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[Cancer Research 62, 1471-1476, March 1, 2002]
© 2002 American Association for Cancer Research

Immunology

Cellular Processing of a Multibranched Lysine Core with Tumor Antigen Peptides and Presentation of Peptide Epitopes Recognized by Cytotoxic T Lymphocytes on Antigen-presenting Cells1

Seisuke Ota, Toshiro Ono, Akimichi Morita, Akiko Uenaka, Mine Harada and Eiichi Nakayama2

Departments of Immunology [S. O., T. O., A. U., E. N.] and Internal Medicine [S. O., M. H.], Okayama University Graduate School of Medicine and Dentistry, Okayama 700-8558, and Department of Dermatology, Nagoya City University Medical School, Nagoya 467-8601 [A.M.], Japan


   ABSTRACT
Top
 ABSTRACT
INTRODUCTION
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 REFERENCES
 
We showed that pRL1a multiple antigen peptide (MAP)-sensitized dendritic cell (DC) and P815 cell lysis by pRL1a-specific B-24 CTL was blocked by incubating target cells at 4°C during sensitization. The finding suggested that pRL1a MAP was mostly internalized in DC and P815 cells and produced pRL1a peptide epitopes for presentation with H-2Ld. Furthermore, we showed that sensitization with pRL1a MAP was inhibited by the addition of chloroquine, cycloheximide, and brefeldin A to the culture, but not by the addition of inhibitors for lysosomal proteases or proteasome. Inhibition of sensitization by the addition of chloroquine to the culture suggested the requirement of acidification of the endosomal compartment for pRL1a MAP processing. Inhibition of sensitization by the addition of cycloheximide and brefeldin A to the culture indicated the requirement of newly generated MHC class I antigen molecules and the involvement of transport of the peptide MHC class I complex from the endoplasmic reticulum to the Golgi. The findings suggested that pRL1a MAP in the endosomal compartment leaked to the cytosol, and degraded, and the pRL1a peptide produced was presented by the MHC class I pathway.


   INTRODUCTION
Top
 ABSTRACT
 INTRODUCTION
MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 REFERENCES
 
Tumor antigen peptides recognized by T cells have been identified in various types of murine and human tumors (1, 2, 3, 4, 5) . Those peptides have been used as tumor vaccines in murine model tumors (6, 7, 8, 9) and in clinical trials (10, 11, 12, 13) . From those studies, it is generally accepted that the tumor antigen peptides are weakly immunogenic compared with the antigen peptides derived from infectious agents (14 , 15) , which were easily shown to be effective for prophylaxis of the diseases. This could be attributable to low antigenicity of the tumor antigen peptide itself and to the sensitivity to proteases of the peptide resulting in rapid degradation. Various kinds of adjuvants (16 , 17) , cytokines (18 , 19) , and DCs3 (11 , 20) have been used to augment immunogenicity of the tumor antigen peptides but have showed limited success.

MAP is a multibranched lysine core with antigenic peptides. Tam (21) and Posnett et al. (22) showed that MAP enhanced specific antibody responses. They also showed that lipid-conjugated MAP containing the CTL epitope of HIV proteins induced both humoral and peptide-specific CTL responses (23 , 24) . Using the antigen peptide pRL1a present on a murine leukemia RL male 1 that was recognized by CTL in association with H-2Ld (25) , we previously showed that immunization with pRL1a MAP, but not pRL1a peptide itself, efficiently induced in vitro CTL generation and inhibited in vivo RL male 1 tumor growth (9) . In this study, we showed the internalization and cellular processing of pRL1a MAP in DCs and mastocytoma P815 cells, and presentation of the antigen peptide pRL1a on the cell surface.


   MATERIALS AND METHODS
Top
 ABSTRACT
 INTRODUCTION
 MATERIALS AND METHODS
RESULTS
 DISCUSSION
 REFERENCES
 
Mice.
Female BALB/c mice were bred in our Animal Center for Medical Research and used when between 8 and 12 weeks of age.

Tumors and Cell Lines.
RL male 1 is a BALB/c radiation-induced leukemia (26) . P815 is a DBA/2 methylcholanthrene-induced mastocytoma (27) .

Inhibitors.
CHQ, an acidification blocker of the endosomal compartment (28) , CHX, a protein synthesis inhibitor (29) , and BFA, a protein trafficking inhibitor from the ER to the Golgi (30) , were purchased from Sigma Chemical Co. (St. Louis, MO). Proteasome inhibitor LLnL (31) was purchased from Calbiochem-Novabiochem Corporation (La Jolla, CA). Proteasome inhibitors lactacystin (32) and MG132 (31) , and lysosomal protease inhibitor E64d (33) were provided by Drs. H. Udono and T. Yamano (Department of Molecular Medicine, Nagasaki University School of Medicine, Nagasaki, Japan).

