Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

Adjuvant properties of IFN-γ and GM-CSF in the scFv6.C4 DNA vaccine against CEA-expressing tumors

Gene Therapy volume 30, pages 41–50 (2023)Cite this article

Subjects

A Correction to this article was published on 20 March 2023

This article has been updated

Abstract

Tumor-associated carcinoembryonic antigen (CEA) is a natural target for vaccines against colorectal cancers. Our previous experience with a DNA vaccine with scFv6.C4, a CEA surrogate, showed a CEA-specific immune response with 40% of tumor-free mice after challenge with B16F10-CEA and 47% with MC38-CEA cells. These percentages increased to 63% after using FrC as an adjuvant. To further enhance the vaccine efficacy, we tested GM-CSF and IFNγ as adjuvants. C57BL/6J-CEA2682 mice were immunized 4 times with uP-PS/scFv6.C4, uP-PS/scFv6.C4 + uP-IFNγ, or uP-PS/scFv6.C4 + uP-GMCSF. After one week, the mice were challenged with MC38-CEA, and tumor growth was monitored over 100 days. Immunization with scFv6.C4 and scFv6.C4 + GM-CSF resulted in a gradual increase in the anti-CEA antibody titer, while scFv6.C4 + IFNγ immunization led to a rapid and sustained increase in the titer. The addition of IFNγ also induced higher CD4 + and CD8 + responses. When challenged, almost 80% of the scFv6.C4 + IFNγ-vaccinated mice did not develop tumors, while the others had a significant tumor growth delay. The probability of being tumor-free was 2700% higher using scFv6.C4 + IFNγ than scFv6.C4. The addition of GM-CSF had no additional effect on tumor protection. DNA immunization with scFv6.C4 + IFNγ, but not GM-CSF, increased the antitumor effect via readily sustained specific humoral and cytotoxic responses to CEA.

This is a preview of subscription content, access via your institution

Access options

Rent or buy this article

Get just this article for as long as you need it

$39.95

Learn more

Prices may be subject to local taxes which are calculated during checkout

Fig. 1: Humoral response and protection against MC38CEA tumor challenge by scFv6.C4 + IFNγ and scFv6.C4 + GM-CSF DNA immunization.
Fig. 2: Evaluation of the humoral response by immunocytochemistry using sera from the scFv6.C4 + IFNγ- or scFv6.C4 + GM-CSF-immunized mice.
Fig. 3: Cellular response of the scFv6.C4 + IFNγ-immunized animals determined by proliferation assay.
Fig. 4: CTL activity in CD8 + cells from the scFv6.C4 + INFγ- immunized animals by LDH activity assay.

Change history

References

  1. Siegel RL, Miller KD, Jemal A. Cancer statistics, 2020. CA Cancer J Clin. 2020;70:7–30.

    Article  PubMed  Google Scholar 

  2. Denapoli PMA, Zanetti BF, Dos Santos AA, de Moraes JZ, Han SW. Preventive DNA vaccination against CEA-expressing tumors with anti-idiotypic scFv6.C4 DNA in CEA-expressing transgenic mice. Cancer Immunol Immunother. 2016;66:333–42.

    Article  PubMed  Google Scholar 

  3. Zanetti BF, Ferreira CP, de Vasconcelos JRC, Han SW. scFv6.C4 DNA vaccine with fragment C of Tetanus toxin increases protective immunity against CEA-expressing tumor. Gene Ther. 2019;26:441–54.

    Article  CAS  PubMed  Google Scholar 

  4. Pignatari GC, Takeshita D, Parise CB, Soares FA, de Moraes JZ, Han SW. Carcinoembryonic antigen (CEA) mimicry by an anti-idiotypic scFv isolated from anti-Id 6.C4 hybridoma. J Biotechnol. 2007;127:615–25.

    Article  CAS  PubMed  Google Scholar 

  5. de Moraes JZ, Carneiro CR, Buchegger F, Mach JP, Lopes JD. Induction of an immune response through the idiotypic network with monoclonal anti-idiotype antibodies in the carcinoembryonic antigen system. J Cell Biochem. 1992;50:324–35.

