Our laboratory has been investigating the immunobiology of the Neisserial porin PorB for the last two decades. We first began by investigating the use of this protein as a potential anti-Neisserial candidate, but through this work we found that this protein had unique immune stimulatory abilities above and beyond its own potential use as a vaccine. We found that PorB can be used as an immune adjuvant for vaccines and that this ability was directly related to its ability to stimulate antigen presenting cells (APC) and increase expression of the costimulatory molecule CD86 and class II MHC. Moreover, we found that the ability of PorB to stimulate and activate antigen presenting cells was directly due to an interaction with the pattern recognition receptor, TOLL-like Receptor 2 (TLR2) and requires the TLR adaptor protein MyD88. We have further shown, as described in the preliminary data section and in that TLR2 needs to heterodimerize with TLR1 (and not TLR6) in order for PorB to stimulate these APCs. Utilizing this body of information, the next logical step for studying the immune stimulating adjuvant activity of PorB is to compare this activity to other known vaccine adjuvants. This will allow for the better use of PorB as adjuvant, by understanding what type of immune response it induces as compared to these other adjuvants. Moreover, we believe it is essential to continue to investigate the adjuvant potential of PorB, as it is currently the only TLR2 ligand being investigated as a vaccine adjuvant. We shall compare the immune stimulation by PorB to panel of other adjuvants, including the only licensed adjuvant, Alum (inflammasomes mediated immune activation, other TLR ligands, including CpG DNA (TLR9), and monophosphoryl lipid A (MPL)(TLR4), and QS-21. We shall examine the ability of these adjuvants to induce immune responses to a test antigen, ovalbumin, which we have used previously as described in the Preliminary Data section. We shall measure antibody levels induced, isotypes and IgG subtypes induced, antibody affinity, CD4 T cell response and ability to induce APC migration and lymph node specific T cell stimulation. Moreover, we shall compare the ability of these different adjuvants to protect mice from infection with Listeria which secretes OVA. This is a well characterized model and has been used previously to demonstrate that the presence of OVA specific CD4 and CD8 T cells are necessary for protection of mice from infection with this pathogen when it is constructed to secrete OVA. We shall obtain this strain from Dr. Darren Higgins in the Dept. of Microbiology at the Harvard Medical School, who has significant expertise in this model and will act as a consultant on this proposal. Even though the protection induced by most licensed vaccines are due to the induction of neutralizing or functional antibodies, we feel it would extremely important to begin to definitively compare the ability of these adjuvants, including PorB, to not only induce functional antibodies, but to potentially induce a protective CD8 CTL response.

Public Health Relevance

We have shown that the Neisserial porin, PorB, is a potent immune adjuvant and works via recognition by TLR2 and induction of antigen presenting cell activation. This proposal will examine the adjuvant activity of PorB and compare this activity to other currently investigated adjuvants (including Alum, CpG DNA, MPL and QS-21). We shall examine induction of both humoral responses and CD8 T cell responses and the ability of the vaccines to prevent Listeria infection in mice. Moreover, we shall further examine the mechanism of PorB adjuvant activity via the use of MyD88 floxed mice bred with mice expressing cell specific cre recombinase.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
5R01AI040944-17
Application #
8521047
Study Section
Vaccines Against Microbial Diseases (VMD)
Program Officer
Leitner, Wolfgang W
Project Start
1997-04-01
Project End
2015-08-31
Budget Start
2013-09-01
Budget End
2014-08-31
Support Year
17
Fiscal Year
2013
Total Cost
$530,513
Indirect Cost
$216,600
Name
Boston Medical Center
Department
Type
DUNS #
005492160
City
Boston
State
MA
Country
United States
Zip Code
02118
Kattner, Christof; Toussi, Deana N; Zaucha, Jan et al. (2014) Crystallographic analysis of Neisseria meningitidis PorB extracellular loops potentially implicated in TLR2 recognition. J Struct Biol 185:440-7
Massari, Paola; Wetzler, Lee M (2012) Analysis of parameters associated with prevention of cellular apoptosis by pathogenic Neisseriae and purified porins. Methods Mol Biol 799:319-41
Massari, Paola; Gunawardana, Jay; Liu, Xiuping et al. (2010) Meningococcal porin PorB prevents cellular apoptosis in a toll-like receptor 2- and NF-kappaB-independent manner. Infect Immun 78:994-1003
Wetzler, Lee M (2010) Innate immune function of the neisserial porins and the relationship to vaccine adjuvant activity. Future Microbiol 5:749-58
Liu, Xiuping; Wetzler, Lee M; Nascimento, Laura Oliveira et al. (2010) Human airway epithelial cell responses to Neisseria lactamica and purified porin via Toll-like receptor 2-dependent signaling. Infect Immun 78:5314-23
MacLeod, Heather; Bhasin, Navneet; Wetzler, Lee M (2008) Role of protein tyrosine kinase and Erk1/2 activities in the Toll-like receptor 2-induced cellular activation of murine B cells by neisserial porin. Clin Vaccine Immunol 15:630-7
Liu, Xiuping; Wetzler, Lee M; Massari, Paola (2008) The PorB porin from commensal Neisseria lactamica induces Th1 and Th2 immune responses to ovalbumin in mice and is a potential immune adjuvant. Vaccine 26:786-96
Ganley-Leal, Lisa M; Liu, Xiuping; Wetzler, Lee M (2006) Toll-like receptor 2-mediated human B cell differentiation. Clin Immunol 120:272-84
Agarwal, Sarika; King, Carol A; Klein, Ellen K et al. (2005) The gonococcal Fur-regulated tbpA and tbpB genes are expressed during natural mucosal gonococcal infection. Infect Immun 73:4281-7
Massari, Paola; King, Carol A; MacLeod, Heather et al. (2005) Improved purification of native meningococcal porin PorB and studies on its structure/function. Protein Expr Purif 44:136-46

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