The JI
HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
QUICK SEARCH:   [advanced]


     
 

This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow Request Permissions
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Turner, M. S.
Right arrow Articles by Finn, O. J.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Turner, M. S.
Right arrow Articles by Finn, O. J.
Right arrowPubmed/NCBI databases
*Substance via MeSH
The Journal of Immunology, 2007, 178: 2787-2793.
Copyright © 2007 by The American Association of Immunologists, Inc.

Lack of Effective MUC1 Tumor Antigen-Specific Immunity in MUC1-Transgenic Mice Results from a Th/T Regulatory Cell Imbalance That Can Be Corrected by Adoptive Transfer of Wild-Type Th Cells1

Michael S. Turner, Patrice A. Cohen and Olivera J. Finn2

Department of Immunology, University of Pittsburgh School of Medicine, Pittsburgh, PA 15213


   Abstract
Top
 Abstract
Introduction
 Materials and Methods
 Results
 Discussion
 Disclosures
 References
 
Glycoprotein tumor Ag MUC1 is overexpressed on the majority of epithelial adenocarcinomas. CTLs that recognize MUC1 and can kill tumor cells that express this molecule have been found in cancer patients, yet they are present in low frequency and unable to eradicate MUC1+ tumors. Patients also make anti-MUC1 Abs but predominantly of the IgM isotype reflecting the lack of effective MUC1-specific Th responses. Mice transgenic for the human MUC1 gene (MUC1-Tg) are similarly hyporesponsive to MUC1. We used a vaccine consisting of dendritic cells loaded with a long synthetic MUC1 peptide to investigate the fate and function of MUC1-specific CD4+ Th elicited in wild-type (WT) or MUC1-Tg mice or adoptively transferred from vaccinated WT mice. We show that hyporesponsiveness of MUC1-Tg mice to this vaccine is a result of insufficient expansion of Th cells, while at the same time their regulatory T cells are efficiently expanded to the same extent as in WT mice and exert a profound suppression on MUC1-specific B and T cell responses in vivo. Adoptive transfer of WT Th cells relieved this suppression and enhanced T and B cell responses to subsequent MUC1 immunization. Our data suggest that the balance between Th and regulatory T cells is a critical parameter that could be modulated to improve the response to cancer vaccines.


   Introduction
Top
 Abstract
 Introduction
Materials and Methods
 Results
 Discussion
 Disclosures
 References
 
The epithelial mucin MUC1 is a large transmembrane glycoprotein that is expressed on the apical surface of healthy ductal epithelia and also on a broad range of adenocarcinomas (1, 2, 3, 4, 5). The ability of Abs and T cells to distinguish between normal and tumor-specific glycoforms of MUC1 makes this molecule an attractive immunological target for the treatment of many cancers of epithelial origin (6). Evidence indicates that the presence of MUC1-specific serum Abs correlates favorably with protection against MUC1-associated malignancies (7, 8, 9). However, in many cancer patients with MUC1+ tumors, MUC1-specific immunity does not halt the progression of developing adenocarcinomas. Studies in animal models have shown that vaccines capable of inducing robust MUC1-specific immune responses offer protection against MUC1+ tumors (10, 11, 12, 13, 14). The ability to induce similarly effective immunity to MUC1 in humans could provide treatment for the majority of adenocarcinomas. Clinical trials of MUC1-based vaccines in cancer patients have so far elicited only weak CTL responses and low titer Abs of predominantly IgM isotype (15, 16, 17, 18, 19), suggesting that there is a problem in generating effective MUC1-specific T cell help. As CD8+ T cells require help from CD4+ T cells to establish effective CTL memory (20, 21, 22, 23, 24, 25), the lack of effective MUC1-specific Th cells in cancer patients is considered to be a major obstacle in generating effective antitumor immunity.

We have previously reported that one of the mechanisms responsible for weak MUC1-specific Th responses is that the full-length MUC1 secreted by tumor cells is inefficiently processed and presented by APCs through the MHC class II pathway (26). Another reason we found failure of Th cells in cancer patients is that the aberrantly glycosylated tumor MUC1 has a negative effect on the ability of APCs to skew Th responses to the desired Th1 type (27). Both of these problems can be overcome by a well-designed vaccine. However, experiments comparing vaccine-induced anti-MUC1 responses in wild-type (WT)3 mice and MUC1-transgenic (Tg) mice have also shown reduced Th responses even when MUC1 is efficiently processed and presented by properly matured APCs (13, 26, 28, 29, 30). There is thus a possibility that the presence of MUC1 as a self-Ag has a negative influence on the MUC1-specific helper T cell repertoire. Furthermore, the recently appreciated importance of regulatory T cells (Tregs) in down-regulating immune responses to tumors (31, 32, 33, 34) suggested that they may also be influenced by the presence of MUC1 and differentially regulating anti-MUC1 responses in WT and MUC1-Tg mice.

In our previous study (13), a dendritic cell (DC)-based immunization protocol revealed distinctly the difference in the magnitude of MUC1-specific CD4+ T cell responses between MUC1-Tg and WT mice. Therefore, we used the same DC vaccine in the current study to better understand how MUC1-specific CD4+ T cells are regulated in MUC1-Tg mice compared with WT mice and how this affects MUC1-specific CTL and Ab responses. We show that, while MUC1-Tg mice fail to mount effective Th responses to MUC1 vaccines, their Tregs are expanded to the same extent as in WT mice and function equally by suppressing MUC1-specific B and T cell responses in vivo. Adoptive transfer of MUC1-primed Th cells from WT mice restores MUC1-specific Th responses in MUC1-Tg mice and enhances their T and B cell response to the MUC1 vaccine.


   Materials and Methods
Top
 Abstract
 Introduction
 Materials and Methods
Results
 Discussion
 Disclosures
 References
 
Mice and cell lines

C57BL/6 mice were purchased from The Jackson Laboratory and C57BL/6 MUC1-Tg mice from Dr. S. Gendler (Mayo Clinic, Scottsdale, AZ). All mice were maintained in a specific pathogen-free environment at the University of Pittsburgh Cancer Institute and treated in accordance with the guidelines of the Institutional Animal Care and Use Committee of the University of Pittsburgh.

The RMA-MUC1 cell line, previously described (13), was generated by transfection of RMA lymphoma cells with full-length MUC1 cDNA. All mouse cells were cultured in complete DMEM (cDMEM) containing 10% FBS, penicillin and streptomycin, L-glutamine, sodium pyruvate, nonessential amino acids, HEPES buffer, and 2-ME, plus other additives, as indicated.

