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Comparative Study
. 2013 Aug;173(2):230-41.
doi: 10.1111/cei.12101.

RGD capsid modification enhances mucosal protective immunity of a non-human primate adenovirus vector expressing Pseudomonas aeruginosa OprF

A Krause  1 , W Z Whu, J Qiu, D Wafadari, N R Hackett, A Sharma, R G Crystal, S Worgall
Affiliations
Comparative Study

RGD capsid modification enhances mucosal protective immunity of a non-human primate adenovirus vector expressing Pseudomonas aeruginosa OprF

A Krause et al. Clin Exp Immunol. 2013 Aug.

Abstract

Replication-deficient adenoviral (Ad) vectors of non-human serotypes can serve as Ad vaccine platforms to circumvent pre-existing anti-human Ad immunity. We found previously that, in addition to that feature, a non-human primate-based AdC7 vector expressing outer membrane protein F of P. aeruginosa (AdC7OprF) was more potent in inducing lung mucosal and protective immunity compared to a human Ad5-based vector. In this study we analysed if genetic modification of the AdC7 fibre to display an integrin-binding arginine-glycine-aspartic acid (RGD) sequence can further enhance lung mucosal immunogenicity of AdC7OprF. Intratracheal immunization of mice with either AdC7OprF.RGD or AdC7OprF induced robust serum levels of anti-OprF immunoglobulin (Ig)G up to 12 weeks that were higher compared to immunization with the human vectors Ad5OprF or Ad5OprF.RGD. OprF-specific cellular responses in lung T cells isolated from mice immunized with AdC7OprF.RGD and AdC7OprF were similar for T helper type 1 (Th1) [interferon (IFN)-γ in CD8(+) and interleukin (IL)-12 in CD4(+)], Th2 (IL-4, IL-5 and IL-13 in CD4(+)) and Th17 (IL-17 in CD4(+)). Interestingly, AdC7OprF.RGD induced more robust protective immunity against pulmonary infection with P. aeruginosa compared to AdC7OprF or the control Ad5 vectors. The enhanced protective immunity induced by AdC7OprF.RGD was maintained in the absence of alveolar macrophages (AM) or CD1d natural killer T cells. Together, the data suggest that addition of RGD to the fibre of an AdC7-based vaccine is useful to enhance its mucosal protective immunogenicity.

Keywords: RGD; adenovirus; lung; mucosal immunity; pseudomonas.

