Skip to main content
Log in

Evaluating combinations of costimulatory antibody–ligand fusion proteins for targeted cancer immunotherapy

Cancer Immunology, Immunotherapy Aims and scope Submit manuscript

Abstract

Combinatory strategies are becoming of increasing interest in cancer immunotherapy. Costimulation by individual members of the immunoglobulin-like (Ig)- and TNF superfamily have already shown promising antitumor potential, thus prompting the exploration of their synergistic abilities in combinatorial approaches. Here, we pursued a targeted strategy with antibody-fusion proteins composed of a tumor-directed antibody and the extracellular domain of the costimulatory ligand B7.1, 4-1BBL, OX40L, GITRL or LIGHT, respectively. Costimulatory activity was assessed in an experimental setting where initial T cell activation was induced by a bispecific antibody (tumor-related antigen × CD3). Advantage of combined targeted costimulation was shown for either B7.1 or 4-1BBL with OX40L, GITRL, LIGHT and 4-1BBL in terms of T cell proliferation and IFN-γ release. Since encouraging results were obtained by the combination of B7.1 and 4-1BBL, we adapted the model system for a time-shift setting. Here, enhanced proliferation and granzyme B expression as well as reduced PD-1 expression on the T cell population demonstrated the benefit of costimulation-assisted restimulation. Finally, the antitumor potential of this combinatorial setting was confirmed in vivo in a lung metastasis mouse model. Thus, combinatorial approaches with costimulatory antibody–ligand fusion proteins seem a promising strategy to be further investigated for cancer immunotherapy.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5

Abbreviations

GITRL:

Glucocorticoid-induced TNF receptor ligand

LIGHT:

Homologous to lymphotoxins, shows inducible expression and competes with herpes simplex virus glycoprotein D for herpesvirus entry mediator (HVEM), a receptor expressed by T lymphocytes

TNFSF:

Tumor necrosis factor superfamily

mAb:

Monoclonal antibody

FAP:

Fibroblast activation protein

References

  1. Pardoll D, Drake C (2012) Immunotherapy earns its spot in the ranks of cancer therapy. J Exp Med 209(2):201–209. doi:10.1084/jem.20112275

    Article  PubMed  CAS  Google Scholar 

  2. Schaer DA, Murphy JT, Wolchok JD (2012) Modulation of GITR for cancer immunotherapy. Curr Opin Immunol 24(2):217–224. doi:10.1016/j.coi.2011.12.011

    Article  PubMed  CAS  Google Scholar 

  3. Melero I, Martinez-Forero I, Dubrot J, Suarez N, Palazón A, Chen L (2009) Palettes of vaccines and immunostimulatory monoclonal antibodies for combination. Clin Cancer Res 15(5):1507–1509. doi:10.1158/1078-0432.CCR-08-2931

    Article  PubMed  CAS  Google Scholar 

  4. Takeda K, Kojima Y, Uno T, Hayakawa Y, Teng MW, Yoshizawa H, Yagita H, Gejyo F, Okumura K, Smyth MJ (2010) Combination therapy of established tumors by antibodies targeting immune activating and suppressing molecules. J Immunol 184(10):5493–5501. doi:10.4049/jimmunol.0903033

    Article  PubMed  CAS  Google Scholar 

  5. Serghides L, Bukczynski J, Wen T, Wang C, Routy JP, Boulassel MR, Sekaly RP, Ostrowski M, Bernard NF, Watts TH (2005) Evaluation of OX40 ligand as a costimulator of human antiviral memory CD8 T cell responses: comparison with B7.1 and 4–1BBL. J Immunol 175(10):6368–6377

    PubMed  CAS  Google Scholar 

  6. Pan PY, Zang Y, Weber K, Meseck ML, Chen SH (2002) OX40 ligation enhances primary and memory cytotoxic T lymphocyte responses in an immunotherapy for hepatic colon metastases. Mol Ther 6(4):528–536

    Article  PubMed  CAS  Google Scholar 

  7. Uno T, Takeda K, Kojima Y, Yoshizawa H, Akiba H, Mittler RS, Gejyo F, Okumura K, Yagita H, Smyth MJ (2006) Eradication of established tumors in mice by a combination antibody-based therapy. Nat Med 12(6):693–698

    Article  PubMed  CAS  Google Scholar 

  8. Kocak E, Lute K, Chang X, May KF Jr, Exten KR, Zhang H, Abdessalam SF, Lehman AM, Jarjoura D, Zheng P, Liu Y (2006) Combination therapy with anti-CTL antigen-4 and anti-4-1BB antibodies enhances cancer immunity and reduces autoimmunity. Cancer Res 66(14):7276–7284

