Comparative Clinical Pathology

, Volume 22, Issue 3, pp 497–506

Clinical implication of nucleophosmin gene mutation and Flt-3 internal tandem duplication in a cohort of Egyptian AML patients

  • Mervat M. Khorshied
  • Wael A. Said
  • Hebat Allah M. Shaaban
Original Article

Abstract

Nucleophosmin (NPM) gene mutations are the most frequent genetic abnormality in adult AML. NPM gene mutation (NPM1) leads to aberrant localization of the NPM protein into the cytoplasm. As NPM1 mutation is frequently associated with FMS-like tyrosine kinase 3—internal tandem duplication (FLT3-ITD) that appears to abrogate its favorable prognostic effect. This study aimed at detecting the frequency of NPM1 exon-12 gene mutation and FLT3-ITD in 62 de novo AML patients by reverse-transcriptase polymerase chain reaction and immunocytochemical staining. Twenty age- and sex-matched healthy volunteers were included in the current study as a control group. NPM1 mutation was detected in 30/62 (48.3%) of cases, while 27/62 (43.5%) of cases were FLT3-ITD-positive. All the control subjects were negative for the studied genetic mutations. Immunostaining for NPM revealed cytoplasmic positivity (NPMc+) in 32/62 (51.6%) of case. NPM1 mutation was significantly higher in patients with normal karyotype, FAB-M4 subtype, low expression of CD34 and favorable response to induction therapy. FLT3-ITD was higher among female patients and was associated with poor response to induction therapy. Patients harboring both mutations showed unfavorable response as the presence of FLT3-ITD abolished the favorable effect of NPM1. In conclusion, all AML cases should be screened prior to therapy for both mutations as two important prognostic markers that can be valuable in predicting the response to therapy, in addition to their role in monitoring minimal residual disease and early detection of relapse. Furthermore, they represent potential therapeutic targets.

