Hedgehog/GLI1 activation leads to leukemic transformation of myelodysplastic syndrome in vivo and GLI1 inhibition results in antitumor activity

Abstract

Myelodysplastic syndromes (MDSs) are stem cell disorders with risk of transformation to acute myeloid leukemia (AML). Gene expression profiling reveals transcriptional expression of GLI1, of Hedgehog (Hh) signaling, in poor-risk MDS/AML. Using a murine model of MDS we demonstrated that constitutive Hh/Gli1 activation accelerated leukemic transformation and decreased overall survival. Hh/Gli1 activation resulted in clonal expansion of phenotypically defined granulocyte macrophage progenitors (GMPs) and acquisition of self-renewal potential in a non-self-renewing progenitor compartment. Transcriptome analysis of GMPs revealed enrichment in gene signatures of self-renewal pathways, operating via direct Gli1 activation. Using human cell lines we demonstrated that in addition to canonical Hh signaling, GLI1 is activated in a Smoothened-independent manner. GLI1 knockdown or inhibition with GANT61 resulted in decreased proliferation and clonogenic potential. Our data suggest that GLI1 activation is frequent in MDS during disease progression and inhibition of GLI1 is an attractive therapeutic target for a subset of patients.

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References

  1. 1.

    Tefferi A, Vardiman JW. Myelodysplastic syndromes. N Engl J Med. 2009;361:1872–85.

    CAS  Article  Google Scholar 

  2. 2.

    Garcia-Manero G. Myelodysplastic syndromes: 2015 Update on diagnosis, risk-stratification and management. Am J Hematol. 2015;90:831–41.

    Article  Google Scholar 

  3. 3.

    Greenberg P, Cox C, LeBeau MM, Fenaux P, Morel P, Sanz G, et al. International scoring system for evaluating prognosis in myelodysplastic syndromes. Blood. 1997;89:2079–88.

    CAS  PubMed  PubMed Central  Google Scholar 

  4. 4.

    Malcovati L, Germing U, Kuendgen A, Della Porta MG, Pascutto C, Invernizzi R, et al. Time-dependent prognostic scoring system for predicting survival and leukemic evolution in myelodysplastic syndromes. J Clin Oncol. 2007;25:3503–10.

    Article  Google Scholar 

  5. 5.

    Meggendorfer M, de Albuquerque A, Nadarajah N, Alpermann T, Kern W, Steuer K, et al. Karyotype evolution and acquisition of FLT3 or RAS pathway alterations drive progression of myelodysplastic syndrome to acute myeloid leukemia. Haematologica. 2015;100:e487–90.

    Article  Google Scholar 

  6. 6.

    Makishima H, Yoshizato T, Yoshida K, Sekeres MA, Radivoyevitch T, Suzuki H, et al. Dynamics of clonal evolution in myelodysplastic syndromes. Nat Genet. 2017;49:204–12.

    CAS  Article  Google Scholar 

  7. 7.

    Jiang Y, Dunbar A, Gondek LP, Mohan S, Rataul M, O’Keefe C, et al. Aberrant DNA methylation is a dominant mechanism in MDS progression to AML. Blood. 2009;113:1315–25.

    CAS  Article  Google Scholar 

  8. 8.

    Fenaux P, Giagounidis A, Selleslag D, Beyne-Rauzy O, Mufti G, Mittelman M, et al. A randomized phase 3 study of lenalidomide versus placebo in RBC transfusion-dependent patients with low-/intermediate-1-risk myelodysplastic syndromes with del5q. Blood. 2011;118:3765–76.

    CAS  Article  Google Scholar 

  9. 9.

    Krug U, Rollig C, Koschmieder A, Heinecke A, Sauerland MC, Schaich M, et al. Complete remission and early death after intensive chemotherapy in patients aged 60 years or older with acute myeloid leukaemia: a web-based application for prediction of outcomes. Lancet. 2010;376:2000–8.

    CAS  Article  Google Scholar 

  10. 10.

    Walter RB, Othus M, Borthakur G, Ravandi F, Cortes JE, Pierce SA, et al. Prediction of early death after induction therapy for newly diagnosed acute myeloid leukemia with pretreatment risk scores: a novel paradigm for treatment assignment. J Clin Oncol. 2011;29:4417–23.

    Article  Google Scholar 

  11. 11.

    McMillan R, Matsui W. Molecular pathways: the Hedgehog signaling pathway in cancer. Clin Cancer Res. 2012;18:4883–8.