    Article  PubMed  Google Scholar 

  6. de Moraes JZ, Gesztesi JL, Westermann P, Le Doussal JM, Lopes JD, Mach JP. Anti-idiotypic monoclonal antibody AB3, reacting with the primary antigen (CEA), can localize in human colon-carcinoma xenografts as efficiently as AB1. Int J Cancer. 1994;57:586–91.

    Article  PubMed  Google Scholar 

  7. Geethadevi A, Jadhav K, Kumar G, Parashar D scFv6.C4 DNA vaccine with fragment C of tetanus toxin increases protective immunity against CEA-expressing tumor. Gene Ther. 2020;26:441–54. https://doi.org/10.1038/s41434-020-0161-9.

  8. Coleman DL, Chodakewitz JA, Bartiss AH, Mellors JW. Granulocyte-macrophage colony-stimulating factor enhances selective effector functions of tissue-derived macrophages. Blood. 1988;72:573–8.

    Article  CAS  PubMed  Google Scholar 

  9. Inaba K, Inaba M, Romani N, Aya H, Deguchi M, Ikehara S, et al. Generation of large numbers of dendritic cells from mouse bone marrow cultures supplemented with granulocyte/macrophage colony-stimulating factor. J Exp Med. 1992;176:1693–702.

    Article  CAS  PubMed  Google Scholar 

  10. Steinman RM, Witmer-Pack M, Inaba K. Dendritic cells: antigen presentation, accessory function and clinical relevance. Adv Exp Med Biol. 1993;329:1–9.

    Article  CAS  PubMed  Google Scholar 

  11. Clive KS, Tyler JA, Clifton GT, Holmes JP, Mittendorf EA, Ponniah S, et al. Use of GM-CSF as an adjuvant with cancer vaccines: beneficial or detrimental? Expert Rev Vaccines. 2010;9:519–25.

    Article  CAS  PubMed  Google Scholar 

  12. Kass E, Panicali DL, Mazzara G, Schlom J, Greiner JW Granulocyte / macrophage-colony stimulating factor produced by recombinant avian poxviruses enriches the regional lymph nodes with antigen-presenting cells and acts as an immunoadjuvant granulocyte / macrophage-colony stimulating factor produced by recomb. Cancer Res. 2001;27:206–14.

  13. Aarts WM, Schlom J, Hodge JW. Vector-based vaccine / cytokine combination therapy to enhance induction of immune responses to a self-antigen and antitumor activity. Am Assoc Cancer Res. 2002;62:5770–7.

    CAS  Google Scholar 

  14. Lima J, Jenkins C, Guerrero A, Triozzi PL, Shaw DR, Strong TV. A DNA vaccine encoding genetic fusions of carcinoembryonic antigen (CEA) and granulocyte/macrophage colony-stimulating factor (GM-CSF). Vaccine. 2005;23:1273–83.

    Article  CAS  PubMed  Google Scholar 

  15. Hallermalm K, Johansson S, Bråve A, Ek M, Engström G, Boberg A, et al. Pre-clinical evaluation of a CEA DNA prime/protein boost vaccination strategy against colorectal cancer. Scand J Immunol. 2007;66:43–51.

    Article  CAS  PubMed  Google Scholar 

  16. Schwegler C, Dorn-Beineke A, Nittka S, Stocking C, Neumaier M. Monoclonal anti-idiotype antibody 6G6.C4 fused to GM-CSF is capable of breaking tolerance to carcinoembryonic antigen (CEA) in CEA-transgenic mice. Cancer Res. 2005;65:1925–33.

    Article  CAS  PubMed  Google Scholar 

  17. Hodge JW, Poole DJ, Aarts WM, Yafal AG, Gritz L, Schlom J. Modified vaccinia virus ankara recombinants are as potent as vaccinia recombinants in diversified prime and boost vaccine regimens to elicit therapeutic antitumor responses. Cancer Res. 2003;63:7942–9.