Peptide Synthesis.
pRL1a peptide (IPGLPLSL) was synthesized by standard solid-phase methods using N-(9-fluorenyl)methoxycarbonyl (Fmoc) chemistry using a peptide synthesizer (model 430A; Perkin-Elmer Applied Biosystems, Foster City, CA). pRL1a MAP is an octabranched ß-alanyl hydroxyl lysine with pRL1a peptides (Fig. 1)Citation synthesized using Fmoc MAP Resin (Perkin-Elmer Applied Biosystems). The Mr of pRL1a MAP is 7930, ~80% of which is a pRL1a peptide component, with the remaining 20% being a lysine core. The peptide was purified by reversed-phase HPLC.



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  		Fig. 1. Structure of pRL1a MAP. pRL1a MAP was synthesized using eight-branched ß-alanyl MAP resin. The Mr is 7930, about 80% of which is a pRL1a peptide component.

 
CTL Generation by in Vivo and in Vitro Sensitization with the Peptide.
BALB/c mice were given injections of 120 µg of pRL1a MAP or 100 µg of pRL1a into the footpad twice at an interval of 2 weeks. After 2 weeks, spleen cells were obtained and stimulated in vitro with the indicated concentrations of pRL1a MAP or pRL1a in 24-well tissue culture plates (Falcon 3047, Becton Dickinson, Lincoln Park, NJ) for 5 days at 37°C in a 5% CO2 atmosphere. The medium used was RPMI 1640 (Life Technologies, Inc. Laboratories, Grand Island, NY) supplemented with 10% FCS, 2 mM L-glutamine, 100 units/ml of penicillin, 100 µg/ml of streptomycin, and 50 µM 2-mercaptoethanol.

Establishment and Maintenance of CTL Clone B-24.
pRL1a-specific CTL clone B-24 was established from RL male 1-bearing BALB/c spleen cells (34) and maintained by repetitive stimulation with mitomycin C-treated RL male 1 cells (5 x 105) and splenic feeder cells (5 x 106) in the presence of human recombinant interleukin-2 (5 units/ml; Takeda Pharmaceutical, Tokyo, Japan) in 24-well culture plates.

Cytotoxicity Assay.
Cytotoxicity was determined by a 3.5-h 51Cr-release assay as described previously (9) .

For inhibition, P815 cells, which had been maintained in a serum-free AIM-V (Life Technologies, Inc. Laboratories) medium were 51Cr labeled and used as targets. They were preincubated with inhibitors for 45 min at 37°C before adding pRL1a MAP or pRL1a peptide. Assays were performed in AIM-V medium. Inhibitors were present throughout the assay.

Generation of DCs.
Bone marrow-derived DCs were generated as described previously (35) . Briefly, bone marrow cells were cultured for 10 days in the RPMI 1640 supplemented with recombinant GM-CSF (500 units/ml), provided by Dr. T. Sudo (Basic Research Laboratories, Toray Industries, Inc., Kanagawa, Japan). The aggregates were subcultured in fresh medium with recombinant GM-CSF (500 units/ml) and used as targets in the 51Cr-release assay.

Immunofluorescence Staining.
pRL1a MAP was labeled with FITC (Boehringer Mannheim, Indianapolis, IN) according to the manufacturer’s directions. Free FITC molecules were removed by reversed-phase HPLC. The cells were stained with LysoTracker Red (Molecular probes, Eugene, OR) at 37°C for 30 min to indicate the endosomes in red. Intracellular staining was analyzed by immunofluorescence confocal microscopy (LSM510; Carl Zeiss, Jena, Germany).


   RESULTS
Top
 ABSTRACT
 INTRODUCTION
 MATERIALS AND METHODS
 RESULTS
DISCUSSION
 REFERENCES
 
Generation of CTLs in BALB/c Spleen Cells by Sensitization with pRL1a MAP but not with pRL1a.
pRL1a (IPGLPLSL) is the H-2Ld-binding peptide on BALB/c RL male 1 leukemia recognized by CTLs (25) . Generation of CTLs in BALB/c spleen cells was investigated by in vivo and in vitro sensitization with pRL1a MAP or by the original pRL1a peptide. Spleen cells were obtained from BALB/c mice that had been given injections of pRL1a MAP or pRL1a into the footpad twice and stimulated in vitro with pRL1a MAP or pRL1a, respectively. Cytotoxicity was assayed on 51Cr-labeled P815 target cells pulsed with 1 x 10-5 M pRL1a. As shown in Table 1Citation , sensitization with pRL1a MAP, but not pRL1a, generated cytotoxicity. No generation of cytotoxicity was observed with in vivo pRL1a MAP sensitization alone. The cytotoxicity was blocked by anti-CD8 mAb and anti-H-2Ld mAb, but not anti-CD4 mAb, anti-H-2Kd mAb, or anti-H-2Dd mAb (data not shown). The results confirmed the previous findings published by Manki et al. (9) .