    CAS  Article  Google Scholar 

  12. 12.

    Denef N, Neubuser D, Perez L, Cohen SM. Hedgehog induces opposite changes in turnover and subcellular localization of patched and smoothened. Cell. 2000;102:521–31.

    CAS  Article  Google Scholar 

  13. 13.

    Ruiz i, Altaba A. Catching a Gli-mpse of Hedgehog. Cell. 1997;90:193–6.

    Article  Google Scholar 

  14. 14.

    Borycki A, Brown AM, Emerson CP Jr. Shh and Wnt signaling pathways converge to control Gli gene activation in avian somites. Development. 2000;127:2075–87.

    CAS  PubMed  Google Scholar 

  15. 15.

    Ringuette R, Atkins M, Lagali PS, Bassett EA, Campbell C, Mazerolle C, et al. A Notch-Gli2 axis sustains Hedgehog responsiveness of neural progenitors and Muller glia. Dev Biol. 2016;411:85–100.

    CAS  Article  Google Scholar 

  16. 16.

    Liu Z, Li T, Reinhold MI, Naski MC. MEK1-RSK2 contributes to Hedgehog signaling by stabilizing GLI2 transcription factor and inhibiting ubiquitination. Oncogene. 2014;33:65–73.

    Article  Google Scholar 

  17. 17.

    Johnson RW, Nguyen MP, Padalecki SS, Grubbs BG, Merkel AR, Oyajobi BO, et al. TGF-beta promotion of Gli2-induced expression of parathyroid hormone-related protein, an important osteolytic factor in bone metastasis, is independent of canonical Hedgehog signaling. Cancer Res. 2011;71:822–31.

    CAS  Article  Google Scholar 

  18. 18.

    Lim Y, Gondek L, Li L, Wang Q, Ma H, Chang E, et al. Integration of Hedgehog and mutant FLT3 signaling in myeloid leukemia. Sci Transl Med. 2015;7:291ra296.

    Article  Google Scholar 

  19. 19.

    Dagklis A, Demeyer S, De Bie J, Radaelli E, Pauwels D, Degryse S, et al. Hedgehog pathway activation in T-cell acute lymphoblastic leukemia predicts response to SMO and GLI1 inhibitors. Blood. 2016;128:2642–54.

    CAS  Article  Google Scholar 

  20. 20.

    Hahn H, Wicking C, Zaphiropoulous PG, Gailani MR, Shanley S, Chidambaram A, et al. Mutations of the human homolog of Drosophila patched in the nevoid basal cell carcinoma syndrome. Cell. 1996;85:841–51.

    CAS  Article  Google Scholar 

  21. 21.

    Johnson RL, Rothman AL, Xie J, Goodrich LV, Bare JW, Bonifas JM, et al. Human homolog of patched, a candidate gene for the basal cell nevus syndrome. Science. 1996;272:1668–71.

    CAS  Article  Google Scholar 

  22. 22.

    Xavier-Ferrucio JM, Pericole FV, Lopes MR, Latuf-Filho P, Barcellos KS, Dias AI, et al. Abnormal Hedgehog pathway in myelodysplastic syndrome and its impact on patients’ outcome. Haematologica. 2015;100:e491–3.

    CAS  Article  Google Scholar 

  23. 23.

    Martinelli G, Oehler VG, Papayannidis C, Courtney R, Shaik MN, Zhang X, et al. Treatment with PF-04449913, an oral smoothened antagonist, in patients with myeloid malignancies: a phase 1 safety and pharmacokinetics study. Lancet Haematol. 2015;2:e339–46.

    Article  Google Scholar 

  24. 24.

    Lancet JE, Komrokji RS, Sweet KL, Duong VH, McGraw KL, Zhang L, et al. Phase 2 trial of smoothened (SMO) inhibitor PF-04449913 (PF-04) in refractory myelodysplastic syndromes (MDS). Blood. 2016;128:3174.

    Google Scholar 

  25. 25.

    Lin YW, Slape C, Zhang Z, Aplan PD. NUP98-HOXD13 transgenic mice develop a highly penetrant, severe myelodysplastic syndrome that progresses to acute leukemia. Blood. 2005;106:287–95.

    CAS  Article  Google Scholar 

  26. 26.

    Mao J, Ligon KL, Rakhlin EY, Thayer SP, Bronson RT, Rowitch D, et al. A novel somatic mouse model to survey tumorigenic potential applied to the Hedgehog pathway. Cancer Res. 2006;66:10171–8.

