| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
|
|
|
|||||||
|
|||||||||
*
Institute of Medical Microbiology, Immunology and Hygiene, Technical University of Munich, Munich, Germany;
Institute of Immunology, Vilnius, Lithuania; and
Department of Immunology, Institute of Medical Microbiology and Hygiene, University of Freiburg, Freiburg, Germany
 |
Abstract |
|---|
 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
|
Introduction |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
Alternatively, a CD4 Th-independent pathway of DC activation exists that operates via recognition of molecular patterns associated with microbial pathogens. Pathogen derived molecular pattern ligands include bacterial cell wall components like LPS (11), double-stranded RNA (12), and bacterial CpG-DNA (13). As member of the innate immune system, DC express pattern recognition receptors that on binding of pathogen-derived ligands initiate signaling cascades activating the DC to provide T cell costimulation and cytokine release (11). Prototypic for pattern recognition are responses to LPS, which triggers a signaling complex when toll-like receptor 4 or toll-like receptor 2 interacts with LPS bound to CD14 (14, 15). Professional APC function of DC may thus be induced indirectly via CD40-CD40L engagement with Th cells or directly via recognition of pathogen-derived molecular pattern ligands.
CpG-DNA serves as a profound Th1-polarizing adjuvant for proteinaceous Ags (16, 17, 18, 19). In addition to strongly promoting humoral responses, CpG-DNA was shown to allow priming of CTL specific for exogenously given Ags (20, 21). The pronounced adjuvanticity of CpG-DNA may be, at least in part, explained by its ability to directly activate immature DC to transit to professional APC (22, 23). CpG-DNA triggers DC to up-regulate costimulatory molecules (CD40, CD80, CD86), MHC class II, and to produce cytokines such as IL-12 both in vitro (22) and in vivo (23).4
Here we addressed the question of whether MHC class I-restricted CD8 T cell peptides may be rendered immunogenic when presented in vivo by CpG-DNA-activated DC. By infecting immunized H-2b mice with lymphocytic choriomeningitis virus (LCMV), we tested whether induced CTL responses are paralleled by protective antiviral immunity. To clarify the mechanisms of CpG-DNA adjuvanticity, we analyzed the fate of OVA-derived peptide SIINFEKL in vivo and report here the strikingly exclusive ability of DC from draining lymph nodes (LN) to present the injected peptide. Finally, we analyzed cellular requirements for CpG-DNA-mediated CTL induction and describe that CpG-DNA allows Th cell-independent CTL responses to MHC class I-restricted T cell epitopes.
|
Materials and Methods |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
L-Phosphatidylcholine (type XI-E), L-phosphatidyl-DL-glycerol, cholesterol, and IFA were purchased from Sigma (Munich, Germany). OVA peptide 257–264 (SIINFEKL) and LCMV glycoprotein 33–41 peptide (KAVYNFATM = gp33) were custom synthesized by Research Genetics (Huntsville, AL). For technical reasons (to prevent dimer formation), the original cysteine at the anchor position 41 in the gp33 had been replaced by methionine. Phosphorothioate-stabilized oligodeoxynucleotides (ODN) were synthesized by TibMolBiol (Berlin, Germany). CpG-ODN (=ODN 1668, containing a CG-motif marked with bold letters: 5'-TCC-ATG-ACG-TTC-CTG-ATG-CT) and control GpC-ODN (inverted CG = ODN 1720: 5'-TCC-ATG- AGC-TTC-CTG-ATG-CT) were taken from Ref. 24 . Human rIL-2 was donated by Eurocetus (Amsterdam, The Netherlands).
Mice
Female C57BL/6 mice were purchased from Harlan Winkelmann
(Borchen, Germany). CD40L knockout mice (C57BL/6J-Tnfsf5<tm1Imx>)
were bought from The Jackson Laboratory (Bar Harbor, ME) and MHC
class II-deficient (B6
A
0/A0) mice
(25) were kindly provided by Dr. I. Förster
(Technical University Munich, Munich, Germany). All animals were housed
under specific pathogen-free conditions and were used at 8–12 wk
of age.
Cell lines and in vitro culture medium
EL-4 (H-2b) thymoma cells were purchased from American Type Culture Collection (Rockville, MD). B3Z, a somatic T cell hybrid generated by fusing the OVA/Kb-specific cytotoxic clone, B3, with a lacZ-inducible derivative of BW5147 fusion partner (26), was kindly provided by Dr. B. L. Kelsall (National Institutes of Health, Bethesda, MD). Cells were cultured in Clicks/RPMI (Biochrom, Berlin, Germany) supplemented with 10% (v/v) heat-inactivated FCS (Biochrom), 5 x 10-5 M 2-ME, and antibiotics (100 IU/ml penicillin G and 100 IU/ml streptomycin sulfate) at 37°C and 5% CO2.