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  		Table 1 Generation of CTL in BALB/c spleen cells by in vivo and in vitro sensitization with pRL1a MAP, but not pRL1a

In vivo sensitization was performed by injecting 120 µg of pRL1a MAP or 100 µg of pRL1a into the footpad twice at an interval of 2 weeks. Two weeks later, spleen cells were obtained and incubated with pRL1a MAP or pRL1a for 5 days and used as effector cells. The target cells were pRL1a (10 µM)-pulsed P815 cells. The cytotoxicity was determined by a 3.5-h 51Cr-release assay.

 
Presentation of pRL1a Peptide on DCs and P815 by in Vitro Sensitization with pRL1a MAP.
Presentation of pRL1a peptide on DCs and P815 by in vitro sensitization with pRL1a MAP was investigated for lysis of the target cells by pRL1a-specific B-24 CTL in serum-free medium. As shown in Fig. 2Citation , lysis was observed with both DCs and P815 cells.



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  		Fig. 2. pRL1a MAP-sensitized target cell lysis by B-24 CTL. 51Cr-labeled DCs (A) and P815 cells (B) were sensitized with pRL1a MAP (1 x 10-6 M) and pRL1a (1 x 10-5 M) for 2 h at 37°C. In (C), 51Cr-labeled P815 cells were sensitized with pRL1a MAP or pRL1a at the indicated concentrations for 2 h at 37°C. The cells were washed and used as targets for pRL1a-specific B-24 CTLs in a 3.5-h 51Cr-release assay. E:T ratio was 20 in C. Peptide sensitization and assays were done in serum-free medium.

 
Furthermore, the peptide dose required to sensitize P815 target cells for lysis by B-24 CTL was examined. For optimal sensitization for lysis, 1 x 10-7 M pRL1a MAP was necessary. In contrast, as low as 1 x 10-9 M pRL1a was sufficient for sensitization.

Abrogation of pRL1a MAP-sensitized DC and P815 Target-Cell Lysis by Incubating at 4°C.
Labeled DCs and P815 target cells were incubated with pRL1a MAP for 1 h at 4°C and were used as targets for B-24 CTLs in serum-free medium. As shown in Fig. 3Citation , lysis was abrogated. No abrogation was observed with pRL1a-sensitized DC or P815 target cell lysis. The results suggested that internalization was necessary for presentation of pRL1a peptide on DCs and P815 by sensitization with pRL1a MAP.



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  		Fig. 3. Abrogation of target cell lysis by sensitization with pRL1a MAP at 4°C. 51Cr-labeled DCs and P815 cells were sensitized with pRL1a MAP (1 x 10-6 M; A and C) or pRL1a (1 x 10-5 M; B and D) for 1 h at 37°C or 4°C. The cells were washed and used as targets for pRL1a-specific B-24 CTLs in a 3.5-h 51Cr-release assay. Peptide sensitization and assays were done in serum-free medium.

 
After this, the effect of various inhibitors on the MHC class I presentation pathway was investigated with P815.

The Effect of CHQ on pRL1a MAP-sensitized P815 Target Cell Lysis.
The effect of CHQ on pRL1a MAP-sensitized P815 target cell lysis by B-24 CTL was investigated in serum-free medium. As shown in Fig. 4Citation , CHQ blocked pRL1a MAP but not pRL1a-sensitized P815 lysis.



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  		Fig. 4. Inhibition of pRL1a MAP-sensitized P815 target cell lysis by CHQ. P815 cells were sensitized with pRL1a MAP (1 x 10-6 M) or pRL1a (1 x 10-5 M) for 1 h at 37°C and were 51Cr labeled, washed, and used as targets for pRL1a-specific B-24 CTLs in 3.5-h 51Cr-release assay. The indicated doses of CHQ were present from 45 min before incubation with the peptides, during sensitization, and throughout assay. Treatment with chemicals, peptide sensitization, and assays were done in serum-free medium.

 
The Effect of Inhibitors for Lysosomal Proteases or Proteasomes on Processing of pRL1a MAP.
The involvement of the lysosomal proteases and proteasomes for processing pRL1a MAP was investigated using various kinds of inhibitor. First, we examined the effect of lysosomal protease inhibitor E64d on pRL1a MAP-sensitized P815 target cell lysis by B-24 CTL in serum-free medium. As shown in Fig. 5Citation , no specific inhibition of target cell lysis was observed. Then, we examined the effect of LLnL, lactacystin, and MG132 on lysis. These proteasomal inhibitors differ in their mechanisms of action. LLnL and MG132 were also shown to be inhibitors for cathepsin B and calpains, respectively. None of them inhibited lysis.



