Background Dengue is caused by 4 antigenically distinct serotypes of dengue virus (DENV1–4). Takeda’s live attenuated tetravalent dengue vaccine (TAK-003) candidate is composed of an attenuated DENV2 and chimeric viruses containing prM/E of DENV1, 3 and 4 on the DENV2 backbone. The multicolor FluoroSpot (MCF) assay enables quantitation of serotype-specific and cross-reactive individual memory B cells (MBCs) secreting DENV-specific antibodies in a polyclonal mixture. Methods Using the MCF assay, we determined the type-specific and cross-reactive MBC response in peripheral blood mononuclear cells collected pre- and postvaccination from 7 macaques and 15 randomly selected individuals who received TAK-003 (8 DENV seronegative and 7 DENV seropositive) in a phase 2 clinical trial in Singapore (DEN-205 study). Results Preexisting DENV-specific MBC responses were detected only in seropositive vaccine recipients at day 0. Following vaccination, both type-specific and cross-reactive MBCs to all 4 DENV serotypes were observed in all macaques and clinical trial participants. The proportion of type-specific MBCs was higher than cross-reactive MBCs and remained stable between day 30 and 360 post vaccination. Conclusions These results demonstrate that, unlike primary or secondary natural DENV infection, tetravalent vaccination elicits tetravalent type-specific MBCs, and thus all 4 components of TAK-003 contribute to the DENV-specific MBC response following vaccination. Clinical Trials Registration NCT02425098.
Antibodies capable of activating the complement system (CS) when bound with antigen are referred to as “complement-fixing antibodies” and are involved in protection against Flaviviruses. A complement-fixing antibody test has been used in the past to measure the ability of dengue virus (DENV)-specific serum antibodies to activate the CS. As originally developed, the test is time-consuming, cumbersome, and has limited sensitivity for DENV diagnosis. Here, we developed and characterized a novel multiplex anti-DENV complement-fixing assay based on the Luminex platform to quantitate serum antibodies against all four serotypes (DENV1-4) that activate the CS based on their ability to fix the complement component 1q (C1q). The assay demonstrated good reproducibility and showed equivalent performance to a DENV microneutralization assay that has been used to determine DENV serostatus. In non-human primates, antibodies produced in response to primary DENV1-4 infection induced C1q fixation on homologous and heterologous serotypes. Inter-serotype cross-reactivity was associated with homology of the envelope protein. Interestingly, the antibodies produced following vaccination against Zika virus fixed C1q on DENV. The anti-DENV complement fixing antibody assay represents an alternative approach to determine the quality of functional antibodies produced following DENV natural infection or vaccination and a biomarker for dengue serostatus, while providing insights about immunological cross-reactivity among different Flaviviruses.
Background Antibody-driven complement system (CS) activation has been associated with protection against symptomatic dengue virus (DENV) infection. Aggregation, opsonization, lysis, and phagocytosis are mechanisms triggered by antibody-antigen immunocomplexes following fixation of the component 1q (C1q) and activation of the classical pathway. As a result, DENV neutralization and clearance are facilitated, whereas antibody-dependent enhancement of infection is inhibited. We investigated the ability of antibodies produced in response to Takeda’s dengue vaccine candidate, TAK-003, to fix C1q and activate CS. Methods Serum samples were collected from seronegative and seropositive participants in a phase 2 clinical trial (DEN-203; ClinicalTrials.gov: NCT01511250), pre- and post-vaccination. Samples were evaluated for the presence of complement-fixing antibodies (CFAs) against DENV using a Luminex multiplex-based immunoassay. Results TAK-003 elicited production of CFAs against all 4 DENV serotypes, which persisted for 1 year post-vaccination, irrespective of baseline serostatus. CFA levels were correlated with neutralizing antibody titers and virus-binding total IgG and IgG1 concentrations. Furthermore, efficiency of CFA fixation was greater in samples with higher polyclonal IgG avidity. Conclusion These results indicate that antibodies produced after TAK-003 vaccination are functional in both activating CS and neutralizing virus infection by all DENV serotypes, which may contribute to efficacy of TAK-003.
Background A safe and effective vaccine against dengue is needed to address an unmet medical need that affects a large portion of the world’s population. Takeda’s live attenuated tetravalent dengue vaccine candidate (TAK-003) has shown protection in an ongoing Phase 3 efficacy trial. TAK-003 contains an attenuated dengue type 2 virus (DENV-2), and 3 genetically modified viruses in which the structural proteins from each of the serotypes 1, 3 and 4 have been placed into the DENV-2 backbone. Exploratory immunological assessments have been a part of the TAK-003 clinical development plan to better understand the mechanisms of action of TAK-003, and to identify immune response signatures that may correlate with protection. Methods Cellular and humoral immune responses elicited by vaccination in dengue-naïve and dengue-exposed individuals were measured across several clinical trials. For the humoral response, several methods were used to measure the magnitude and characteristics of the antibodies following vaccination with TAK-003 including studies of neutralizing antibodies, antibodies that bind to the viral components of the vaccine, the affinity and complement fixing capabilities of antibodies specific to structural proteins, and additionally the level of antibodies specific to nonstructural protein 1 (NS1). Results A multifunctional cellular immune response was found following vaccination that primarily targeted nonstructural proteins in the DENV-2 backbone and was cross reactive to epitopes found in the other serotypes. The vaccine elicited neutralizing antibodies with high tetravalent seropositivity rates among participants. Further assessment of this response revealed that it consists of serotype-specific and cross-reactive neutralizing antibodies against all four serotypes. In addition, sera from vaccinated individuals neutralized genotypically diverse dengue strains. In addition to antibodies specific to structural components, antibodies to DENV-2 NS1 that were cross reactive to the NS1 proteins of the other serotypes were found. Conclusion The breadth of the cellular and humoral immune responses elicited by TAK-003 in vaccine recipients across a wide age range living in different endemicities aligns with the response profile expected of a multivalent live vaccine. Disclosures Ralph Braun, PhD, Takeda Pharmaceuticals (Employee) Mayuri Sharma, PhD, Takeda Pharmaceuticals (Employee) Christina DeMaso, n/a, Takeda Pharmaceuticals (Employee) Allan Parker, n/a, Takeda Pharmaceuticals (Employee) David Dominguez, n/a, Takeda Pharmaceuticals (Employee) Heather Watkins, n/a, Takeda Pharmaceuticals (Employee) Hansi Dean, PhD, Takeda Pharmaceuticals (Consultant) Lovkesh Karwal, n/a, Takeda Pharmaceuticals (Employee) Eduardo Nascimento, n/a, Takeda Pharmaceuticals (Employee) Nicole Messere, n/a, Takeda Pharmaceuticals (Employee) Isamu Tsuji, n/a, Takeda Pharmaceuticals (Employee) Melissa Zahralban-Steele, n/a, Takeda Pharmaceuticals (Employee) Jeffrey R. Currier, PhD, Takeda Pharmaceuticals (Scientific Research Study Investigator) Heather Friberg-Robertson, PhD, Takeda Pharmaceuticals (Scientific Research Study Investigator)
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