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.

  • Original Article
  • Published:

Gene Therapy

Ectopic retroviral expression of LMO2, but not IL2Rγ, blocks human T-cell development from CD34+ cells: implications for leukemogenesis in gene therapy

Leukemia volume 21, pages 754–763 (2007)Cite this article

Abstract

The occurrence of leukemia in a gene therapy trial for SCID-X1 has highlighted insertional mutagenesis as an adverse effect. Although retroviral integration near the T-cell acute lymphoblastic leukemia (T-ALL) oncogene LIM-only protein 2 (LMO2) appears to be a common event, it is unclear why LMO2 was preferentially targeted. We show that of classical T-ALL oncogenes, LMO2 is most highly transcribed in CD34+ progenitor cells. Upon stimulation with growth factors typically used in gene therapy protocols transcription of LMO2, LYL1, TAL1 and TAN1 is most prominent. Therefore, these oncogenes may be susceptible to viral integration. The interleukin-2 receptor gamma chain (IL2Rγ), which is mutated in SCID-X1, has been proposed as a cooperating oncogene to LMO2. However, we found that overexpressing IL2Rγ had no effect on T-cell development. In contrast, retroviral overexpression of LMO2 in CD34+ cells caused severe abnormalities in T-cell development, but B-cell and myeloid development remained unaffected. Our data help explain why LMO2 was preferentially targeted over many of the other known T-ALL oncogenes. Furthermore, during T-cell development retrovirus-mediated expression of IL2Rγ may not be directly oncogenic. Instead, restoration of normal IL7-receptor signaling may allow progression of T-cell development to stages where ectopic LMO2 expression causes aberrant thymocyte growth.

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

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

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

Figure 1
Figure 2
Figure 3
Figure 4

Similar content being viewed by others

References

  1. Gaspar HB, Parsley KL, Howe S, King D, Gilmour KC, Sinclair J et al. Gene therapy of X-linked severe combined immunodeficiency by use of a pseudotyped gamma retroviral vector. Lancet 2004; 364: 2181–2187.

    Article  CAS  Google Scholar 

  2. Hacein-Bey-Abina S, Fischer A, Cavazzana-Calvo M . Gene therapy of X-linked severe combined immunodeficiency. Int J Hematol 2002; 76: 295–298.

    Article  CAS  Google Scholar 

  3. Hacein-Bey-Abina S, Le Deist F, Carlier F, Bouneaud C, Hue C, De Villartay JP et al. Sustained correction of X-linked severe combined immunodeficiency by ex vivo gene therapy. N Engl J Med 2002; 346: 1185–1193.

    Article  CAS  Google Scholar 

  4. Aiuti A, Slavin S, Aker M, Ficara F, Deola S, Mortellaro A et al. Correction of ADA-SCID by stem cell gene therapy combined with nonmyeloablative conditioning. Science 2002; 296: 2410–2413.

    Article  CAS  Google Scholar 

  5. Hacein-Bey-Abina S, von Kalle C, Schmidt M, Le Deist F, Wulffraat N, McIntyre E et al. A serious adverse event after successful gene therapy for X-linked severe combined immunodeficiency. N Engl J Med 2003; 348: 255–256.

    Article  Google Scholar 

  6. Marshall E . Gene therapy. Second child in French trial is found to have leukemia. Science 2003; 299: 320.

    Article  CAS  Google Scholar 

  7. Check E . Gene therapy put on hold as third child develops cancer. Nature 2005; 433: 561.

    Google Scholar 

  8. Hacein-Bey-Abina S, Von Kalle C, Schmidt M, McCormack MP, Wulffraat N, Leboulch P et al. LMO2-associated clonal T cell proliferation in two patients after gene therapy for SCID-X1. Science 2003; 302: 415–419.

    Article  CAS  Google Scholar 

  9. Dave UP, Jenkins NA, Copeland NG . Gene therapy insertional mutagenesis insights. Science 2004; 303: 333.

    Article  Google Scholar 

  10. Woods NB, Bottero V, Schmidt M, von Kalle C, Verma IM . Gene therapy: therapeutic gene causing lymphoma. Nature 2006; 440: 1123.

    Article  CAS  Google Scholar 

  11. Foot AB, Oakhill A, Kitchen C . Acute monoblastic leukemia of infancy in Klinefelter's syndrome. Cancer Genet Cytogenet 1992; 61: 99–100.

    Article  CAS  Google Scholar 

  12. Lessard J, Sauvageau G . Bmi-1 determines the proliferative capacity of normal and leukaemic stem cells. Nature 2003; 423: 255–260.

    Article  CAS  Google Scholar 

  13. Hussey DJ, Nicola M, Moore S, Peters GB, Dobrovic A . The (4;11)(q21;p15) translocation fuses the NUP98 and RAP1GDS1 genes and is recurrent in T-cell acute lymphocytic leukemia. Blood 1999; 94: 2072–2079.

    CAS  Google Scholar 

  14. Woods NB, Bottero V, Schmidt M, von Kalle C, Verma IM . Gene therapy: is IL2RG oncogenic in T-cell development?: X-SCID transgene leukaemogenicity (reply). Nature 2006; 443: E6–E7.

