Oral Administration of Influenza Vaccine in Combination with the Adjuvants LT-K63 and LT-R72 Induces Potent Immune Responses Comparable to or Stronger than Traditional Intramuscular Immunization

Abstract

Mucosal immunization strategies are actively being pursued in the hopes of improving the efficacy of vaccines against the influenza virus. Our group investigated the oral immunization of mice via intragastric gavage with influenza hemagglutinin (HA) combined with mutant Escherichia coli heat-labile enterotoxins K63 (LT-K63) and R72 (LT-R72). These oral immunizations resulted in potent serum antibody and HA inhibition titers, in some cases stronger than those obtained with traditional intramuscular administration, in addition to HA-specific immunoglobulin A in the saliva and nasal secretions. This study demonstrates that it may be possible to develop effective oral influenza vaccines.Influenza is a serious human disease exhibiting high mortality in vulnerable populations such as the very young and the very old, as well as causing significant morbidity in the general population (17). The social and economic costs associated with yearly influenza outbreaks are high (7). Formalin-inactivated whole-virus and split-virus vaccines administered intramuscularly (i.m.) are commercially available to control the spread and severity of influenza (15,38). These prophylactic vaccines, although important agents in controlling influenza, suffer from a number of shortcomings that limit their efficacy and acceptability. Notably, inactivated whole-virus and split-virus vaccines are known to activate CD8 ϩ cytotoxic T-lymphocyte responses only sporadically, have poor cross-reactivity to antigenic variants, and produce poor secretory immunoglobulin A (IgA) responses (4,7,17,24,34,36). In addition, injection site reactogenicity and weak immune responses can be a problem in very young children (18,19). Significant efforts are currently being pursued to improve the vaccines' efficacy and tolerability primarily through the development of mucosally active influenza vaccines (2,7,10,33,40). Oral immunization is considered by many to be a highly desirable form of vaccination, although numerous obstacles make oral immunization using subunit antigens a significant challenge (3, 6, 11). Many approaches have been investigated to develop viable orally active influenza vaccines (3,21,29,30). Mucosal adjuvants, primarily Escherichia coli heat-labile enterotoxin (LT) and cholera toxin (CT), are the most commonly employed vaccine enhancers (11,12). Although potent mucosal adjuvants, LT and CT are toxic in humans at doses useful for adjuvanticity due to their ADPribosyltransferase activity (28). The nontoxic B subunit of CT (CTB) has also been investigated; however, studies have indicated that small amounts of the whole CT are required for sufficient adjuvant potency, inhibiting the potential of CTB in humans (44,45,46). Our group has investigated the mutant LT toxins LT-K63 and LT-R72, which demonstrate extremely low (LT-R72) to undetectable (LT-K63) levels of ADP-ribosyltransferase activity yet maintain potent mucosal adjuvant activity, demonstrating that ADP-ribosyltransferase activity may not be linked to the adjuvant activity (2, 13, 16). In thi...

Effects of different routes of administration on the immunogenicity of the Tat protein and a Tat-derived peptide

“…18,19 At the same time, the oral administration is gaining interest for its high compliance and the capability of inducing mucosal responses. 28 However, to our knowledge, neither IM nor oral administration of the Tat protein have ever been compared to the ID route.…”

“…This may be of particular relevance for the development of vaccines against HIV/AIDS or other infectious diseases, which are transmitted mostly by mucosal routes. 34 The immunogenicity of oral administration, which may be further boosted by proper adjuvants or different vaccination schedules, 28,35 could be due to the binding properties of Tat. Indeed, some studies have shown that the oral administration of antigens capable of binding to mucosal surface induces strong immune responses.…”

Distribution of intranasal instillations in mice: effects of volume, time, body position, and anesthesia

“…In published studies, volumes of substances intranasally instilled into mice range from 5 l (22) to 100 l (3), with little justification for the chosen volumes. Intranasal delivery is often carried out after intraperitoneal (9, 19) or inhalation (5, 14, 15) anesthesia but has also been performed with fully awake mice (1,6,17). The position of the mouse during intranasal delivery has also varied between studies, with horizontal (13) and head-down supine (12) positions having been used.…”

Enhancement of intranasal vaccination in mice with deglycosylated chain A ricin by LTR72, a novel mucosal adjuvant

“…Furthermore, i.n. vaccination with influenza vaccine and LTR72 induced a high IgG2a response in the serum [15]. Intranasal vaccination with ovalbumin in the presence of LTR72 increased to about 30-and 50-fold the IgA levels in the nasal washings and in the serum of mice, respectively, comparable to IgA responses stimulated i.n.…”

“…Combined i.n. vaccination of mice with influenza vaccine and LTR72 significantly enhanced the antibody response compared with the response elicited by vaccine given by the traditional parenteral route [15]. Intranasal vaccination also induced mucosal IgA antibody responses, while systemic vaccination did not.…”