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:

Acute Leukemias

The novel calicheamicin-conjugated CD22 antibody inotuzumab ozogamicin (CMC-544) effectively kills primary pediatric acute lymphoblastic leukemia cells

Leukemia volume 26, pages 255–264 (2012)Cite this article

Abstract

We investigated whether the newly developed antibody (Ab) -targeted therapy inotuzumab ozogamicin (CMC-544), consisting of a humanized CD22 Ab linked to calicheamicin, is effective in pediatric primary B-cell precursor acute lymphoblastic leukemia (BCP-ALL) cells in vitro, and analyzed which parameters determine its efficacy. CMC-544 induced dose-dependent cell kill in the majority of BCP-ALL cells, although IC50 values varied substantially (median 4.8 ng/ml, range 0.1–1000 ng/ml at 48 h). The efficacy of CMC-544 was highly dependent on calicheamicin sensitivity and CD22/CMC-544 internalization capacity of BCP-ALL cells, but hardly on basal and renewed CD22 expression. Although CD22 expression was essential for uptake of CMC-544, a repetitive loop of CD22 saturation, CD22/CMC-544 internalization and renewed CD22 expression was not required to achieve intracellular threshold levels of calicheamicin sufficient for efficient CMC-544-induced apoptosis in BCP-ALL cells. This is in contrast to studies with the comparable CD33 immunotoxin gemtuzumab ozogamicin (Mylotarg) in acute myeloid leukemia (AML) patients, in which complete and prolonged CD33 saturation was required for apoptosis induction. These data suggest that CMC-544 treatment may result in higher response rates in ALL compared with response rates obtained in AML with Mylotarg, and that therefore clinical studies in ALL, preferably with multiple low CMC-544 dosages, are warranted.

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
Figure 5
Figure 6
Figure 7

Similar content being viewed by others

References

  1. Johnston WT, Lightfoot TJ, Simpson J, Roman E . Childhood cancer survival: a report from the United Kingdom Childhood Cancer Study. Cancer Epidemiol 2010; 34: 659–666.

    Article  Google Scholar 

  2. Pui CH, Evans WE . Treatment of acute lymphoblastic leukemia. N Engl J Med 2006; 354: 166–178.

    Article  CAS  Google Scholar 

  3. Pieters R, Carroll WL . Biology and treatment of acute lymphoblastic leukemia. Hematol Oncol Clin North Am 2010; 24: 1–18.

    Article  Google Scholar 

  4. Pui CH, Campana D, Pei D, Bowman WP, Sandlund JT, Kaste SC et al. Treating childhood acute lymphoblastic leukemia without cranial irradiation. N Engl J Med 2009; 360: 2730–2741.

    Article  CAS  Google Scholar 

  5. Elliott P . Pathogenesis of cardiotoxicity induced by anthracyclines. Semin Oncol 2006; 33 (Suppl 8): S2–S7.

    Article  CAS  Google Scholar 

  6. Lipshultz SE . Exposure to anthracyclines during childhood causes cardiac injury. Semin Oncol 2006; 33 (Suppl 8): S8–S14.

    Article  CAS  Google Scholar 

  7. Hamann PR, Hinman LM, Hollander I, Beyer CF, Lindh D, Holcomb R et al. Gemtuzumab ozogamicin, a potent and selective anti-CD33 antibody-calicheamicin conjugate for treatment of acute myeloid leukemia. Bioconjug Chem 2002; 13: 47–58.

    Article  CAS  Google Scholar 

  8. Sievers EL, Larson RA, Stadtmauer EA, Estey E, Lowenberg B, Dombret H et al. Efficacy and safety of gemtuzumab ozogamicin in patients with CD33-positive acute myeloid leukemia in first relapse. J Clin Oncol 2001; 19: 3244–3254.

    Article  CAS  Google Scholar 

  9. Zein N, Sinha AM, McGahren WJ, Ellestad GA . Calicheamicin gamma 1I: an antitumor antibiotic that cleaves double-stranded DNA site specifically. Science 1988; 240: 1198–1201.

