Plasmid Vectors Encoding Cholera Toxin or the Heat-Labile Enterotoxin fromEscherichia coliAre Strong Adjuvants for DNA Vaccines

Abstract

DNA vaccines are widely recognized for their ability to elicit cellular as well as humoral immune responses to microbial antigens (23). An important characteristic of DNA vaccines is that they can mimic characteristics of live vaccines due to their ability to induce the de novo production of microbial antigens in both transiently transfected nonimmune cells and antigenpresenting cells in the vaccine inoculation site and nearby draining lymph nodes (12,14,25). The promise of intramuscularly injected DNA vaccines has yet to be realized in the clinic, since a number of recent human trials investigating this approach have not yielded the strength and breadth of responses obtained previously in animal models (4,26,49). This may be in part due to the need for better delivery systems, since a recent human trial employing particle-mediated, intracellular delivery ("gene gun") of a hepatitis B DNA vaccine to the epidermis resulted in 100% seroconversion to protective titers and the induction of strong cellular responses (38). In addition to improved delivery, any DNA vaccine, with or without a specific delivery system, can potentially benefit from the inclusion of an adjuvant to augment the strength or quality of a given response. To this end, several investigators have demonstrated the ability to modulate the responses to both naked DNA and particle-based DNA vaccines by inclusion of vectors encoding a variety of cytokines and chemokines (23,24,36,40,46).Our efforts to identify a safe and potent adjuvant for particle-mediated DNA vaccines have recently focused on cholera toxin (CT) and the related Escherichia coli heat-labile enterotoxin (LT). CT and LT are two of the most powerful adjuvants known, due to their ability to elicit robust mucosal and systemic responses via the mucosal and parenteral routes (37, 50). CT and LT are members of the AB 5 class of bacterial toxins and are 80% homologous in their primary structure with essentially identical tertiary structures. These toxins exist as hexamers in which the pentameric B subunit oligomer contains the cellular receptor binding function and is linked to a single A subunit containing an ADP ribosyltransferase activity that is associated with both toxicity and adjuvant effects. Proteolytic cleavage of a trypsin-sensitive site in the A subunit is required for activation of enzymatic activity that is contained in the N-terminal A 1 subunit. This activity results in the ADP ribosylation of several G proteins involved in signal transduction. ADP ribosylation of G s , the regulator of adenylate cyclase, results in the permanent activation of adenylate cyclase and accumulation of cellular cyclic AMP (cAMP) levels.Although CT and LT are closely related molecules, important differences exist in the adjuvant effects that they exhibit (3,37,50). Use of CT as an adjuvant generally results in the induction of antigen-specific Th2 cytokine production, while LT elicits both Th1 and Th2 cytokines (30,51). This immuno-* Corresponding author. Mailing address: PowderJect Vaccines, Inc., 5...

Immunogenicity of combination DNA vaccines for Rift Valley fever virus, tick-borne encephalitis virus, Hantaan virus, and Crimean Congo hemorrhagic fever virus

“…The adjuvants did not improve the antibody responses elicited to the RVFV +NSm vaccine, which was not entirely unexpected. In earlier studies, influences of both sets of vectors on antibody responses were antigen dependent and ranged from no effect to sharp reductions in the immunoglobulin G1 (IgG1)-to-IgG2a ratios [28]. Here, we did not assess the antibody isotypes in the sera of vaccinated mice, thus we do not know if the adjuvants shifted the responses from the normal Th2-type response that we observe after gene gun vaccination of mice toward a more Th1-type response.…”

“…Co-administration of these genetic adjuvants has been shown to augment the Th1 cytokine responses (gamma interferon) in mice to multiple viral antigens when co-delivered with DNA vaccines. In addition, both adjuvants also increased antibody responses and Th2 cytokine responses (interleukin 4) to certain antigens tested [28].…”

“…The DNA loads were 0.5-1 g/mg of gold. For studies involving the genetic adjuvants, the RVFV +NSm DNA vaccine was mixed with plasmids individually encoding the alpha and beta subunits of cholera toxin (CT) or the alpha and beta subunits of a labile enterotoxin from E. coli (LT) [28] in a ratio of 2:1:1 and the mixture was precipitated onto gold beads. The DNA-coated gold was dried on the inside walls of Tefzel tubing, which was then cut into 0.5 in sections.…”

Preclinical and clinical progress of particle-mediated DNA vaccines for infectious diseases

“…In contrast, this review shows that DNA vaccines administered by PMED, even in the absence of heterologous boosting or adjuvants, have translated well from the mouse and bench to preclinical large animal species, including nonhuman primates and pigs, and to human clinical trials. New improvements in the PMED technology [72], vector design [25,76,77], including co-formulation of PMED DNA vaccines with adjuvants [78], are in progress to further enhance the potency of particle-mediated DNA vaccines. These eVorts are expected to accelerate the commercialization of PMED as an eVective approach for vaccination against infectious diseases.…”

Multi-antigenic DNA immunization using herpes simplex virus type 2 genomic fragments

“…Previous work has found that plasmids expressing Cholera Toxin (CT) and E. coli Heat Labile Toxin (LT) can greatly enhance cellular immune responses of standard DNA vaccines delivered by PMED. 13,14 To explore if these adjuvants, termed DNA Encoded Immunostimulators (DEIs), also enhanced SGV immunogenicity, experiments were performed to examine the activity of SGV/DEI formulations. These formulas were very potent and may be useful for the development of a vaccine requiring strong cellular responses in humans.…”