    CAS  Article  Google Scholar 

  27. 27.

    Gao J, Graves S, Koch U, Liu S, Jankovic V, Buonamici S, et al. Hedgehog signaling is dispensable for adult hematopoietic stem cell function. Cell Stem Cell. 2009;4:548–58.

    CAS  Article  Google Scholar 

  28. 28.

    Hofmann I, Stover EH, Cullen DE, Mao J, Morgan KJ, Lee BH, et al. Hedgehog signaling is dispensable for adult murine hematopoietic stem cell function and hematopoiesis. Cell Stem Cell. 2009;4:559–67.

    CAS  Article  Google Scholar 

  29. 29.

    Torchia EC, Boyd K, Rehg JE, Qu C, Baker SJ. EWS/FLI-1 induces rapid onset of myeloid/erythroid leukemia in mice. Mol Cell Biol. 2007;27:7918–34.

    CAS  Article  Google Scholar 

  30. 30.

    Beauchamp E, Bulut G, Abaan O, Chen K, Merchant A, Matsui W, et al. GLI1 is a direct transcriptional target of EWS-FLI1 oncoprotein. J Biol Chem. 2009;284:9074–82.

    CAS  Article  Google Scholar 

  31. 31.

    Zwerner JP, Joo J, Warner KL, Christensen L, Hu-Lieskovan S, Triche TJ, et al. The EWS/FLI1 oncogenic transcription factor deregulates GLI1. Oncogene. 2008;27:3282–91.

    CAS  Article  Google Scholar 

  32. 32.

    Joo J, Christensen L, Warner K, States L, Kang HG, Vo K. et al. GLI1 is a central mediator of EWS/FLI1 signaling in Ewing tumors. PLoS ONE. 2009;4:e7608

    Article  Google Scholar 

  33. 33.

    Kobune M, Iyama S, Kikuchi S, Horiguchi H, Sato T, Murase K, et al. Stromal cells expressing hedgehog-interacting protein regulate the proliferation of myeloid neoplasms. Blood Cancer J. 2012;2:e87.

    CAS  Article  Google Scholar 

  34. 34.

    Zou J, Hong Y, Tong Y, Wei J, Qin Y, Shao S, et al. Sonic hedgehog produced by bone marrow-derived mesenchymal stromal cells supports cell survival in myelodysplastic syndrome. Stem Cells Int. 2015;2015:957502.

    Article  Google Scholar 

  35. 35.

    Kang HJ, Kim YI, Kim HC, Jae HJ, Hur S, Chung JW. Does establishing a safety margin reduce local recurrence in subsegmental transarterial chemoembolization for small nodular hepatocellular carcinomas? Korean J Radiol. 2015;16:1068–78.

    Article  Google Scholar 

  36. 36.

    Cortes JE, Heidel FH, Heuser M, Fiedler W, Smith BD, Robak T, et al. A phase 2 randomized study of low dose Ara-C with or without glasdegib (PF-04449913) in untreated patients with acute myeloid leukemia or high-risk myelodysplastic syndrome. Blood. 2016;128:99.

    Google Scholar 

  37. 37.

    Seto M, Ohta M, Asaoka Y, Ikenoue T, Tada M, Miyabayashi K, et al. Regulation of the hedgehog signaling by the mitogen-activated protein kinase cascade in gastric cancer. Mol Carcinog. 2009;48:703–12.

    CAS  Article  Google Scholar 

  38. 38.

    Brechbiel J, Miller-Moslin K, Adjei AA. Crosstalk between hedgehog and other signaling pathways as a basis for combination therapies in cancer. Cancer Treat Rev. 2014;40:750–9.

    CAS  Article  Google Scholar 

  39. 39.

    Chaudhry P, Singh M, Triche TJ, Guzman M, Merchant AA. GLI3 repressor determines Hedgehog pathway activation and is required for response to SMO antagonist glasdegib in AML. Blood. 2017;129:3465–75.

    CAS  Article  Google Scholar 

  40. 40.

    Peacock CD, Wang Q, Gesell GS, Corcoran-Schwartz IM, Jones E, Kim J, et al. Hedgehog signaling maintains a tumor stem cell compartment in multiple myeloma. Proc Natl Acad Sci USA. 2007;104:4048–53.

    CAS  Article  Google Scholar 

  41. 41.