Keywords

Nucleophosmin FLT3-ITD AML RT-PCR Immunohistochemical staining 

References

  1. Aly R, Shahin D, Azmy E (2011) Prognostic significance of FLT3 internal tandem duplication in Egyptian acute myeloid leukemia and normal cytogenetics. Comp Clin Pathol pp. 1–7. doi:10.1007/s00580-011-1223-6 Key: citeulike:9441245Google Scholar
  2. Ahmad F, Mandava S, Das BR (2009) Mutations of NPM1 gene in de novo acute myeloid leukemia: determination of incidence, distribution pattern and identification of two novel mutations in Indian population. Hematol Oncol 27(2):90–97PubMedCrossRefGoogle Scholar
  3. Alcalay M, Tiacci E, Bergomas R, Bigerna B et al (2005) Acute myeloid leukemia bearing cytoplasmic nucleophosmin (NPMc+ AML) shows a distinct gene expression profile characterized by up regulation of genes involved in stem cell maintenance. Blood 106:899–902PubMedCrossRefGoogle Scholar
  4. Bacher U, Kern W, Schoch C et al (2006) Evaluation of complete disease remission in acute myeloid leukemia: a prospective study based on cytomorphology, interphase fluorescence in situ hybridization, and immunophenotyping during follow-up in patients with acute myeloid leukemia. Cancer 106(4):839–847PubMedCrossRefGoogle Scholar
  5. Beran M, Luthra R, Kantarjian H, Estey E (2004) FLT3 mutation and response to intensive chemotherapy in young adult and elderly patients with normal karyotype. Leuk Res 28:547–550PubMedCrossRefGoogle Scholar
  6. Bienz M, Ludwig M, Oppliger Leibundgut E et al (2005) Risk assessment in patients with acute myeloid leukemia and a normal karyotype. Clin Cancer Res 11:1416–1424PubMedCrossRefGoogle Scholar
  7. Boissel N, Renneville A, Biggio V, Philippe N, Thomas X, Cayuela J, Terre C, Tigaud I et al (2005) Prevalence, clinical profile, and prognosis of NPM mutations in AML with normal karyotype. Blood 106:3618–3620PubMedCrossRefGoogle Scholar
  8. Boonthimat C, Thongnoppakhun W, Auewarakul CU (2008) Nucleophosmin mutation in Southeast Asian acute myeloid leukemia: eight novel variants, FLT3 coexistence and prognostic impact of NPM1/FLT3 mutations. Haematologica 93(10):1565–1569PubMedCrossRefGoogle Scholar
  9. Byrd JC, Mrózek K, Dodge RK et al (2002) Pretreatment cytogenetic abnormalities are predictive of induction success, cumulative incidence of relapse, and overall survival in adult patients with de novo acute myeloid leukemia: results from Cancer and Leukemia Group B (CALGB 8461). Blood 100:4325–4336PubMedCrossRefGoogle Scholar
  10. Calvo KL, Ojeda MJ, Ammatuna E, Lavorgna S, Ottone T, Targovnik HM, Lo-Coco F, Noguera NI (2009) Detection of the nucleophosmin gene mutations in acute myelogenous leukemia through RT-PCR and polyacrylamide gel electrophoresis. Eur J Haematol 82(1):69–72PubMedCrossRefGoogle Scholar
  11. Cazzaniga G, Dell’Oro M, Mecucci C, Giarin E, Masetti R, Falini B, Rossi V, Locatelli F, Martelli M, Basso G, Pession A (2005) Nucleophosmin mutations in childhood acute myelogenous leukemia with normal karyotype. Blood 106:1419–1422PubMedCrossRefGoogle Scholar
  12. Chang H, Salma F, Yi QL, Patterson B, Brein B, Minden MD (2004) Prognostic relevance of immunophenotyping in 379 patients with acute myeloid leukemia. Leuk Res 28:43–48PubMedCrossRefGoogle Scholar
  13. Chen W, Rassidakis GZ, Medeiros LJ (2006) Nucleophosmin gene mutations in acute myeloid leukemia. Arch Pathol Lab Med 130(11):1687–1692PubMedGoogle Scholar
  14. Chou WC, Tang JL, Lin LI, Yao M, Tsay W, Chen CY, Wu SJ, Huang CF, Chiou RJ, Tseng MH, Lin DT, Lin KH, Chen YC, Tien HF (2006) Nucleophosmin mutations in de novo acute myeloid leukemia: the age-dependent incidences and the stability during disease evolution. Cancer Res 15;66(6):3310–3316Google Scholar
  15. De Propris MS, Raponi S, Diverio D, Milani ML, Meloni G, Falini B, Foà R, Guarini A (2011) High CD33 expression levels in acute myeloid leukemia cells carrying the nucleophosmin (NPM1) mutation. Haematologica 96(10):1548–1551PubMedCrossRefGoogle Scholar
  16. Döhner K, Döhner H (2008) Molecular characterization of acute myeloid leukemia. Haematologica 93(7):976–982PubMedCrossRefGoogle Scholar