Peptide-liposome formation
Liposomes were manufactured by a rehydration entrapment method (27). Briefly, 18 mg phosphatidylcholine, 2 mg phosphatidylglycerol, and 5 mg cholesterol (2:0.2:1) were suspended in 5 ml chloroform in a 250-ml round bottom flask. The mixture was rotary evaporated under reduced pressure until a thin lipid film formed on the flask wall. Residual chloroform was removed by vacuum desiccation. The peptide (3 mg) was solubilized in a minimal volume of water and diluted to 1 ml with PBS. This solution was slowly added to the dried lipid and hand shaken until the lipids were resuspended. The mixture was then filter extruded through a 0.2-µm pore size Anotop 10 Plus syringe mount filter (Whatman, Maidstone, England).
Immunization and 51Cr release assay
For priming of cytotoxic T cells, 0.3 mg of respective peptide
entrapped in liposomes or solubilized in PBS were injected into both
fore footpads of mice. As an adjuvant CpG-ODN was coinjected (10
nmol/mouse). After 4–5 days, draining LN were removed and a
single-cell suspension was prepared by pressing the LN through a
screen. LN cells were cultured at 1–2 x
106/ml in 24-well plates in medium conditioned
with 10 U/ml rIL-2 for additional 4–5 days, and then
51Cr release assay was performed. For in vitro
restimulation of primed lymphocytes 2 wk after immunization, spleen
cell suspensions were prepared (5 x 106/ml)
and cocultured with SIINFEKL-labeled (0.5 µM) irradiated (15 Gy)
spleen cells (2 x 106/ml) with 10 U/ml
rIL-2 for 7 days, and then 51Cr release assay was
performed. In some experiments, CD4- or CD8-positive T cells were
depleted from the expanded lymphocyte population before measuring lytic
activity. This was done by labeling lymphocytes with magnetic beads
coated with the anti-CD4 or anti-CD8 Abs (Dynal, Hamburg,
Germany) allowing negative selection of the CD4+
or CD8+ T cell subset, respectively. The
efficiency of negative selection was >90% as judged by FACS analysis.
The 51Cr release assay was performed as follows.
EL-4 target cells (2 x 106) were labeled
with 200 µCi
Na251CrO4
(Amersham Pharmacia Biotech, Freiburg, Germany) for 1 h at 37°C
and washed; one-half of 51Cr-labeled cells was
incubated with peptide solution (0.1 µM SIINFEKL or 1 µM KAVYNFATM)
for an additional 30 min. Peptide-untreated cells served as a
specificity control. After extensive washing, 100 µl
(103 cells) of target cells were added to the
same volume of replicate serial dilutions of effector CTL. After 4
h of incubation at 37°C and 5% CO2, 100 µl
of culture supernatant were removed from each well and 
-irradiation
was measured. Specific lysis was calculated according to the formula:
% specific lysis = [cpm (sample) - cpm (spontaneous
release)/cpm (maximal release) - cpm (spontaneous release) ] x
100. Spontaneous release ranged between 5 and 15%.
Preparation of DC
For preparation of DC, draining LN were removed and collected into ice-cold HBSS (Life Technologies, Karlsruhe, Germany). LN were digested for 1 h at 37°C using collagenase type Ia purchased from Sigma. Single-cell suspensions were prepared, and clumps were removed using a 100-µm pore size filter (Becton Dickinson, Heidelberg, Germany). Immediately after collagenase treatment LN cells were washed in Ca2+-free HBSS; from this point onwards the cell suspension was always handled in buffers containing 2 mM EDTA. For positive selection of dendritic cells, single-cell suspensions were incubated with magnetic beads coated with the anti-CD11c Ab (Miltenyi, Bergisch Gladbach, Germany). Selection of DC was performed using MS+ separation columns and MiniMACS according to the protocol supplied by the manufacturer (Miltenyi). The selection ensued >80% pure CD11c+ cell population as judged by FACS analysis.