    Article  PubMed  CAS  Google Scholar 

  9. Curran MA, Kim M, Montalvo W, Al-Shamkhani A, Allison JP (2011) Combination CTLA-4 blockade and 4–1BB activation enhances tumor rejection by increasing T-cell infiltration, proliferation, and cytokine production. PLoS ONE 6(4):e19499. doi:10.1371/journal.pone.0019499

    Article  PubMed  CAS  Google Scholar 

  10. Hirano F, Kaneko K, Tamura H, Dong H, Wang S, Ichikawa M, Rietz C, Flies DB, Lau JS, Zhu G, Tamada K, Chen L (2005) Blockade of B7–H1 and PD-1 by monoclonal antibodies potentiates cancer therapeutic immunity. Cancer Res 65(3):1089–1096

    PubMed  CAS  Google Scholar 

  11. Driessens G, Kline J, Gajewski TF (2009) Costimulatory and coinhibitory receptors in anti-tumor immunity. Immunol Rev 229(1):126–144. doi:10.1111/j.1600-065X.2009.00771.x

    Article  PubMed  CAS  Google Scholar 

  12. Melero I, Hervas-Stubbs S, Glennie M, Pardoll DM, Chen L (2007) Immunostimulatory monoclonal antibodies for cancer therapy. Nat Rev Cancer 7(2):95–106

    Article  PubMed  CAS  Google Scholar 

  13. Murphy KA, Lechner MG, Popescu FE, Bedi J, Decker SA, Hu P, Erickson JR, O’Sullivan MG, Swier L, Salazar AM, Olin MR, Epstein AL, Ohlfest JR (2012) An in vivo immunotherapy screen of costimulatory molecules identifies Fc-OX40L as a potent reagent for the treatment of established murine gliomas. Clin Cancer Res 18(17):4657–4668. doi:10.1158/1078-0432.CCR-12-0990

    Article  PubMed  CAS  Google Scholar 

  14. Lechner MG, Russell SM, Bass RS, Epstein AL (2011) Chemokines, costimulatory molecules and fusion proteins for the immunotherapy of solid tumors. Immunotherapy 3(11):1317–1340. doi:10.2217/imt.11.115

    Article  PubMed  CAS  Google Scholar 

  15. Kanagavelu SK, Snarsky V, Termini JM, Gupta S, Barzee S, Wright JA, Khan WN, Kornbluth RS, Stone GW (2012) Vaccine. 30(4):691–702. doi:10.1016/j.vaccine.2011.11.088

  16. Sharma RK, Schabowsky RH, Srivastava AK, Elpek KG, Madireddi S, Zhao H, Zhong Z, Miller RW, Macleod KJ, Yolcu ES, Shirwan H (2010) 4–1BB ligand as an effective multifunctional immunomodulator and antigen delivery vehicle for the development of therapeutic cancer vaccines. Cancer Res 70(10):3945–3954. doi:10.1158/0008-5472.CAN-09-4480

    Article  PubMed  CAS  Google Scholar 

  17. Wyzgol A, Müller N, Fick A, Munkel S, Grigoleit GU, Pfizenmaier K, Wajant H (2009) Trimer stabilization, oligomerization, and antibody-mediated cell surface immobilization improve the activity of soluble trimers of CD27L, CD40L, 41BBL, and glucocorticoid-induced TNF receptor ligand. J Immunol 183(3):1851–1861. doi:10.4049/jimmunol.0802597

    Article  PubMed  CAS  Google Scholar 

  18. Ascierto PA, Simeone E, Sznol M, Fu YX, Melero I (2010) Clinical experiences with anti-CD137 and anti-PD1 therapeutic antibodies. Semin Oncol 37(5):508–516. doi:10.1053/j.seminoncol.2010.09.008

    Article  PubMed  CAS  Google Scholar 

  19. Liu A, Hu P, Khawli LA, Epstein AL (2006) B7.1/NHS76: a new costimulator fusion protein for the immunotherapy of solid tumors. J Immunother 29(4):425–435

    Article  PubMed  Google Scholar 

  20. Liu R, Jiang W, Yang M, Guo H, Zhang Y, Wang J, Zhu H, Shi R, Fan D, Yang C, Zhu Z, Xie Y, Xiong D (2010) Efficient inhibition of human B-cell lymphoma in SCID mice by synergistic antitumor effect of human 4–1BB ligand/anti-CD20 fusion proteins and anti-CD3/anti-CD20 diabodies. J Immunother 33(5):500–509. doi:10.1097/CJI.0b013e3181d75c20

