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Successful xenografts of AML3 samples in immunodeficient NOD/shi-SCID IL2Rγ−/− mice

Leukemia volume 26, pages 2432–2435 (2012)Cite this article

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Xenotransplantation of human acute myeloid leukemia (AML) cells into immunodeficient mice has allowed to identify leukemia-initiating cells (LIC).1 Recent studies of phenotypic and genetic tracking of LIC subclones in leukemia patients underscored the use of xenografts to study AML patient’s clonal LIC heterogeneity and/or clonal dominance and thus address issues such as patient relapses.2 Unfortunately, identification of LIC in all AML patients and in all AML subtypes has not been achieved. Lack of successful engraftment has been attributed to clinical and biological prognostic features, antibody-mediated immune clearance and the rarity of LIC in AML.3, 4, 5 AML3 leukemia has always featured among the good prognostic AML subtypes, and we and others have reported their lack or weak engraftment in NOD-SCID mice.1, 5, 6, 7 Improved cell separation/preparation techniques and increased immunodeficiency of mouse models were shown to increase engraftment efficiency of cancer-initiating cells.8, 9 AML3 is a rare leukemia and has thus not yet been reported in this new improved setting. We therefore reinvestigated the issue of LIC in the AML3 subtype using NOD/shi-SCID IL2Rγ−/− (NOG) mice.

Out of these eight AML3 samples, two samples (Pat. 1 and Pat. 2) successfully engrafted (Figures 1b and c, Supplementary Figure S1A and Table 1), with hCD45+CD33+CD19− levels reaching more than 50% at 12 weeks and 20 weeks, respectively, along with immunophenotypic features (CD34, CD38, CD33, CD44 and CXCR4), cytological and molecular (PML-RARA positivity by real-time quantitative PCR and interphase fluorescence in situ hybridization analysis) features similar to the AML3 patients’ leukemic cell characteristics at diagnosis (Table 1, Figure 1c and Supplementary Figure S1A and Supplementary Table S1). Two other samples (Pat. 3 and Pat. 4) were also assessed as positive leukemic engrafters, based on the phenotype criteria, hCD45+CD33+CD19−.10 Unfortunately, despite hCD45+ cells bearing cytology and immunophenotype characteristics similar to those of the AML3 cells’ characteristics at diagnosis, AML3 could not be confirmed in these two xenografts by molecular analysis (PML-RAR detection). Xenografts of Pat. 3 samples died before analysis, and hCD45+ purified from BM and spleen cells of xenografts of Pat. 4 samples yielded too low numbers for successful molecular assays (Table 1). Furthermore, engraftment kinetics in these two xenografts were poor (Figure 1b). In three samples (Pat. 6, 7 and 8), only normal human hematopoietic engraftment (CD45+CD19+ cells >75% and CD45+CD33+ cells >20%) was noted, and one AML3 patient sample (Pat. 5) did not engraft after 16 weeks (Table 1). Thus, in the NOG setting, at least two AML3 samples were confirmed of having successfully engrafted, and allowed to further pursue the experiments to validate the AML3 leukemic subtype (that is, sensitivity to all-trans-retinoic acid (ATRA)) and the existence of AML3 LIC in these patients’ samples.

Table 1 Xenograft samples after first transplantation and serial transplantations
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Acknowledgements

This work was supported by grants from Cancéropôle Ile de France, Association Recherche contre le Cancer, INSERM and Université Paris Diderot, Université Paris XI, Association Laurette Fugain, Association Cent pour Cent la vie and Cancéropole, the Fondation Institut Gustave Roussy, Fondation de France. Satyananda Patel was a recipient of a postdoctoral fellowship from Cancéropôle Ile de France. We thank members of the Institut Gustave Roussy, specifically the animal facility and the flow cytometry facility, and Elisabeth Savariau of the photography laboratory of the Institute of Hematology. We are also grateful to M Ito (Kawasaki, Japan) for the NOG mice.

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Author notes

  1. S Patel, Y Zhang, C Chomienne and F Louache: These authors contributed equally to this work.

  2. These authors are co-senior authors.

Authors and Affiliations

  1. Institut Gustave Roussy, IFR54, Villejuif, France

    S Patel, Y Zhang & F Louache

  2. Institut National de la Santé et de la Recherche Médicale (INSERM), U1009, Villejuif, France

    S Patel, Y Zhang & F Louache

  3. Université Paris Sud, Villejuif, France

    S Patel, Y Zhang & F Louache

  4. INSERM U940, Hôpital Saint Louis, Paris, France

    S Patel, B Cassinat, F Zassadowski, N Ferré, D Bonnet & C Chomienne

  5. Département d’Hématologie, Université Paris-Diderot, Sorbonne Paris Cité, IUH, UMRS940, Paris, France

    B Cassinat, F Zassadowski, N Ferré, D Bonnet & C Chomienne

  6. AP-HP, Hôpital Saint-Louis, Unité de Biologie Cellulaire, Paris, France

    B Cassinat, F Zassadowski & C Chomienne

  7. AP-HP, Hôpital Saint Louis, Laboratoire d'Hématologie, Paris, France

    W Cuccuini & J M Cayuela

  8. AP-HP, Hôpital Avicenne, Service d’Hématologie Clinique, Bobigny, France

    P Fenaux

  9. Haematopoietic Stem Cell Laboratory, London Research Institute, London, UK

    D Bonnet

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Correspondence to C Chomienne or F Louache.

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Patel, S., Zhang, Y., Cassinat, B. et al. Successful xenografts of AML3 samples in immunodeficient NOD/shi-SCID IL2Rγ−/− mice. Leukemia 26, 2432–2435 (2012). https://doi.org/10.1038/leu.2012.154

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