Presentation assay
Presentation of SIINFEKL was assayed by measurement of induced lacZ activity in SIINFEKL/Kb-specific T cell hybridoma (B3Z) transfected with a LacZ reporter under the transcriptional control of IL-2 gene promoter elements (26) after coincubation with subpopulations of LN cells. Briefly, 12 and 24 h after SIINFEKL injection, the draining LN were harvested, and LN cells were separated into CD11c+ and CD11c- fractions as described above. Graded numbers of fractionated cells were incubated with 104 B3Z cells/well in 96-well plate at 37°C/5% CO2 overnight. On the next day, individual cultures were washed with 100 µl PBS and lysed by addition of 100 µl Z buffer (100 mM 2-ME, 9 mM MgCl2, 0.125% Nonidet P-40, 0.15 mM chlorophenol red ß-galactoside (Calbiochem, San Diego, CA) in PBS). After 4 h incubation at 37°C, 50 µl stop buffer (300 mM glycine and 15 mM EDTA in water) were added to each well, and absorption of individual wells was read using a 96-well Emex plate reader (Molecular Devices, Sunnyvale, CA). The absorption wavelength was 570 nm, with 650 nm as the reference wavelength. Induction index was calculated by dividing induced activity by background.
Flow cytometry
Cells were washed in PBS containing 2% FCS (PBS/FCS) and first
incubated for 10 min at 4°C with anti-FcRII/III mAb to block
unspecific binding of the following Ab reagents. All mAbs used were
purchased from PharMingen (Hamburg, Germany). FITC- and PE-labeled mAbs
(used at 5–20 µg/ml) included Abs against MHC class II (clone 2G9),
CD11c (clone N418), CD80/B7-1 (clone 1G10), CD86/B7-2 (clone GL1), and
CD40 (clone 3/23). Isotype controls included purified rat IgG2a and
hamster IgG. After incubation with mAbs for 30 min at 4°C, cells were
washed with PBS/FCS. FACS analysis was performed on a flow cytometer
FACSCalibur (Becton Dickinson), acquiring 10,000 events. FACS data were
analyzed using CellQuest FACS software.
Protection of mice from replication of LCMV
Mice were immunized once s.c. at the tail base with 0.3 mg gp33 plus 10 nmol CpG-ODN 1668, gp33 emulsified in IFA, CpG-ODN 1668 alone, or gp33 alone or were mock treated. Two weeks later, mice were infected i.v. with 200 PFU of LCMV, and viral titers in the spleens were determined after 4 days in a virus plaque assay, as described previously (28). LCMV was quantified with an immunological focus assay in 24- or 96-well plates. The LCMV-WE virus strain used in this study was originally obtained form Rolf Zinkernagel (University Hospital Zurich, Zurich, Switzerland) and was grown on L929 fibroblast cells with a low multiplicity of infection.
|
Results |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
The use of CpG-DNA as an adjuvant appears to function by virtue of
its direct DC-activating qualities (22, 23). We were
interested to determine whether CpG-ODN-mediated activation of DC in
vivo would render MHC class I-restricted peptides immunogenic for CD8
CTL precursors. C57BL/6 mice were challenged with SIINFEKL, either
encapsulated in liposomes or as an aqueous solution, together with the
CpG-ODN 1668 or the control GpC-ODN 1720 (inverted CG). Mice
immunized with either SIINFEKL or CpG-ODN 1668 alone served as
controls. Fig. 1
A depicts a
representative experiment of at least four performed to show that
immunization with SIINFEKL, encapsulated in liposomes or even as an
aqueous solution, generated peptide-specific CTL in the draining LN
provided the inoculum contained CpG-ODN as an adjuvant. LN cells of
mice immunized with SIINFEKL alone, SIINFEKL plus GpC-ODN 1720 or with
CpG-ODN 1668 alone failed to display CTL responses. Fig. 1B depicts a representative experiment of two performed to
show that SIINFEKL-specific cytolytic activity is mediated by
CD8+ T cells. These data suggested that in
draining LN of wild-type mice, the peptide SIINFEKL activates CD8 CTL
precursors in the presence of the immunostimulatory CpG-ODN
1668.
|
A, within 24 h LN
draining the injection site developed peptide-specific CTL which could
be detected after in vitro IL-2-driven expansion. Activated CTL
precursors were present in draining LN for up to 9 days. Based on
comparison of lytic units generated per culture, activation of CTL
precursors peaked at days 3–4 (data not shown).
|
B, strong SIINFEKL-specific lytic activity was generated,
implying systemic dissemination of activated CTL. This conclusion is
also supported by the induction of protective immunity toward LCMV
after immunization with an MHC class I peptide from LCMV (see
below). Finally, we tested the effective dose range of the adjuvant CpG-ODN 1668. SIINFEKL-specific CTL could be generated in the draining LN in mice challenged locally with Ag plus 10–1 nmol CpG-ODN 1668 (data not shown).