    CAS  PubMed  Google Scholar 

  18. Luo Y, O’Hagan D, Zhou H, Singh M, Ulmer J, Reisfeld RA, et al. Plasmid DNA encoding human carcinoembryonic antigen (CEA) adsorbed onto cationic microparticles induces protective immunity against colon cancer in CEA-transgenic mice. Vaccine. 2003;21:1938–47.

    Article  CAS  PubMed  Google Scholar 

  19. Samanci A, Yi Q, Fagerberg J, Strigard K, Smith G, Ruden U, et al. Pharmacological administration of granulocyte/macrophage-colony-stimulating factor is of significant importance for the induction of a strong humoral and cellular response in patients immunized with recombinant carcinoembryonic antigen. Cancer Immunol Immunother. 1998;47:131–42.

    Article  CAS  PubMed  Google Scholar 

  20. Ullenhag GJ, Frödin JE, Jeddi-Tehrani M, Strigård K, Eriksson E, Samanci A, et al. Durable carcinoembryonic antigen (CEA)-specific humoral and cellular immune responses in colorectal carcinoma patients vaccinated with recombinant CEA and granulocyte/macrophage colony-stimulating factor. Clin Cancer Res. 2004;10:3273–81.

    Article  CAS  PubMed  Google Scholar 

  21. Duggan MC, Jochems C, Donahue RN, Richards J, Karpa V, Foust E, et al. A phase I study of recombinant (r) vaccinia-CEA(6D)-TRICOM and rFowlpox-CEA(6D)-TRICOM vaccines with GM-CSF and IFN-α-2b in patients with CEA-expressing carcinomas. Cancer Immunol Immunother. 2016;65:1353–64.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  22. Marshall JL, Gulley JL, Arlen PM, Beetham PK, Tsang KY, Slack R, et al. Phase I study of sequential vaccinations with fowlpox-CEA(6D)-TRICOM alone and sequentially with vaccinia-CEA(6D)-TRICOM, with and without granulocyte-macrophage colony-stimulating factor, in patients with carcinoembryonic antigen-expressing carcinomas. J Clin Oncol. 2005;23:720–31.

    Article  CAS  PubMed  Google Scholar 

  23. Geynisman DM, Zha Y, Kunnavakkam R, Aklilu M, Catenacci DV, Polite BN, et al. A randomized pilot phase I study of modified carcinoembryonic antigen (CEA) peptide (CAP1-6D)/montanide/GM-CSF-vaccine in patients with pancreatic adenocarcinoma. J Immunother cancer. 2013;1:8.

    Article  PubMed  PubMed Central  Google Scholar 

  24. Farrar M, Schreiber R. The molecular cell biology of interferon-gamma and its receptor. Annu Rev Immunol. 1993;11:571–611.

    Article  CAS  PubMed  Google Scholar 

  25. Ikeda H, Old LJ, Schreiber RD. The roles of IFNγ in protection against tumor development and cancer immunoediting. Cytokine Growth Factor Rev. 2002;13:95–109.

    Article  CAS  PubMed  Google Scholar 

  26. Miller CHT, Maher SG, Young HA. Clinical use of Interferon- γ. Ann N Y Acad Sci. 2009;79:69–79.

    Article  Google Scholar 

  27. Song K, Chang Y, Prud’homme GJ. Regulation of T-helper-1 versus T-helper-2 activity and enhancement of tumor immunity by combined DNA-based vaccination and nonviral cytokine gene transfer. Gene Ther. 2000;7:481–92.

    Article  CAS  PubMed  Google Scholar 

  28. Sacramento CB, Cantagalli VD, Grings M, Carvalho LP, Baptista-Silva JCC, Beutel A, et al. Granulocyte-macrophage colony-stimulating factor gene based therapy for acute limb ischemia in a mouse model. J Gene Med. 2009;11:345–53.