    Article  CAS  Google Scholar 

  15. Shou Y, Ma Z, Lu T, Sorrentino BP . Unique risk factors for insertional mutagenesis in a mouse model of XSCID gene therapy. Proc Natl Acad Sci USA 2006; 103: 11730–11735.

    Article  CAS  Google Scholar 

  16. Pike-Overzet K, de Ridder D, Weerkamp F, Baert MR, Verstegen MM, Brugman MH et al. Gene therapy: is IL2RG oncogenic in T-cell development? Nature 2006; 443: E6–E7, (E5 discussion).

    Article  Google Scholar 

  17. Dik WA, Pike-Overzet K, Weerkamp F, de Ridder D, de Haas EF, Baert MR et al. New insights on human T cell development by quantitative T cell receptor gene rearrangement studies and gene expression profiling. J Exp Med 2005; 201: 1715–1723.

    Article  CAS  Google Scholar 

  18. van Hennik PB, Verstegen MM, Bierhuizen MF, Limon A, Wognum AW, Cancelas JA et al. Highly efficient transduction of the green fluorescent protein gene in human umbilical cord blood stem cells capable of cobblestone formation in long-term cultures and multilineage engraftment of immunodeficient mice. Blood 1998; 92: 4013–4022.

    CAS  Google Scholar 

  19. Wognum AW, Visser TP, Peters K, Bierhuizen MF, Wagemaker G . Stimulation of mouse bone marrow cells with kit ligand, FLT3 ligand, and thrombopoietin leads to efficient retrovirus-mediated gene transfer to stem cells, whereas interleukin 3 and interleukin 11 reduce transduction of short- and long-term repopulating cells. Hum Gene Ther 2000; 11: 2129–2141.

    Article  CAS  Google Scholar 

  20. Cavazzana-Calvo M, Hacein-Bey S, de Saint Basile G, Gross F, Yvon E, Nusbaum P et al. Gene therapy of human severe combined immunodeficiency (SCID)-X1 disease. Science 2000; 288: 669–672.

    Article  CAS  Google Scholar 

  21. Irizarry RA, Hobbs B, Collin F, Beazer-Barclay YD, Antonellis KJ, Scherf U et al. Exploration, normalization, and summaries of high density oligonucleotide array probe level data. Biostatistics 2003; 4: 249–264.

    Article  Google Scholar 

  22. Bolstad BM, Irizarry RA, Astrand M, Speed TP . A comparison of normalization methods for high density oligonucleotide array data based on variance and bias. Bioinformatics 2003; 19: 185–193.

    Article  CAS  Google Scholar 

  23. de Ridder D, Staal FJ, van Dongen JJ, Reinders MJ . Maximum significance clustering of oligonucleotide microarrays. Bioinformatics 2006; 22: 326–331.

    Article  CAS  Google Scholar 

  24. Kinsella TM, Nolan GP . Episomal vectors rapidly and stably produce high-titer recombinant retrovirus. Hum Gene Ther 1996; 7: 1405–1413.

    Article  CAS  Google Scholar 

  25. Weerkamp F, Baert MR, Brugman MH, Dik WA, de Haas EF, Visser TP et al. Human thymus contains multipotent progenitors with T/B lymphoid, myeloid, and erythroid lineage potential. Blood 2006; 107: 3131–3137.

    Article  CAS  Google Scholar 

  26. Schroder AR, Shinn P, Chen H, Berry C, Ecker JR, Bushman F . HIV-1 integration in the human genome favors active genes and local hotspots. Cell 2002; 110: 521–529.

    Article  CAS  Google Scholar 

  27. Mitchell RS, Beitzel BF, Schroder AR, Shinn P, Chen H, Berry CC et al. Retroviral DNA integration: ASLV, HIV, and MLV show distinct target site preferences. PLoS Biol 2004; 2: E234.

    Article  Google Scholar 

  28. Ng YY, van Kessel B, Lakhorst HM, Baert MR, van den Burg CM, Bloem AC et al. Gene expression profiling of CD34+ cells from various hematopoietic stem-cell sources reveals functional differences in stem-cell activity. J Leukoc Biol 2004; 75: 314–323.

    Article  CAS  Google Scholar 

  29. Spits H, Blom B, Jaleco AC, Weijer K, Verschuren MC, van Dongen JJ et al. Early stages in the development of human T, natural killer and thymic dendritic cells. Immunol Rev 1998; 165: 75–86.

    Article  CAS  Google Scholar 

  30. Weerkamp F, de Haas EF, Naber BA, Comans-Bitter WM, Bogers AJ, van Dongen JJ et al. Age-related changes in the cellular composition of the thymus in children. J Allergy Clin Immunol 2005; 115: 834–840.

    Article  Google Scholar 

  31. Puel A, Ziegler SF, Buckley RH, Leonard WJ . Defective IL7R expression in T(-)B(+)NK(+) severe combined immunodeficiency. Nat Genet 1998; 20: 394–397.