Immature DC constitutively present SIINFEKL whereas CpG-DNA activates Ag-presenting DC
In situ, DC are believed to initiate primary CTL responses in a
two-step process. After sampling and processing Ag as immature DC, they
must subsequently mature into professional APC via activation
(7). DC activation may be brought about by engagement with
CD40L-positive Th cells (8, 9, 10) or via direct recognition
of pathogen-derived molecular pattern ligands (11). We
analyzed whether immature DC within draining LN take up SIINFEKL and
present the peptide in a H-2Kb-restricted
fashion. CD11c+ DC from LN draining a local site
of SIINFEKL challenge were positively selected and probed for
H-2Kb-restricted peptide presentation. This was
accomplished by using the SIINFEKL/Kb-reactive T
cell hybridoma B3Z, which is transfected with a LacZ reporter under the
transcriptional control of IL-2 gene promoter elements
(26). Within 12 h after s.c. challenge with soluble
SIINFEKL, positively selected DC from draining LN effectively presented
SIINFEKL, in that the T cell hybridoma became activated (Fig. 3). The DC minus LN cell population,
however, was strongly impoverished for the ability to activate the
hybridoma. By 24 h after injection, SIINFEKL presentation by DC
from the draining LN began to decline. After injection of CpG-ODN 1668
alone, DC were unable to activate the hybridoma. On the other hand,
challenge with SIINFEKL plus CpG-ODN 1668 generated slightly reduced
SIINFEKL presentation when tested after 12–24 h. Whether this reflects
activation of DC known to curtail the capacity of DC to take up
extracellular ligands (29) needs to be analyzed. These
data implied that SIINFEKL presentation was transient, DC derived, and
CpG-ODN independent.
|
). DC activation is CpG-motif dependent,
because ODN lacking a CpG-motif fail to activate DC (22, 23). Together, these data suggested that within the draining LN
immature DC constitutively take up and present SIINFEKL. It is the
CpG-DNA inoculation, however, that causes their activation and transit
to professional APC.
|
Activation of Ag-presenting immature DC can be brought about by
CD40L+ Th cells or by direct DC recognition of
pathogen derived ligands. We analyzed whether CTL induction by peptides
plus CpG-ODN was independent of T cell help. CTL induction by SIINFEKL
plus CpG-ODN 1668 was tested in wild-type C57BL/6 mice and compared
with that of haplotype matched
CD40L-/- mice or Th cell-deficient MHC class
II-/- mice. Fig. 5 shows that the magnitudes of SIINFEKL
specific CTL responses in CD40L-/- and MHC
class II-/- mice were similar to those in
H-2Kb wild-type mice. These data suggests that
CpG-DNA allowed induction of Th cell-independent CTL responses toward
MHC class I-restricted T cell peptides.
|
Murine LCMV infections are controlled by LCMV-specific CD8 CTL via
perforin-dependent cytolysis of virus-infected cells (30).
A major MHC class I-restricted T cell epitope for
H-2b mice is the aa 33–41 peptide (gp33) derived
from LCMV glycoprotein (31). This T cell epitope injected
in IFA has been shown to induce protective immunity (32).
We used this model system to determine whether CpG-ODN 1668 renders the
MHC class I-restricted peptide gp33 immunogenic for CTL and whether the
immunized mice are protected against LCMV infection. Draining LN of
C57BL/6 mice challenged s.c. with an aqueous solution of gp33 plus
CpG-ODN 1668 generated gp33-specific CTL (Fig. 6). The draining LN of mice immunized
with gp33 alone, however, failed to do so. These data demonstrate that
CpG-DNA used as an adjuvant conveyed immunogenicity to the MHC class
I-restricted T cell epitope gp33.
|
, mice immunized once with gp33 plus
CpG-ODN 1668 were protected from LCMV infection similar in magnitude to
mice immunized once with gp33 in IFA. Mice immunized with gp33 alone or
CpG-ODN 1668 alone, however, remained unprotected. These data
demonstrate that like IFA, CpG-DNA acts as an adjuvant for the T cell
peptide gp33 allowing for priming of CTL and induction of protective
antiviral immunity.