Generation of bone marrow DCs (BMDCs) and vaccination of mice

BMDCs were generated as described previously (13). Briefly, BM cells removed from the femurs and tibiae of C57BL/6 mice were cultured in cDMEM containing 10 ng/ml each of GM-CSF and IL-4 (a gift from Immunex, Seattle, WA). Cells were fed on days 2, 4, and 6 by adding 5 ml of cDMEM containing 10 ng/ml GM-CSF and IL-4. On day 7 of culture, cells were harvested and purified by density centrifugation over Nycoprep 1.068 (Accurate Chemical) gradient. Purified DCs were loaded for 4–6 h in polypropylene tubes with 20–50 µg/ml synthetic MUC1 100-mer. Mice were immunized s.c. in the right flank with 200 µl of PBS containing 105 MUC1-loaded DCs plus 20 µg of soluble MUC1 100-mer. Boosts were administered similarly at 2-wk intervals. The MUC1 100-mer synthetic peptide represents five tandem repeats of an unglycosylated peptide (GVTSAPDTRPAPGSTAPPAH), which is found tandemly repeated in the extracellular region of MUC1. MUC1 100-mer was synthesized by the solid-phase on a Pioneer peptide synthesizer (Applied Biosystems) using a FMOC synthesis protocol in the Peptide Synthesis Facility, University of Pittsburgh. The crude peptides were analyzed, characterized, and purified by Gel filtration (G-25 column) and reversed-phase HPLC (486 and 600E by Waters), and the correct mass was confirmed by Electrospray Mass Spectroscopy (Quattro II; Fisons).

T cell cultures and in vitro stimulation

Lymph node (LN) and spleen cells were prepared by mechanical disruption and RBC lysis and stimulated in vitro with MUC1 100-mer-loaded BMDCs in the presence of 20 U/ml murine IL-2 (Immunex). Cultures were fed every 2 days with cDMEM containing 20 U/ml murine IL-2. T cell function was tested on day 5 for cytotoxicity and on day 7 for cytokine production, as described below.

Flow cytometry

Cell surface FCR was blocked with anti-CD16/CD32 mAb (BD Biosciences). Without washing, blocked cells were then surface stained for 30 min with the indicated Abs (BD Biosciences) diluted 1/50 in FACS buffer. After washing, stained cells were analyzed on a LSR II Flow Cytometer (BD Biosciences). FACSorting was performed with a FACSAria (BD Biosciences).

Intracellular staining for FoxP3 was performed ex vivo using a mouse FoxP3 staining kit (eBioscience). For intracellular cytokine staining, LN or spleen cells were stimulated in vitro for 7 days with MUC1-loaded DCs, then restimulated with PMA/ionomycin for 6 h in the presence of Golgi Plug (BD Biosciences). After restimulation, cells were stained for surface markers CD3, CD4, or CD8 as described above, then treated with Cytofix/Cytoperm (BD Biosciences) to allow intracellular staining with anti-IFN-{gamma} Ab (BD Biosciences).

Serum Ig ELISA

Mice were bled from the carotid artery immediately after sacrifice, and the serum was separated by centrifugation. Immulon 4HBX plates (Fisher Scientific) were coated overnight with PBS containing 20 µg/ml MUC1 100-mer or 2.5% BSA. Plates were then washed with PBS and blocked with 2.5% BSA. Sera were diluted 1/100 in 2.5% BSA and added to the ELISA plate in triplicates. Plates were washed five times with 200 µl of Tween 20 (0.1% in PBS), then HRP-conjugated anti-mouse IgG Ab (Southern Biotechnology Associates) was added diluted 1/500 in 2.5% BSA. After washing again in Tween 20, 100 µl of tetramethylbenzidine substrate (BD Biosciences) was added for 30 min, and the reaction was then stopped with 2 N H2SO4. Absorbance was read at 450 nm. All ELISA data show the average of triplicate wells with SD.

51Cr release cytotoxicity assay

Responder LN/spleen cells were stimulated for 5 days in vitro with MUC1-loaded DCs, then added to RMA and RMA-MUC1 target cells (13) (E:T = 10:1) that were prelabeled with radioactive sodium chromate (Amersham Biosciences). After 4–6 h, supernatants were harvested and analyzed on a Cobra II auto gamma counter (PerkinElmer). Specific lysis was calculated as (lysis – spontaneous lysis) ÷ (total lysis – spontaneous lysis). All CTL assays were performed in triplicate. Data show the averages and SDs of triplicate wells.

Adoptive T cell transfers

Donor mice were immunized with MUC1-loaded DCs, as described above. Two weeks after immunization, CD4+ T lymphocytes were purified from LN and spleen by MACS using CD4 (L3T4) Microbeads (Miltenyi Biotec). For the Treg depletion experiment, the Treg isolation kit (Miltenyi Biotec) was used to negatively purify total CD4+ T cells and CD4+CD25– cells. Mice were injected via the lateral tail vein with 200 µl of PBS containing 5 x 106–1 x 107 T cells.

Statistical analysis

The statistical significance of our results was calculated by an unpaired t test. The two-tailed p values were determined with the statistical program GraphPad INSTAT version 3 Software. Values of p < 0.05 were considered significant.


   Results
Top
 Abstract
 Introduction
 Materials and Methods
 Results
Discussion
 Disclosures
 References
 
DC-MUC1 vaccine elicits MUC1-specific Tregs that limit the function of MUC1-specific T cells

WT and MUC1-Tg mice were immunized s.c. with DCs that had been loaded in vitro with the synthetic MUC1 100-mer peptide. Two weeks after vaccination, inguinal LNs, spleens, and sera were collected and analyzed for the presence of MUC1-specific immune responses. As expected, immunization of WT mice with MUC1-loaded DCs induced substantial numbers of IFN-{gamma}-producing CD4+ and CD8+ T cells (Fig. 1, A and C), CTLs that efficiently lysed the MUC1+ tumor cell line RMA-MUC1 (Fig. 1D), and high levels of MUC1-specific serum IgG (Fig. 1B). MUC1-Tg mice in contrast responded to the MUC1-pulsed DC vaccine with low numbers of CD4+ T cells (Fig. 1A). The absence of MUC1-specific IgG in the sera of these mice after immunization (Fig. 1B) indicates an inability of their MUC1-specific B cells to undergo isotype class switching and is an important sign of the absence of functional MUC1-specific T cell help. In addition, the weak MUC1-specific Th cell response in MUC1-Tg mice correlated with weak MUC1-specific CD8 T cell responses, as measured by both IFN-{gamma} production (Fig. 1C) and lysis of MUC1+ tumors (Fig. 1D).


