Vaccines are well-known for their effects on inducing antibody responses, but cellular immunity can be equally important in protection from infectious diseases, such as dengue. In our paper, published in NPJ Vaccines, we describe the long-term T cell responses induced by vaccination with a tetravalent dengue vaccine (TAK-003), and how these likely impact how effective the vaccine is in protecting children and adolescents from dengue.

What is dengue and why is it a problem?

Dengue is a virus transmitted by mosquitoes which are primarily found in tropical and subtropical regions of the world. As the world warms due to climate change, it is likely that these mosquitoes will reproduce more and expand into new areas, including higher altitude and temperate regions, exposing millions more people to potential dengue infection.¹ While the majority of people who catch dengue only develop mild symptoms, the disease has the potential to be life-threatening, with approximately 22,000 deaths occurring every year, mostly in children.^2,3 

Four serotypes of the dengue virus exist, and while primary infection with one serotype is often mild, a subsequent infection with a different serotype increases the risk for severe disease, at least in part due to antigen-dependent enhancement.^4,5 This complex immune response has posed a challenge for effective vaccine development, with the ideal vaccine eliciting responses against all four serotypes simultaneously to maximise protection.

Why are T cell responses important?

T cell responses have been found to play a vital role in protection against natural infection from dengue, with both CD4 and CD8 T cells contributing to viral clearance and reduced risk of severe infection.^6-9 As such, an ideal dengue vaccine would stimulate these T cell responses, as well as inducing neutralizing antibodies, to maximise the reduction in disease risk.

What was our clinical study about?

TAK-003 is a live-attenuated, tetravalent dengue vaccine developed to protect against all four serotypes of dengue. In the pivotal phase 3 study, TAK-003 was shown to provide protection against virologically confirmed dengue and consequent hospitalization in children and adolescents aged 4–16 years living in dengue-endemic regions (TIDES; NCT02747927).^10-14

As part of the early clinical development program of TAK-003, vaccination was shown to induce potent and durable T cell responses against all 4 dengue serotypes, predominantly mediated by interferon gamma (IFN-γ)-producing CD8 T cells.^15-17 In the current study (NCT02948829), specifically focused on analysis of T cells, we further evaluated T cell responses induced by TAK-003 in 200 children aged 4–16 years living in Panama and The Philippines, where dengue is endemic. The study was designed to be similar in setting and participant characteristics as the pivotal phase 3 efficacy study in terms of age, region and enrolment timeframe, to maximize clinical applicability of the findings.

What did we do?

Using blood samples taken from the children and adolescents, we investigated the response rates, magnitude, kinetics and functionality of T cell responses after vaccination with TAK-003 up to 3 years post vaccination. In the study, participants received 2 doses of TAK-003 administered 3 months apart. Further analyses assessed neutralizing antibody (NAb) and T cell responses in TAK-003 recipients who developed breakthrough dengue, and any age/regional effects on T cell responses.

What did we find?

TAK-003 elicited durable T cell responses, which remained elevated through 3 years post-vaccination, irrespective of baseline serostatus. One month after the second TAK-003 dose, T cell response rates measured by IFN-γ ELISPOT assays were 96% in baseline seropositive and 99% in baseline seronegative TAK-003 recipients, with similar responses in both countries and across age groups (Fig. 1). 

Figure 1. T cell response rates against DENV induced by TAK-003. The figure shows the percentage of participants with a positive T cell response based on the IFN-γ ELISPOT assay through the study. The red bars represent participants who were seropositive at baseline, and the blue bars participants who were seronegative at baseline. Vaccine doses were administered on Days 1 and 90.

When we evaluated responses to each dengue serotype separately, we saw a similar pattern, with 79.8%, 90.2%, 77.3%, and 74.0% of participants having serotype-specific T cell responses against dengue viruses 1, 2, 3, and 4, respectively, one month after the second dose. Response rates were similar in baseline seropositive and seronegative participants (Fig. 2).

Figure 2. Specific T cell response rates against individual DENV serotypes induced by TAK-003. The top panel shows responses against each serotype through the study in baseline seropositive participants. Responses in baseline seronegative participants are shown in the bottom panel.

When evaluated against dengue serotype 2 non-structural (NS) proteins, the CD4 and CD8 T cell responses elicited by TAK-003 were multifunctional (i.e. secretion of ≥2 cytokines of IFN-γ, TNF-α, and IL-2). CD8 T cells targeting NS1, NS3, and NS5 typically secreted both IFN-γ and TNF-α, whereas CD4 T cells typically secreted 2 or more of IFN-γ, TNF-α, and IL-2. Responses were similar in baseline seropositive and seronegative participants.

During the study, 12 participants contracted dengue confirmed by PCR testing, with most cases being mild. All participants who developed dengue had both T cell and neutralizing antibody responses post-vaccination, and 9 of the cases had lower than average T cell responses. Infection with dengue generally resulted in an increase in the magnitude of both T cell and antibody responses, irrespective of baseline serostatus or causative serotype.

Why is our research important?

Our study was specifically designed to evaluate T cell responses after vaccination with TAK-003 and complements the efficacy, safety, and immunogenicity findings from the pivotal phase 3 efficacy study, which was performed in a similar population (i.e. children and adolescents aged 4 to 16 years living in dengue-endemic regions). Our data provide insight into the mechanistic basis of vaccine-induced immunity to dengue from TAK-003, and highlight the importance of long-term cellular and humoral responses in disease protection. Analysis of the small number of breakthrough dengue cases showed both humoral and cellular responses against all four serotypes, regardless of the causative serotype. In natural infections, strong T cell responses have been associated with prevention of severe dengue and in line with this, nearly all the VCD cases in the current study were mild. In addition, repeat infections with different dengue serotypes are believed to induce distinct immune responses profiles compared with primary infections, which is believed to be basis for the mild symptoms associated with tertiary and later infections. We hypothesize that individuals with breakthrough infections post-vaccination with TAK-003 may still derive indirect benefit of vaccination against future infections.

 Key takeaway messages

·       TAK-003 elicited durable specific T cell responses against all 4 dengue serotypes irrespective of baseline serostatus

·       The CD4 and CD8 T cell responses were multifunctional (i.e. secretion of ≥2 cytokines of IFN-γ, TNF-α, and IL-2) and persisted for up to 3 years post-vaccination

Acknowledgments

This study was sponsored by Takeda Pharmaceuticals International AG. We thank the trial participants, their parents/guardians, and the study investigators and members of the DEN-313 study team. Medical writing support was provided by Jennifer Engelmoer, PhD, of Sula Communications, and funded by Takeda Pharmaceuticals International AG. Material has been reviewed by the Walter Reed Army Institute of Research.  There is no objection to its presentation and/or publication. The opinions or assertions contained herein are the private views of the authors, and are not to be construed as official, or as reflecting true views of the Department of the Army or the Department of Defense. The investigators have adhered to the policies for protection of human subjects as prescribed in AR 70–25.

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