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  		Fig. 5. P815 cells were sensitized with pRL1a MAP (1 x 10-6 M) or pRL1a (1 x 10-5 M) for 1 h at 37°C and were 51Cr labeled, washed, and used as targets for pRL1a-specific B-24 CTLs in a 3.5-h 51Cr-release assay. The indicated doses of E64d (A), or LLnL, lactacystin, or MG132 (B) were present from 45 min before incubation with the peptides, during sensitization, and throughout assay. E:T ratio was 20:1. Treatment with chemicals, peptide sensitization, and assays were done in serum-free medium.

 
The Effect of CHX and BFA on pRL1a MAP-sensitized P815 Target Cell Lysis.
The effects on pRLa MAP-sensitized P815 target cell lysis by B-24 CTLs of CHX and BFA, inhibitors, respectively, of protein synthesis and membranous and/or vesicular flow from the ER to the Golgi, were studied in serum-free medium. As shown in Fig. 6Citation , CHX and BFA dose-dependently inhibited lysis. No inhibition was observed with lysis of pRL1a-sensitized P815 target cells. The results suggested that pRL1a generated from pRL1a MAP binds newly synthesized H-2Ld in ER and is transported through the Golgi to the cell surface.



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  		Fig. 6. P815 cells were sensitized with pRL1a MAP (1 x 10-6 M) or pRL1a (1 x 10-5 M) for 1 h at 37°C and were 51Cr labeled, washed, and used as targets for pRL1a-specific B-24 CTLs in a 3.5-h 51Cr-release assay. The indicated doses of CHX (A) or BFA (B) were present from 45 min before incubation with the peptides, during sensitization, and throughout assay. E:T ratio was 20:1 in A and 10:1 in B. Treatment with chemicals, peptide sensitization, and assays were done in serum-free medium.

 
P815 Lysis by B-24 CTLs after Various Sensitization Periods.
P815 cells were incubated with pRL1a MAP for various time periods and used as targets for B-24 CTLs. The assay was carried out in the presence of BFA to block further processing in serum-free medium. As shown in Fig. 7Citation , 30 min was sufficient for optimal sensitization for lysis. In contrast, less than 10 min was sufficient for pRL1a sensitization.



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  		Fig. 7. Kinetic analysis of presentation of pRL1a from pRL1a MAP. 51Cr-labeled P815 cells were incubated with pRL1a MAP (1 x 10-6 M) or pRL1a (1 x 10-5 M) for various time periods and used as targets for pRL1a-specific B-24 CTLs in a 3.5-h 51Cr-release assay. The assay was carried out in the presence of 5 µg/ml BFA. Peptide sensitization and assays were done in serum-free medium.

 
Confocal Microscopy of the Intracellular Localization of pRL1a MAP in P815 cells.
P815 cells were incubated with FITC-conjugated pRL1a MAP and analyzed by confocal microscopy. As shown in Fig. 8Citation , FITC-conjugated pRL1a MAP was colocalized in lysosomes 30 min after incubation and was accumulated in the vesicles. At the later time points, cytoplasmic localization was gradually increased.



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  		Fig. 8. Confocal microscopy. P815 cells were incubated with FITC-conjugated pRL1a MAP together with (A) or without (B) LysoTracker Red. FITC-pRL1a MAP (in A, green) and (in B, red). Incubation time of A was 30 min.

 

   DISCUSSION
Top
 ABSTRACT
 INTRODUCTION
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
REFERENCES
 
In this study, we showed that pRL1a MAP-sensitized DC and P815 cell lysis by pRL1a specific B-24 CTL was blocked by incubating at 4°C during sensitization. pRL1a MAP was a Mr 7930 molecule composed of a branched lysine core with eight pRL1a peptides and could easily release pRL1a peptide epitopes by degradation. However, the findings suggested that pRL1a MAP was mostly internalized in DCs and P815 cells and produced pRL1a peptide epitopes for presentation with H-2Ld. Furthermore, we showed that sensitization with pRL1a MAP was inhibited by the addition of CHQ, CHX, and BFA to the culture, but not by the addition of inhibitors for lysosomal proteases or proteasome. Inhibition of sensitization by the addition of CHQ to the culture suggested that acidification of the endosomal compartment for pRL1a MAP processing was required. Inhibition of sensitization by the addition of CHX and BFA to the culture indicated the requirement of newly generated MHC class I antigen molecules and the involvement of transport of the peptide MHC class I complex from the ER to the Golgi. The findings suggested that pRL1a MAP in the endosomal compartment leaked to the cytosol and degraded, and that the pRL1a peptide produced was presented by the MHC class I pathway.