Figure 1
View larger version (21K):
[in this window]
[in a new window]

  		FIGURE 1. MUC1-Tg mice are hyporesponsive to immunization with MUC1-loaded DCs. Two weeks after s.c. immunization with MUC1-loaded DCs, LNs and sera were isolated from WT and MUC1-Tg mice (four mice per treatment group), pooled, and tested for reactivity to MUC1. A and C, IFN-{gamma} production. LN cells were stimulated in vitro with MUC1-loaded DCs for 6 days, then restimulated for 6 h with MUC1-loaded DCs, PMA, and ionomycin in the presence of brefeldin A and stained for surface CD4 and CD8 and intracellular IFN-{gamma}. B, MUC1-specific serum IgG. Sera from immunized mice were incubated on ELISA plates coated with MUC1 100-mer, then blotted with anti-mouse IgG (OD, OD). D, Specific lysis of the MUC1+ tumor RMA-MUC1. Pooled LN cells were incubated for 4 h with 51Cr-labeled RMA or RMA-MUC1 targets in triplicate.

 
We reasoned that, if MUC1-Tg mice had an inherent problem with MUC1-specific Th cells due to their development in the presence of MUC1 as a self-Ag, they should mount improved responses to the MUC1 vaccine if they first received an adoptive transfer of Th cells from WT mice. We transferred CD4+ T cells from WT mice into MUC1-Tg mice, which were then immunized and boosted with MUC1-loaded DCs. The WT donor mice were vaccinated with MUC1-loaded DCs 2 wk before cell transfer to expand the numbers of MUC1-specific cells within the pool of transferred CD4+ T cells.

Contrary to our expectations, adoptive transfer of MUC1-primed CD4+ T cells from WT mice failed to enhance the ability of MUC1-Tg mice to respond to subsequent immunization with MUC1-loaded DCs. In fact, there was consistently a small reduction in the already weak cellular (Fig 2, A and B) and humoral (Fig. 2C) immune responses to the DC-MUC1 vaccine. The full suppressive capacity of these WT CD4+ T cells was revealed when they were transferred into WT mice, resulting in reduced numbers of IFN-{gamma}-producing CD8+ and CD4+ T cells in the LN draining the vaccine site (Fig. 2D), and a substantially reduced capacity to lyse the MUC1+ tumor RMA-MUC1 (Fig. 2E) compared with nonvaccinated WT mice. In addition, there were significantly lower levels of MUC1-specific IgG in the sera of these mice (Fig. 2F), indicating that B cell function was also affected.


Figure 2
View larger version (20K):
[in this window]
[in a new window]

  		FIGURE 2. CD4 T cells primed by MUC1-pulsed DCs suppress MUC1-specific immune responses. MUC1-Tg (A–C) and WT (D–F) mice were immunized s.c. with MUC1-loaded DCs, following I.V. injection of PBS ({blacksquare}) or 5 x 106 CD4+ T cells from MUC1-immunized WT donors ({square}). Two weeks after immunization, LNs and sera were pooled (four mice per group) and tested for immune responses: A and D, IFN-{gamma} production; LN cells were stimulated in vitro for 6 h in the presence of brefeldin A, then stained for surface CD8, CD4, and intracellular IFN-{gamma}. B and E, Specific lysis of RMA-MUC1 tumor cells; LN cells were incubated with 51Cr-labeled RMA or RMA-MUC1 targets for 4 h in triplicate. C and F, MUC1-specific serum Ig ELISA; sera from immunized mice were incubated on ELISA plates coated with MUC1 100-mer, then blotted with polyclonal anti-mouse Ig. This adoptive transfer experiment was performed on three separate occasions with similar results. Data shown are from one representative experiment.

 
Given the ability of CD4+ Tregs to suppress Ag-specific immune responses (33, 35, 36), we wondered whether the immunosuppressive effect of the adoptive transfer might be due to the presence (and potential expansion by the DC vaccine) of Tregs within the population of transferred WT CD4+ T cells. To test this, WT mice were again vaccinated once with MUC1-loaded DCs and the numbers of Tregs in the draining LN compared with those in age-matched unimmunized mice. The immunized mice exhibited an increased frequency of FoxP3+ cells within the CD4+ T cell population (Fig. 3A). When CD3+CD4+CD25high T cells, which expressed FoxP3, were isolated from MUC1-immunized WT mice, they effectively suppressed the production of IL-2 by CD8+ T cells responding to APC-presenting MUC1 (data not shown), confirming that the FoxP3+CD4+CD25+ T cells expanded in MUC1-immunized mice were functionally suppressive Tregs. Induction of Tregs was Ag driven, as increased numbers of FoxP3+ CD4+ T cells were observed only in mice that were immunized with MUC1-loaded DCs but not with control unloaded DCs (data not shown).


Figure 3
View larger version (27K):
[in this window]
[in a new window]

  		FIGURE 3. In vivo expansion of Tregs by MUC1-loaded DCs in WT and MUC1-Tg mice. Age-matched WT (A) and MUC1-Tg (B) mice were immunized and boosted s.c. with MUC1-loaded DCs. Two weeks after boosting, LN and spleen cells were isolated and stained on the surface for CD3 and CD4, and intracellularly for FoxP3. The frequency of FoxP3+ cells is expressed as a percentage of gated CD4+CD3+ cells. Differences between groups: WT Immunized vs Naive, p = 0.0573; MUC1-Tg Immunized vs Naive, p = 0.0568; Immunized WT vs MUC1-Tg p = 0.255. (C) Representative histograms showing FoxP3 staining in CD3+CD4+ cells.

