Adjuvant Activity of the Catalytic A1 Domain of Cholera Toxin for Retroviral Antigens Delivered by GeneGun

Abstract

Most DNA-encoded adjuvants enhance immune responses to DNA vaccines in small animals but are less effective in primates. Here, we characterize the adjuvant activity of the catalytic A1 domain of cholera toxin (CTA1) for human immunodeficiency virus (HIV) and simian immunodeficiency virus (SIV) antigens in mice and macaques delivered by GeneGun. The inclusion of CTA1 with SIVmac239 Gag dramatically enhanced anti-Gag antibody responses in mice. The adjuvant effects of CTA1 for the secreted antigen HIV gp120 were much less pronounced than those for Gag, as the responses to gp120 were high in the absence of an adjuvant. CTA1 was a stronger adjuvant for Gag than was granulocyte-macrophage colony-stimulating factor (GM-CSF), and it also displayed a wider dose range than GM-CSF in mice. In macaques, CTA1 modestly enhanced the antibody responses to SIV Gag but potently primed for a recombinant Gag protein boost. The results of this study show that CTA1 is a potent adjuvant for SIV Gag when delivered by GeneGun in mice and that CTA1 provides a potent GeneGun-mediated DNA prime for a heterologous protein boost in macaques.Most DNA vaccines have translated poorly from mice to primates. The exact reasons for this loss of potency are not fully elucidated. This topic is discussed in detail in references 11, 13, 31, and 32. Two strategies to improve the immunogenicity of DNA vaccines for cellular and/or humoral immune responses have evolved in recent years. One utilizes enhanced delivery of the DNA using ballistic particle bombardment with the GeneGun (14) or in vivo electroporation (3). The other main strategy entails the use of plasmid mixtures that combine the DNA vaccine of interest with a plasmid that expresses an adjuvant. The best-studied DNA adjuvants are based on cytokines (9,25,29,30,48) or chemokines (33,53,54). Although several of these adjuvant approaches have shown great promise in small animal models, most have been disappointing in humans and nonhuman primates (11,13,32).Cholera toxin (CT) and the highly related heat-labile enterotoxin of Escherichia coli (LT) have been found to be powerful mucosal immunogens and adjuvants (reviewed in reference 34). In mice, antibody (Ab) responses to CT and bystander antigens last at least 2 years (35, 50). We previously showed that the adjuvant properties of CT can be incorporated into DNA vaccines delivered intramuscularly (i.m.) (7). CT and LT are AB5 enterotoxins produced by Vibrio cholerae and E. coli, respectively. CT and LT consist of catalytic A subunits (separated into A1 and A2 subdomains) anchored in rings of five identical B subunits (43). Their enzymatic active sites reside within the A1 subdomains, while the A2 subdomains anchor the A subunits into the B pentamers (43). The B pentamers of CT and LT bind to gangliosides on cell membranes and facilitate their entry into lysosomes (23). The A1 subdomains are cleaved from the A2 subdomains in the Golgi complex or the endoplasmic reticulum (ER), and then the A1 subdomains exploit host protein retention and degradatio...

Balance of cellular and humoral immunity determines the level of protection by HIV vaccines in rhesus macaque models of HIV infection

“…Others and we have shown that IL-12 is an excellent adjuvant for the induction of Th1 responses (38)(39)(40)(41)(42)(43)(44) as well as for augmenting ADCC activity in situ (45)(46)(47). We have also shown that LTA1 and the related enzymatically active A1 domain of cholera toxin (CTA1) are potent genetic adjuvants capable of augmenting antibody responses in mice (48)(49)(50).…”

The catalytic A1 domains of cholera toxin and heat-labile enterotoxin are potent DNA adjuvants that evoke mixed Th1/Th17 cellular immune responses

“…Cholera Toxin (CT), heat-labile enterotoxin (LT) and their enzymatically active A1 domains have also shown great promise as DNA vaccine adjuvants. 6,19,21,40,41 CT and LT are closely related AB5 enterotoxins produced by Vibrio cholera and E. coli, respectively and are well known mucosal immunogens and adjuvants (reviewed in ref. 42 ).…”

“…Unfortunately, the CT and LT holotoxins cannot be used as mucosal adjuvants in humans since they are extremely toxic when administered at mucosal sites even at very small doses. 56 Several methods have been used to minimize toxicity including toxin-inactivating mutations, 51,[57][58][59] immunizing at sites distant from the mucosa, such as the skin, [60][61][62][63][64] or by re-targeting the A1 subunits using plasmid DNA 40,41 or other proteins 47,48 absent the promiscuous native B subunit. CTA1, expressed as the native bacterial DNA sequence as a DNA adjuvant, enhanced both cellular and antibody responses to HIV Bal -gp120 when delivered intramuscularly (i.m.).…”

“…41 Human codon-optimization enhanced the in vitro biological activity of CTA1 by 250-fold and enhanced antibody responses to SIV gag when administered by gene gun. 40 Cellular responses, however, were absent when delivered by the gene gun. 40 Much of the promise of DNA based adjuvants lies in their abilities to "tune" immune responses in directions better suited to protect vaccinees from target infectious agents.…”

“…40 Cellular responses, however, were absent when delivered by the gene gun. 40 Much of the promise of DNA based adjuvants lies in their abilities to "tune" immune responses in directions better suited to protect vaccinees from target infectious agents. Enabling this outcome requires more in depth profiling of the immune responses induced by the selected adjuvants.…”