It was shown previously that high antigen concentration (36) or interaction of synthetic polymeric particles with phagosomal membranes (37, 38, 39) facilitated antigen delivery to the cytosol by inducing artificial leakage from the endocytic compartment. On the other hand, Rodriguez et al. (40) reported that DCs developed a unique membrane transport pathway linking the lumens of endocytic compartments and the cytosol. The transport was specific to internalized antigens and selective for the size of the molecules. The optimal concentration of pRL1a MAP for sensitization of DCs and P815 cells was 1 x 10-6 to 1 x 10-7 M, being similar to that of, e.g., the OVA immune complex (40) and outer membrane protein A from Klebsiella pneumoniae (41) , with which selective transport of the internalized molecules to the cytosol for MHC class I presentation was recently demonstrated. On the other hand, the concentration of OVA causing artificial leakage (39 , 42 , 43) was 100- to 200-fold higher than the concentration of pRL1a MAP used in this study. Furthermore, pRL1a MAP may interact with the endosomal membranes because of its simple structure and could be easily transported to the cytosol because of its small size.

The absence of inhibition of pRL1a MAP processing by the inhibitors of lysosomal proteases suggested that no peptide degradation occurred in the endosomal compartment, although acidification was required. Acidic conditions may be necessary for solving aggregated pRL1a MAP molecules as demonstrated with the OVA immune complex (40) . Rather, the findings suggested that pRL1a was produced from pRL1a MAP in the cytosol. The absence of inhibition of the processing by the addition of proteasome inhibitors in the culture was consistent with the findings of others (44, 45, 46, 47, 48) . The mechanism for the epitope peptide pRL1a generation from pRL1a MAP in the cytosol is unknown. Lysine adjacent to the COOH termius of the epitope peptide could provide the target site for proteases with trypsin-like activity present in the cytosol. The epitope peptide would be subsequently produced by deleting lysine. The epitope production could be facilitated by the presence of eight repeats of the pRL1a peptide in a single pRL1a MAP molecule.

Tumor antigen peptides recognized by CD8 and CD4 T cells have been defined on various types of tumors (4 , 5) . It has been generally accepted from the experimental and clinical studies that tumor antigen peptides were weakly immunogenic. A tumor vaccine that could efficiently elicit a protective immune response needs to be developed. It was shown that the use of adjuvant and lipid membrane with the antigen peptides augmented immunogenicity. In this study, we showed that MAP could be another molecule to augment immunogenicity with potentially no adverse effects observed in the clinical use of adjuvant. Immunization with pRL1a MAP efficiently generated a specific CTL response in BALB/c mice and protected the mice from RL male 1 tumor growth (9) . This study showed the processing of MAP in the MHC class I pathway and elucidated the basis for the effect of MAP as a tumor vaccine.


   ACKNOWLEDGMENTS
 
We thank Drs. Heiichiro Udono and Taketoshi Yamano for providing the proteasome inhibitors, and Dr. Tetsuo Sudo for the GM-CSF. We also thank Junko Mizuuchi for her help in the preparation of the manuscript and Midori Isobe for her excellent technical assistance.


   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 Supported in part by a Grant-in-Aid for Scientific Research on Priority Areas (C) from the Ministry of Education, Culture, Sports, Science, and Technology, Japan. Back

2 To whom requests for reprints should be addressed, at Department of Immunology, Okayama University Graduate School of Medicine and Dentistry, 2-5-1 Shikata-cho, Okayama 700-8558, Japan. Phone: 81-86-235-7187; Fax: 81-86-235-7193; E-mail: nakayama{at}md.okayama-u.ac.jp. Back

3 The abbreviations used are: DC, dendritic cell; MAP, multiple antigen peptide; CHQ, chloroquine; LLnL, N-acetyl-Leu-Leu-norleucinal; CHX, cycloheximide; BFA, brefeldin A; ER, endoplasmic reticulum; HPLC, high-performance liquid chromatography; GM-CSF, granulocyte macrophage colony-stimulating factor; mAb, monoclonal antibody; OVA, ovalbumin. Back

Received 9/19/01. Accepted 1/ 3/02.


