccine production and not necessarily on the ability of the vaccine to protect from B. pertussis
It is widely accepted that the method applied to wP vaccines to evaluate the potency of vaccine lots is inappropriate for the evaluation of acellular formulations (26
). In particular, the conventional intracerebral mouse protection test has not been found to predict clinical performance when applied to aP vaccines (13
To overcome these problems, a mouse intranasal challenge model has been proposed (11
). This method was able to discriminate among aP vaccines that showed a different efficacy in clinical trials (24
) and to reveal differences between the mouse lung clearance activities of bi- or tri-component aP pertussis vaccines (24
). Furthermore, as also shown in our results, this method was able to discriminate between the lung clearance induced by vaccination with 1/10 of a vaccine dose and with the full dose. An international collaborative study group is now working to formulate recommendations and guidelines for manufacturers to use this method as a possible assay to contribute to the determination of the potency of aP vaccines (17
The specific goal of this study was to evaluate the CMI in mice to better understand the immunogenicity mechanisms induced by aP vaccines. Growing evidence suggests a relevant role of CMI in protection induced by antipertussis vaccines in children (3
). As parameters of CMI responses, two assays were chosen: the T-cell proliferation assay and an assessment of Th1 cytokine IFN-γ by ELISA.
Th1 cytokines are associated with protection in various B. pertussis
infection models, and in particular, in humans, protection from pertussis after infection or vaccination is determined by the presence or by the induction of Th1 cytokines such as interleukin-12 (IL-12) and IFN-γ (3
Also, in mice the clearance of B. pertussis
or protection induced by wP vaccines is dependent on the production of appropriate Th1 cytokines. Mills and coworkers demonstrated the induction of elevated amounts of IFN-γ after vaccination with wP vaccines in mice, particularly when lymphocytes were activated by killed B. pertussis
). These authors showed also that protection induced by aP vaccines was associated with the secretion of Th2 cytokines, such as IL-5 (35
). But, depending on the mouse strain and on the antigen used to activate lymphocytes, appreciable amounts of IFN-γ were also induced by aP vaccines (35
). In addition, the same authors also describe a contribution of IL-12 in vaccine efficacy in the case of aP vaccines (28
). Therefore, we thought it would be relevant to measure IFN-γ induction in aP-vaccinated mice, a parameter which could be also used to measure the efficacy of wP cell vaccines.
The results obtained indicate that aP vaccine preparations which had previously passed the routine immunogenicity test and were able to induce the clearance of B. pertussis
from the lungs, following intranasal challenge in mice, may behave differently when inducing a CMI response. Indeed, one vaccine preparation was unable and the other two showed a borderline ability to prime mouse T cells to proliferate and to induce IFN-γ secretion when cultured in the presence of PT, the essential constituent of all aP vaccines. These results were in part supported by the results obtained in a recent study on the evaluation of efficacy of Japanese Biken and Takeda aP vaccines. The authors did not find a correlation between the protective effects induced by aP vaccine preparations, measured by respiratory infection protection test, and CMI responses (45
In conclusion, the results of this study showed that immunization with aP vaccine preparations may not induce an equal CMI response to B. pertussis
antigens in mice, especially to PT, which is a critical requirement for long-lasting protection (5
). The differences in CMI induction cannot be ascribed to specific vaccine formulation, such as differences in PT inactivation or in protein contents, because the same proportion of vaccine preparations studied, produced either by GSK or CB, was able to induce a positive CMI response. Whether and to what extent this observation is relevant to the assessment of protective capacity of the vaccine preparations in children is not clear at the moment, also in consideration of the low number of vaccine preparations tested.
Considering the critical role of the axis IL-12-IFN-γ for protection from pertussis (28
), our results suggest that testing the induction of IFN-γ, as a parameter of CMI responses, could be an additional tool for the evaluation of the immune response induced by aP vaccines. As in the case of the intranasal protection assay, a working group including control laboratories, manufacturers, and research groups could be established to set up and standardize the IFN-γ assay and determine parameters of acceptance for aP vaccine preparations.