    Article  CAS  Google Scholar 

  10. Zwaan CM, Reinhardt D, Zimmerman M, Hasle H, Stary J, Stark B et al. Salvage treatment for children with refractory first or second relapse of acute myeloid leukaemia with gemtuzumab ozogamicin: results of a phase II study. Br J Haematol 2008; 148: 768–776.

    Article  Google Scholar 

  11. DiJoseph JF, Popplewell A, Tickle S, Ladyman H, Lawson A, Kunz A et al. Antibody-targeted chemotherapy of B-cell lymphoma using calicheamicin conjugated to murine or humanized antibody against CD22. Cancer Immunol Immunother 2005; 54: 11–24.

    Article  CAS  Google Scholar 

  12. DiJoseph JF, Armellino DC, Boghaert ER, Khandke K, Dougher MM, Sridharan L et al. Antibody-targeted chemotherapy with CMC-544: a CD22-targeted immunoconjugate of calicheamicin for the treatment of B-lymphoid malignancies. Blood 2004; 103: 1807–1814.

    Article  CAS  Google Scholar 

  13. Tedder TF, Tuscano J, Sato S, Kehrl JH . CD22, a B lymphocyte-specific adhesion molecule that regulates antigen receptor signaling. Annu Rev Immunol 1997; 15: 481–504.

    Article  CAS  Google Scholar 

  14. Cesano A, Gayko U . CD22 as a target of passive immunotherapy. Semin Oncol 2003; 30: 253–257.

    Article  CAS  Google Scholar 

  15. Dijoseph JF, Dougher MM, Armellino DC, Evans DY, Damle NK . Therapeutic potential of CD22-specific antibody-targeted chemotherapy using inotuzumab ozogamicin (CMC-544) for the treatment of acute lymphoblastic leukemia. Leukemia 2007; 21: 2240–2245.

    Article  CAS  Google Scholar 

  16. DiJoseph JF, Goad ME, Dougher MM, Boghaert ER, Kunz A, Hamann PR et al. Potent and specific antitumor efficacy of CMC-544, a CD22-targeted immunoconjugate of calicheamicin, against systemically disseminated B-cell lymphoma. Clin Cancer Res 2004; 10: 8620–8629.

    Article  CAS  Google Scholar 

  17. Advani A, Coiffier B, Czuczman MS, Dreyling M, Foran J, Gine E et al. Safety, pharmacokinetics, and preliminary clinical activity of inotuzumab ozogamicin, a novel immunoconjugate for the treatment of B-cell non-Hodgkin's lymphoma: results of a phase I study. J Clin Oncol 2010; 28: 2085–2093.

    Article  CAS  Google Scholar 

  18. van der Velden VH, Boeckx N, Jedema I, te Marvelde JG, Hoogeveen PG, Boogaerts M et al. High CD33-antigen loads in peripheral blood limit the efficacy of gemtuzumab ozogamicin (Mylotarg) treatment in acute myeloid leukemia patients. Leukemia 2004; 18: 983–988.

    Article  CAS  Google Scholar 

  19. van Der Velden VH, te Marvelde JG, Hoogeveen PG, Bernstein ID, Houtsmuller AB, Berger MS et al. Targeting of the CD33-calicheamicin immunoconjugate Mylotarg (CMA-676) in acute myeloid leukemia: in vivo and in vitro saturation and internalization by leukemic and normal myeloid cells. Blood 2001; 97: 3197–3204.

    Article  CAS  Google Scholar 

  20. Zwaan CM, Reinhardt D, Jurgens H, Huismans DR, Hahlen K, Smith OP et al. Gemtuzumab ozogamicin in pediatric CD33-positive acute lymphoblastic leukemia: first clinical experiences and relation with cellular sensitivity to single agent calicheamicin. Leukemia 2003; 17: 468–470.

    Article  CAS  Google Scholar 

  21. Gudowius S, Recker K, Laws HJ, Dirksen U, Troger A, Wieczorek U et al. Identification of candidate target antigens for antibody-based immunotherapy in childhood B-cell precursor ALL. Klin Padiatr 2006; 218: 327–333.

    Article  CAS  Google Scholar 

  22. de Vries JF, von dem Borne PA, van Luxemburg-Heijs SA, Heemskerk MH, Willemze R, Falkenburg JH et al. Differential activation of the death receptor pathway in human target cells induced by cytotoxic T lymphocytes showing different kinetics of killing. Haematologica 2007; 92: 1671–1678.