   REFERENCES
Top
 ABSTRACT
 INTRODUCTION
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 REFERENCES
 

  1. Boon T., Cerottini J-C., Van den Eynde B., van der Bruggen P., Van Pel A. Tumor antigens recognized by T lymphocytes. Annu. Rev. Immunol., 12: 337-365, 1994.[Medline]
  2. Jaffee E. M., Pardoll D. M. Murine tumor antigens: is it worth the search?. Curr. Opin. Immunol., 8: 622-627, 1996.[Medline]
  3. Boon T., Old L. J. Tumor antigens. Curr. Opin. Immunol., 9: 681-683, 1997.[Medline]
  4. Van den Eynde B., van der Bruggen P. T cell defined tumor antigens. Curr. Opin. Immunol., 9: 684-693, 1997.[Medline]
  5. Rosenberg S. A. A new era for cancer immunotherapy based on the genes that encode cancer antigens. Immunity, 10: 281-287, 1999.[Medline]
  6. Mayordomo J. I., Zorina T., Storkus W. J., Zitvogel L., Celluzzi C., Falo L. D., Melief C. J., Ildstad S. T., Kast W. M., DeLeo A. B., Lotze M. T. Bone marrow-derived dendritic cells pulsed with synthetic tumour peptides elicit protective and therapeutic antitumour immunity. Nat. Med., 1: 1297-1302, 1995.[Medline]
  7. Nagata Y., Furugen R., Hiasa A., Ikeda H., Ohta N., Furukawa K., Nakamura H., Furukawa K., Kanematsu T., Shiku H. Peptides derived from a wild-type murine proto-oncogene c-erbB-2/HER2/neu can induce CTL and tumor suppression in syngeneic hosts. J. Immunol., 159: 1336-1343, 1997.[Abstract]
  8. Ikeda H., Ohta N., Furukawa K., Miyazaki H., Wang L., Kuribayashi K., Old L. J., Shiku H. Mutated mitogen-activated protein kinase: a tumor rejection antigen of mouse sarcoma. Proc. Natl. Acad. Sci. USA, 94: 6375-6379, 1997.[Abstract/Free Full Text]
  9. Manki A., Ono T., Uenaka A., Seino Y., Nakayama E. Vaccination with multiple antigen peptide as rejection antigen peptide in murine leukemia. Cancer Res., 58: 1960-1964, 1998.[Abstract/Free Full Text]
  10. Rosenberg S. A., Yang J. C., Schwartzentruber D. J., Hwu P., Marincola F. M., Topalian S. L., Restifo N. P., Dudley M. E., Schwarz S. L., Spiess P. J., Wunderlich J. R., Parkhurst M. R., Kawakami Y., Seipp C. A., Einhorn J. H., White D. E. Immunologic and therapeutic evaluation of a synthetic peptide vaccine for the treatment of patients with metastatic melanoma. Nat. Med., 4: 321-327, 1998.[Medline]
  11. Thurner B., Haendle I., Roder C., Dieckmann D., Keikavoussi P., Jonuleit H., Bender A., Maczek C., Schreiner D., von den Driesch P., Brocker E. B., Steinman R. M., Enk A., Kampgen E., Schuler G. Vaccination with MAGE-3A1 peptide-pulsed mature, monocyte-derived dendritic cells expands specific cytotoxic T cells and induces regression of some metastases in advanced stage IV melanoma. J. Exp. Med., 190: 1669-1678, 1999.[Abstract/Free Full Text]
  12. Marchand M., van Baren N., Weynants P., Brichard V., Dreno B., Tessier M. H., Rankin E., Parmiani G., Arienti F., Humblet Y., Bourlond A., Vanwijck R., Lienard D., Beauduin M., Dietrich P. Y., Russo V., Kerger J., Masucci G., Jäger E., De Greve J., Atzpodien J., Brasseur F., Coulie P. G., van der Bruggen P., Boon T. Tumor regressions observed in patients with metastatic melanoma treated with an antigenic peptide encoded by gene MAGE-3 and presented by HLA-A1. Int. J. Cancer, 80: 219-230, 1999.[Medline]
  13. Jäger E., Gnjatic S., Nagata Y., Stockert E., Jäger D., Karbach J., Neumann A., Rieckenberg J., Chen Y-T., Ritter G., Hoffman E., Arand M., Old L. J., Knuth A. Induction of primary NY-ESO-1 immunity: CD8+ T lymphocyte and antibody responses in peptide-vaccinated patients with NY-ESO-1+ cancers. Proc. Natl. Acad. Sci. USA, 97: 12198-12203, 2000.[Abstract/Free Full Text]
  14. Harty J. T., Bevan M. J. CD8+ T cells specific for a single nonamer epitope of Listeria monocytogenes are protective in vivo. J. Exp. Med., 175: 1531-1538, 1992.[Abstract/Free Full Text]
  15. Blake N., Haigh T., Shaka’a G., Croom-Carter D., Rickinson A. The importance of exogenous antigen in priming the human CD8+ T cell response: Lessons from the EBV nuclear antigen EBNA 1. J. Immunol., 165: 7078-7087, 2000.[Abstract/Free Full Text]
