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Myelodysplastic syndrome

Mutation-specific signaling profiles and kinase inhibitor sensitivities of juvenile myelomonocytic leukemia revealed by induced pluripotent stem cells

Leukemia (2018) | Download Citation

Abstract

Juvenile myelomonocytic leukemia (JMML) is an uncommon myeloproliferative neoplasm driven by Ras pathway mutations and hyperactive Ras/MAPK signaling. Outcomes for many children with JMML remain dismal with current standard-of-care cytoreductive chemotherapy and hematopoietic stem cell transplantation. We used patient-derived induced pluripotent stem cells (iPSCs) to characterize the signaling profiles and potential therapeutic vulnerabilities of PTPN11-mutant and CBL-mutant JMML. We assessed whether MEK, JAK, and PI3K/mTOR kinase inhibitors (i) could inhibit myeloproliferation and aberrant signaling in iPSC-derived hematopoietic progenitors with PTPN11 E76K or CBL Y371H mutations. We detected constitutive Ras/MAPK and PI3K/mTOR signaling in PTPN11 and CBL iPSC-derived myeloid cells. Activated signaling and growth of PTPN11 iPSCs were preferentially inhibited in vitro by the MEKi PD0325901 and trametinib. Conversely, JAK/STAT signaling was selectively activated in CBL iPSCs and abrogated by the JAKi momelotinib and ruxolitinib. The PI3Kδi idelalisib and mTORi rapamycin inhibited signaling and myeloproliferation in both PTPN11 and CBL iPSCs. These findings demonstrate differential sensitivity of PTPN11 iPSCs to MEKi and of CBL iPSCs to JAKi, but similar sensitivity to PI3Ki and mTORi. Clinical investigation of mutation-specific kinase inhibitor therapies in children with JMML may be warranted.

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References

  1. 1.

    Locatelli F, Niemeyer CM. How I treat juvenile myelomonocytic leukemia. Blood. 2015;125:1083–90.

  2. 2.

    Chang TY, Dvorak CC, Loh ML. Bedside to bench in juvenile myelomonocytic leukemia: insights into leukemogenesis from a rare pediatric leukemia. Blood. 2014;124:2487–97.

  3. 3.

    Loh ML. Recent advances in the pathogenesis and treatment of juvenile myelomonocytic leukaemia. Br J Haematol. 2011;152:677–87.

  4. 4.

    Emanuel PD, Bates LJ, Castleberry RP, Gualtieri RJ, Zuckerman KS. Selective hypersensitivity to granulocyte-macrophage colony-stimulating factor by juvenile chronic myeloid leukemia hematopoietic progenitors. Blood. 1991;77:925–9.

  5. 5.

    Woods WG, Barnard DR, Alonzo TA, Buckley JD, Kobrinsky N, Arthur DC, et al. Prospective study of 90 children requiring treatment for juvenile myelomonocytic leukemia or myelodysplastic syndrome: a report from the Children’s Cancer Group. J Clin Oncol. 2002;20:434–40.

  6. 6.

    Locatelli F, Nollke P, Zecca M, Korthof E, Lanino E, Peters C, et al. Hematopoietic stem cell transplantation (HSCT) in children with juvenile myelomonocytic leukemia (JMML): results of the EWOG-MDS/EBMT trial. Blood. 2005;105:410–9.

  7. 7.

    Yabe M, Ohtsuka Y, Watanabe K, Inagaki J, Yoshida N, Sakashita K, et al. Transplantation for juvenile myelomonocytic leukemia: a retrospective study of 30 children treated with a regimen of busulfan, fludarabine, and melphalan. Int J Hematol. 2015;101:184–90.

  8. 8.

    Niemeyer CM, Arico M, Basso G, Biondi A, Cantu Rajnoldi A, Creutzig U, et al. Chronic myelomonocytic leukemia in childhood: a retrospective analysis of 110 cases. European Working Group on Myelodysplastic Syndromes in Childhood (EWOG-MDS). Blood. 1997;89:3534–43.

  9. 9.

    Flotho C, Kratz CP, Bergstrasser E, Hasle H, Stary J, Trebo M, et al. Genotype-phenotype correlation in cases of juvenile myelomonocytic leukemia with clonal RAS mutations. Blood. 2008;111:966–7. Author reply 967-8

  10. 10.

