Chris Minion at Iowa State University for providing the BioRobotics MicroGRID II printer used for printing microarray slides. largely evidenced by its history of such use [2]. Though the United States and 103 other countries co-signed an agreement to terminate biological weapons programs in 1972, remains listed as a Tier 1 Select Agent. Currently, there is no FDA-approved vaccine against to civilian or military populations. As is predominantly an extracellular pathogen, many vaccination strategies have focused on developing strong humoral immunity, characterized by neutralizing antibodies against the V antigen and opsonizing antibodies against the F1 capsule [5C8]. In mice, anti-F1 IgG antibody titers 100,000 have been shown to correlate with protection [5]. Additionally, monoclonal antibody (mAb) 7.3, sodium 4-pentynoate which binds to the V antigen, has been shown to neutralize in vitro and provides passive protection against lethal challenge [6, 9]. The United States Army Medical Research Institute of Infectious Diseases (USAMRIID) developed a recombinant fusion protein, F1-V, comprised of full length F1 and V antigens, and shows promise as a target antigen for [10C13]. This vaccine has long shown promise as a vaccine candidate against both pneumonic and bubonic plague in rodents [10C12]. In fact, it was recently shown that F1-V adjuvanted with aluminum hydroxide (alum) using an IM/SC prime-boost regimen provided complete protection against intranasal challenge with virulent CO92 in mice, guinea pigs, and macaques [13]. Additionally, serum collected from immunized macaques conferred passive protection to mice. Despite such success, this vaccine formulation is most often administered as a prime-boost regimen; in the event of mass exposure, it would be ideal to use single dose formulations that can provide rapid immunity in as short a time frame as possible and that can maintain long-lived protective immunity. Cyclic dinucleotides (CDNs), a class of Rabbit Polyclonal to Pim-1 (phospho-Tyr309) small molecule adjuvants, are recognized as microbial-associated molecular patterns (MAMPs) by the pattern recognition receptor (PRR) STING (STimulator of INterferon Genes) [14], resulting in the phosphorylation of transcription factors NFfor at least 280 days post-immunization (DPI) [37]. In addition to long-lived protective immunity, it is also important that vaccines induce rapid (i.e., 14 days) protective immunity to counter acute outbreaks of disease. Designing next-generation vaccine platforms that provide rapid and long-lived immunity against highly lethal pathogens will likely require novel approaches sodium 4-pentynoate including developing new adjuvants or vaccine regimen with combination adjuvants to enhance both rapid and long-lived protective immunity. Herein, the design and evaluation of a combination nanovaccine, comprising of F1-V-containing polyanhydride nanoparticles and a non-canonical CDG CDN adjuvant (containing 2,5C3 C5 phosphate linkages) is described. This nanovaccine formulation synergistically combines the adjuvant properties of polyanhydride nanoparticles and CDNs to induce rapid immune responses and facilitate the induction of long-lived protective immunity against pneumonic plague. 2.?Material and methods 2.1. Materials sodium 4-pentynoate Chemicals used for CPTEG and CPH diacid and polymer synthesis included 1,6-dibromohexane, triethylene 4-fusion protein F1-V (NR-4526) were obtained from the Biodefense and Emerging Infections Repository (Manassas, VA). 2.2. Polyanhydride synthesis CPTEG and CPH diacids were synthesized as previously described [28, 35]. 20:80 CPTEG:CPH copolymer synthesis was performed using melt polycondensation [28]. Copolymer composition and molecular weight were approximated using end group evaluation of 1H NMR (DXR 500) spectra. 2.3. Nanoparticle synthesis 10% (w/w) F1-V-loaded 20:80 CPTEG:CPH nanoparticles had been synthesized using display nanoprecipitation, as described [30] previously. Quickly, F1-V and 20:80 CPTEG:CPH copolymer was dissolved in methylene chloride at 2 mg/mL and 20 mg/mL, respectively, sonicated at 30 Hz for 30 s around, and poured into pentane chilled to ?20 C at.