 
Tregs are induced equally in highly MUC1-responsive WT mice and hyporesponsive MUC1-Tg mice

Very little is known to date about vaccine-induced Tregs, and we wondered whether DC-MUC1-induced Tregs would expand differently in the presence of MUC1 in MUC1-Tg mice compared with WT mice. One possibility was that fewer Tregs would be generated in MUC1-Tg mice than in WT mice because the Th cell response in MUC1-Tg mice is suboptimal and there would be less need for regulation than in WT mice, whereas the vigorous Th response could presumably require higher numbers of Tregs. Alternatively, we could postulate increased numbers of MUC1-specific Tregs in MUC1-Tg mice before vaccination, due to the presence of self-MUC1, that would then be further expanded with the MUC1 vaccine. Surprisingly, we found no difference in the level of expansion of FoxP3+ Tregs between MUC1-Tg mice and WT mice (Fig. 3). Furthermore, these cells were equally immunosuppressive; when CD4+ T cells from immunized MUC1-Tg mice were adoptively transferred into WT mice, they suppressed MUC1-specific T and B cell responses by the same degree as CD4+ T cells from WT mice. Regardless of the donor mouse strain, WT mice receiving CD4+ T cells from either MUC1-Tg or WT donors exhibited reduced numbers of IFN-{gamma}+CD4+ T cells (Fig. 4A). As seen before, a modest reduction in measured Th1 responders translated functionally into significantly reduced MUC1-specific serum IgG (Fig. 4B), and impaired IFN-{gamma} production by CD8+ T cells (Fig. 4C) was accompanied by a reduced ability to lyse MUC1-expressing tumor cells (Fig. 4D).


Figure 4
View larger version (23K):
[in this window]
[in a new window]

  		FIGURE 4. CD4+ T cells from immunized WT and MUC1-Tg mice suppress MUC1-specific immune responses when transferred into WT recipients. WT and MUC1-Tg donor mice were immunized with MUC1-loaded DCs. Two weeks later, their CD4+ T cells were purified and adoptively transferred to WT recipients (5 x 106 cells/mouse). Recipient mice were immunized and boosted with MUC1-pulsed DCs. Two weeks postboost, LN and blood samples were tested for MUC1-specific responses: A and C, IFN-{gamma} production by CD4+ and CD8+ T cells, measured by Intracellular Cytokine Staining. D, MUC1-specific lysis of RMA-MUC1 tumor. B, MUC1-specific serum IgG1 ELISA. Data are from one experiment, representative of three separate experiments performed with similar results.

 
Adoptive immunotherapy with Treg-depleted WT Th cells improves MUC1-specific immunity in MUC1-Tg mice

The data presented so far indicate that, although MUC1-Tg mice have hyporesponsive MUC1-specific Th cells, their repertoire of Tregs is fully competent and responds strongly to MUC1 peptides presented by DCs. Furthermore, the expansion of Tregs in WT donors by immunization with MUC1-loaded DCs (Fig. 3A) explains why the initial transfers of WT CD4+ T cells into MUC1-Tg mice resulted in suppression of their immune response (Fig. 2) rather than the improvement we had expected. These results suggested that the response to the MUC1 vaccine is affected by the balance of MUC1-specific Th and Tregs present at the time of immunization. To confirm this, we performed another cell transfer experiment using immunized WT donors and tested the effect of eliminating CD4+CD25+ double-positive Tregs from the transferred CD4+ population. CD4+ T cells were purified from MUC1-immunized WT donors and divided into two batches, and CD25+ cells were depleted from one batch by MACS. MUC1-Tg mice receiving adoptive transfers of either whole CD4+ T cells or CD4+CD25– T cells were vaccinated with MUC1-loaded DCs and subsequently evaluated for MUC1-specific immune responses. This time, MUC1-Tg mice that received purified WT Th cells exhibited improved MUC1-specific cellular and humoral immune responses compared with MUC1-Tg mice that received total CD4+ T cells (Fig. 5). Increased numbers of IFN-{gamma}+CD4+ T cells were detected in the draining LN of MUC1-Tg mice that received the purified WT Th cells before vaccination (Fig. 5A). This correlated with increases in IFN-{gamma}+CD8+ T cells (Fig. 5B) and higher levels of circulating MUC1-specific IgG (Fig. 5C). This result indicates that the adoptive transfer of purified Th cells counterbalanced the immunosuppressive action of Tregs and provided help for both CD8+ T cells and B cells responding to the MUC1 vaccine.


Figure 5
View larger version (9K):
[in this window]
[in a new window]

  		FIGURE 5. Treg-depleted WT Th cells improve immune responses of MUC1-Tg mice to vaccination with MUC1-loaded DCs. MUC1-Tg mice were injected i.v. with CD4+ T cells ({diamondsuit}), or CD4+ T cells depleted of CD25+ Tregs ({blacktriangleup}) from WT mice immunized with MUC1-loaded DCs. All recipient mice were then immunized with MUC1-loaded DCs and boosted 2 wk later. Serum and LN cells were harvested 10 days postboost. A and B, IFN-{gamma} production by LN T cells: LN cells from adoptively transferred and immunized mice were stimulated in vitro with MUC1-loaded DCs for 7 days, then restimulated for 6 h with PMA and ionomycin in the presence of brefeldin A. Cells were then stained for surface CD3, plus CD4 or CD8, and intracellular IFN-{gamma}. Data represent the percentage of IFN-{gamma}+ cells in the CD3+CD4+ or CD3+CD8+ gates (A and B, respectively). C, Sera of mice tested for presence of MUC1-specific IgG by ELISA. Differences were observed between treatment groups were significant: A, p = 0.049; B, p = 0.029; C, p = 0.0458. Results were reproducible in repeat experiments.

 

   Discussion
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
Disclosures
 References
 
The hyporesponsiveness of MUC1-specific Th in cancer patients and MUC1-Tg mice is considered to be a major obstacle to establishing effective protective immunity against MUC1+ tumors. We reasoned that this should be easily corrected by transferring CD4+ T cells from MUC1-immunized WT mice, which respond strongly. Unexpectedly, there was no improvement in the response of MUC1-Tg mice after adoptive transfer. Indeed, there was a modest, yet consistent reduction in their cellular and humoral responses. The full suppressive capacity of the donor WT CD4+ T cells was revealed when they were transferred into WT mice, which normally respond strongly to MUC1. The degree of suppression was different in WT and MUC1-Tg recipients because, even in the absence of such an adoptive transfer, MUC1-Tg mice already exhibit reduced Th responses to MUC1. With no "room" for further suppression, the transferred WT CD4+ T cells appeared simply to fail to improve the poor response in MUC1-Tg mice. We concluded that MUC1-specific Tregs were expanded in the WT donor mice by immunizing them before harvesting their CD4+ T cells for transfer. This resulted in suppression of immune responses in the recipient mice.