|
|
Discussion |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
MHC class I-restricted T cell epitopes (Ag) admixed with CFA or IFA
have been shown to prime peptide-specific CTL responses (32, 34), implying presentation of T cell epitopes by APC. To address
the question of whether upon s.c. challenge soluble T cell epitopes
become presented by DC within draining LN, DC from such LN were scored
ex vivo for H-2Kb-restricted peptide
presentation. Within 12 h after peptide challenge, positively
selected immature DC presented the OVA peptide SIINFEKL, activating the
SIINFEKL-specific, H-2Kb-restricted T cell
hybridoma, whereas the DC negative population did not (Fig. 3). Thus,
it appears that uptake and presentation of peptide are constitutive
functions of phenotypically immature DC (38). Others have
pointed out a MHC class I presentation pathway for cytosolic delivery
of exogenous Ags in DC (39, 40, 41). Injection of peptide
alone failed to induce productive CTL responses. Whether T cell
peptides presented by immature DC are ignored by Ag-reactive T cells or
induce tolerance (34, 42) needs to be analyzed. In
contrast, coadministration of CpG-DNA activated Ag-presenting immature
DC to transit to professional APC as defined by up-regulation of
costimulatory molecules (CD80, CD86, CD40) and MHC class II (Fig. 4).
The evidence for DC maturation is also supported by priming of
peptide-reactive precursor CTL (Fig. 1) and by induction of cytokine
production.4 The effects of CpG-ODN on immature DC within
LN were CpG-specific, because control ODN were inactive.
Recently, a pivotal role of CD40 ligation in DC activation has been identified (6, 7, 43). CD40 ligation by CD40L-positive Th cells or cross-linking by anti-CD40 mAb caused Ag-presenting immature DC to transit to professional APC. Once matured DC could activate CTL precursors in a Th cell-independent fashion (8, 9, 10). Here we describe that similar to CD40 ligation, CpG-DNA also activates in situ Ag-presenting immature DC to transit to professional APC. There are additional parallels between DC CD40 ligation and pathogen pattern recognition receptor engagement. Pathogen-derived ligands like CpG-DNA and LPS or Th cell-dependent CD40L-CD40 interaction initiate in APC signal transduction via activation of c-Jun NH2-terminal kinase and p38, but not via the extracellular signal-related kinase (44, 45, 46). We propose that CD40 or the CpG-DNA receptor, despite being different receptors, trigger similar intracellular signal pathways. These similarities therefore mimic each other in functional outcome, i.e., in the activation/maturation of immature DC. One would thus predict a number of potential uses for CpG-ODN as adjuvants or immunomodulating agents (47, 48, 49, 50). As shown here, CpG-ODN renders T cell epitopes immunogenic by activating Ag-presenting immature DC. Because activation of Ag-presenting DC by CpG-DNA bypasses the need for Th cells, productive CTL responses can be initiated merely by MHC class I-restricted T cell epitopes. This attribute could be helpful in cases where the Th cell compartment is compromised. Additionally, CpG-ODN represents a single, easily produced, and chemically defined reagent.
To determine whether the potential benefits may be of practical significance, we vaccinated H-2b mice with a mixture of MHC class I-restricted LCMV-derived gp33 peptide plus CpG-ODN. LCMV infections are controlled by LCMV-specific CTL (30). This model system allowed us to analyze whether CpG-ODN not only aids peptide-specific CTL induction but also confers antiviral protection. Indeed, mice challenged once with gp33 peptide admixed with CpG-ODN 1668 generated primary CTL responses and were also protected from a subsequent LCMV infection. During the completion of these studies Oxenius et al. (51) arrived at a similar conclusion by priming and boosting H-2b mice with CpG-ODN admixed with gp33.
The nature of the Ag is believed to control the induction of CD8 or CD4 T cell immunity, in that endogenous Ags are presented by MHC class I molecules whereas exogenous and endocytosed Ags are presented by MHC class II molecules (1). DC, however, have the capacity to generate MHC class I-restricted T cell epitopes from soluble proteins via TAP-dependent pathways. High rate macropinocytosis and cell type-specific routes may deliver exogenous material to the cytosol and may account, at least in part, for the cross-priming capacity of DC (38, 39, 40, 41). Here we show that DC from LN draining a site of soluble SIINFEKL-peptide challenge retain their immature phenotype but effectively present SIINFEKL in a H-2Kb-restricted fashion. DC-depleted LN cells, however, did not present SIINFEKL. The exclusivity of cell type for the ability to present SIINFEKL contradicts a simple loading of empty MHC class I heavy chains. Whether DC indeed are specialized to take up and to sample, via macropinocytosis, extracellular peptides to deliver them in the MHC class I presentation pathway needs to be clarified.