  17. Döhner K, Schlenk RF, Habdank M, Scholl C, Rücker FG, Corbacioglu A et al (2005) Mutant nucleophosmin (NPM1) predicts favorable prognosis in younger adults with acute myeloid leukemia and normal cytogenetics: interaction with other gene mutations. Blood 106:3740–3746PubMedCrossRefGoogle Scholar
  18. Facchetti F, Pileri SA, Agostinelli C, Martelli MP, Paulli M, Venditti A, Martelli MF, Falini B (2009) Cytoplasmic nucleophosmin is not detected in blastic plasmacytoid dendritic cell neoplasm. Haematologica 94(2):285–288PubMedCrossRefGoogle Scholar
  19. Falini B, Mecucci C, Tiacci E, Alcalay M, Rosati R, Pasqualucci L et al (2005) Cytoplasmic nucleophosmin in acute myelogenous leukemia with a normal karyotype. GIMEMA Acute Leukemia Working Party. N Engl J Med 352:254–266PubMedCrossRefGoogle Scholar
  20. Falini B, Nicoletti I, Martelli FM, Mecucci M (2007) Acute myeloid leukemia carrying cytoplasmic/mutated nucleophosmin (NPMc_AML): biological and clinical features. Blood 109:874–885PubMedCrossRefGoogle Scholar
  21. Falini B, Martelli MP, Mecucci C, Liso A, Bolli N, Bigerna B et al (2008) Cytoplasmic mutated nucleophosmin is stable in primary leukemic cells and in a xenotransplant model of NPMc+ acute myeloid leukemia in SCID mice. Haematologica 93:775–779PubMedCrossRefGoogle Scholar
  22. Falini B, Gionfriddo I, Cecchetti F, Ballanti S, Pettirossi V, Martelli MP (2011) Acute myeloid leukemia with mutated nucleophosmin (NPM1): any hope for a targeted therapy? Blood Rev 25(6):247–254PubMedCrossRefGoogle Scholar
  23. Gale R, Green C, Allen C, Mead AJ, Burnett AK, Hills RK et al (2008) The impact of FLT3 internal tandem duplication mutant level, number, size, and interaction with NPM1 mutations in a large cohort of young adult patients with acute myeloid leukemia. Blood 111(5):2776–2784PubMedCrossRefGoogle Scholar
  24. Giles F, Schiffer C, Kantarjian H et al (2004) Phase 1 study of PKC412, an oral FLT3 kinase inhibitor, in sequential and concomitant combinations with daunorubicin and cytarabine (DA) induction and high-dose cytarabine (HDAra-C) consolidation in newly diagnosed patients with AML. Blood 104:78a, Abstract 262Google Scholar
  25. Haferlach C, Mecucci C, Schnittger S, Kohlmann A, Mancini M, Cuneo A et al (2009) AML with mutated NPM1 carrying a normal or aberrant karyotype show overlapping biologic, pathologic, immunophenotypic, and prognostic features. Blood 114(14):3024–3032PubMedCrossRefGoogle Scholar
  26. Kainz B, Heintel D, Marculescu R et al (2002) Variable prognostic value of FLT3 internal tandem duplications in patients with de novo AML and a normal karyotype, t(15;17), t(8;21) or inv(16). Hematol J 3:283–289PubMedCrossRefGoogle Scholar
  27. Kiyoi H, Naoe T (2006) Biology, clinical relevance, and molecularly targeted therapy in acute leukemia with FLT3 mutation. Int J Hematol 83(4):301–308PubMedCrossRefGoogle Scholar
  28. Kottaridis PD, Gale RE, Frew ME et al (2001) The presence of a FLT3 internal tandem duplication in patients with acute myeloid leukemia (AML) adds important prognostic information to cytogenetic risk group and response to the first cycle of chemotherapy: analysis of 854 patients from the United Kingdom Medical Research Council AML 10 and 12 trials. Blood 98:1752–1759PubMedCrossRefGoogle Scholar
  29. Liso L, Castiglione F, Cappuccio A, Stracci F, Schlenk R et al (2008) A one-mutation mathematical model can explain the age incidence of acute myeloid leukemia with mutated nucleophosmin (NPM1). Haematologica 93(8):1219–1226PubMedCrossRefGoogle Scholar
  30. Mrózek K, Marcucci G, Paschka P, Whitman S, Bloomfield C (2007) Clinical relevance of mutations and gene-expression changes in adult acute myeloid leukemia with normal cytogenetics: are we ready for prognostically prioritized molecular classification? Blood 109:431–448PubMedCrossRefGoogle Scholar
  31. Ohno R, Asou N, Ohnishi K (2003) Treatment of acute promyelocytic leukemia: strategy toward further increase of cure rate. Leukemia 17:1454–1463PubMedCrossRefGoogle Scholar
  32. Ottone T, Ammatuna E, Lavorgna S, Noguera N, Buccisano F et al (2008) An Allele-Specific RT-PCR Assay to Detect Type A Mutation of the Nucleophosmin-1 Gene in Acute Myeloid Leukemia. J Mol Diagn 10(3):212–216PubMedCrossRefGoogle Scholar