  16. Singh M., O’Hagen D. Advances in vaccine adjuvants. Nat. Biotechnol., 17: 1075-1081, 1999.[Medline]
  17. Schmidt W., Buschle M., Zauner W., Kirlappos H., Mechtler K., Trska B., Birnstiel M. L. Cell-free tumor antigen peptide-based cancer vaccines. Proc. Natl. Acad. Sci. USA, 94: 3262-3267, 1997.[Abstract/Free Full Text]
  18. Jäger E., Ringhoffer M., Dienes H-P., Arand M., Karbach J., Jäger D., Ilsemann C., Hagedorn M., Oesch F., Knuth A. Granulocyte-macrophage-colony-stimulating factor enhances immune responses to melanoma-associated peptides in vivo. Int. J. Cancer, 67: 54-62, 1996.[Medline]
  19. Rodolfo M., Colombo M. P. Interleukin-12 as an adjuvant for cancer immunotherapy. Methods, 19: 114-120, 1999.[Medline]
  20. Celluzzi C. M., Mayordomo J. I., Storkus W. J., Lotze M. T., Falo L. D. Peptide-pulsed dendritic cells induce antigen-specific CTL-mediated protective tumor immunity. J. Exp. Med., 183: 283-287, 1996.[Abstract/Free Full Text]
  21. Tam J. P. Synthetic peptide vaccine design: synthesis and properties of a high-density multiple antigenic peptide system. Proc. Natl. Acad. Sci. USA, 85: 5409-5413, 1988.[Abstract/Free Full Text]
  22. Posnett D. N., McGrath H., Tam J. P. A novel method for producing anti-peptide antibodies. Production of site-specific antibodies to the T cell antigen receptor ß-chain. J. Biol. Chem., 263: 1719-1725, 1988.[Abstract/Free Full Text]
  23. Huang W., Nardelli B., Tam J. P. Lipophilic multiple antigen peptide system for peptide immunogen and synthetic vaccine. Mol. Immunol., 31: 1191-1199, 1994.[Medline]
  24. Nardelli B., Tam J. P. Cellular immune responses induced by in vivo priming with a lipid-conjugated multimeric antigen peptide. Immunology, 79: 355-361, 1993.[Medline]
  25. Uenaka A., Ono T., Akisawa T., Wada H., Yasuda T., Nakayama E. Identification of a unique antigen peptide pRL1 on BALB/c RL1 leukemia recognized by cytotoxic T lymphocytes and its relation to the Akt oncogene. J. Exp. Med., 180: 1599-1607, 1994.[Abstract/Free Full Text]
  26. Nakayama E., Shiku H., Takahashi T., Oettgen H. F., Old L. J. Definition of a unique cell surface antigen of mouse leukemia RL1 by cell-mediated cytotoxicity. Proc. Natl. Acad. Sci. USA, 76: 3486-3490, 1979.[Abstract/Free Full Text]
  27. Dunn T. B., Potter M. A transplantable mast-cell neoplasm in the mouse. J. Natl. Cancer Inst. (Bethesda), 18: 587-595, 1957.
  28. Ziegler H. K., Unanue E. R. Decrease in macrophage antigen catabolism caused by ammonia and chloroquine is associated with inhibition of antigen presentation to T cells. Proc. Natl. Acad. Sci. USA, 79: 175-178, 1982.[Abstract/Free Full Text]
  29. Blanar M. A., Baldwin A. S., Flavell R. A., Sharp P. A. A {gamma}-interferon-induced factor that binds the interferon response sequence of the MHC class I gene, H-2Kb. EMBO J., 8: 1139-1144, 1989.[Medline]
  30. Yewdell J. W., Bennink J. R. Brefeldin A specifically inhibits presentation of protein antigens to cytotoxic T lymphocytes. Science (Wash. DC), 244: 1072-1075, 1989.[Abstract/Free Full Text]
  31. Rock K. L., Gramm C., Rothstein L., Clark K., Stein R., Dick L., Hwang D., Goldberg A. L. Inhibitors of the proteasome block the degradation of most cell proteins and the generation of peptides presented on MHC class I molecules. Cell, 78: 761-771, 1994.[Medline]
  32. Fenteany G., Standaert R. F., Lane W. S., Choi S., Corey E. J., Schreiber S. L. Inhibition of proteasome activities and subunit-specific amino-terminal threonine modification by lactacystin. Science (Wash. DC), 268: 726-731, 1995.[Abstract/Free Full Text]
  33. Kumar A., Shafiq S., Wadgaonkar R., Stracher A. The effect of protease inhibitors, leupeptin and E64d, on differentiation of C2C12 myoblasts in tissue culture. Cell Mol. Biol., 38: 477-483, 1992.[Medline]