    Matsuda K, Shimada A, Yoshida N, Ogawa A, Watanabe A, Yajima S, et al. Spontaneous improvement of hematologic abnormalities in patients having juvenile myelomonocytic leukemia with specific RAS mutations. Blood. 2007;109:5477–80.

  11. 11.

    Kotecha N, Flores NJ, Irish JM, Simonds EF, Sakai DS, Archambeault S, et al. Single-cell profiling identifies aberrant STAT5 activation in myeloid malignancies with specific clinical and biologic correlates. Cancer Cell. 2008;14:335–43.

  12. 12.

    Chang T, Krisman K, Theobald EH, Xu J, Akutagawa J, Lauchle JO, et al. Sustained MEK inhibition abrogates myeloproliferative disease in Nf1 mutant mice. J Clin Invest. 2013;123:335–9.

  13. 13.

    Lyubynska N, Gorman MF, Lauchle JO, Hong WX, Akutagawa JK, Shannon K, et al. A MEK inhibitor abrogates myeloproliferative disease in Kras mutant mice. Sci Transl Med. 2011;3:76ra27.

  14. 14.

    Lauchle JO, Kim D, Le DT, Akagi K, Crone M, Krisman K, et al. Response and resistance to MEK inhibition in leukaemias initiated by hyperactive Ras. Nature. 2009;461:411–4.

  15. 15.

    Akutagawa J, Huang TQ, Epstein I, Chang T, Quirindongo-Crespo M, Cottonham CL, et al. Targeting the PI3K/Akt pathway in murine MDS/MPN driven by hyperactive Ras. Leukemia. 2016;30:1335–43.

  16. 16.

    Goodwin CB, Li XJ, Mali RS, Chan G, Kang M, Liu Z, et al. PI3K p110delta uniquely promotes gain-of-function Shp2-induced GM-CSF hypersensitivity in a model of JMML. Blood. 2014;123:2838–42.

  17. 17.

    Bunda S, Qin K, Kommaraju K, Heir P, Ohh M. Juvenile myelomonocytic leukaemia-associated mutation in Cbl promotes resistance to apoptosis via the Lyn-PI3K/AKT pathway. Oncogene. 2015;34:789–97.

  18. 18.

    Gandre-Babbe S, Paluru P, Aribeana C, Chou ST, Bresolin S, Lu L, et al. Patient-derived induced pluripotent stem cells recapitulate hematopoietic abnormalities of juvenile myelomonocytic leukemia. Blood. 2013;121:4925–9.

  19. 19.

    Mulero-Navarro S, Sevilla A, Roman AC, Lee DF, D’Souza SL, Pardo S, et al. Myeloid dysregulation in a human induced pluripotent stem cell model of PTPN11-associated juvenile myelomonocytic leukemia. Cell Rep. 2015;13:504–15.

  20. 20.

    Arber DA, Orazi A, Hasserjian R, Thiele J, Borowitz MJ, Le Beau MM, et al. The2016 revision to the World Health Organization classification of myeloid neoplasms and acute leukemia. Blood. 2016;127:2391–405.

  21. 21.

    Archambeault S, Flores NJ, Yoshimi A, Kratz CP, Reising M, Fischer A, et al. Development of an allele-specific minimal residual disease assay for patients with juvenile myelomonocytic leukemia. Blood. 2008;111:1124–7.

  22. 22.

    Loh ML, Sakai DS, Flotho C, Kang M, Fliegauf M, Archambeault S, et al. Mutations in CBL occur frequently in juvenile myelomonocytic leukemia. Blood. 2009;114:1859–63.

  23. 23.

    Stadtfeld M, Hochedlinger K. Induced pluripotency: history, mechanisms, and applications. Genes Dev. 2010;24:2239–63.

  24. 24.

    Mills JA, Paluru P, Weiss MJ, Gadue P, French DL. Hematopoietic differentiation of pluripotent stem cells in culture. Methods Mol Biol. 2014;1185:181–94.

  25. 25.

    Byrska-Bishop M, VanDorn D, Campbell AE, Betensky M, Arca PR, Yao Y, et al. Pluripotent stem cells reveal erythroid-specific activities of the GATA1 N-terminus. J Clin Invest. 2015;125:993–1005.

  26. 26.

    Chou ST, Byrska-Bishop M, Tober JM, Yao Y, Vandorn D, Opalinska JB, et al. Trisomy 21-associated defects in human primitive hematopoiesis revealed through induced pluripotent stem cells. Proc Natl Acad Sci USA. 2012;109:17573–8.