In the early years of their characterization, Tregs were originally thought to be anergic because they appeared to lack classical features of T cell activation such as proliferation and cytokine production (35, 37). However, recent studies have shown that their proliferation can be induced by different subsets of DCs (38, 39, 40, 41, 42, 43). The data presented here indicate that DCs presenting MUC1 peptides can simultaneously stimulate potent MUC1-specific immunity and induce FoxP3+ Tregs. In WT mice, where MUC1 is a foreign Ag, the net outcome of immunization with MUC1-loaded DCs is effective MUC1-specific immunity because the vaccine-induced MUC1-specific Tregs are balanced by an efficient MUC1-specific Th cell response. The concomitant induction of MUC1-specific Th and Tregs in WT mice may therefore be part of a normal, balanced immune response to DCs presenting a foreign Ag (in this case, MUC1), where Tregs are necessary to bring the induced immune response to a quiescent state (44, 45, 46). In MUC1-Tg mice, however, endogenous MUC1 expression renders Th cells hyporesponsive to MUC1-based vaccines, so the Th cells fail to keep pace with Tregs, which our data show respond strongly to DCs presenting MUC1. In this situation, the Treg/Th balance is skewed in favor of Tregs and results in suppression of MUC1-specific CTLs and Ab responses (Fig. 1).

Recent studies have identified different subsets of DCs that have different abilities to activate Th and Tregs and can induce either immunity or tolerance (47, 48, 49, 50, 51). By targeting the MUC1 Ag to the appropriate population of DCs, one might be able to dictate the nature and outcome of the DC-T cell interaction to favor the induction of Th cells over Tregs, thereby promoting the establishment of effective tumor-specific memory CTLs. Our data show that this is especially important where the tumor Ag is also a self-Ag and the effector T cell repertoire may be adversely affected, for example, by being limited to low-affinity T cells (52). What our data show, and what is very significant, is that Tregs are apparently not under the influence of this "self-tolerance" since they are induced equally efficiently in the absence or in the presence of the self-Ag MUC1. We do not yet understand how this happens and are in the process of exploring the selection of these populations using a MUC1-specific TCR Tg mouse where both Treg and Th cells bare the same TCR (M. S. Turner and O. J. Finn, manuscript in preparation).

There is mounting evidence that Tregs play a role in the suppression of antitumor immunity (34). The presence of Tregs in cancer patients correlates inversely with strength of the tumor-specific immune response and prognosis (32, 53, 54), and depletion of Tregs has been shown to enhance the antitumor immune response (31, 33). Our data show that MUC1-specific Tregs respond to MUC1 presented by DCs and suppress the stimulation of effector T cells by those same DCs. Nevertheless, the lack of effective MUC1-specific CTL responses in MUC1-Tg mice seems to be due primarily to a deficiency in MUC1-specific T cell help, rather than an increased frequency of Tregs. This was born out by the fact that MUC1-specific T and B cell responses were improved in MUC1-Tg mice by the adoptive transfer of functional WT Th cells alone, without depleting Tregs from the host. This indicates that MUC1-specific Th cells play a fundamental role in generating effective MUC1-specific CTLs and suggests that, if effective MUC1-specific T cell help can be induced or provided exogenously, it would be possible to change the host environment from one that is refractory to MUC1 vaccines to one that favors the induction of effective MUC1-specific antitumor immunity. This is supported by a recent study in which adoptive transfer of CD4+ Th cells provided help for adoptively transferred melanoma-specific CTLs (31).

WT mice were chosen as donors for the adoptive transfer experiments because their repertoire of MUC1-specific Th cells is not affected by the expression of self-Ag MUC1, as in MUC1-Tg mice. Therefore, we reasoned that the WT Th repertoire would contain more high-affinity MUC1-specific Th cells, which would restore MUC1-specific Th function to the MUC1-Tg recipients. This was indeed the case. Humans, like MUC1-Tg mice, are hyporesponsive to MUC1. Therefore, transferring a high-affinity MUC1-specific repertoire from WT mice may not be the same as transferring cells from the same or another human. However, the adoptive transfer of MUC1-expanded CD4+CD25– T cells from WT mice to MUC1-Tg mice to raise the level of MUC1-specific T cell help is a proof of principle that we believe can be applied to the clinic where patients would receive transfers of autologous Treg-depleted Th cells that are expanded to large numbers in vitro by stimulation with MUC1. Restoring the balance to the MUC1-specific immune response via the provision of a defined component, such as functional Th cells, may be a more feasible clinical approach to treating malignancies than global depletion of the patients’ Tregs, which may be fraught with side effects such as autoimmunity (55, 56, 57, 58, 59, 60).

Furthermore, our data show that when Th and Tregs are transferred together, Tregs dominate over Th cells and therefore suppress immune responses of the recipient. The reason for this may be their preferential survival in the recipient, as suggested by a recently published observation that Tregs express higher levels of the antiapoptotic molecule Bcl-2, causing them to be less susceptible to apoptosis than their Th counterparts (61). These findings have obvious implications for the design of adoptive cell transfer approaches, where it would clearly be prudent to deplete Tregs from the population of transferred cells.


   Disclosures
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
 Disclosures
References
 
The authors have no financial conflict of interest.


   Footnotes
 
The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

1 This work was supported by National Institutes of Health Grant R01-CA56103. Back

2 Address correspondence and reprint requests to Dr. Olivera J. Finn, Department of Immunology, University of Pittsburgh School of Medicine, Pittsburgh, PA 15213. Back

3 Abbreviations used in this paper: WT, wild type; BMDC, bone marrow DC; cDMEM, complete DMEM; DC, dendritic cell; LN, lymph node; Tg, transgenic; Treg, regulatory T cell. Back

Received for publication September 5, 2006. Accepted for publication December 14, 2006.


   References
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
 Disclosures
 References
 

