[PMC free article] [PubMed] [Google Scholar] [7] Sharma A, Tandon M, Ahi YS, Bangari DS, Vemulapalli R, Mittal SK. 10 months post-HAd-GFP inoculation, na?ve- or HAdV-primed animals were vaccinated i.m. with 108 PFU of HAd-H5HA [HAdV-C5 vector expressing hemagglutinin (HA) of H5N1 influenza computer virus]. There was a significant continual decrease in vector immunity titers with time, thereby leading to significant continual increases in the levels of HA-specific humoral and cell-mediated immune responses. In addition, significant improvement in protection efficacy against challenge with an antigenically heterologous H5N1 computer virus was observed in HAdV-primed animals at 6 months and onwards. These results indicate that this annual immunization with the same AdV vector GRS may be effective due to a significant decline in vector immunity. Keywords: Adenoviral vectors, vector immunity, longevity of adenoviral vector immunity, prevalence of vector immunity, human adenoviral vector, avian influenza INTRODUCTION Adenovirus (AdV) vector-based vaccines induce excellent humoral and cell-mediated immune (CMI) responses[1C5] due to the adjuvant-like effect of Ad vectors in stimulating the innate immune system through both Toll-like receptor (TLR)-dependent and TLR-independent pathways [6, 7]. Ad vector-based influenza vaccines have shown excellent potential in both animal BAY 11-7085 models [8C10] and clinical trials in humans [11C14]. Our immunogenicity and protective efficacy BAY 11-7085 studies in mice demonstrate that Ad vector-based vaccines provide complete protection against challenge with both homologous and antigenically unique strains of influenza viruses [9, 15]. There is a high incidence of AdV infections in the general population due to the circulation of more than 60 human AdV (HAdV) serotypes. The development of Ad-specific neutralizing antibodies, popularly known as pre-existing vector immunity in the majority of individuals [16C18] is usually a potential concern for Ad vector-based vaccine efficacy. HAdV neutralizing antibody titers in humans in the U.S. was found to be in BAY 11-7085 the range of 256C512 in 16% of the samples [16]. In Sub-Saharan children, a median HAdV-C5 neutralizing antibody titer of 512 was observed [19]. However, it is unclear what levels of vector immunity may have a significant unfavorable impact on the development of effective immune BAY 11-7085 responses. As well, since the use of AdV vectors as vaccines would often require repeated immunization, each immunization might at least temporarily induce or boost vector immunity, making it important to understand the rate of decline of vector immunity with time. We have evaluated the role of HAd-C5-neutralizing antibodies or vector immunity in impacting the immunogenicity and protection efficacy of a HAdV-C5 vector (HAd-HA-NP) expressing the HA and NP genes of A/Vietnam/1203/04 (H5N1) influenza computer virus [20]. The mouse groups were primed either intranasally (i.n.) or intramuscularly (i.m.) with varying doses of HAdV-C5, and following the development of vector immunity, the animal groups were immunized with HAd-HA-NP via the i.n. or i.m. route. The immunogenicity and protection results suggested that moderate levels of vector immunity [520 virus-neutralization (VN) titer] did not adversely impact the protective efficacy of the vaccine. Further increases in vector immunity (up to 2240 VN titers) were overcome by either increasing the vaccine dose by 5 or using an alternate route of vaccination. In the presence of exceptionally high levels of vector immunity (~3040 VN titers), immunization with a 5 vaccine dose still resulted in approximately 3.3C3.7 logs reduction in lung titers of the challenge virus. Canarypox virus-based vaccines are routinely used in pet animals on an annual basis suggesting that the development of immunity against a canarypox vector due BAY 11-7085 to yearly exposure does not negatively impact the vaccine efficacy [21, 22]. In this manuscript, using a mouse model, we resolved whether anti-AdV immunity declines sufficiently in a 12 months to permit annual vaccination with AdV vector-based vaccines. We found that there was a continual decline in the vector immunity with time, leading to a significant increase in humoral and cell-mediated immune responses against the target immunogen. In addition, effective immunogenicity and protection was observed in HAdV-C5-primed animal groups immunized with a HAd vector (HAd-H5HA) expressing the HA gene of A/Hong Kong/156/97(H5N1) (HK/156)] at 6-month and onwards. This study should be useful for determining the practical power of AdV vector-based vaccines for human use. MATERIAL AND METHODS Cell lines and viruses All cell lines were grown in minimum essential medium (MEM) (Life Technologies, Gaithersburg, MD) with 10% fetal calf serum (Hyclone, Logan, UT) and 50 g/ml gentamycin. 293 (human embryonic kidney cells expressing HAdV-C5 E1 proteins) [23] and BHH2C (bovine-human hybrid clone 2C) [24].