Dengue virus (DENV) infection is a major public health concern, being one of the most important mosquito-borne viral diseases. Despite ongoing global research efforts, no effective vaccine is currently available. In this study, we investigated the use of dengue virus type 2 (DENV-2) domain III (DIII) as a potential subunit immunogen utilizing the Nanopatch microneedle delivery system in a mouse model. The Nanopatch (NP) is a novel vaccine delivery system with an array of thousands of microprojections that targets dry-coated vaccines to the skin. The skin has a higher density of antigen presenting cells (APCs) as compared to muscle tissue, where APCs are responsible for the uptake of vaccines. Targeting vaccines directly to the skin delivers vaccines more effectively and therefore provide a dose-sparing effect with a lesser amount of vaccine needed for efficient stimulation of an immune response. Other advantages include eliminating the need for the costly cold chain as well as the requirement of healthcare personnel to administer the vaccine.
DIII of the envelope protein (E) is the smallest dengue viral domain that contains epitopes known to elicit protective antibodies. Corresponding to the C-terminal domain of the soluble ectodomain of E, DIII has an immunoglobulin-like fold and has been implicated in both receptor binding and membrane fusion. To produce high levels of recombinant DIII, we established a stable Chinese Hamster Ovary (CHO) cell line using puromycin and glutamine synthetase double selection. To assist in cell selection and analysis of protein expression and purification levels, we established a novel quantitative capture enzyme-linked immunosorbent assay (ELISA) for the detection of DIII.
Following confirmation of DIII expression and purification, AG129 mice were immunized via the NP. Mice were immunized with three doses of vaccine, 21 days apart. DIII specific antibody titres were assessed by ELISA and neutralizing responses analyzed by the plaque reduction neutralization test (PRNT).