  1. Ohno, T., R. Aihara, Y. Kamiyama, E. Mochiki, T. Asao, H. Kuwano. 2006. Prognostic significance of combined expression of MUC1 and adhesion molecules in advanced gastric cancer. Eur. J. Cancer 42: 256-263. [Medline]
  2. Singh, A. P., S. C. Chauhan, S. Bafna, S. L. Johansson, L. M. Smith, N. Moniaux, M. F. Lin, S. K. Batra. 2006. Aberrant expression of transmembrane mucins, MUC1 and MUC4, in human prostate carcinomas. Prostate 66: 421-429. [Medline]
  3. Baldus, S. E., J. R. Wienand, J. P. Werner, S. Landsberg, U. Drebber, F. G. Hanisch, H. P. Dienes. 2005. Expression of MUC1, MUC2 and oligosaccharide epitopes in breast cancer: prognostic significance of a sialylated MUC1 epitope. Int. J. Oncol. 27: 1289-1297. [Medline]
  4. Hebbar, V., G. Damera, G. P. Sachdev. 2005. Differential expression of MUC genes in endometrial and cervical tissues and tumors. BMC Cancer 5: 124[Medline]
  5. Turner, M. S., J. R. McKolanis, R. K. Ramanathan, D. C. Whitcomb, O. J. Finn. 2003. Mucins in gastrointestinal cancers. Cancer Chemother. Biol. Response Modif. 21: 259-274. [Medline]
  6. Vlad, A. M., J. C. Kettel, N. M. Alajez, C. A. Carlos, O. J. Finn. 2004. MUC1 immunobiology: from discovery to clinical applications. Adv. Immunol. 82: 249-293. [Medline]
  7. Cramer, D. W., L. Titus-Ernstoff, J. R. McKolanis, W. R. Welch, A. F. Vitonis, R. S. Berkowitz, O. J. Finn. 2005. Conditions associated with antibodies against the tumor-associated antigen MUC1 and their relationship to risk for ovarian cancer. Cancer Epidemiol. Biomarkers Prev. 14: 1125-1131. [Abstract/Free Full Text]
  8. Jerome, K. R., A. D. Kirk, G. Pecher, W. W. Ferguson, O. J. Finn. 1997. A survivor of breast cancer with immunity to MUC-1 mucin, and lactational mastitis. Cancer Immunol. Immunother. 43: 355-360. [Medline]
  9. von Mensdorff-Pouilly, S., A. A. Verstraeten, P. Kenemans, F. G. Snijdewint, A. Kok, G. J. Van Kamp, M. A. Paul, P. J. Van Diest, S. Meijer, J. Hilgers. 2000. Survival in early breast cancer patients is favorably influenced by a natural humoral immune response to polymorphic epithelial mucin. J. Clin. Oncol. 18: 574-583. [Abstract/Free Full Text]
  10. Kagan, E., G. Ragupathi, S. S. Yi, C. A. Reis, J. Gildersleeve, D. Kahne, H. Clausen, S. J. Danishefsky, P. O. Livingston. 2005. Comparison of antigen constructs and carrier molecules for augmenting the immunogenicity of the monosaccharide epithelial cancer antigen Tn. Cancer Immunol. Immunother. 54: 424-430. [Medline]
  11. Lees, C. J., V. Apostolopoulos, B. Acres, I. Ramshaw, A. Ramsay, C. S. Ong, I. F. McKenzie. 2000. Immunotherapy with mannan-MUC1 and IL-12 in MUC1 transgenic mice. Vaccine 19: 158-162. [Medline]
  12. Snyder, L. A., T. J. Goletz, G. R. Gunn, F. F. Shi, M. C. Harris, K. Cochlin, C. McCauley, S. G. McCarthy, P. J. Branigan, D. M. Knight. 2006. A MUC1/IL-18 DNA vaccine induces anti-tumor immunity and increased survival in MUC1 transgenic mice. Vaccine 24: 3340-3352. [Medline]
  13. Soares, M. M., V. Mehta, O. J. Finn. 2001. Three different vaccines based on the 140-amino acid MUC1 peptide with seven tandemly repeated tumor-specific epitopes elicit distinct immune effector mechanisms in wild-type versus MUC1-transgenic mice with different potential for tumor rejection. J. Immunol. 166: 6555-6563. [Abstract/Free Full Text]
  14. Sorensen, A. L., C. A. Reis, M. A. Tarp, U. Mandel, K. Ramachandran, V. Sankaranarayanan, T. Schwientek, R. Graham, J. Taylor-Papadimitriou, M. A. Hollingsworth, et al 2006. Chemoenzymatically synthesized multimeric Tn/STn MUC1 glycopeptides elicit cancer-specific anti-MUC1 antibody responses and override tolerance. Glycobiology 16: 96-107. [Abstract/Free Full Text]
  15. Gilewski, T., S. Adluri, G. Ragupathi, S. Zhang, T. J. Yao, K. Panageas, M. Moynahan, A. Houghton, L. Norton, P. O. Livingston. 2000. Vaccination of high-risk breast cancer patients with mucin-1 (MUC1) keyhole limpet hemocyanin conjugate plus QS-21. Clin. Cancer Res. 6: 1693-1701. [Abstract/Free Full Text]
  16. Pecher, G., A. Haring, L. Kaiser, E. Thiel. 2002. Mucin gene (MUC1) transfected dendritic cells as vaccine: results of a phase I/II clinical trial. Cancer Immunol. Immunother. 51: 669-673. [Medline]
  17. Ramanathan, R. K., K. M. Lee, J. McKolanis, E. Hitbold, W. Schraut, A. J. Moser, E. Warnick, T. Whiteside, J. Osborne, H. Kim, et al 2005. Phase I study of a MUC1 vaccine composed of different doses of MUC1 peptide with SB-AS2 adjuvant in resected and locally advanced pancreatic cancer. Cancer Immunol. Immunother. 54: 254-264. [Medline]
  18. Rochlitz, C., R. Figlin, P. Squiban, M. Salzberg, M. Pless, R. Herrmann, E. Tartour, Y. Zhao, N. Bizouarne, M. Baudin, B. Acres. 2003. Phase I immunotherapy with a modified vaccinia virus (MVA) expressing human MUC1 as antigen-specific immunotherapy in patients with MUC1-positive advanced cancer. J. Gene Med. 5: 690-699. [Medline]
  19. von Mensdorff-Pouilly, S., E. Petrakou, P. Kenemans, K. van Uffelen, A. A. Verstraeten, F. G. Snijdewint, G. J. van Kamp, D. J. Schol, C. A. Reis, M. R. Price, et al 2000. Reactivity of natural and induced human antibodies to MUC1 mucin with MUC1 peptides and N-acetylgalactosamine (GalNAc) peptides. Int. J. Cancer 86: 702-712. [Medline]
  20. Janssen, E. M., N. M. Droin, E. E. Lemmens, M. J. Pinkoski, S. J. Bensinger, B. D. Ehst, T. S. Griffith, D. R. Green, S. P. Schoenberger. 2005. CD4+ T cell help controls CD8+ T cell memory via TRAIL-mediated activation-induced cell death. Nature 434: 88-93. [Medline]