    Article  CAS  Google Scholar 

  32. Roifman CM, Zhang J, Chitayat D, Sharfe N . A partial deficiency of interleukin-7R alpha is sufficient to abrogate T-cell development and cause severe combined immunodeficiency. Blood 2000; 96: 2803–2807.

    CAS  Google Scholar 

  33. Li Z, Dullmann J, Schiedlmeier B, Schmidt M, von Kalle C, Meyer J et al. Murine leukemia induced by retroviral gene marking. Science 2002; 296: 497.

    Article  CAS  Google Scholar 

  34. Calmels B, Ferguson C, Laukkanen MO, Adler R, Faulhaber M, Kim HJ et al. Recurrent retroviral vector integration at the Mds1/Evi1 locus in nonhuman primate hematopoietic cells. Blood 2005; 106: 2530–2533.

    Article  CAS  Google Scholar 

  35. Weerkamp F, Pike-Overzet K, Staal FJ . T-sing progenitors to commit. Trends Immunol 2006; 27: 125–131.

    Article  CAS  Google Scholar 

  36. Thrasher AJ, Gaspar HB, Baum C, Modlich U, Schambach A, Candotti F et al. Gene therapy: X-SCID transgene leukaemogenicity. Nature 2006; 443: E5–E6, (discussion E6–E7).

    Article  CAS  Google Scholar 

  37. Larson RC, Osada H, Larson TA, Lavenir I, Rabbitts TH . The oncogenic LIM protein Rbtn2 causes thymic developmental aberrations that precede malignancy in transgenic mice. Oncogene 1995; 11: 853–862.

    CAS  Google Scholar 

  38. Neale GA, Rehg JE, Goorha RM . Ectopic expression of rhombotin-2 causes selective expansion of CD4-CD8- lymphocytes in the thymus and T-cell tumors in transgenic mice. Blood 1995; 86: 3060–3071.

    CAS  Google Scholar 

  39. Allman D, Sambandam A, Kim S, Miller JP, Pagan A, Well D et al. Thymopoiesis independent of common lymphoid progenitors. Nat Immunol 2003; 4: 168–174.

    Article  CAS  Google Scholar 

  40. Weng AP, Ferrando AA, Lee W, Morris JPt, Silverman LB, Sanchez-Irizarry C et al. Activating mutations of NOTCH1 in human T cell acute lymphoblastic leukemia. Science 2004; 306: 269–271.

    Article  CAS  Google Scholar 

  41. Trono D . Virology. Picking the right spot. Science 2003; 300: 1670–1671.

    Article  CAS  Google Scholar 

  42. Baum C, von Kalle C, Staal FJ, Li Z, Fehse B, Schmidt M et al. Chance or necessity? Insertional mutagenesis in gene therapy and its consequences. Mol Ther 2004; 9: 5–13.

    Article  CAS  Google Scholar 

Download references

Acknowledgements

We thank Dr Langerak, Dr van Velden and Dr Dik for critically reading the manuscript. In addition, we are grateful for the primers and probes made available by Dr Dik. We also thank E de Haas for performing the sorting and Dr Kwee Yong, Department of Haematology, University College London, for supplying the PBSC. This work was supported in part by the 5th and 6th EU Framework program (Contract Nos. QLK3-CT-2001-0427 (INHERINET) and LSHB-CT-2004-005242 (CONSERT), as well as by the Translational Gene Therapy Research Programme of ZonMw – the Netherlands Organization for Health Research and Development. AJT is supported by the Wellcome Trust.

Author information

Authors and Affiliations

  1. Department of Immunology, Erasmus University Medical Center, Rotterdam, The Netherlands

    K Pike-Overzet, F Weerkamp, M R M Baert, J J M van Dongen & F J T Staal

  2. Information and Communication Theory Group, Faculty of Electrical Engineering, Mathematics and Computer Science, Delft University of Technology, Delft, The Netherlands

    D de Ridder & M J T Reinders

  3. Department of Hematology, Erasmus University Medical Center, Rotterdam, The Netherlands

    M M A Verstegen, M H Brugman & G Wagemaker

  4. Molecular Immunology Unit, Institute of Child Health, University College London, London, UK

    S J Howe & A J Thrasher

Authors
  1. K Pike-Overzet
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. D de Ridder
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. F Weerkamp
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. M R M Baert
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. M M A Verstegen
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. M H Brugman
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. S J Howe
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. M J T Reinders
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. A J Thrasher
    View author publications

    You can also search for this author in PubMed Google Scholar

  10. G Wagemaker
    View author publications

    You can also search for this author in PubMed Google Scholar

  11. J J M van Dongen
    View author publications

    You can also search for this author in PubMed Google Scholar

  12. F J T Staal
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to F J T Staal.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Pike-Overzet, K., de Ridder, D., Weerkamp, F. et al. Ectopic retroviral expression of LMO2, but not IL2Rγ, blocks human T-cell development from CD34+ cells: implications for leukemogenesis in gene therapy. Leukemia 21, 754–763 (2007). https://doi.org/10.1038/sj.leu.2404563

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/sj.leu.2404563

Keywords

This article is cited by

Search

Advanced search

Quick links