pestis antigens (Ags), the outer capsule protein (F1-Ag), which is believed Luminespib nmr to help avoid phagocytosis [4] and [5], and the low calcium response (LcrV) protein, V-Ag, which has been implicated in mediating a suppressive effect upon Th1 cells via the stimulation of IL-10 [6]. These individual vaccine candidates are protective against bubonic and pneumonic plague [7] and [8]; however, these vaccines, when applied in combination or in a fusion form, act synergistically in conferring

protection [9], [10], [11] and [12]. Although the observed protective immunity is largely Ab-dependent, Y. pestis is an intracellular pathogen, and new data have shown that during early infection events cellular immunity can contribute to effective CP-673451 ic50 protective immunity against plague [13], [14] and [15]. Lymphotactin (LTN; XCL1) is a member of the chemokine superfamily and classified into the C chemokine family as a single C motif-1 chemokine in both mice and humans [16] and [17]. LTN is produced mainly by CD8+ T cells and NK cells and has chemotactic

activity for lymphocytes, CD4+ and CD8+ T cells, and NK cells upon binding to its specific receptor, XC chemokine receptor-1 (XCR1) [18], [19], [20], [21] and [22]. In addition, Boismenu et al. reported that TCRγδ TCR+ intraepitheral lymphocytes (IELs) also produce LTN and induce innate and adaptive immunity via chemotaxis for T cells and NK cells [19] and [23]. Thus, we hypothesize below that LTN can enhance recruitment of lymphocytes to react to the encoded plague DNA vaccines, which should result in improved vaccine efficacy when given either by the mucosal or parenteral routes similar to that previously shown [24]. To develop an effective vaccine

against pneumonic plague, we constructed LTN-based DNA vaccines that co-express V-Ag or F1-V fusion protein, using a bicistronic eukaryotic expression vector, and assessed their vaccine efficacy against pneumonic plague challenge. This is the first example of using an immunization approach with LTN DNA vaccines for plague. These DNA vaccines did effectively prime and, with subsequent nasal F1-Ag protein boosts, were able to confer variable protection against pneumonic plague. Thus, the LTN DNA vaccine can be used to prime for protection against plague. To develop the lymphotactin (LTN) DNA vaccines, the LTN cDNA was PCR-amplified from pGT146-mLTN (Invivogen, San Diego, CA) as a template similar to that previously described [25]. Primers contained restriction sites for HindIII at the 5′-teminus and BamHI at the 3′-terminus. After TA cloning (TOPO cloning kit, Invitrogen Corp., Carlsbad, CA) and verification of the PCR products’ sequence, the LTN fragment was excised from the TA vector and inserted into the pBudCE4.1 vector (Invitrogen Corp.) cut with HindIII and BamHI, resulting in the plasmid pBud-LTN.