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Construction, expression, and function of 6B11ScFv–mIL-12, a fusion protein that attacks human ovarian carcinoma

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Abstract

We previously produced an anti-idiotypic monoclonal antibody, 6B11, which mimics ovarian cancer antigen CA166-9 and induces cellular and humoral immunity. Here, to enhance the immunogenicity of 6B11, we constructed the 6B11ScFv–mIL-12 fusion protein (FP), by fusing single-chain fragment of 6B11 variable region (6B11ScFv) with mouse interleukin-12 (mIL-12), which was expressed in eukaryotic 293EBNA cells transfected with pSBI vectors. A binding activity assay showed 6B11ScFv–mIL-12 to have activities of both 6B11 and mIL-12—it specifically bound both ovarian monoclonal antibody COC166-9 and rabbit anti-mouse IL-12 antibody. The immune activity assay showed 6B11ScFv–mIL-12 to promote proliferation of lymphocytes stimulated by phytohemagglutinin, increase the absolute numbers and percentages of CD3−/CD56+ natural killer cells and CD3+/CD56+ natural killer T cells among peripheral lymphocytes, and increase interferon-γ. The FP was specifically cytotoxic to the CA166-9+ ovarian cancer cell lines HOC1A and SKOV3 and inhibited growth of ID8 subcutaneous tumors in C57BL/6J mice. This study provides an experimental basis for clinical use of 6B11ScFv–mIL-12 in ovarian cancer therapy. To our knowledge, this is the first report of a fusion protein from an anti-idiotypic antibody and IL-12.

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References

  1. Rebecca S, Deepa N, Ahmedin J. Cancer Statistics, 2012. CA Cancer J Clin. 2012;62:10–29.

    Article  Google Scholar 

  2. de Cerio AL, Zabalegui N, Rodríguez-Calvillo M, Inogés S, Bendandi M. Anti-idiotype antibodies in cancer treatment. Oncogene. 2007;26:3594–602.

    Article  PubMed  Google Scholar 

  3. Yin X, Yan X, Yang Q, Cao H, Liang H. Antitumor mechanism of recombinant murine interleukin-12 vaccine. Cancer Biother Radiopharm. 2010;25:263–8.

    Article  CAS  PubMed  Google Scholar 

  4. Kayashima H, Toshima T, Okano S, Taketomi A, Harada N, Yamashita Y, et al. Intratumoral neo-adjuvant immunotherapy using IL-12 and dendritic cells is an effective strategy to control recurrence of murine hepatocellular carcinoma in immunosuppressed mice. J Immunol. 2010;185:698–708.

    Article  CAS  PubMed  Google Scholar 

  5. Qian HN, Feng J, Cui H, Fu TY, Wei P, Fu ZY. Generation and characterization of three monoclonal antibodies to human ovarian epithelial adenocarcinomas. Chin Med J (Engl). 1989;102:839–43.

    CAS  Google Scholar 

  6. Qian HN, Lu WY. Anti-idiotypic monoclonal antibodies against anti-ovarian carcinoma monoclonal antibody COC166-9 generation and application. Chin Med J (Engl). 1994;107:99–103.

    CAS  Google Scholar 

  7. Yang W, Feng J, Chang X, Fu T, Ye X, Zhang H, et al. Cytotoxic effects of T cells induced by fusion protein 6B11-pulsed dendritic cells on ovarian carcinoma cells. Gynecol Oncol. 2007;105:238–43.

    Article  CAS  PubMed  Google Scholar 

  8. Li W, Cui H, Meng FQ, Chang XH, Zhang G, Liu B, et al. New T cell epitopes identified from an anti-idiotypic antibody mimicking ovarian cancer associated antigen. Cancer Immunol Immunother. 2008;57:143–54.

    Article  CAS  PubMed  Google Scholar 

  9. Redfern CH, Guthrie TH, Bessudo A, Densmore JJ, Holman PR, Janakiraman N, et al. Phase II trial of idiotype vaccination in previously treated patients with indolent non-Hodgkin’s lymphoma resulting in durable clinical responses. J Clin Oncol. 2006;24:3107–12.

    Article  CAS  PubMed  Google Scholar 

  10. Reinis M. Drug evaluation: FavId, a patient-specific idiotypic vaccine for non-Hodgkin’s lymphoma. Curr Opin Mol Ther. 2007;9:291–8.

    CAS  PubMed  Google Scholar 

  11. LaCasce AS, Freedman AS. Antibody and immunomodulatory agents in the treatment of indolent non-Hodgkin’s lymphoma. Semin Hematol. 2008;45:85–9.

    Article  CAS  PubMed  Google Scholar 

  12. Wang X, Ko EC, Peng L, Gillies SD, Ferrone S. Human high molecular weight melanoma-associated antigen mimicry by mouse anti-idiotypic monoclonal antibody MK2-23: enhancement of immunogenicity of anti-idiotypic monoclonal antibody MK2-23 by fusion with interleukin-2. Cancer Res. 2005;65:6976–83.

    Article  CAS  PubMed  Google Scholar 

  13. Chong G, Bhatnagar A, Cunningham D, Cosgriff TM, Harper PG, Steward W, et al. Phase III trial of 5-fluorouracil and leucovorin plus either 3H1 anti-idiotype monoclonal antibody or placebo in patients with advanced colorectal cancer. Ann Oncol. 2006;17:437–42.

    Article  CAS  PubMed  Google Scholar 

  14. Saha A, Chatterjee SK, Foon KA, Celis E, Bhattacharya-Chatterjee M. Therapy of established tumors in a novel murine model transgenic for human carcinoembryonic antigen and HLA-A2 with a combination of anti-idiotype vaccine and CTL peptides of carcinoembryonic antigen. Cancer Res. 2007;67:2881–92.

    Article  CAS  PubMed  Google Scholar 

  15. Ullenhag GJ, Spendlove I, Watson NF, Indar AA, Dube M, Robins RA, et al. A neo-adjuvant/adjuvant randomized trial of colorectal cancer patients vaccinated with an anti-idiotypic antibody, 105AD7, mimicking CD55. Clin Cancer Res. 2006;12:7389–96.

    Article  CAS  PubMed  Google Scholar 

  16. Fernandez LE, Gabri MR, Guthmann MD, Gomez RE, Gold S, Fainboim L, et al. NGcGM3 ganglioside: a privileged target for cancer vaccines. Clin Dev Immunol. 2010;2010:814397.

    Article  PubMed Central  PubMed  Google Scholar 

  17. Neninger E, Díaz RM, de la Torre A, Rives R, Díaz A, Saurez G, et al. Active immunotherapy with 1E10 anti-idiotype vaccine in patients with small cell lung cancer. Report of a Phase I trial. Cancer Biol Ther. 2007;6:145–50.

    Article  CAS  PubMed  Google Scholar 

  18. Alfonso S, Diaz RM, de la Torre A, Santiesteban E, Aguirre F, Pérez K, et al. 1E10 anti-idiotype vaccine in non-small cell lung cancer: experience in stage IIIb/IV patients. Cancer Biol Ther. 2007;6:1847–52.

    Article  CAS  PubMed  Google Scholar 

  19. Ladjemi MZ, Jacot W, Chardès T, Pèlegrin A, Navarro-Teulon I. Anti-HER2 vaccines: new prospects for breast cancer therapy. Cancer Immunol Immunother. 2010;59:1295–312.

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  20. Mohanty K, Saha A, Pal S, Mallick P, Chatterjee SK, Foon KA, et al. Anti-tumor immunity induced by anti-idiotype antibody mimicking human Her-2/neu. Breast Cancer Res Treat. 2007;104:1–11.

    Article  CAS  PubMed  Google Scholar 

  21. Saha A, Chatterjee SK. Dendritic cells pulsed with an anti-idiotype antibody mimicking Her-2/neu induced protective antitumor immunity in two lines of Her-2/neu transgenic mice. Cell Immunol. 2010;263:9–21.

    Article  CAS  PubMed  Google Scholar 

  22. Alvarez-Rueda N, Ladjemi MZ, Béhar G, Corgnac S, Pugnière M, Roquet F, et al. A llama single domain anti-idiotypic antibody mimicking HER2 as a vaccine: immunogenicity and efficacy. Vaccine. 2009;27:4826–33.

    Article  CAS  PubMed  Google Scholar 

  23. Ladjemi MZ, Chardes T, Corgnac S, Garambois V, Morisseau S, Robert B, et al. Vaccination with human anti-trastuzumab anti-idiotype scFv reverses HER2 immunological tolerance and induces tumor immunity in MMTV.f.huHER2 (Fo5) mice. Breast Cancer Res. 2011;13:R17.

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  24. Wagner U, Köhler S, Reinartz S, Giffels P, Huober J, Renke K, et al. Immunological consolidation of ovarian carcinoma recurrences with monoclonal anti-idiotype antibody ACA125: immune responses and survival in palliative treatment. Clin Cancer Res. 2001;7:1154–62.

    CAS  PubMed  Google Scholar 

  25. Reinartz S, Köhler S, Schlebusch H, Krista K, Giffels P, Renke K, et al. Vaccination of patients with advanced ovarian carcinoma with the anti-idiotype ACA125: immunological response and survival (Phase Ib/II). Clin Cancer Res. 2004;10:1580–7.

    Article  CAS  PubMed  Google Scholar 

  26. Pfisterer J, du Bois A, Sehouli J, Loibl S, Reinartz S, Reuss A, et al. The anti-idiotypic antibody abagovomab in patients with recurrent ovarian cancer. A phase I trial of the AGO-OVAR. Ann Oncol. 2006;17:1568–77.

    Article  CAS  PubMed  Google Scholar 

  27. Grisham RN, Berek J, Pfisterer J, Sabbatini P. Abagovomab: an anti-idiotypic CA-125 targeted immunotherapeutic agent for ovarian cancer. Immunotherapy. 2011;3:153–62.

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  28. McCormick AL, Thomas MS, Heath AW. Enhancement of immunogenicity of recombinant antigens by production of a cytokine-antigen fusion protein for vaccination. Biochem Soc Trans. 1997;25(2):297S.

    CAS  PubMed  Google Scholar 

  29. Schwegler Christian, Dorn-Beineke Alexandra, Nittka Stefanie, Stocking Carol, Neumaier Michael. 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 

  30. Reinartz Silke, Hombach Andreas, Köhler Siegmund, Schlebusch Harald, Wallwiener Diethelm, Abken Hinrich, et al. Interleukin-6 fused to an anti-idiotype antibody in a vaccine increases the specific humoral immune response against CA125+ (MUC-16) ovarian cancer. Cancer Res. 2003;63:3234–40.

    CAS  PubMed  Google Scholar 

  31. Hammerstrom AE, Cauley DH, Atkinson BJ, Sharma P. Cancer immunotherapy: sipuleucel-T and beyond. Pharmacotherapy. 2011;31:813–28.

    Article  PubMed Central  PubMed  Google Scholar 

  32. Kim TS, DeKruyff RH, Rupper R, Maecker HT, Levy S, Umetsu DT. An ovalbumin-IL-12 fusion protein is more effective than ovalbumin plus free recombinant IL-12 in inducing a T helper cell type 1-dominated immune response and inhibiting antigen-specific IgE production. J Immunol. 1997;158:4137–44.

    CAS  PubMed  Google Scholar 

  33. Cooper AM, Khader SA. IL-12p40: an inherently agonistic cytokine. Trends Immunol. 2007;28:33–8.

    Article  CAS  PubMed  Google Scholar 

  34. Lieschke GJ, Rao PK, Gately MK, Mulligan RC. Bioactive murine and human interleukin-12 fusion proteins which retain antitumor activity in vivo. Nat Biotechnol. 1997;15:35–40.

    Article  CAS  PubMed  Google Scholar 

  35. Peng LS, Penichet ML, Morrison SL. A single-chain IL-12IgG3 antibody fusion protein retains antibody specificity and IL-12 bioactivity and demonstrates antitumor activity. J Immunol. 1999;163:250–8.

    CAS  PubMed  Google Scholar 

Download references

Acknowledgments

This work was supported by the National Natural Science Foundation of China (NSFC, Grant No. 30901748) and the National High Technology Research and Development Program (863 Program, Grant No. 2012AA02A507).

Conflict of interest

The authors have declared that no conflict of interests exists.

Ethical standard

We declare that all human and animal studies have been approved by the Institutional Review Board of Peking University People’s Hospital when we applied for the National Natural Science Foundation of China project and therefore been performed in accordance with the ethical standards laid down in the 1964 Declaration of Helsinki and its later amendments. Human peripheral blood mononuclear cells (PBMCs) were provided by the blood donation volunteers of the Blood Center of the Beijing Red Cross, and all volunteers gave their informed written consent prior to their inclusion in the study. Details that might disclose the identity of the subject under study have been omitted.

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    Authors and Affiliations

    1. Key Laboratory of Cell Proliferation and Regulation Biology, College of Life Science, Beijing Normal University, No. 19, Xin Jie Kou Wai Street, HaiDian District, Beijing, 100875, People’s Republic of China

      Hongyan Cheng & Jianli Sang

    2. Gynecology Oncology Center of Peking University People’s Hospital, No. 11, South Avenue, Xi Zhi Men, Xicheng District, Beijing, 100044, People’s Republic of China

      Hongyan Cheng, Xue Ye, Xiaohong Chang, Ruiqiong Ma, Xu Cong, Yidong Niu, Menglei Zhang, Kai Liu & Heng Cui

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    1. Hongyan Cheng
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    Heng Cui and Jianli Sang have contributed equally to this work.

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    Cheng, H., Ye, X., Chang, X. et al. Construction, expression, and function of 6B11ScFv–mIL-12, a fusion protein that attacks human ovarian carcinoma. Med Oncol 32, 130 (2015). https://doi.org/10.1007/s12032-015-0586-y

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