  34. Yokoi T., Uenaka A., Ono T., Onizuka S., Inoue H., Nakayama E. Diversity of epitopes recognized by cytotoxic T lymphocytes that are specific for rejection antigen peptide pRL1a presented on BALB/c leukemia RL1. Int. Immunol., 9: 1195-1201, 1997.[Abstract/Free Full Text]
  35. Inaba K., Inaba M., Romani N., Aya H., Deguchi M., Ikehara S., Muramatsu S., Steinman R. M. Generation of large numbers of dendritic cells from mouse bone marrow cultures supplemented with granulocyte/macrophage colony-stimulating factor. J. Exp. Med., 176: 1693-1702, 1992.[Abstract/Free Full Text]
  36. Norbury C. C., Hewlett L. J., Prescott A. R., Shastri N., Watts C. Class I MHC presentation of exogenous soluble antigen via macropinocytosis in bone marrow macrophages. Immunity, 3: 783-791, 1995.[Medline]
  37. Kovacsovics-Bankowski M., Clark K., Benacerraf B., Rock K. L. Efficient major histocompatibility complex class I presentation of exogenous antigen upon phagocytosis by macrophages. Proc. Natl. Acad. Sci. USA, 90: 4942-4946, 1993.[Abstract/Free Full Text]
  38. Kovacsovics-Bankowski M., Rock K. L. A phagosome-to-cytosol pathway for exogenous antigens presented on MHC class I molecules. Science (Wash. DC), 267: 243-246, 1995.[Abstract/Free Full Text]
  39. Reis e Sousa C., Germain R. N. Major histocompatibility complex class I presentation of peptides derived from soluble exogenous antigen by a subset of cells engaged in phagocytosis. J. Exp. Med., 182: 841-851, 1995.[Abstract/Free Full Text]
  40. Rodriguez A., Regnault A., Kleijmeer M., Ricciardi-Castagnoli P., Amigorena S. Selective transport of internalized antigens to the cytosol for MHC class I presentation in dendritic cells. Nat. Cell Biol., 1: 362-368, 1999.[Medline]
  41. Jeannin P., Renno T., Goetsch L., Miconnet I., Aubry J. P., Delneste Y., Herbault N., Baussant T., Magistrelli G., Soulas C., Romero P., Cerottini J. C., Bonnefoy J. Y. OmpA targets dendritic cells, induces their maturation and delivers antigen into the MHC class I presentation pathway. Nat. Immunol., 1: 502-509, 2000.[Medline]
  42. Moore M. W., Carbone F. R., Bevan M. J. Introduction of soluble protein into the class I pathway of antigen processing and presentation. Cell, 54: 777-785, 1988.[Medline]
  43. Oh Y. K., Harding C. V., Swanson J. A. The efficiency of antigen delivery from macrophage phagosomes into cytoplasm for MHC class I-restricted antigen presentation. Vaccine, 15: 511-518, 1997.[Medline]
  44. Sijts A. J., Villanueva M. S., Pamer E. G. CTL epitope generation is tightly linked to cellular proteolysis of a Listeria monocytogenes antigen. J. Immunol., 156: 1497-1503, 1996.[Abstract]
  45. Vinitsky A., Anton L. C., Snyder H. L., Orlowski M., Bennink J. R., Yewdell J. W. The generation of MHC class I-associated peptides is only partially inhibited by proteasome inhibitors: involvement of nonproteasomal cytosolic proteases in antigen processing?. J. Immunol., 159: 554-564, 1997.[Abstract]
  46. Luckey C. J., King G. M., Marto J. A., Venketeswaran S., Maier B. F., Crotzer V. L., Colella T. A., Shabanowitz J., Hunt D. F., Engelhard V. H. Proteasomes can either generate or destroy MHC class I epitopes: evidence for nonproteasomal epitope generation in the cytosol. J. Immunol., 161: 112-121, 1998.[Abstract/Free Full Text]
  47. Anton L. C., Snyder H. L., Bennink J. R., Vinitsky A., Orlowski M., Porgador A., Yewdell J. W. Dissociation of proteasomal degradation of biosynthesized viral proteins from generation of MHC class I-associated antigenic peptides. J. Immunol., 160: 4859-4868, 1998.[Abstract/Free Full Text]
  48. Brouwenstijn N., Serwold T., Shastri N. MHC class I molecules can direct proteolytic cleavage of antigenic precursors in the endplasmic reticulum. Immunity, 15: 95-104, 2001.[Medline]



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J. F. Fonteneau, D. G. Kavanagh, M. Lirvall, C. Sanders, T. L. Cover, N. Bhardwaj, and M. Larsson
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