    Feldmann G, Dhara S, Fendrich V, Bedja D, Beaty R, Mullendore M, et al. Blockade of hedgehog signaling inhibits pancreatic cancer invasion and metastases: a new paradigm for combination therapy in solid cancers. Cancer Res. 2007;67:2187–96.

    CAS  Article  Google Scholar 

  42. 42.

    Bar EE, Chaudhry A, Lin A, Fan X, Schreck K, Matsui W, et al. Cyclopamine-mediated hedgehog pathway inhibition depletes stem-like cancer cells in glioblastoma. Stem Cells. 2007;25:2524–33.

    CAS  Article  Google Scholar 

  43. 43.

    Zhao C, Chen A, Jamieson CH, Fereshteh M, Abrahamsson A, Blum J, et al. Hedgehog signalling is essential for maintenance of cancer stem cells in myeloid leukaemia. Nature. 2009;458:776–9.

    CAS  Article  Google Scholar 

  44. 44.

    Wang Y, Krivtsov AV, Sinha AU, North TE, Goessling W, Feng Z, et al. The Wnt/beta-catenin pathway is required for the development of leukemia stem cells in AML. Science. 2010;327:1650–3.

    CAS  Article  Google Scholar 

  45. 45.

    Jamieson CH, Ailles LE, Dylla SJ, Muijtjens M, Jones C, Zehnder JL, et al. Granulocyte-macrophage progenitors as candidate leukemic stem cells in blast-crisis CML. N Engl J Med. 2004;351:657–67.

    CAS  Article  Google Scholar 

  46. 46.

    Merchant A, Joseph G, Wang Q, Brennan S, Matsui W. Gli1 regulates the proliferation and differentiation of HSCs and myeloid progenitors. Blood. 2010;115:2391–6.

    CAS  Article  Google Scholar 

  47. 47.

    Sekulic A, Migden MR, Oro AE, Dirix L, Lewis KD, Hainsworth JD, et al. Efficacy and safety of vismodegib in advanced basal-cell carcinoma. N Engl J Med. 2012;366:2171–9.

    CAS  Article  Google Scholar 

  48. 48.

    Zou J, Zhou Z, Wan L, Tong Y, Qin Y, Wang C. et al. Targeting the sonic Hedgehog-Gli1 pathway as a potential new therapeutic strategy for myelodysplastic syndromes. PLoS ONE. 2015;10:e0136843

    Article  Google Scholar 

  49. 49.

    Wellbrock J, Latuske E, Kohler J, Wagner K, Stamm H, Vettorazzi E, et al. Expression of Hedgehog pathway mediator GLI represents a negative prognostic marker in human acute myeloid leukemia and its inhibition exerts antileukemic effects. Clin Cancer Res. 2015;21:2388–98.

    CAS  Article  Google Scholar 

  50. 50.

    Long B, Wang LX, Zheng FM, Lai SP, Xu DR, Hu Y, et al. Targeting GLI1 suppresses cell growth and enhances chemosensitivity in CD34+ enriched acute myeloid leukemia progenitor cells. Cell Physiol Biochem. 2016;38:1288–1302.

    CAS  Article  Google Scholar 

  51. 51.

    Gerstung M, Pellagatti A, Malcovati L, Giagounidis A, Porta MG, Jadersten M, et al. Combining gene mutation with gene expression data improves outcome prediction in myelodysplastic syndromes. Nat Commun. 2015;6:5901.

    CAS  Article  Google Scholar 

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Acknowledgements

We would like to acknowledge Dr. David Huso, previously an Associate Professor in the Department of Molecular and Comparative Pathobiology at Johns Hopkins, for preparing the histology slides; and Dr. Hao Zhang, Research Associate in the Department of Molecular Microbiology and Immunology at Johns Hopkins, for helping to sort the murine bone marrow for cell subpopulations. This work was supported by grants from the National Institute of Health (K08 HL136894) (LPG) and Edward P. Evans Foundation (LPG). Flow cytometry and microarray analysis was performed with the support of the Sidney Kimmel Comprehensive Cancer Center Core Facilities (P30 CA006973).

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Correspondence to William Matsui or Lukasz P. Gondek.

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Lau, B.W., Huh, K., Madero-Marroquin, R. et al. Hedgehog/GLI1 activation leads to leukemic transformation of myelodysplastic syndrome in vivo and GLI1 inhibition results in antitumor activity. Oncogene 38, 687–698 (2019). https://doi.org/10.1038/s41388-018-0431-9

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