The data described here define the adjuvanticity of CpG-ODN by their ability to activate Ag-presenting immature DC to transit to professional APC. Operationally, similar to CD40 ligation by Th cells or by anti-CD40 mAb, activated Ag-presenting DC initiate CTL responses in the absence of T help. Our data endorse CpG-DNA as an inexpensive and promising adjuvant for vaccination protocols directed toward infectious and perhaps tumor-associated diseases.
|
Acknowledgments |
|---|
|
Footnotes |
|---|
2 Address correspondence and reprint requests to Dr. H. Wagner, Institute of Medical Microbiology, Immunology and Hygiene, Technical University of Munich, Trogerstrasse 9, 81675 Munich, Germany. E-mail address:
3 Abbreviations used in this paper: DC, dendritic cells; CD40L, CD40 ligand; B3Z, the SIINFEKL/Kb-specific T cell hybridoma; LCMV, lymphocytic choriomeningitis virus; gp33, LCMV peptide KAVYNFATM; LN, lymph nodes; ODN, oligodeoxynucleotides.
4 T. Sparwasser, R. M. Vabulas, B. Villmow, G. B. Lipford, and H. Wagner. Bacterial CpG-DNA activates dendritic cells in vivo: T helper cell independent cytotoxic T cell responses to soluble proteins. Submitted for publication.
Received for publication September 13, 1999. Accepted for publication December 17, 1999.
|
References |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
-mediated shock. Eur. J. Immunol. 27:1671.[Medline]
This article has been cited by other articles:
|
|
 |
|
  C. Berchtold, K. Panthel, S. Jellbauer, B. Kohn, E. Roider, M. Partilla, J. Heesemann, S. Endres, C. Bourquin, and H. Russmann Superior Protective Immunity against Murine Listeriosis by Combined Vaccination with CpG DNA and Recombinant Salmonella enterica Serovar Typhimurium Infect. Immun., December 1, 2009; 77(12): 5501 - 5508. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 A. Ziegler, R. Heidenreich, H. Braumuller, H. Wolburg, S. Weidemann, R. Mocikat, and M. Rocken EpCAM, a human tumor-associated antigen promotes Th2 development and tumor immune evasion Blood, April 9, 2009; 113(15): 3494 - 3502. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 J. P. Leonard, B. K. Link, C. Emmanouilides, S. A. Gregory, D. Weisdorf, J. Andrey, J. Hainsworth, J. A. Sparano, D. E. Tsai, S. Horning, et al. Phase I Trial of Toll-Like Receptor 9 Agonist PF-3512676 with and Following Rituximab in Patients with Recurrent Indolent and Aggressive Non Hodgkin's Lymphoma Clin. Cancer Res., October 15, 2007; 13(20): 6168 - 6174. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
A. R. M. Kraft, F. Krux, S. Schimmer, C. Ohlen, P. D. Greenberg, and U. Dittmer CpG oligodeoxynucleotides allow for effective adoptive T-cell therapy in chronic retroviral infection Blood, April 1, 2007; 109(7): 2982 - 2984. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. Gunzer, H. Riemann, Y. Basoglu, A. Hillmer, C. Weishaupt, S. Balkow, B. Benninghoff, B. Ernst, M. Steinert, T. Scholzen, et al. Systemic administration of a TLR7 ligand leads to transient immune incompetence due to peripheral-blood leukocyte depletion Blood, October 1, 2005; 106(7): 2424 - 2432. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 L. Bevaart, H. H. Van Ojik, A. W. Sun, T. H. Sulahian, J. H. W. Leusen, G. J. Weiner, J. G. J. van de Winkel, and M. J. Van Vugt CpG oligodeoxynucleotides enhance Fc{gamma}RI-mediated cross presentation by dendritic cells Int. Immunol., August 1, 2004; 16(8): 1091 - 1098. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S. Rothenfusser, V. Hornung, M. Ayyoub, S. Britsch, A. Towarowski, A. Krug, A. Sarris, N. Lubenow, D. Speiser, S. Endres, et al. CpG-A and CpG-B oligonucleotides differentially enhance human peptide-specific primary and memory CD8+ T-cell responses in vitro Blood, March 15, 2004; 103(6): 2162 - 2169. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 N. Chaput, N. E. C. Schartz, F. Andre, J. Taieb, S. Novault, P. Bonnaventure, N. Aubert, J. Bernard, F. Lemonnier, M. Merad, et al. Exosomes as Potent Cell-Free Peptide-Based Vaccine. II. Exosomes in CpG Adjuvants Efficiently Prime Naive Tc1 Lymphocytes Leading to Tumor Rejection J. Immunol., February 15, 2004; 172(4): 2137 - 2146. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