    Article  PubMed  Google Scholar 

  21. Burckhart T, Thiel M, Nishikawa H, Wüest T, Müller D, Zippelius A, Ritter G, Old L, Shiku H, Renner C (2010) Tumor-specific crosslinking of GITR as costimulation for immunotherapy. J Immunother 33(9):925–934. doi:10.1097/CJI.0b013e3181f3cc87

    Article  PubMed  CAS  Google Scholar 

  22. Hornig N, Kermer V, Frey K, Diebolder P, Kontermann RE, Müller D (2012) Combination of a bispecific antibody and costimulatory antibody-ligand fusion proteins for targeted cancer immunotherapy. J Immunother 35(5):418–429. doi:10.1097/CJI.0b013e3182594387

    Article  PubMed  CAS  Google Scholar 

  23. Zhang N, Sadun RE, Arias RS, Flanagan ML, Sachsman SM, Nien YC, Khawli LA, Hu P, Epstein AL (2007) Targeted and untargeted CD137L fusion proteins for the immunotherapy of experimental solid tumors. Clin Cancer Res 13(9):2758–2767

    Article  PubMed  CAS  Google Scholar 

  24. Müller D, Trunk G, Sichelstiel A, Zettlitz K, Quintanilla M, Kontermann RE (2008) Murine endoglin-specific single-chain Fv fragments for the analysis of vascular targeting strategies in mice. J Immunol Methods 339(1):90–98. doi:10.1016/j.jim.2008.08.008

    Article  PubMed  Google Scholar 

  25. Völkel T, Müller R, Kontermann RE (2004) Isolation of endothelial cell-specific human antibodies from a novel fully synthetic scFv library. Biochem Biophys Res Commun 317(2):515–521

    Article  PubMed  Google Scholar 

  26. Gerspach J, Muller D, Munkel S, Selchow O, Nemeth J, Noack M, Petrul H, Menrad A, Wajant H, Pfizenmaier K (2006) Restoration of membrane TNF-like activity by cell surface targeting and matrix metalloproteinase-mediated processing of a TNF prodrug. Cell Death Differ 13(2):273–284

    Article  PubMed  CAS  Google Scholar 

  27. Liao KW, Lo YC, Roffler SR (2000) Activation of lymphocytes by anti-CD3 single-chain antibody dimers expressed on the plasma membrane of tumor cells. Gene Ther 7(4):339–347

    Article  PubMed  CAS  Google Scholar 

  28. Cochran WG, Cox GM (1992) Experimental designs, 2nd edn. Wiley, Oxford

    Google Scholar 

  29. Croft M (2003) Co-stimulatory members of the TNFR family: keys to effective T-cell immunity? Nat Rev Immunol 3(8):609–620

    Article  PubMed  CAS  Google Scholar 

  30. Bodmer JL, Schneider P, Tschopp J (2002) The molecular architecture of the TNF superfamily. Trends Biochem Sci 27(1):19–26

    Article  PubMed  CAS  Google Scholar 

  31. Bremer E, de Bruyn M, Wajant H, Helfrich W (2009) Targeted cancer immunotherapy using ligands of the tumor necrosis factor super-family. Curr Drug Targets 10(2):94–103

    Article  PubMed  CAS  Google Scholar 

  32. Wajant H, Gerspach J, Pfizenmaier K (2005) Tumor therapeutics by design: targeting and activation of death receptors. Cytokine Growth Factor Rev 16(1):55–76

    Article  PubMed  CAS  Google Scholar 

  33. Müller N, Wyzgol A, Münkel S, Pfizenmaier K, Wajant H (2008) Activity of soluble OX40 ligand is enhanced by oligomerization and cell surface immobilization. FEBS J 275(9):2296–2304. doi:10.1111/j.1742-4658.2008.06382.x

    Article  PubMed  Google Scholar 

  34. Chattopadhyay K, Lazar-Molnar E, Yan Q, Rubinstein R, Zhan C, Vigdorovich V, Ramagopal UA, Bonanno J, Nathenson SG, Almo SC (2009) Sequence, structure, function, immunity: structural genomics of costimulation. Immunol Rev 229(1):356–386. doi:10.1111/j.1600-065X.2009.00778.x