  33. Quentmeier H, Martelli MP, Dirks WG, Bolli N, Liso A, MacLeod RA et al (2005) Cell line OCI/AML3 bears exon-12 NPM gene mutation-A and cytoplasmic expression of nucleophosmin. Leukemia 19:1760–1767PubMedCrossRefGoogle Scholar
  34. Rau R, Brown P (2009) Nucleophosmin (NPM1) mutations in adult and childhood acute leukemia: towards definition of a new leukemia entity. Hematol Oncol 27(4):171–181PubMedCrossRefGoogle Scholar
  35. Schlenk RF, Dohner K, Krauter J, Frohling S, Cobacioglu A et al (2009) Mutations and treatment outcome in cytogenetically normal acute myeloid leukemia. N Engl J Med 358(18):1909–1918CrossRefGoogle Scholar
  36. Scholl S, Krause C, Loncarevic IF, Müller R, Kunert C, Wedding U, Sayer HG, Clement JH, Höffken K (2005) Specific detection of Flt3 point mutations by highly sensitive real-time polymerase chain reaction in acute myeloid leukemia. J Lab Clin Med 145(6):295–304PubMedCrossRefGoogle Scholar
  37. Schneider F, Hoster E, Unterhalt M, Schneider S, Dufour A, Benthaus T, Mellert G, Zellmeier E, Bohlander SK, Feuring-Buske M, Buske C, Braess J, Fritsch S, Heinecke A, Sauerland MC, Berdel WE, Buechner T, Woermann BJ, Hiddemann W, Spiekermann K (2009) NPM1 but not FLT3-ITD mutations predict early blast cell clearance and CR rate in patients with normal karyotype AML (NK-AML) or highrisk myelodysplastic syndrome (MDS). Blood 21;113(21):5250–5253Google Scholar
  38. Schnittger S, Schoch C, Kern W, Mecucci C, Tschulik C, Martelli MF, Haferlach T, Hiddemann W, Falini B (2005) Nucleophosmin gene mutations are predictors of favorable prognosis in acute myelogenous leukemia with a normal karyotype. Blood 106(12):3733–3739PubMedCrossRefGoogle Scholar
  39. Schnittger S, Kern W, Tschulik C, Weiss T, Dicker F, Falini B, Haferlach C, Haferlach T (2009) Minimal residual disease levels assessed by NPM1 mutation-specific RQ-PCR provide important prognostic information in AML. Blood 114(11):2220–2231PubMedCrossRefGoogle Scholar
  40. Suzuki T, Kiyoi H, Oziki K, Tomita A (2005) Clinical characteristics and prognostic implication of NPM1 mutation in acute myeloid leukemia. Blood 106(8):2854–2861PubMedCrossRefGoogle Scholar
  41. Tan A, Westerman D, Carney D, Seymour J, Juneja S, Dobrovic A (2008) Detection of NPM1 exon 12 mutations and FLT3—internal tandem duplications by high resolution melting analysis in normal karyotype acute myeloid leukemia. J Hematol Oncol 1:10PubMedCrossRefGoogle Scholar
  42. Thiede C, Koch S, Creutzig E et al (2006) Prevalence and prognostic impact of NPM1 mutations in 1485 adult patients with acute myeloid leukemia (AML). Blood 107:4011–4020PubMedCrossRefGoogle Scholar
  43. Verhaak RG, Goudswaard CS, van Putten W, Bijl MA, Sanders MA, Hugens W et al (2005) Mutations in nucleophosmin NPM1 in acute myeloid leukemia (AML): association with other gene abnormalities and previously established gene expression signatures and their favorable prognostic significance. Blood 106:3747–3754PubMedCrossRefGoogle Scholar
  44. Weisberg E, Ray A, Nelson E, Adamia S, Barrett R, Sattler M, Zhang C, Daley JF, Frank D, Fox E, Griffin JD (2011) Reversible resistance induced by FLT3 inhibition: a novel resistance mechanism in mutant FLT3-expressing cells. PLoS One 6(9):e25351PubMedCrossRefGoogle Scholar
  45. Whitman SP, Archer KJ, Feng L et al (2001) Absence of the wild-type allele predicts poor prognosis in adult de novo acute myeloid leukemia with normal cytogenetics and the internal tandem duplication of FLT3: a cancer and leukemia group B study. Cancer Res 61:7233–7239PubMedGoogle Scholar

Copyright information

© Springer-Verlag London Limited 2012

Authors and Affiliations

  • Mervat M. Khorshied
    • 1
    • 4
  • Wael A. Said
    • 2
  • Hebat Allah M. Shaaban
    • 3
  1. 1.Department of Clinical Pathology, Faculty of MedicineCairo UniversityCairoEgypt
  2. 2.Department of Oncology, Faculty of MedicineCairo UniversityCairoEgypt
  3. 3.Department of Cytopathology, National Cancer InstituteCairo UniversityCairoEgypt
  4. 4.GizaEgypt

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