  27. 27.

    Tasian SK, Doral MY, Borowitz MJ, Wood BL, Chen IM, Harvey RC, et al. Aberrant STAT5 and PI3K/mTOR pathway signaling occurs in human CRLF2-rearranged B-precursor acute lymphoblastic leukemia. Blood. 2012;120:833–42.

  28. 28.

    Tasian SK, Teachey DT, Li Y, Shen F, Harvey RC, Chen IM, et al. Potent efficacy of combined PI3K/mTOR and JAK or ABL inhibition in murine xenograft models of Ph-like acute lymphoblastic leukemia. Blood. 2017;129:177–87.

  29. 29.

    Kotecha N, Krutzik PO, Irish JM. Web-based analysis and publication of flow cytometry experiments. Curr Protoc Cytom. 2010; Chapter 10: Unit10 17. PMID 20578106https://www.ncbi.nlm.nih.gov/pubmed/?term=20578106A

  30. 30.

    Yoshida N, Yagasaki H, Xu Y, Matsuda K, Yoshimi A, Takahashi Y, et al. Correlation of clinical features with the mutational status of GM-CSF signaling pathway-related genes in juvenile myelomonocytic leukemia. Pediatr Res. 2009;65:334–40.

  31. 31.

    Niemeyer CM, Kang MW, Shin DH, Furlan I, Erlacher M, Bunin NJ, et al. Germline CBL mutations cause developmental abnormalities and predispose to juvenile myelomonocytic leukemia. Nat Genet. 2010;42:794–800.

  32. 32.

    Hyakuna N, Muramatsu H, Higa T, Chinen Y, Wang X, Kojima S. Germline mutation of CBL is associated with moyamoya disease in a child with juvenile myelomonocytic leukemia and Noonan syndrome-like disorder. Pediatr Blood Cancer. 2015;62:542–4.

  33. 33.

    Caye A, Strullu M, Guidez F, Cassinat B, Gazal S, Fenneteau O, et al. Juvenile myelomonocytic leukemia displays mutations in components of the RAS pathway and the PRC2 network. Nat Genet. 2015;47:1334–40.

  34. 34.

    Stieglitz E, Troup CB, Gelston LC, Haliburton J, Chow ED, Yu KB, et al. Subclonal mutations in SETBP1 confer a poor prognosis in juvenile myelomonocytic leukemia. Blood. 2015;125:516–24.

  35. 35.

    Jessen WJ, Miller SJ, Jousma E, Wu J, Rizvi TA, Brundage ME, et al. MEK inhibition exhibits efficacy in human and mouse neurofibromatosis tumors. J Clin Invest. 2013;123:340–7.

  36. 36.

    Liu W, Yu WM, Zhang J, Chan RJ, Loh ML, Zhang Z, et al. Inhibition of the Gab2/PI3K/mTOR signaling ameliorates myeloid malignancy caused by Ptpn11 (Shp2) gain-of-function mutations. Leukemia. 2017;31:1415–22.

  37. 37.

    Tasian SK, Teachey DT, Rheingold SR. Targeting the PI3K/mTOR pathway in pediatric hematologic malignancies. Front Oncol. 2014;4:108.

  38. 38.

    Teachey DT, Greiner R, Seif A, Attiyeh E, Bleesing J, Choi J, et al. Treatment with sirolimus results in complete responses in patients with autoimmune lymphoproliferative syndrome. Br J Haematol. 2009;145:101–6.

  39. 39.

    Bride KL, Vincent T, Smith-Whitley K, Lambert MP, Bleesing JJ, Seif AE, et al. Sirolimus is effective in relapsed/refractory autoimmune cytopenias: results of a prospective multi-institutional trial. Blood. 2016;127:17–28.

  40. 40.

    Verstovsek S, Mesa RA, Gotlib J, Levy RS, Gupta V, DiPersio JF, et al. A double-blind, placebo-controlled trial of ruxolitinib for myelofibrosis. N Engl J Med. 2012;366:799–807.

  41. 41.

    Vannucchi AM, Kiladjian JJ, Griesshammer M, Masszi T, Durrant S, Passamonti F, et al. Ruxolitinib versus standard therapy for the treatment of polycythemia vera. N Engl J Med. 2015;372:426–35.

  42. 42.

