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Recombinant complexes of antigen with stress proteins are potent CD8 T-cell-stimulating immunogens

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Abstract

Heat shock proteins (Hsp) of the Hsp70/90 families facilitate cellular immune responses to antigenic peptides or proteins bound to them and have therefore been used as vaccine vehicles. We developed an expression system in which chimeric proteins with an Hsp-capturing, viral J domain fused to diverse antigen-encoding sequences form stable complexes with eukaryotic (Hsp70, Hsp73) or bacterial (DnaK) stress proteins and accumulate to high steady-state levels. J domains from different species (viruses/SV40, bacteria/Chlamydia trachomatis or plants/Arabidopsis thaliana) efficiently capture murine or human stress proteins in this system, thus making different J domains available for vaccine production. A novel expression and purification method was developed to produce native Hsp/antigen complexes in transfectants. These purified Hsp/antigen complexes efficiently elicited antigen-specific CD8 T cell responses in mice when delivered as vaccines without adjuvants. In situ complex formation of antigen with Hsp was critical for CD8 T cell priming. Because the described expression system supports the flexible design of multivalent vaccines, it is an attractive strategy to elicit CD8 T cell responses either to recombinant proteins or to selected antigenic domains of these molecules.

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Abbreviations

Ad:

adenovirus

APC:

antigen-presenting cells

DC:

dendritic cell

eGFP (or G):

enhanced green fluorescent protein

FCM:

flow cytometry

Flu (or F):

influenza A

HBsAg (or S):

hepatitis B surface antigen

HBV:

hepatitis B virus

Hsp:

heat shock protein

LPS:

lipopolysaccharide

ODN:

oligodeoxynucleotide

OVA:

ovalbumin

Pol (or P):

hepatitis B polymerase

SV40:

simian virus 40

T-Ag (or T):

large tumor antigen of SV40

wt:

wild type

References

  1. Mayer MP, Bukau B (2005) Hsp70 chaperones: cellular functions and molecular mechanism. Cell Mol Life Sci 62:670–684

    Article  PubMed  CAS  Google Scholar 

  2. Reimann J, Schirmbeck R (2004) DNA vaccines expressing antigens with a stress protein-capturing domain display enhanced immunogenicity. Immunol Rev 199:54–67

    Article  PubMed  CAS  Google Scholar 

  3. Chiang HL, Terlecky SR, Plant CP, Dice JF (1989) A role for a 70-kilodalton heat shock protein in lysosomal degradation of intracellular proteins. Science 246:382–385

    Article  PubMed  CAS  Google Scholar 

  4. Brodsky JL, Pipas JM (1998) Polyomavirus T antigens: molecular chaperones for multiprotein complexes. J Virol 72:5329–5334

    PubMed  CAS  Google Scholar 

  5. Schirmbeck R, Kwissa M, Fissolo N, Elkholy S, Riedl P, Reimann J (2002) Priming polyvalent immunity by DNA vaccines expressing chimeric antigens with a stress protein-capturing, viral J-domain. FASEB J 16:1108–1110

    PubMed  CAS  Google Scholar 

  6. Asea A (2005) Stress proteins and initiation of immune response: chaperokine activity of hsp72. Exerc Immunol Rev 11:34–45

    PubMed  Google Scholar 

  7. Enomoto Y, Bharti A, Khaleque AA, Song B, Liu C, Apostolopoulos V, Xing PX, Calderwood SK, Gong J (2006) Enhanced immunogenicity of heat shock protein 70 peptide complexes from dendritic cell-tumor fusion cells. J Immunol 177:5946–5955

    PubMed  CAS  Google Scholar 

  8. Binder RJ, Srivastava PK (2005) Peptides chaperoned by heat-shock proteins are a necessary and sufficient source of antigen in the cross-priming of CD8+ T cells. Nat Immunol 6:593–599

    Article  PubMed  CAS  Google Scholar 

  9. Srivastava PK (2005) Immunotherapy for human cancer using heat shock protein–peptide complexes. Curr Oncol Rep 7:104–108

