We evaluated the safety and efficacy of the mast cell activator compound 48/80 (C48/80) when used as an adjuvant delivered intradermally (ID) with recombinant anthrax protective antigen (rPA) in comparison with two well-known adjuvants. Mice were vaccinated in the ear pinnae with rPA or rPA + C48/80, CpG oligodeoxynucleotides (CpG), or cholera toxin (CT). All adjuvants induced similar increases in serum anti-rPA IgG and lethal toxin neutralizing antibodies. C48/80 induced a balanced cytokine production (Th1/Th2/Th17) by antigen-restimulated splenocytes, minimal injection site inflammation, and no antigen-specific IgE. Histological analysis demonstrated that vaccination with C48/80 reduced the number of resident mast cells and induced an injection-site neutrophil influx within 24 hours. Our data demonstrate that C48/80 is a safe and effective adjuvant, when used by the intradermal route, to induce protective antibody and balanced Th1/Th2/Th17 responses. KeywordsCompound 48/80; adjuvant safety; intradermal vaccination IntroductionImmunization programs have led to the eradication of smallpox, the near eradication of polio, and the control of other infectious diseases, including measles, mumps, rubella, and diphtheria. While there are many success stories, HIV, pandemic Influenza, and a variety of emerging Corresponding author: Herman F. Staats, Ph.D., Department of Pathology, Box 3712, DUMC, Durham, NC 27710, Telephone: 919-684-8823, herman.staats@duke.edu. Publisher's Disclaimer: This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final citable form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain. NIH Public Access Author ManuscriptVaccine. Author manuscript; available in PMC 2010 June 2. NIH-PA Author ManuscriptNIH-PA Author Manuscript NIH-PA Author Manuscript infectious diseases (West Nile Virus, Dengue, etc.) provide active reminders that safe and effective vaccines continue to be needed to combat infectious diseases.Adjuvants are substances that, when combined with vaccine antigens, enhance the induction of the desired immune responses [1,2]. For example, the addition of adjuvants to vaccines may increase vaccine potency by enhancing the magnitude of antibody or cellular responses induced, reducing the time to seroprotection or selectively inducing CD4+ Th1, Th2, Th17, or CD8+ T cell responses [1,2]. The mechanism of action of adjuvants varies depending on the adjuvant used, but the end result is thought to include the activation and migration of dendritic cells as well as the expression of antigen presenting molecules, providing superior induction of antigen-specific T and B cell responses [1,2]. Despite increasing information on the mechanism of action ...

Recent studies have reported that mast cells play a role in the trafficking of dendritic cells and lymphocytes to lymph nodes draining sites of infection. To test the hypothesis that mast cell activators could be used as vaccine adjuvants, C3H/HeN mice were intradermally vaccinated in the ear on days 0 & 21 with 0.5 μg recombinant anthrax protective antigen (rPA) alone or combined with 30 μg compound 48/80 (C 48/80), a mast cell activator. CpG oligodeoxynucleotide (CpG ODN, 1 μg) was used as a control. Ear swelling was monitored 24 hours after immunization to measure injection site inflammation. Mice immunized with rPA alone had an average ear thickness (mm) of 0.0059 while mice immunized with rPA + C 48/80 had an ear thickness of 0.0426 (p < 0.001 vs rPA alone). Mice immunized with rPA + CpG ODN had an ear thickness of 0.1519 (p < 0.001 vs rPA), significantly greater than swelling induced by C 48/80 (p < 0.001). Day 42 serum from mice immunized with PA alone had serum anti-PA IgG titers of 1:27,238 while mice immunized with PA + C 48/80 had anti-PA titers of 1:10,568,984 (p < .001 vs PA alone). Similar titers were detected in mice vaccinated with rPA plus CpG ODN. Antibodies induced with rPA + C 48/80 neutralized anthrax lethal toxin. Our findings suggest that mast cell activators provide effective vaccine adjuvant activity when delivered intradermally while inducing less injection site inflammation than CpG ODN.