B. Spies, H. Hochrein, M. Vabulas, K. Huster, D. H. Busch, F. Schmitz, A. Heit, and H. Wagner Vaccination with Plasmid DNA Activates Dendritic Cells via Toll-Like Receptor 9 (TLR9) but Functions in TLR9-Deficient Mice J. Immunol., December 1, 2003; 171(11): 5908 - 5912. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 P. Lundberg, P. Welander, X. Han, and E. Cantin Herpes Simplex Virus Type 1 DNA Is Immunostimulatory In Vitro and In Vivo J. Virol., October 15, 2003; 77(20): 11158 - 11169. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
P. Daftarian, S. Ali, R. Sharan, S. F. Lacey, C. La Rosa, J. Longmate, C. Buck, R. F. Siliciano, and D. J. Diamond Immunization with Th-CTL Fusion Peptide and Cytosine-Phosphate-Guanine DNA in Transgenic HLA-A2 Mice Induces Recognition of HIV-Infected T Cells and Clears Vaccinia Virus Challenge J. Immunol., October 15, 2003; 171(8): 4028 - 4039. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
L. Beloeil, M. Tomkowiak, G. Angelov, T. Walzer, P. Dubois, and J. Marvel In Vivo Impact of CpG1826 Oligodeoxynucleotide on CD8 T Cell Primary Responses and Survival J. Immunol., September 15, 2003; 171(6): 2995 - 3002. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
D. Stober, I. Jomantaite, R. Schirmbeck, and J. Reimann NKT Cells Provide Help for Dendritic Cell-Dependent Priming of MHC Class I-Restricted CD8+ T Cells In Vivo J. Immunol., March 1, 2003; 170(5): 2540 - 2548. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 I. Drexler, C. Staib, W. Kastenmuller, S. Stevanovic', B. Schmidt, F. A. Lemonnier, H.-G. Rammensee, D. H. Busch, H. Bernhard, V. Erfle, et al. Identification of vaccinia virus epitope-specific HLA-A*0201-restricted T cells and comparative analysis of smallpox vaccines PNAS, January 7, 2003; 100(1): 217 - 222. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
A. R. M. Olbrich, S. Schimmer, K. Heeg, K. Schepers, T. N. M. Schumacher, and U. Dittmer Effective Postexposure Treatment of Retrovirus-Induced Disease with Immunostimulatory DNA Containing CpG Motifs J. Virol., October 11, 2002; 76(22): 11397 - 11404. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
K. Heckelsmiller, K. Rall, S. Beck, A. Schlamp, J. Seiderer, B. Jahrsdorfer, A. Krug, S. Rothenfusser, S. Endres, and G. Hartmann Peritumoral CpG DNA Elicits a Coordinated Response of CD8 T Cells and Innate Effectors to Cure Established Tumors in a Murine Colon Carcinoma Model J. Immunol., October 1, 2002; 169(7): 3892 - 3899. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 B. Jahrsdorfer, R. Jox, L. Muhlenhoff, K. Tschoep, A. Krug, S. Rothenfusser, G. Meinhardt, B. Emmerich, S. Endres, and G. Hartmann Modulation of malignant B cell activation and apoptosis by bcl-2 antisense ODN and immunostimulatory CpG ODN J. Leukoc. Biol., July 1, 2002; 72(1): 83 - 92. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S. Zwaveling, S. C. F. Mota, J. Nouta, M. Johnson, G. B. Lipford, R. Offringa, S. H. van der Burg, and C. J. M. Melief Established Human Papillomavirus Type 16-Expressing Tumors Are Effectively Eradicated Following Vaccination with Long Peptides J. Immunol., July 1, 2002; 169(1): 350 - 358. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. Gierynska, U. Kumaraguru, S.-K. Eo, S. Lee, A. Krieg, and B. T. Rouse Induction of CD8 T-Cell-Specific Systemic and Mucosal Immunity against Herpes Simplex Virus with CpG-Peptide Complexes J. Virol., June 5, 2002; 76(13): 6568 - 6576. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