    Article  CAS  PubMed  Google Scholar 

  29. Corbett TH, Griswold DPJ, Roberts BJ, Peckham JC, Schabel FMJ. Tumor induction relationships in development of transplantable cancers of the colon in mice for chemotherapy assays, with a note on carcinogen structure. Cancer Res. 1975;35:2434–9.

    CAS  PubMed  Google Scholar 

  30. Tompkins WA, Watrach AM, Schmale JD, Schultz RM, Harris JA. Cultural and antigenic properties of newly established cell strains derived from adenocarcinomas of the human colon and rectum. J Natl Cancer Inst. 1974;52:1101–10.

    Article  CAS  PubMed  Google Scholar 

  31. Eades-Perner AM, van der Putten H, Hirth A, Thompson J, Neumaier M, von Kleist S, et al. Mice transgenic for the human carcinoembryonic antigen gene maintain its spatiotemporal expression pattern. Cancer Res. 1994;54:4169–76.

    CAS  PubMed  Google Scholar 

  32. Parise CB, Lisboa B, Takeshita D, Sacramento CB, de Moraes JZ, Han SW. Humoral immune response after genetic immunization is consistently improved by electroporation. Vaccine. 2008;26:3812–7.

    Article  CAS  PubMed  Google Scholar 

  33. Goldstein MJ, Mitchell EP. Carcinoembryonic antigen in the staging and follow-up of patients with colorectal cancer. Cancer Invest. England. 2005;23:338–51.

    Google Scholar 

  34. Duffy MJ. Carcinoembryonic antigen as a marker for colorectal cancer: is it clinically useful? Clin Chem. 2001;47:624–30.

    Article  CAS  PubMed  Google Scholar 

  35. Panina-Bordignon P, Tan A, Termijtelen A, Corradin G, Lanzavecchia A. Universally immunogenic T cell epitopes:promiscuous binding to human MHC class II and promiscuous recognition by T cells. Eur J Immunol. 1989;19:2237–42.

    Article  CAS  PubMed  Google Scholar 

  36. Demotz S, Lanzavecchia A, Eisel U, Niemann H, Widmann C, Corradin G. Delineation of several DR-restricted tetanus toxin T cell epitopes. J Immunol. 1989;142:394–402.

    Article  CAS  PubMed  Google Scholar 

  37. Finkelman FD, Katona IM, Mosmann TR, Coffman RL. IFN-gamma regulates the isotypes of Ig secreted during in vivo humoral immune responses. J Immunol. 1988;140:1022–7.

    Article  CAS  PubMed  Google Scholar 

  38. Kawano Y, Noma T, Yata J. Regulation of human IgG subclass production by cytokines. IFN-gamma and IL-6 act antagonistically in the induction of human IgG1 but additively in the induction of IgG2. J Immunol. 1994;153:4948–58.

    Article  CAS  PubMed  Google Scholar 

  39. Conaghan PJ, Ashraf SQ, Tytherleigh MG, Wilding JL, Tchilian E, Bicknell D, et al. Targeted killing of colorectal cancer cell lines by a humanised IgG1 monoclonal antibody that binds to membrane-bound carcinoembryonic antigen. Br J Cancer. 2008;98:1217–25.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  40. Blumenthal RD, Osorio L, Hayes MK, Horak ID, Hansen HJ, Goldenberg DM. Carcinoembryonic antigen antibody inhibits lung metastasis and augments chemotherapy in a human colonic carcinoma xenograft. Cancer Immunol Immunother. 2005;54:315–27.

    Article  CAS  PubMed  Google Scholar 

  41. Sharkey RM, Juweid M, Shevitz J, Behr T, Dunn R, Swayne LC, et al. Evaluation of a complementarity-determining region-grafted (humanized) anti-carcinoembryonic antigen monoclonal antibody in preclinical and clinical studies. Cancer Res. 1995;55:5935s–5945s.

    CAS  PubMed  Google Scholar 

  42. Greiner JW, Zeytin H, Anver MR, Schlom J. Vaccine-based therapy directed against carcinoembryonic antigen demonstrates antitumor activity on spontaneous intestinal tumors in the absence of autoimmunity. Cancer Res. 2002;62:6944–51.

