Protection by Interleukin-12 Adjuvanted Intranasal Whole-cell Killed Vaccine of Yersinia pestis Against Plague

is Mediated by Antibodies and Fc gamma Receptors

Devender Kumar and Dennis W. Metzger
Center for Immunology and Microbial Disease, Albany Medical College, Albany, NY 12208

CONCLUSIONS
We thank Michelle Wyland-O'Brien, Sharon Salmon, and Shannan Zilles for
assistance in the ABSL-3 lab.
We thank Yili Lin of the AMC Immunology Core for help with histology.
The authors received financial support from the DOD-ONR (Award no. N00014-06-
1-1176).
ABSTRACT RESULTS
Fig. 1. Intranasal vaccination with iYp vaccine plus IL-12
protects BALB/c mice from pneumonic plague
Fig. 2. 10
7
CFU of intranasal iYp vaccine have limited toxicity
Fig. 3. IL-12 enhances the antibody titers against intranasal iYp
whole-cell vaccine
METHODS
Vaccine: 0.25% paraformaldehyde inactivated Y. pestis CO92 (iYp)
Adjuvant: Interleukin-12
Animal model: BALB/c mice
Vaccine dose: 10
7
CFU
Vaccine route: Intranasal
Vaccine volume: 30 l/mouse
Vaccination protocol:
Intranasally-administered inactivated whole-cell Y. pestis can
be used as a safe and potent vaccine against primary
pneumonic plague in mice.
Vaccination generates robust levels of serum and lung
antibodies.
IL-12 is an effective mucosal adjuvant for intranasal
vaccination against pneumonic plague
Antibodies are capable of protection against pneumonic
plague and this protection is mediated through Fc receptors
CD4 and CD8 T-cells are dispensable for protection against
pneumonic plague when mice are vaccinated with whole-cell
killed vaccine.
Acknowledgements

Plague caused by Yersinia pestis is a rapidly developing and
devastating disease that causes 50-100% mortality when there is a
delay in treatment or no treatment. Y. pestis is classified as Category A
agent by Centers for Disease Control and Prevention and is also a re-
emerging infectious disease threat. With fulminate pathology followed
by rapid death, poor understanding of protective immunemechanisms,
and no vaccine available, the disease is a real hazard. The only
licensed inactivated Plague vaccine is unavailable as it did not provide
protection against pneumonic plague and was reactogenic. F1/LcrV-
based vaccines provide protection in cynomolgus macaques, but not in
African green monkeys for unknown reasons, and there is skepticism
whether these vaccines will provide protection in humans.
We have evaluated an intranasal whole killed cell vaccine (iYp) of
Y. pestis CO92 strain adjuvanted with Interleukin-12 (IL-12) for safety,
immunogenicity, efficacy, and mechanisms of immunity in a mouse
model of pneumonic plague. Intranasal delivery of 10
7
CFU of iYp +
0.5g of IL-12 on 0 and 21 days protected 100% of mice against a
lethal intranasal Y. pestis CO92 challenge. The iYp + IL-12 was well
tolerated, safe, immunogenic, and efficacious in mice by intranasal
route and did not cause systemic adverse reactions. The levels of
serum and lung antibodies against Y. pestis correlated with increased
survival, decreased bacterial burdens, and protection from lung
pathology. Protection mediated by iYp + IL-12 was not dependent on
CD4 and CD8 T-cell subsets as depletion of these cells before
challenge did not reduce survival of vaccinated mice. Fc gamma
receptor (FcR) KO mice had reduced protection against plague
following vaccination with iYp + IL-12. Furthermore, protection was
induced in nave mice by passive transfer of immune sera, and
protection was lost in FcR KO mice. Our study shows a vital role for
antibodies in protection against plague. The study suggests the
existence of novel protective molecules in iYp that could ultimately lead
to development of new generation plague vaccine.
INTRODUCTION
Plague is an exceptionally virulent and zoonotic disease.
Y. pestis is classified as a Category A select agent by the CDC, and
could to be used as a biological weapon.
Pneumonic plague has a high fatality rate even if antibiotic treatment
is started within 20 hours of infection.
Plague vaccine USP does not prevent primary pneumonic plague in
humans or mice.
No effective vaccine is available.
Interleukin-12 (IL-12) is an extremely potent mucosal adjuvant that
enhances humoral and cell-mediated immunity.
Immune mechanisms responsible for protection are still unknown.
A.









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C.
Fig. 4. Survival induced by iYp plus IL-12 is correlated with
reduced bacterial organ burden.
Fig. 5. Survival of iYp plus IL-12 vaccinated mice correlates
with reduced lung pathology.
iYp + IL-12 PBS
Fig. 6. FcR are required for protection against pneumonic
plague in passive and active immunity.

A.









B.
Fig. 7. Depletion of CD4 and / or CD8 T cells does not affect
survival of iYp plus IL-12 vaccinated mice
Unstained Rat IgG treatment
Anti-mCD4 GK 1.5
+
Anti-mCD8 53-6.72
S
p
l e
e
n
L
u
n
g
CD8 Tri-color (CT-CD8a)
C
D
4
P
E
- C
y
7
( R
M
4
- 5
)22.29 %
9.33 %
0.29 %
0.64 %
22.25 %
8.58 %
0.31 %
0.98 %
B.


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