    Article  PubMed  CAS  Google Scholar 

  35. Kober J, Leitner J, Klauser C, Woitek R, Majdic O, Stöckl J, Herndler-Brandstetter D, Grubeck-Loebenstein B, Reipert BM, Pickl WF, Pfistershammer K, Steinberger P (2008) The capacity of the TNF family members 4–1BBL, OX40L, CD70, GITRL, CD30L and LIGHT to costimulate human T cells. Eur J Immunol 38(10):2678–2688. doi:10.1002/eji.200838250

    Article  PubMed  CAS  Google Scholar 

  36. Yu P, Lee Y, Liu W, Chin RK, Wang J, Wang Y, Schietinger A, Philip M, Schreiber H, Fu YX (2004) Priming of naive T cells inside tumors leads to eradication of established tumors. Nat Immunol 5(2):141–149

    Article  PubMed  CAS  Google Scholar 

  37. Melero I, Bach N, Hellström KE, Aruffo A, Mittler RS, Chen L (1998) Amplification of tumor immunity by gene transfer of the co-stimulatory 4–1BB ligand: synergy with the CD28 co-stimulatory pathway. Eur J Immunol 28(3):1116–1121

    Article  PubMed  CAS  Google Scholar 

  38. Li G, Wu X, Zhang F, Li X, Sun B, Yu Y, Yin A, Deng L, Yin J, Wang X (2010) Triple expression of B7–1, B7–2 and 4–1BBL enhanced antitumor immune response against mouse H22 hepatocellular carcinoma. J Cancer Res Clin Oncol 137(4):695–703. doi:10.1007/s00432-010-0905-9

    Article  PubMed  Google Scholar 

  39. Habib-Agahi M, Jaberipour M, Searle PF (2009) 4–1BBL costimulation retrieves CD28 expression in activated T cells. Cell Immunol 256(1–2):39–46. doi:10.1016/j.cellimm.2009.01.003

    Article  PubMed  CAS  Google Scholar 

  40. Ronchetti S, Nocentini G, Bianchini R, Krausz LT, Migliorati G, Riccardi C (2007) Glucocorticoid-induced TNFR-related protein lowers the threshold of CD28 costimulation in CD8 + T cells. J Immunol 179(9):5916–5926

    PubMed  CAS  Google Scholar 

  41. Tamada K, Shimozaki K, Chapoval AI, Zhu G, Sica G, Flies D, Boone T, Hsu H, Fu YX, Nagata S, Ni J, Chen L (2000) Modulation of T-cell-mediated immunity in tumor and graft-versus-host disease models through the LIGHT co-stimulatory pathway. Nat Med 6(3):283–289

    Article  PubMed  CAS  Google Scholar 

  42. Fife BT, Bluestone JA (2008) Control of peripheral T-cell tolerance and autoimmunity via the CTLA-4 and PD-1 pathways. Immunol Rev 224:166–182. doi:10.1111/j.1600-065X.2008.00662.x

    Article  PubMed  CAS  Google Scholar 

  43. Dong H, Strome SE, Salomao DR, Tamura H, Hirano F, Flies DB, Roche PC, Lu J, Zhu G, Tamada K, Lennon VA, Celis E, Chen L (2002) Tumor-associated B7–H1 promotes T-cell apoptosis: a potential mechanism of immune evasion. Nat Med 8(8):793–800

    PubMed  CAS  Google Scholar 

  44. Topalian SL, Drake CG, Pardoll DM (2012) Targeting the PD-1/B7-H1(PD-L1) pathway to activate anti-tumor immunity. Curr Opin Immunol 24(2):207–212. doi:10.1016/j.coi.2011.12.009

    Article  PubMed  CAS  Google Scholar 

Download references

Acknowledgments

This work was supported by a Grant from the Deutsche Krebshilfe (108743). We would like to thank Robert Lindner for technical support on the high‐performance liquid chromatography analysis.

Conflict of interest

The authors declare that they have no conflict of interest.

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to Dafne Müller.

Electronic supplementary material

Below is the link to the electronic supplementary material.

Supplementary material 1 (PDF 125 kb)

Rights and permissions

Reprints and permissions

About this article

Cite this article

Hornig, N., Reinhardt, K., Kermer, V. et al. Evaluating combinations of costimulatory antibody–ligand fusion proteins for targeted cancer immunotherapy. Cancer Immunol Immunother 62, 1369–1380 (2013). https://doi.org/10.1007/s00262-013-1441-7

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00262-013-1441-7

Keywords

Navigation