K. S. Kawamura, R.-C. Su, L. T. Nguyen, A. R. Elford, P. S. Ohashi, and J. Gariepy In Vivo Generation of Cytotoxic T Cells from Epitopes Displayed on Peptide-Based Delivery Vehicles J. Immunol., June 1, 2002; 168(11): 5709 - 5715. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. Gursel, D. Verthelyi, I. Gursel, K. J. Ishii, and D. M. Klinman Differential and competitive activation of human immune cells by distinct classes of CpG oligodeoxynucleotide J. Leukoc. Biol., May 1, 2002; 71(5): 813 - 820. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
T. Decker, S. Hipp, R. J. Kreitman, I. Pastan, C. Peschel, and T. Licht Sensitization of B-cell chronic lymphocytic leukemia cells to recombinant immunotoxin by immunostimulatory phosphorothioate oligodeoxynucleotides Blood, February 15, 2002; 99(4): 1320 - 1326. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
I. Miconnet, S. Koenig, D. Speiser, A. Krieg, P. Guillaume, J.-C. Cerottini, and P. Romero CpG Are Efficient Adjuvants for Specific CTL Induction Against Tumor Antigen-Derived Peptide J. Immunol., February 1, 2002; 168(3): 1212 - 1218. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
G. G. Chikh, S. Kong, M. B. Bally, J.-C. Meunier, and M.-P. M. Schutze-Redelmeier Efficient Delivery of Antennapedia Homeodomain Fused to CTL Epitope with Liposomes into Dendritic Cells Results in the Activation of CD8+ T Cells J. Immunol., December 1, 2001; 167(11): 6462 - 6470. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
Y. Kawarada, R. Ganss, N. Garbi, T. Sacher, B. Arnold, and G. J. Hammerling NK- and CD8+ T Cell-Mediated Eradication of Established Tumors by Peritumoral Injection of CpG-Containing Oligodeoxynucleotides J. Immunol., November 1, 2001; 167(9): 5247 - 5253. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
T. De Smedt, E. Butz, J. Smith, R. Maldonado-Lopez, B. Pajak, M. Moser, and C. Maliszewski CD8{alpha}- and CD8{alpha}+ subclasses of dendritic cells undergo phenotypic and functional maturation in vitro and in vivo J. Leukoc. Biol., June 1, 2001; 69(6): 951 - 958. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. Bauer, V. Redecke, J. W. Ellwart, B. Scherer, J.-P. Kremer, H. Wagner, and G. B. Lipford Bacterial CpG-DNA Triggers Activation and Maturation of Human CD11c-, CD123+ Dendritic Cells J. Immunol., April 15, 2001; 166(8): 5000 - 5007. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
B. Jahrsdörfer, G. Hartmann, E. Racila, W. Jackson, L. Mühlenhoff, G. Meinhardt, S. Endres, B. K. Link, A. M. Krieg, and G. J. Weiner CpG DNA increases primary malignant B cell expression of costimulatory molecules and target antigens J. Leukoc. Biol., January 1, 2001; 69(1): 81 - 88. [Abstract] [Full Text]
|
|
|
|
|
|
T. L. Warren, S. K. Bhatia, A. M. Acosta, C. E. Dahle, T. L. Ratliff, A. M. Krieg, and G. J. Weiner APC Stimulated by CpG Oligodeoxynucleotide Enhance Activation of MHC Class I-Restricted T Cells J. Immunol., December 1, 2000; 165(11): 6244 - 6251. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
H. Weighardt, C. Feterowski, M. Veit, M. Rump, H. Wagner, and B. Holzmann Increased Resistance Against Acute Polymicrobial Sepsis in Mice Challenged with Immunostimulatory CpG Oligodeoxynucleotides Is Related to an Enhanced Innate Effector Cell Response J. Immunol., October 15, 2000; 165(8): 4537 - 4543. [Abstract] [Full Text] [PDF]
|
|
| |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
, ) or EL-4 cells pulsed with the SIINFEKL (•, 
) was
scored in a standard 51Cr release assay. Symbols represent
CTL derived from individual mice. B. Before testing lytic
activity in 51Cr release assay, SIINFEKL plus
CpG-ODN-induced CTL were depleted of CD4+ or
CD8+ cells. Targets were EL-4 cells (
) or EL-4 cells pulsed with the SIINFEKL (•, 
) was
scored in a standard 51Cr release assay. Symbols represent
CTL derived from individual mice. The experiment has been repeated
twice. B, Systemic dissemination of primed CTL. Mice were
immunized with 0.3 mg SIINFEKL plus 10 nmol CpG-ODN 1668 in the fore
footpads; 2 wk after immunization, splenocytes were isolated and
restimulated in vitro for 7 days as described in Materials and
Methods. Lytic activity against EL-4 cells (
, average virus titer in each group.
·····, virus detection limit in
the assay.