  21. Janssen, E. M., E. E. Lemmens, T. Wolfe, U. Christen, M. G. von Herrath, S. P. Schoenberger. 2003. CD4+ T cells are required for secondary expansion and memory in CD8+ T lymphocytes. Nature 421: 852-856. [Medline]
  22. Schoenberger, S. P., R. E. Toes, E. I. van der Voort, R. Offringa, C. J. Melief. 1998. T cell help for cytotoxic T lymphocytes is mediated by CD40-CD40L interactions. Nature 393: 480-483. [Medline]
  23. Shedlock, D. J., H. Shen. 2003. Requirement for CD4 T cell help in generating functional CD8 T cell memory. Science 300: 337-339. [Abstract/Free Full Text]
  24. Sun, J. C., M. J. Bevan. 2003. Defective CD8 T cell memory following acute infection without CD4 T cell help. Science 300: 339-342. [Abstract/Free Full Text]
  25. Sun, J. C., M. A. Williams, M. J. Bevan. 2004. CD4+ T cells are required for the maintenance, not programming, of memory CD8+ T cells after acute infection. Nat. Immunol. 5: 927-933. [Medline]
  26. Hiltbold, E. M., A. M. Vlad, P. Ciborowski, S. C. Watkins, O. J. Finn. 2000. The mechanism of unresponsiveness to circulating tumor antigen MUC1 is a block in intracellular sorting and processing by dendritic cells. J. Immunol. 165: 3730-3741. [Abstract/Free Full Text]
  27. Carlos, C. A., H. F. Dong, O. M. Howard, J. J. Oppenheim, F. G. Hanisch, O. J. Finn. 2005. Human tumor antigen MUC1 is chemotactic for immature dendritic cells and elicits maturation but does not promote Th1 type immunity. J. Immunol. 175: 1628-1635. [Abstract/Free Full Text]
  28. Chen, D., S. Koido, Y. Li, S. Gendler, J. Gong. 2000. T cell suppression as a mechanism for tolerance to MUC1 antigen in MUC1 transgenic mice. Breast Cancer Res. Treat. 60: 107-115. [Medline]
  29. Tempero, R. M., M. L. VanLith, K. Morikane, G. J. Rowse, S. J. Gendler, M. A. Hollingsworth. 1998. CD4+ lymphocytes provide MUC1-specific tumor immunity in vivo that is undetectable in vitro and is absent in MUC1 transgenic mice. J. Immunol. 161: 5500-5506. [Abstract/Free Full Text]
  30. Rowse, G. J., R. M. Tempero, M. L. VanLith, M. A. Hollingsworth, S. J. Gendler. 1998. Tolerance and immunity to MUC1 in a human MUC1 transgenic murine model. Cancer Res. 58: 315-321. [Abstract/Free Full Text]
  31. Antony, P. A., C. A. Piccirillo, A. Akpinarli, S. E. Finkelstein, P. J. Speiss, D. R. Surman, D. C. Palmer, C. C. Chan, C. A. Klebanoff, W. W. Overwijk, et al 2005. CD8+ T cell immunity against a tumor/self-antigen is augmented by CD4+ T helper cells and hindered by naturally occurring T regulatory cells. J. Immunol. 174: 2591-2601. [Abstract/Free Full Text]
  32. Curiel, T. J., G. Coukos, L. Zou, X. Alvarez, P. Cheng, P. Mottram, M. Evdemon-Hogan, J. R. Conejo-Garcia, L. Zhang, M. Burow, et al 2004. Specific recruitment of regulatory T cells in ovarian carcinoma fosters immune privilege and predicts reduced survival. Nat. Med. 10: 942-949. [Medline]
  33. Shimizu, J., S. Yamazaki, S. Sakaguchi. 1999. Induction of tumor immunity by removing CD25+CD4+ T cells: a common basis between tumor immunity and autoimmunity. J. Immunol. 163: 5211-5218. [Abstract/Free Full Text]
  34. Zou, W.. 2006. Regulatory T cells, tumour immunity and immunotherapy. Nat. Rev. Immunol. 6: 295-307. [Medline]
  35. Takahashi, T., Y. Kuniyasu, M. Toda, N. Sakaguchi, M. Itoh, M. Iwata, J. Shimizu, S. Sakaguchi. 1998. Immunologic self-tolerance maintained by CD25+CD4+ naturally anergic and suppressive T cells: induction of autoimmune disease by breaking their anergic/suppressive state. Int. Immunol. 10: 1969-1980. [Abstract/Free Full Text]
  36. Thornton, A. M., E. M. Shevach. 2000. Suppressor effector function of CD4+CD25+ immunoregulatory T cells is antigen nonspecific. J. Immunol. 164: 183-190. [Abstract/Free Full Text]
  37. Itoh, M., T. Takahashi, N. Sakaguchi, Y. Kuniyasu, J. Shimizu, F. Otsuka, S. Sakaguchi. 1999. Thymus and autoimmunity: production of CD25+CD4+ naturally anergic and suppressive T cells as a key function of the thymus in maintaining immunologic self-tolerance. J. Immunol. 162: 5317-5326. [Abstract/Free Full Text]
  38. Feili-Hariri, M., D. H. Falkner, P. A. Morel. 2002. Regulatory Th2 response induced following adoptive transfer of dendritic cells in prediabetic NOD mice. Eur. J. Immunol. 32: 2021-2030. [Medline]
  39. Feili-Hariri, M., D. H. Falkner, P. A. Morel. 2005. Polarization of naive T cells into Th1 or Th2 by distinct cytokine-driven murine dendritic cell populations: implications for immunotherapy. J. Leukocyte Biol. 78: 656-664. [Abstract/Free Full Text]
  40. Jonuleit, H., E. Schmitt, G. Schuler, J. Knop, A. H. Enk. 2000. Induction of interleukin 10-producing, nonproliferating CD4+ T cells with regulatory properties by repetitive stimulation with allogeneic immature human dendritic cells. J. Exp. Med. 192: 1213-1222. [Abstract/Free Full Text]
  41. Kuwana, M.. 2002. Induction of anergic and regulatory T cells by plasmacytoid dendritic cells and other dendritic cell subsets. Hum. Immunol. 63: 1156-1163. [Medline]
  42. Lutz, M. B., G. Schuler. 2002. Immature, semi-mature and fully mature dendritic cells: which signals induce tolerance or immunity?. Trends Immunol. 23: 445-449. [Medline]
  43. Roncarolo, M. G., M. K. Levings, C. Traversari. 2001. Differentiation of T regulatory cells by immature dendritic cells. J. Exp. Med. 193: F5-F9. [Free Full Text]
  44. Belkaid, Y., C. A. Piccirillo, S. Mendez, E. M. Shevach, D. L. Sacks. 2002. CD4+CD25+ regulatory T cells control Leishmania major persistence and immunity. Nature 420: 502-507. [Medline]