    Article  CAS  Google Scholar 

  23. de Vries JF, Falkenburg JH, Willemze R, Barge RM . The mechanisms of Ara-C-induced apoptosis of resting B-chronic lymphocytic leukemia cells. Haematologica 2006; 91: 912–919.

    CAS  PubMed  Google Scholar 

  24. Den Boer ML, Harms DO, Pieters R, Kazemier KM, Gobel U, Korholz D et al. Patient stratification based on prednisolone-vincristine-asparaginase resistance profiles in children with acute lymphoblastic leukemia. J Clin Oncol 2003; 21: 3262–3268.

    Article  CAS  Google Scholar 

  25. Rogakou EP, Pilch DR, Orr AH, Ivanova VS, Bonner WM . DNA double-stranded breaks induce histone H2AX phosphorylation on serine 139. J Biol Chem 1998; 273: 5858–5868.

    Article  CAS  Google Scholar 

  26. van der Burg M, van Veelen LR, Verkaik NS, Wiegant WW, Hartwig NG, Barendregt BH et al. A new type of radiosensitive T-B-NK+ severe combined immunodeficiency caused by a LIG4 mutation. J Clin Invest 2006; 116: 137–145.

    Article  CAS  Google Scholar 

  27. Freundt EC, Czapiga M, Lenardo MJ . Photoconversion of Lysotracker Red to a green fluorescent molecule. Cell Research 2007; 17: 956–958.

    Article  CAS  Google Scholar 

  28. Ueda K, Cardarelli C, Gottesman MM, Pastan I . Expression of a full-length cDNA for the human ‘MDR1’ gene confers resistance to colchicine, doxorubicin, and vinblastine. Proc Natl Acad Sci U S A 1987; 84: 3004–3008.

    Article  CAS  Google Scholar 

  29. Zaman GJ, Versantvoort CH, Smit JJ, Eijdems EW, de Haas M, Smith AJ et al. Analysis of the expression of MRP, the gene for a new putative transmembrane drug transporter, in human multidrug resistant lung cancer cell lines. Cancer Research 1993; 53: 1747–1750.

    CAS  PubMed  Google Scholar 

  30. Walter RB, Gooley TA, van der Velden VH, Loken MR, van Dongen JJ, Flowers DA et al. CD33 expression and P-glycoprotein-mediated drug efflux inversely correlate and predict clinical outcome in patients with acute myeloid leukemia treated with gemtuzumab ozogamicin monotherapy. Blood 2007; 109: 4168–4170.

    Article  CAS  Google Scholar 

  31. Takeshita A, Shinjo K, Yamakage N, Ono T, Hirano I, Matsui H et al. CMC-544 (inotuzumab ozogamicin) shows less effect on multidrug resistant cells: analyses in cell lines and cells from patients with B-cell chronic lymphocytic leukaemia and lymphoma. Br J Haematol 2009; 146: 34–43.

    Article  CAS  Google Scholar 

  32. Swerts K, De Moerloose B, Dhooge C, Laureys G, Benoit Y, Philippe J . Prognostic significance of multidrug resistance-related proteins in childhood acute lymphoblastic leukaemia. Eur J Cancer 2006; 42: 295–309.

    Article  CAS  Google Scholar 

  33. Den Boer ML, Pieters R, Kazemier KM, Janka-Schaub GE, Henze G, Veerman AJ . Relationship between the intracellular daunorubicin concentration, expression of major vault protein/lung resistance protein and resistance to anthracyclines in childhood acute lymphoblastic leukemia. Leukemia 1999; 13: 2023–2030.

    Article  CAS  Google Scholar 

  34. den Boer ML, Pieters R, Kazemier KM, Rottier MM, Zwaan CM, Kaspers GJ et al. Relationship between major vault protein/lung resistance protein, multidrug resistance-associated protein, P-glycoprotein expression, and drug resistance in childhood leukemia. Blood 1998; 91: 2092–2098.