    CAS  PubMed  Google Scholar 

  43. Saha A, Chatterjee SK, Foon KA, Primus FJ, Sreedharan S, Mohanty K, et al. Dendritic cells pulsed with an anti-idiotype antibody mimicking carcinoembryonic antigen (CEA) can reverse immunological tolerance to CEA and induce antitumor immunity in CEA transgenic mice. Cancer Res. 2004;7:4995–5003.

  44. Chiang CL-L, Benencia F, Coukos G. Whole tumor antigen vaccines. Semin Immunol. 2010;22:132–43.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  45. Joshi VB, Geary SM, Gross BP, Wongrakpanich A, Norian LA, Salem AK. Tumor lysate-loaded biodegradable microparticles as cancer vaccines. Expert Rev Vaccines. 2014;13:9–15.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  46. Tempero MA, Sivinski C, Steplewski Z, Harvey E, Klassen L, Kay HD. Phase II trial of interferon gamma and monoclonal antibody 17-1A in pancreatic cancer: biologic and clinical effects. J Clin Oncol. 1990;8:2019–26.

    Article  CAS  PubMed  Google Scholar 

  47. Wiesenfeld M, O’Connell MJ, Wieand HS, Gonchoroff NJ, Donohue JH, Fitzgibbons RJJ, et al. Controlled clinical trial of interferon-gamma as postoperative surgical adjuvant therapy for colon cancer. J Clin Oncol. 1995;13:2324–9.

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgements

This work was supported by the São Paulo Research Foundation (FAPESP; Grant Numbers: 2012/21861-1 and # 2013/17224-9). BFZ was a recipient of a FAPESP scholarship (2012/21861-1).

Author information

Authors and Affiliations

  1. Research Center for Gene Therapy, Escola Paulista de Medicina, Universidade Federal de São Paulo, São Paulo, SP, Brazil

    Bianca Ferrarini Zanetti & Sang Won Han

  2. Department of Microbiology, Immunology, and Parasitology, Escola Paulista de Medicina, Universidade Federal de São Paulo, São Paulo, SP, Brazil

    Camila Pontes Ferreira & José Ronnie Carvalho Vasconcelos

  3. Department of Biosciences, Instituto de Saúde e Sociedade, Universidade Federal de São Paulo, São Paulo, Brazil

    José Ronnie Carvalho Vasconcelos

  4. Department of Biophysics, Escola Paulista de Medicina, Universidade Federal de São Paulo, São Paulo, SP, Brazil

    Sang Won Han

Authors
  1. Bianca Ferrarini Zanetti
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Camila Pontes Ferreira
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. José Ronnie Carvalho Vasconcelos
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Sang Won Han
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

BFZ: conception and design of the study, acquisition of data, analysis and interpretation of the data, drafting the article; CPF: acquisition of data, analysis, and interpretation of data; JRCV: analysis and interpretation of the data; SWH: conception and design of the study, drafting the article, revising it critically for important intellectual content, and final approval of the version to be submitted.

Corresponding author

Correspondence to Sang Won Han.

Ethics declarations

Conflict of interest

The authors declare no competing interests.

Additional information

Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

The original online version of this article was revised: Fig. 2 has been corrected.

Rights and permissions

Springer Nature or its licensor (e.g. a society or other partner) holds exclusive rights to this article under a publishing agreement with the author(s) or other rightsholder(s); author self-archiving of the accepted manuscript version of this article is solely governed by the terms of such publishing agreement and applicable law.

Reprints and Permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Zanetti, B.F., Ferreira, C.P., Vasconcelos, J.R.C. et al. Adjuvant properties of IFN-γ and GM-CSF in the scFv6.C4 DNA vaccine against CEA-expressing tumors. Gene Ther 30, 41–50 (2023). https://doi.org/10.1038/s41434-021-00270-w

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/s41434-021-00270-w

Search

Advanced search

Quick links