    Article  PubMed  CAS  Google Scholar 

  10. Binder RJ, Blachere NE, Srivastava PK (2001) Heat shock protein-chaperoned peptides but not free peptides introduced into the cytosol are presented efficiently by major histocompatibility complex I molecules. J Biol Chem 276:17163–17171

    Article  PubMed  CAS  Google Scholar 

  11. Flechtner JB, Cohane KP, Mehta S, Slusarewicz P, Leonard AK, Barber BH, Levey DL, Andjelic S (2006) High-affinity interactions between peptides and heat shock protein 70 augment CD8+ T lymphocyte immune responses. J Immunol 177:1017–1027

    PubMed  CAS  Google Scholar 

  12. Moroi Y, Mayhew M, Trcka J, Hoe MH, Takechi Y, Hartl FU, Rothman JE, Houghton AN (2000) Induction of cellular immunity by immunization with novel hybrid peptides complexed to heat shock protein 70. Proc Natl Acad Sci U S A 97:3485–3490

    Article  PubMed  CAS  Google Scholar 

  13. Blachere NE, Li Z, Chandawarkar RY, Suto R, Jaikaria NS, Basu S, Udono H, Srivastava PK (1997) Heat shock protein–peptide complexes, reconstituted in vitro, elicit peptide-specific cytotoxic T lymphocyte response and tumor immunity. J Exp Med 186:1315–1322

    Article  PubMed  CAS  Google Scholar 

  14. Zhang H, Huang W (2006) Fusion proteins of Hsp70 with tumor-associated antigen acting as a potent tumor vaccine and the C-terminal peptide-binding domain of Hsp70 being essential in inducing antigen-independent anti-tumor response in vivo. Cell Stress Chaperones 11:216–226

    Article  PubMed  CAS  Google Scholar 

  15. Chen CH, Wang TL, Hung CF, Yang Y, Young RA, Pardoll DM, Wu TC (2000) Enhancement of DNA vaccine potency by linkage of antigen gene to an HSP70 gene. Cancer Res 60:1035–1042

    PubMed  CAS  Google Scholar 

  16. Cho BK, Palliser D, Guillen E, Wisniewski J, Young RA, Chen J, Eisen HN (2000) A proposed mechanism for the induction of cytotoxic T lymphocyte production by heat shock fusion proteins. Immunity 12:263–272

    Article  PubMed  CAS  Google Scholar 

  17. Suzue K, Zhou X, Eisen HN, Young RA (1997) Heat shock fusion proteins as vehicles for antigen delivery into the major histocompatibility complex class I presentation pathway. Proc Natl Acad Sci U S A 94:13146–13151

    Article  PubMed  CAS  Google Scholar 

  18. Riedl P, Fissolo N, Reimann J, Schirmbeck R (2006) A stress protein-facilitated antigen expression system for plasmid DNA vaccines. Methods Mol Med 127:41–53

    PubMed  CAS  Google Scholar 

  19. Kammerer R, Stober D, Riedl P, Oehninger C, Schirmbeck R, Reimann J (2002) Noncovalent association with stress protein facilitates cross-priming of CD8+ T cells to tumor cell antigens by dendritic cells. J Immunol 168:108–117

    PubMed  CAS  Google Scholar 

  20. Schirmbeck R, Riedl P, Kupferschmitt M, Wegenka U, Hauser H, Rice J, Kroger A, Reimann J (2006) Priming protective CD8 T cell immunity by DNA vaccines encoding chimeric, stress protein-capturing tumor-associated antigen. J Immunol 177:1534–1542

    PubMed  CAS  Google Scholar 

  21. Schirmbeck R, Fissolo N, Chaplin P, Reimann J (2003) Enhanced priming of multispecific, murine CD8+ T cell responses by DNA vaccines expressing stress protein-binding polytope peptides. J Immunol 171:1240–1246

    PubMed  CAS  Google Scholar 

  22. Palliser D, Guillen E, Ju M, Eisen HN (2005) Multiple intracellular routes in the cross-presentation of a soluble protein by murine dendritic cells. J Immunol 174:1879–1887