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Figures

Fig. 1
Fig. 1
Lung mucosal administration of adenovirus (Ad)C7 outer membrane protein F (OprF). arginine–glycine–aspartic acid (RGD) induces robust humoral and protective anti-Pseudomonas aeruginosa immunity. C57BL/6 mice were immunized intratracheally with AdC7OprF.RGD, AdC7OprF, AdC5OprF.RGD, AdC5OprF or Ad5Null (all 1010 particle units/mouse). (a) Anti-OprF immunoglobulin (Ig)G titre in serum determined by enzyme-linked immunosorbent assay (ELISA) at 0, 8 and 12 weeks. Limit of detection is indicated by dotted line. *P < 0·05 between AdC7 vectors and Ad5 vectors, all comparisons at 8 weeks; **P < 0·01 between AC7 vectors and Ad5 vectors, all comparisons at 12 weeks. (b) Anti-OprF IgG and (c) anti-OprF IgA in lung epithelial lining fluid at 0, 4 and 8 weeks. (d,e) Protective immunity against pulmonary challenge with agar-encapsulated P. aeruginosa (2 × 106 colony-forming units) at (d) 8 weeks and (e) 12 weeks after immunization. Bacterial counts in lung homogenate were performed after 48 h. Data are shown as means ± standard error of the mean of seven mice per group. *P < 0·05; ***P < 0·001.
Fig. 2
Fig. 2
Intrapleural and intramuscular administration of adenovirus (Ad)C7 outer membrane protein F (OprF). arginine–glycine–aspartic acid (RGD) induce humoral and protective anti-Pseudomonas aeruginosa immunity. AdC7OprF.RGD, AdC7OprF, AdC5OprF.RGD, AdC5OprF or Ad5Null (all 1010 particle units/mouse) were administered either (a,b) intramuscularly or to the (c,d) pleural space of C57BL/6 mice. (a,c) Anti-OprF immunoglobulin (Ig)G in serum. **P < 0·01, ***P < 0·001, between AdC7 vectors and Ad5 vectors, all comparisons for each time-point. (b,d) Protective immunity against pulmonary challenge with agar-encapsulated P. aeruginosa (2 × 106 colony-forming units) 12 weeks after immunization. Bacterial counts in lung homogenate were performed after 48 h. Data are shown as means ± standard error of the mean of seven mice per group. *P < 0·05; **P < 0·01.
Fig. 3
Fig. 3
Infection of dendritic cells with arginine–glycine–aspartic acid (RGD) unmodified and modified adenovirus (Ad)5 outer membrane protein F (OprF) and AdC7OprF. Murine bone marrow-derived DC (day 6 post-bone marrow harvest; a–d) and human monocyte-derived DC (day 6 post-peripheral blood mononuclear cell harvest; e, f) were infected with AdC7OprF.RGD, AdC7OprF, Ad5OprF, Ad5OprF.RGD and AdNull (all 104 particles per cell). (a,b) Adenoviral DNA, evaluated by Taqman real-time polymerase chain reaction, in murine bone marrow-derived dendritic cells 12 h and (b) 48 h following infection. (c,d) OprF protein expression in cell lysates by Western analysis (c) 12 h and (d) 48 h following infection. (e) Adenoviral DNA and (f) OprF protein expression in human monocyte-derived DC 48 h after infection. *P < 0·05; **P < 0·01; ***P < 0·001.
Fig. 4
Fig. 4
Effect of natural killer T cells on adenovirus (Ad)C7 arginine–glycine–aspartic acid (RGD). outer membrane protein F (OprF)-induced immune response. CD1d–/– mice were immunized intratracheally with AdC7RGD.OprF, AdC7OprF or AdNull (1010 particle units). (a) Anti-OprF immunoglobulin (Ig)G in serum after 8 weeks analysed by enzyme-linked immunosorbent assay (ELISA). (b) Protective immunity following challenge with agar-encapsulated Pseudomonas aeruginosa (2 × 106 colony-forming units) after 8 weeks. Bacterial counts in lung were analysed after 48 h. Data are shown as means ± standard error of the mean of six mice/group. *P < 0·05.
Fig. 5
Fig. 5
Effect of lung macrophages on adenovirus (AdC7) arginine–glycine–aspartic acid (RGD). outer membrane protein F (OprF)-induced immunity. (a,b) Clodronate-liposomes were administered intranasally to C57BL/6 mice to deplete macrophages in the respiratory tract. After 24 h, mice were immunized intratracheally with AdC7RGD.OprF. (a) Anti-OprF immunoglobulin (Ig)G in serum after 8 weeks analysed by enzyme-linked immunosorbent assay. (b) To assess protective immunity, mice were challenged after 8 weeks with agar-encapsulated Pseudomonas aeruginosa (2 × 106 colony-forming units). Bacterial counts in lung were analysed after 48 h. Data are shown as means ± standard error of the mean of six mice/group. *P < 0·05. (c) Bone marrow-derived macrophages (5 days post-harvest) were infected with AdC7OprF, AdC7RGD.OprF, Ad5OprF and Ad5RGD.OprF (104 particles per cell). OprF expression was evaluated in cell lysate by Western analysis using anti-OprF serum.
Fig. 6
Fig. 6
Presence of adenovirus (Ad)C7 genome in the lung and mediastinal lymph nodes 2, 7 and 14 days following intratracheal immunization with AdC7 outer membrane protein F (OprF). arginine–glycine–aspartic acid (RGD), AdC7OprF, Ad5OprF, Ad5OprF.RGD (all 104 particles per cell). (a,b) Adenoviral DNA, evaluated by Taqman real-time polymerase chain reaction in (a) total lung tissue and (b) mediastinal lymph nodes. Data are shown as means ± standard error of the mean of three mice/group. *P < 0·05, **P < 0·02 and ***P < 0·01, comparing AdC7OprF and AdC7OprF.RGD vectors to Ad5OprF and Ad5OprF.RGD vectors.
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References

    1. Tenover FC. Mechanisms of antimicrobial resistance in bacteria. Am J Infect Control. 2006;34:S3–10. - PubMed
    1. Doring G, Pier GB. Vaccines and immunotherapy against Pseudomonas aeruginosa. Vaccine. 2008;26:1011–1024. - PubMed
    1. Holder IA. Pseudomonas immunotherapy: a historical overview. Vaccine. 2004;22:831–839. - PubMed
    1. Sedlak-Weinstein E, Cripps AW, Kyd JM, Foxwell AR. Pseudomonas aeruginosa: the potential to immunise against infection. Exp Opin Biol Ther. 2005;5:967–982. - PubMed
    1. Sharma A, Krause A, Worgall S. Recent developments for Pseudomonas vaccines. Hum Vaccin. 2011;7:999–1011. - PMC - PubMed

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