    Padron E, Dezern A, Andrade-Campos M, Vaddi K, Scherle P, Zhang Q, et al. A multi-institution phase I trial of ruxolitinib in patients with chronic myelomonocytic leukemia (CMML). Clin Cancer Res. 2016;22:3746–54.

  43. 43.

    Sachs Z, Been RA, DeCoursin KJ, Nguyen HT, Mohd Hassan NA, Noble-Orcutt KE, et al. Stat5 is critical for the development and maintenance of myeloproliferative neoplasm initiated by Nf1 deficiency. Haematologica. 2016;101:1190–9.

  44. 44.

    Lv K, Jiang J, Donaghy R, Riling CR, Cheng Y, Chandra V, et al. CBL family E3 ubiquitin ligases control JAK2 ubiquitination and stability in hematopoietic stem cells and myeloid malignancies. Genes Dev. 2017;31:1007–23.

  45. 45.

    Loh ML, Tasian SK, Rabin KR, Brown P, Magoon D, Reid JM, et al. A phase 1 dosing study of ruxolitinib in children with relapsed or refractory solid tumors, leukemias, or myeloproliferative neoplasms: a Children’s Oncology Group phase 1 consortium study (ADVL1011). Pediatr Blood Cancer. 2015;62:1717–24.

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Acknowledgements

We thank the human pluripotent stem cell core at the Children’s Hospital of Philadelphia for technical assistance. This work was supported by the National Institutes of Health/National Cancer Institute K08CA184418 (SKT), National Institute of Diabetes and Digestive and Kidney Diseases 5P30DK090969 (MJW) and R01DK100854 (STC), National Heart Lung Blood Institute T32HL0007150 (JAC) and 5RC2HL101606 (MJW), the Leukemia Lymphoma Society 6466-15 (MLL), and Hyundai Hope on Wheels (MLL). MLL is the Benioff Chair of Children’s Health at the University of California, San Francisco and the Deborah and Arthur Ablin Endowed Chair of Pediatric Molecular Oncology. MJW is the Arthur Nienhuis Endowed Chair in Hematology at St Jude Children’s Research Hospital.

Author contributions

SKT designed and directed the study, performed experiments, analyzed data, and wrote the manuscript. JAC designed the study, performed experiments, and analyzed data. SG-B, DP, ALG, and GL performed experiments and analyzed data. MLL provided critical clinical specimens. MLW contributed to study design and oversight. STC and DLF designed and directed the study, analyzed data, and edited the manuscript. All authors reviewed the final version of the manuscript.

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Author notes

    • Jessica A. Casas

    Present address: Department of Pediatrics, Division of Hematology and Oncology, Baylor College of Medicine and Texas Children’s Hospital, Houston, TX, USA

  1. These authors contributed equally: Sarah K. Tasian, Jessica A. Casas.

  2. These authors contributed equally: Deborah L. French, Stella T. Chou.

Affiliations

  1. Division of Oncology and Center for Childhood Cancer Research, Children’s Hospital of Philadelphia, Philadelphia, PA, USA

    • Sarah K. Tasian
  2. Department of Pediatrics, University of Pennsylvania Perelman School of Medicine, Philadelphia, PA, USA

    • Sarah K. Tasian
    •  & Stella T. Chou
  3. Division of Hematology, Children’s Hospital of Philadelphia, Philadelphia, PA, USA

    • Jessica A. Casas
    • , David Posocco
    • , Shilpa Gandre-Babbe
    •  & Stella T. Chou
  4. Center for Cellular and Molecular Therapeutics, Children’s Hospital of Philadelphia, Philadelphia, PA, USA

    • Alyssa L. Gagne
    • , Ge Liang
    •  & Deborah L. French
  5. Division of Hematology-Oncology, San Francisco Benioff Children’s Hospital and School of Medicine, Pediatrics, University of California, San Francisco, CA, USA

    • Mignon L. Loh
  6. Department of Hematology, St Jude Children’s Research Hospital, Memphis, TN, USA

    • Mitchell J. Weiss
  7. Department of Pathology and Laboratory Medicine, University of Pennsylvania Perelman School of Medicine, Philadelphia, PA, USA

    • Deborah L. French

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Conflict of interest

SKT receives research funding from Gilead Sciences for unrelated studies. SKT and MLL receive research funding from the Incyte Corporation for unrelated studies.

Corresponding author

Correspondence to Sarah K. Tasian.

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DOI

https://doi.org/10.1038/s41375-018-0169-y