    CAS  PubMed  Google Scholar 

  35. Jedema I, Barge RM, van der Velden VH, Nijmeijer BA, van Dongen JJ, Willemze R et al. Internalization and cell cycle-dependent killing of leukemic cells by Gemtuzumab Ozogamicin: rationale for efficacy in CD33-negative malignancies with endocytic capacity. Leukemia 2004; 18: 316–325.

    Article  CAS  Google Scholar 

  36. Balaian L, Ball ED . Cytotoxic activity of gemtuzumab ozogamicin (Mylotarg) in acute myeloid leukemia correlates with the expression of protein kinase Syk. Leukemia 2006; 20: 2093–2101.

    Article  CAS  Google Scholar 

  37. Beers SA, French RR, Chan HT, Lim SH, Jarrett TC, Vidal RM et al. Antigenic modulation limits the efficacy of anti-CD20 antibodies: implications for antibody selection. Blood 2010; 115: 5191–5201.

    Article  CAS  Google Scholar 

  38. Walter RB, Raden BW, Kamikura DM, Cooper JA, Bernstein ID . Influence of CD33 expression levels and ITIM-dependent internalization on gemtuzumab ozogamicin-induced cytotoxicity. Blood 2005; 105: 1295–1302.

    Article  CAS  Google Scholar 

  39. Goemans BF, Zwaan CM, Vijverberg SJ, Loonen AH, Creutzig U, Hahlen K et al. Large interindividual differences in cellular sensitivity to calicheamicin may influence gemtuzumab ozogamicin response in acute myeloid leukemia. Leukemia 2008; 22: 2284–2285.

    Article  CAS  Google Scholar 

  40. ten Cate B, Samplonius DF, Bijma T, de Leij LF, Helfrich W, Bremer E . The histone deacetylase inhibitor valproic acid potently augments gemtuzumab ozogamicin-induced apoptosis in acute myeloid leukemic cells. Leukemia 2007; 21: 248–252.

    Article  CAS  Google Scholar 

  41. Brethon B, Yakouben K, Oudot C, Boutard P, Bruno B, Jerome C et al. Efficacy of fractionated gemtuzumab ozogamicin combined with cytarabine in advanced childhood myeloid leukaemia. Br J Haematol 2008; 143: 541–547.

    CAS  PubMed  Google Scholar 

Download references

Acknowledgements

The authors want to acknowledge Dr Patrick Kelly and Dr Erik Vandendries (both employees of Pfizer) for their comments and discussions, and Pfizer (legacy Wyeth) for providing us with CMC-544, calicheamicin and CD22 antibody (G5/44). The authors would like to thank Dr Adriaan Houtsmuller and Nicole Larmonie for their assistance with the confocal microscopy, and Mathilde Broekhuis for her support with the MTT assays.

Author contributions

JFdV designed and performed research, analyzed and interpreted data, and wrote the manuscript; CMZ designed research and critically reviewed the manuscript; MdB and JSAV performed research and analyzed data; MLdB provided patient samples and analytical tools, and critically reviewed the manuscript; JJMvD and VHJvdV designed research, interpreted data and wrote the manuscript.

Author information

Authors and Affiliations

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

    J F de Vries, M De Bie, J S A Voerman, J J M van Dongen & V H J van der Velden

  2. Department of Pediatric Oncology/Hematology, Erasmus MC, Sophia Children's Hospital, Rotterdam, The Netherlands

    C M Zwaan & M L den Boer

Authors
  1. J F de Vries
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. C M Zwaan
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. M De Bie
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. J S A Voerman
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. M L den Boer
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. J J M van Dongen
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. V H J van der Velden
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to V H J van der Velden.

Ethics declarations

Competing interests

The authors declare no conflict of interest.

Additional information

Supplementary Information accompanies the paper on the Leukemia website

Supplementary information

Rights and permissions

Reprints and permissions

About this article

Cite this article

de Vries, J., Zwaan, C., De Bie, M. et al. The novel calicheamicin-conjugated CD22 antibody inotuzumab ozogamicin (CMC-544) effectively kills primary pediatric acute lymphoblastic leukemia cells. Leukemia 26, 255–264 (2012). https://doi.org/10.1038/leu.2011.206

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/leu.2011.206

Keywords

This article is cited by

Search

Advanced search

Quick links