  45. Caramalho, I., T. Lopes-Carvalho, D. Ostler, S. Zelenay, M. Haury, J. Demengeot. 2003. Regulatory T cells selectively express Toll-like receptors and are activated by lipopolysaccharide. J. Exp. Med. 197: 403-411. [Abstract/Free Full Text]
  46. Steinman, R. M., M. C. Nussenzweig. 2002. Avoiding horror autotoxicus: the importance of dendritic cells in peripheral T cell tolerance. Proc. Natl. Acad. Sci. USA 99: 351-358. [Abstract/Free Full Text]
  47. Dubsky, P., H. Ueno, B. Piqueras, J. Connolly, J. Banchereau, A. K. Palucka. 2005. Human dendritic cell subsets for vaccination. J. Clin. Immunol. 25: 551-572. [Medline]
  48. Masteller, E. L., M. R. Warner, Q. Tang, K. V. Tarbell, H. McDevitt, J. A. Bluestone. 2005. Expansion of functional endogenous antigen-specific CD4+CD25+ regulatory T cells from nonobese diabetic mice. J. Immunol. 175: 3053-3059. [Abstract/Free Full Text]
  49. Tarbell, K. V., S. Yamazaki, K. Olson, P. Toy, R. M. Steinman. 2004. CD25+CD4+ T cells, expanded with dendritic cells presenting a single autoantigenic peptide, suppress autoimmune diabetes. J. Exp. Med. 199: 1467-1477. [Abstract/Free Full Text]
  50. Yamazaki, S., T. Iyoda, K. Tarbell, K. Olson, K. Velinzon, K. Inaba, R. M. Steinman. 2003. Direct expansion of functional CD25+CD4+ regulatory T cells by antigen-processing dendritic cells. J. Exp. Med. 198: 235-247. [Abstract/Free Full Text]
  51. Yamazaki, S., M. Patel, A. Harper, A. Bonito, H. Fukuyama, M. Pack, K. V. Tarbell, M. Talmor, J. V. Ravetch, K. Inaba, R. M. Steinman. 2006. Effective expansion of alloantigen-specific Foxp3+CD25+CD4+ regulatory T cells by dendritic cells during the mixed leukocyte reaction. Proc. Natl. Acad. Sci. USA 103: 2758-2763. [Abstract/Free Full Text]
  52. Bos, R., S. van Duikeren, T. van Hall, P. Kaaijk, R. Taubert, B. Kyewski, L. Klein, C. J. Melief, R. Offringa. 2005. Expression of a natural tumor antigen by thymic epithelial cells impairs the tumor-protective CD4+ T cell repertoire. Cancer Res. 65: 6443-6449. [Abstract/Free Full Text]
  53. Woo, E. Y., H. Yeh, C. S. Chu, K. Schlienger, R. G. Carroll, J. L. Riley, L. R. Kaiser, C. H. June. 2002. Cutting edge: regulatory T cells from lung cancer patients directly inhibit autologous T cell proliferation. J. Immunol. 168: 4272-4276. [Abstract/Free Full Text]
  54. Herrem, C. J., T. Tatsumi, K. S. Olson, K. Shirai, J. H. Finke, R. M. Bukowski, M. Zhou, A. L. Richmond, I. Derweesh, M. S. Kinch, W. J. Storkus. 2005. Expression of EphA2 is prognostic of disease-free interval and overall survival in surgically treated patients with renal cell carcinoma. Clin. Cancer Res. 11: 226-231. [Abstract/Free Full Text]
  55. Bluestone, J. A., A. K. Abbas. 2003. Natural versus adaptive regulatory T cells. Nat. Rev. Immunol. 3: 253-257. [Medline]
  56. Frey, O., P. K. Petrow, M. Gajda, K. Siegmund, J. Huehn, A. Scheffold, A. Hamann, A. Radbruch, R. Brauer. 2005. The role of regulatory T cells in antigen-induced arthritis: aggravation of arthritis after depletion and amelioration after transfer of CD4+CD25+ T cells. Arthritis Res. Ther. 7: R291-301. [Medline]
  57. Kumar, V., K. Stellrecht, E. Sercarz. 1996. Inactivation of T cell receptor peptide-specific CD4 regulatory T cells induces chronic experimental autoimmune encephalomyelitis (EAE). J. Exp. Med. 184: 1609-1617. [Abstract/Free Full Text]
  58. Sakaguchi, S., N. Sakaguchi, M. Asano, M. Itoh, M. Toda. 1995. Immunologic self-tolerance maintained by activated T cells expressing IL-2 receptor {alpha}-chains (CD25): breakdown of a single mechanism of self-tolerance causes various autoimmune diseases. J. Immunol. 155: 1151-1164. [Abstract]
  59. Stephens, L. A., D. Gray, S. M. Anderton. 2005. CD4+CD25+ regulatory T cells limit the risk of autoimmune disease arising from T cell receptor crossreactivity. Proc. Natl. Acad. Sci. USA 102: 17418-17423. [Abstract/Free Full Text]
  60. Suri-Payer, E., A. Z. Amar, A. M. Thornton, E. M. Shevach. 1998. CD4+CD25+ T cells inhibit both the induction and effector function of autoreactive T cells and represent a unique lineage of immunoregulatory cells. J. Immunol. 160: 1212-1218. [Abstract/Free Full Text]
  61. Chen, X., T. Murakami, J. J. Oppenheim, O. M. Howard. 2004. Differential response of murine CD4+CD25+ and CD4+CD25– T cells to dexamethasone-induced cell death. Eur. J. Immunol. 34: 859-869. [Medline]



This article has been cited by other articles:


Home page
 Clin. Cancer Res.Home page
S. Gnjatic, N. K. Altorki, D. N. Tang, S.-M. Tu, V. Kundra, G. Ritter, L. J. Old, C. J. Logothetis, and P. Sharma
NY-ESO-1 DNA Vaccine Induces T-Cell Responses That Are Suppressed by Regulatory T Cells
Clin. Cancer Res., March 15, 2009; 15(6): 2130 - 2139.
[Abstract] [Full Text] [PDF]

Home page
 BloodHome page
C. Ding, L. Wang, J. Marroquin, and J. Yan
Targeting of antigens to B cells augments antigen-specific T-cell responses and breaks immune tolerance to tumor-associated antigen MUC1
Blood, October 1, 2008; 112(7): 2817 - 2825.
[Abstract] [Full Text] [PDF]

This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow Request Permissions
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Turner, M. S.
Right arrow Articles by Finn, O. J.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Turner, M. S.
Right arrow Articles by Finn, O. J.
Right arrowPubmed/NCBI databases
*Substance via MeSH

HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS