Skip to main content
SpringerLink
Log in

A truncated apoptin protein variant selectively kills cancer cells

  • PRECLINICAL STUDIES
  • Published:
Investigational New Drugs Aims and scope Submit manuscript

Summary

Apoptin is a nonstructural protein encoded by one of the three open reading frames of the chicken anemia virus genome. It has attracted a great deal of interest due to its ability to induce apoptosis in multiple transformed and malignant mammalian cell lines without affecting primary and non-transformed cells. However, the use of Apoptin as an anticancer drug is restricted by its strong tendency to aggregate. A number of methods to overcome this problem have been proposed, including transduction techniques to deliver the Apoptin gene into tumor cells, but all such methods have certain drawbacks. Here we describe that a truncated variant of Apoptin, lacking residues 1 to 43, is a soluble, non-aggregating protein that maintains most of the biological properties of wild-type Apoptin when transfected into cells. We show that the cytotoxic effect of this variant is also present when it is added exogenously to cancer cells, but not to normal cells. In addition to the interest this protein has attracted as a promising therapeutic strategy, it is also an excellent model to study the structural properties of Apoptin and how they relate to its mechanism of action.

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

Access this article

Log in via an institution

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

Similar content being viewed by others

References

  1. Prasetyo AA, Kamahora T, Kuroishi A et al (2009) Replication of chicken anemia virus (CAV) requires apoptin and is complemented by VP3 of human torque Teno virus (TTV). Virology 385:85–92. doi:10.1016/j.virol.2008.10.043

    Article  CAS  PubMed  Google Scholar 

  2. Maddika S, Mendoza FJ, Hauff K et al (2006) Cancer-selective therapy of the future: apoptin and its mechanism of action. Cancer Biol Ther 5:10–19. doi:10.4161/cbt.5.1.2400

    Article  CAS  PubMed  Google Scholar 

  3. Poon IKH, Oro C, Dias MM et al (2005) A tumor cell-specific nuclear targeting signal within chicken anemia virus VP3/apoptin. J Virol 79:1339–1341. doi:10.1128/JVI.79.2.1339-1341.2005

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  4. Leliveld SR, Zhang Y-H, Rohn JL et al (2003) Apoptin induces tumor-specific apoptosis as a globular multimer. J Biol Chem 278:9042–9051. doi:10.1074/jbc.M210803200

    Article  CAS  PubMed  Google Scholar 

  5. Leliveld SR, Dame RT, Mommaas MA et al (2003) Apoptin protein multimers form distinct higher-order nucleoprotein complexes with DNA. Nucleic Acids Res 31:4805–4813

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  6. Danen-Van Oorschot AAAM, Zhang Y-HH, Leliveld SR et al (2003) Importance of nuclear localization of apoptin for tumor-specific induction of apoptosis. J Biol Chem 278:27729–27736. doi:10.1074/jbc.M303114200

    Article  CAS  PubMed  Google Scholar 

  7. Rollano Peñaloza OM, Lewandowska M, Stetefeld J et al (2014) Apoptins: selective anticancer agents. Trends Mol Med 20:519–528. doi:10.1016/j.molmed.2014.07.003

    Article  PubMed  Google Scholar 

  8. Rohn JL (2002) A tumor-specific kinase activity regulates the viral death protein apoptin. J Biol Chem 277:50820–50827. doi:10.1074/jbc.M208557200

    Article  CAS  PubMed  Google Scholar 

  9. Poon IKH, Oro C, Dias MM et al (2005) Apoptin nuclear accumulation is modulated by a CRM1-recognized nuclear export signal that is active in normal but not in tumor cells. Cancer Res 65:7059–7064. doi:10.1158/0008-5472.CAN-05-1370

    Article  CAS  PubMed  Google Scholar 

  10. Zhou S, Zhang M, Zhang J et al (2012) Mechanisms of apoptin-induced cell death. Med Oncol 29:2985–2991. doi:10.1007/s12032-011-0119-2

    Article  CAS  PubMed  Google Scholar 

  11. Leliveld SR, Dame RT, Rohn JL et al (2004) Apoptin’s functional N- and C-termini independently bind DNA. FEBS Lett 557:155–158

    Article  CAS  PubMed  Google Scholar 

  12. Guelen L, Paterson H, Gäken J et al (2004) TAT-apoptin is efficiently delivered and induces apoptosis in cancer cells. Oncogene 23:1153–1165. doi:10.1038/sj.onc.1207224

    Article  CAS  PubMed  Google Scholar 

  13. Zhang M, Guller S, Huang Y (2007) Method to enhance transfection efficiency of cell lines and placental fibroblasts. Placenta 28:779–782. doi:10.1016/j.placenta.2007.01.012

    Article  CAS  PubMed  Google Scholar 

  14. Castro J, Ribó M, Puig T et al (2012) A cytotoxic ribonuclease reduces the expression level of P-glycoprotein in multidrug-resistant cell lines. Invest New Drugs 30:880–888. doi:10.1007/s10637-011-9636-2

    Article  CAS  PubMed  Google Scholar 

  15. Castro J, Ribó M, Navarro S et al (2011) A human ribonuclease induces apoptosis associated with p21WAF1/CIP1 induction and JNK inactivation. BMC Cancer 11:9. doi:10.1186/1471-2407-11-9

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. Gras SL, Waddington LJ, Goldie KN (2011) Transmission electron microscopy of amyloid fibrils. Methods Mol Biol 752:197–214. doi:10.1007/978-1-60327-223-0_13

    Article  CAS  PubMed  Google Scholar 

  17. Noteborn MH, Todd D, Verschueren CA et al (1994) A single chicken anemia virus protein induces apoptosis. J Virol 68:346–351

    CAS  PubMed  PubMed Central  Google Scholar 

  18. Zhang YH, Leliveld SR, Kooistra K et al (2003) Recombinant apoptin multimers kill tumor cells but are nontoxic and epitope-shielded in a normal-cell-specific fashion. Exp Cell Res 289:36–46. doi:10.1016/S0014-4827(03)00188-5

    Article  CAS  PubMed  Google Scholar 

  19. Ruiz-Martínez S, Pantoja-Uceda D, Castro J et al (2017) Insights into the mechanism of Apoptin’s exquisitely selective anti-tumor action from atomic level characterization of its conformation and dynamics. Arch Biochem Biophys 614:53–64. doi:10.1016/j.abb.2016.12.010

    Article  PubMed  Google Scholar 

  20. Gualfetti PJ, Iwakura M, Lee JC et al (1999) Apparent radii of the native, stable intermediates and unfolded conformers of the alpha-subunit of tryptophan synthase from E. coli, a TIM barrel protein. Biochemistry 38:13367–13378

    Article  CAS  PubMed  Google Scholar 

  21. Rohn JL, Zhang Y-H, Leliveld SR et al (2005) Relevance of apoptin’s integrity for its functional behavior. J Virol 79:1337–1338. doi:10.1128/JVI.79.2.1337-1338.2005

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  22. Pavlakis N, Vogelzang NJ (2006) Ranpirnase-an antitumour ribonuclease: its potential role in malignant mesothelioma. Expert Opin Biol Ther 6:391–399. doi:10.1517/14712598.6.4.391

    Article  CAS  PubMed  Google Scholar 

  23. Zhao J, Gao P, Xiao W et al (2011) A novel human derived cell-penetrating peptide in drug delivery. Mol Biol Rep 38:2649–2656. doi:10.1007/s11033-010-0406-6

    Article  CAS  PubMed  Google Scholar 

  24. Sun J, Yan Y, Wang X-T et al (2009) PTD4-apoptin protein therapy inhibits tumor growth in vivo. Int J Cancer 124:2973–2981. doi:10.1002/ijc.24279

    Article  CAS  PubMed  Google Scholar 

  25. Zhao M, Hu B, Gu Z et al (2013) Degradable polymeric nanocapsule for efficient intracellular delivery of a high molecular weight tumor-selective protein complex. Nano Today 8:11–20. doi:10.1016/j.nantod.2012.12.003

    Article  CAS  Google Scholar 

  26. Ring J, Seifert J, Jesch F, Brendel W (1977) Anaphylactoid reactions due to non-immune complex serum protein aggregates. Monogr Allergy 12:27–35

    CAS  PubMed  Google Scholar 

  27. Benito A, Vilanova M, Ribó M (2008) Intracellular routing of cytotoxic pancreatic-type ribonucleases. Curr Pharm Biotechnol 9:169–179

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgements

We are very grateful to Dr. Malvash Tavassoli (King’s College, UK) for providing us with the Apoptin gene, to Dr. Bruno Beaumelle (CPBS, France) for providing us with K-562 cell line and to Dr. J. L. Corchero (Autonomous University of Barcelona, Spain) for help with the DLS experiment. S. R-M gratefully acknowledges his FPU fellowship from the MEC, Spain.

Author information

Authors and Affiliations

  1. Laboratori d’Enginyeria de Proteïnes, Departament de Biologia, Facultat de Ciències, Universitat de Girona and Institut d’Investigació Biomèdica de Girona Josep Trueta (IdIBGi), Girona, Spain

    Santiago Ruiz-Martínez, Jessica Castro, Maria Vilanova, Marc Ribó & Antoni Benito

  2. Instituto de Química Física “Rocasolano”, Consejo Superior de Investigaciones Científicas, Serrano 119, 28006, Madrid, Spain

    Marta Bruix & Douglas V. Laurents

Authors
  1. Santiago Ruiz-Martínez
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Jessica Castro
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Maria Vilanova
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Marta Bruix
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Douglas V. Laurents
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Marc Ribó
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Antoni Benito
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Antoni Benito.

Ethics declarations

Conflict of interest

The authors declare that they have no conflict of interest.

Funding

The work was supported by grants BFU2009-06935, BIO2013-43517, SAF-2013-49179-C2-2-R, QTC2014-52633-P and UNGI 10-4E-417 from MINECO (Spain) and MPCU2016/18 and SING12/0 from UdG.

Ethical approval

This article does not contain any studies performed by any of the authors involving human or animal participants.

Informed consent

For this type of study, formal consent is not required.

Electronic supplementary material

ESM 1

(DOCX 2694 kb)

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Ruiz-Martínez, S., Castro, J., Vilanova, M. et al. A truncated apoptin protein variant selectively kills cancer cells. Invest New Drugs 35, 260–268 (2017). https://doi.org/10.1007/s10637-017-0431-6

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s10637-017-0431-6

Keywords

Search

Navigation