Nanogram Doses of Alum-Adjuvanted HBs Antigen Induce Humoral Immune Response in Mice When Orally Administered

Abstract

Mucosal immunity elicited by plant-based and other orally administered vaccines can serve as the first line of defense against most pathogens infecting through mucosal surfaces, but it is also considered for systemic immunity against blood-borne diseases such as hepatitis B (HB). Previous oral immunization trials based on multiple administration of high doses of HBs antigen elicited an immune response; however, a reproducible and long-lasting immunization protocol was difficult to design. The objective of this study was to evaluate the effect of dose and timing of orally delivered alum-adsorbed antigen on the magnitude of the anti-HBs humoral response. Mice were immunized orally by gavage intubation or parenterally by intramuscular injection three times, once every 2 weeks, with doses of 5, 50, or 500 ng alum-adjuvanted HBsAg. A low dose (10 ng) of HBsAg was orally administered three times in different time intervals: 2, 4, 6, and 8 weeks. The three consecutive 5-ng oral doses of the antigen induced immune response at the protective level (>or=10 mIU/ml), significantly higher than the reaction elicited by three 50 or 500 ng doses. In contrast, intramuscular delivery of these doses did not differ significantly; however, they induced a five to six times higher immune response than oral immunization. The 8-week period between each of the three oral immunizations appeared to be favorable to the anti-HBs humoral responses compared with the shorter schedules. The results presented here clearly identify the importance of low doses of antigen administered orally in extended intervals for a significantly higher anti-HBs response. This finding provides some indications concerning the strategy of orally administered vaccines, including plant-based ones.

Is An Oral Plant-based Vaccine against Hepatitis B Virus Possible?

“…Orally administered HBsAg also induced S-IgAs, similarly as in other experiments [67,72,80], and their production corresponded with the antigen dosage. S-IgAs may prevent infection of pathogens penetrating mucosa [29].…”

“…Microgram amounts of rHBsAg adjuvanted with various substances induced significant (tens mIU/ml) or much higher (>1000 mIU/ml) response [97,98]. On the other hand, doses of the antigen, adsorbed on a rather mild adjuvant -alum hydroxide, being as low as 5 -10 ng per mouse, were sufficient to elicit antiHBs antibodies at 20 -30 mIU/ml and lower antigen doses administered in longer intervals corresponded to higher responses [80]. Consequently, it was believed that anti-HBs immune response could be enhanced by a controlled 'oneshot' immunisation procedure secured by the delivery of relatively low and strictly defined doses of plant-associated HBsAg in a size-reduced form of lyophilised tissue.…”

“…It is also possible to apply purified HBs antigens as oral vaccines, as proposed in some reports [100]. Immunogenicity of orally administered purified HBsAg and adjuvanted with various substances was reported in several studies [80,97,98]. Purified HBs antigens may probably be used as mucosal vaccines, for instance nasal or oral, in a form of aerosol, drops, syrup or suspension (Fig.…”

“…Concurrently conducted research based solely on oral administration also showed that an extremely high HBsAg dose was not an indispensable factor in evoking systemic immune response [37,66,67,80]. Production of anti-HBs antibodies was elicited at a level slightly above the protective minimum, i.e.…”

Low-dose oral immunization with lyophilized tissue of herbicide-resistant lettuce expressing hepatitis B surface antigen for prototype plant-derived vaccine tablet formulation

“…A 5-ng dose of alum-adsorbed S-HBsAg administered orally to mice over 8 weeks induced a significantly higher response than a 500-ng dose delivered over 2 weeks (Kapusta et al 2010), or the response was higher and reached the ≥10 mIU/ml protective level in human volunteers fed with lettuce on days 1 and 60 (Kapusta et al 1999) than in those fed on days 1, 7, and 30 (Kapusta et al 2001) with 1–2 μg of S-HBsAg per person. Accordingly, low nanogram doses of S-HBsAg elicited immune response both in the mucosa and the periphery, while higher doses stimulated tolerance (Kostrzak et al 2009); primary humoral response was triggered at an oral delivery of 30 ng of fusion antigen TBI-HBsAg to mice and the successive boost by DNA immunization only slightly increased the level of anti-HBs antibodies (Shchelkunov et al 2006).…”