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本帖最后由 风雨不动 于 2012-4-14 15:03 编辑
ABSTRACT 72
Novel dendritic cell receptor-targeted
HBV therapeutic vaccine induces robust
immune responses in HBV chronic
carrier peripheral blood mononuclear
cells ex vivo
RE Déry1, A Ma1, D Wang1, Y Xia1, K Gutfreund2, and
R George1
1 Paladin Biosciences, a Division of Paladin Labs Inc.,
Edmonton, Alberta, Canada; 2 Department of Medicine,
University of Alberta, Edmonton, Alberta, Canada
BACKGROUND: Exposure to viral antigens during
chronic HBV infection leads to functional impairment
or deletion of virus-specific T cells. Therapeutic
vaccines that induce strong host immune responses
may therefore resolve chronic HBV infection. We
developed a therapeutic HBV vaccine which fuses
HBV S1, S2, Core and murine Fc fragment. We have
previously shown that this vaccine targets specific
receptors on dendritic cells (DCs) and elicits ex vivo
antigen-specific cytotoxic and humoral immune
responses in human peripheral blood mononuclear
cells (PBMCs). In addition, it has been shown that the
vaccine generates robust humoral and cell-mediated
immune responses in vivo in animals. The current
study evaluates the ability of the vaccine to elicit
antigen-specific T cell responses in PBMCs derived
from patients with chronic HBV infection.
METHODS: The vaccine was expressed in Sf9 insect
cells and purified by affinity chromatography. Whole
PBMCs or purified T cells from chronically HBV infected
donors were activated ex vivo by co-culturing
with vaccine-loaded mature DC (mDC). Activated
T-cells were re-stimulated using either HBV S1/S2/
Core overlapping peptides or fresh vaccine-loaded
mDC. Resulting T-cell proliferation and production of
IFN-γ, TNF-α, and Granzyme B were measured by flow
cytometry. The HBV-specific cytotoxic lymphocyte
(CTL) response was measured in chronically HBV infected
patient PBMC derived mDC and T-cells ex
vivo, using the real-time cell microelectronic sensor
(RT-CES) system. Vaccine-loaded mDCs were used as
antigen-presenting cells when co-cultured with naïve T
cells, or as target cells when co-cultured with vaccine primed
T cells.
RESULTS: Primary stimulation of chronically HBV infected
donor T cells with vaccine-loaded mDC
resulted in both CD4+ and CD8+ T cell proliferation.
Re-stimulation of vaccine-activated T cells with HBV
S1/S2/Core overlapping peptides induced IFN-g
production in both CD4+ and CD8+ T cells. Restimulation
of activated chronically HBV-infected T
cells with vaccine-loaded mDC significantly increased
IFN-g and TNF-a production. Interestingly, vaccine activation
of PBMCs from chronically HBV-infected
patients stimulated Granzyme B production by
CD4+CD25- cells (Tresp), accompanied by an increase
in Annexin V+ staining on CD4+CD25+ cells (Tregs). In
our dynamic CTL killing assay, co-culture of vaccine activated
HBV chronic carrier T cells with mDCs
resulted in a dose, time and T cell-dependent decrease
in DC viability.
CONCLUSION: The vaccine-dependent expansion
of IFN-g, TNF-a producing CD8+ and CD4+ T cells,
Tresp-induced apoptosis of Tregs in chronically HBVinfected
donor PBMCs as well as CTL killing of target
mDC by vaccine-activated HBV infected donor T cells
suggests that the vaccine can break HBV tolerance
in chronic HBV carriers by inducing strong immune
responses which include functional activation of
Tresp-dependent apoptosis of Tregs. Thus, our HBV
therapeutic vaccine may be a good candidate for the
treatment of chronic HBV infections.
(Financial support from NRC-IRAP Canada and Alberta
Innovates Technology Futures is gratefully acknowledged.)
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发表于 2011-12-16 16:32