    PubMed  CAS  Google Scholar 

  23. Wortmann A, Vohringer S, Engler T, Corjon S, Schirmbeck R, Reimann J, Kochanek S, Kreppel F (2008) Fully detargeted polyethylene glycol-coated adenovirus vectors are potent genetic vaccines and escape from pre-existing anti-adenovirus antibodies. Mol Ther 16:154–162

    Article  PubMed  CAS  Google Scholar 

  24. Riedl P, Bertoletti A, Lopes R, Lemonnier F, Reimann J, Schirmbeck R (2006) Distinct, cross-reactive epitope specificities of CD8 T cell responses are induced by natural hepatitis B surface antigen variants of different hepatitis B virus genotypes. J Immunol 176:4003–4011

    PubMed  CAS  Google Scholar 

  25. Schirmbeck R, Gerstner O, Reimann J (1999) Truncated or chimeric endogenous protein antigens gain immunogenicity for B cells by stress protein-facilitated expression. Eur J Immunol 29:1740–1749

    Article  PubMed  CAS  Google Scholar 

  26. Ye Z, Gan YH (2007) Flagellin contamination of recombinant heat shock protein 70 is responsible for its activity on T cells. J Biol Chem 282:4479–4484

    Article  PubMed  CAS  Google Scholar 

  27. Bausinger H, Lipsker D, Ziylan U, Manie S, Briand JP, Cazenave JP, Muller S, Haeuw JF, Ravanat C, de la SH, Hanau D (2002) Endotoxin-free heat-shock protein 70 fails to induce APC activation. Eur J Immunol 32:3708–3713

    Article  PubMed  CAS  Google Scholar 

  28. Wallin RP, Lundqvist A, More SH, von Bonin A, Kiessling R, Ljunggren HG (2002) Heat-shock proteins as activators of the innate immune system. Trends Immunol 23:130–135

    Article  PubMed  CAS  Google Scholar 

  29. Bendz H, Ruhland SC, Pandya MJ, Hainzl O, Riegelsberger S, Brauchle C, Mayer MP, Buchner J, Issels RD, Noessner E (2007) Human heat shock protein 70 enhances tumor antigen presentation through complex formation and intracellular antigen delivery without innate immune signaling. J Biol Chem 282:31688–31702

    Article  PubMed  CAS  Google Scholar 

  30. Gurunathan S, Klinman DM, Seder RA (2000) DNA vaccines: immunology, application, and optimization. Annu Rev Immunol 18:927–974

    Article  PubMed  CAS  Google Scholar 

  31. Cresswell P, Ackerman AL, Giodini A, Peaper DR, Wearsch PA (2005) Mechanisms of MHC class I-restricted antigen processing and cross-presentation. Immunol Rev 207:145–157

    Article  PubMed  CAS  Google Scholar 

  32. Binder RJ (2006) Heat shock protein vaccines: from bench to bedside. Int Rev Immunol 25:353–375

    Article  PubMed  CAS  Google Scholar 

  33. Yewdell JW, Del Val M (2004) Immunodominance in TCD8+ responses to viruses: cell biology, cellular immunology, and mathematical models. Immunity 21:149–153

    Article  PubMed  CAS  Google Scholar 

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Acknowledgments

We thank K. Ölberger and C. Heilig for outstanding technical assistance. We thank Dr. F. Lemonnier (Institut Pasteur, Paris, France) for the HLA-A*0201 transgenic mice and A. Schreiber (Dept. of Virology, University of Ulm, Germany) for the bacterial expressed, recombinant eGFP-protein. This work was supported by a grant from the Deutsche Forschungsgemeinschaft (SCHI-505/2-4) to R.S.

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Correspondence to Reinhold Schirmbeck.

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Wieland, A., Denzel, M., Schmidt, E. et al. Recombinant complexes of antigen with stress proteins are potent CD8 T-cell-stimulating immunogens. J Mol Med 86, 1067–1079 (2008). https://doi.org/10.1007/s00109-008-0371-x

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  • DOI: https://doi.org/10.1007/s00109-008-0371-x

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