Detection of FLT3 Oncogene Mutations in Acute Myeloid Leukemia Using Conformation Sensitive Gel Electrophoresis
Abstract
:1. Introduction
2. Results and Discussion
2.1. Detection of the FLT3-ITD mutation
2.2. Detection of the Asp835Tyr mutation
2.3. Analysis of correlation with survival
3. Conclusions
4. Experimental Section
4.1. Patient DNA samples
4.2. Genomic DNA amplification
4.3. Mutation detection
Acknowledgments
References
- Ullrich, A; Schlessinger, J. Signal transduction by receptors with tyrosine kinase activity. Cell 1990, 61, 203–212. [Google Scholar]
- Matthews, W; Jordan, CT; Wiegand, GW; Pardoll, D; Lemischka, IR. A receptor tyrosine kinase specific to hematopoietic stem and progenitor cell-enriched populations. Cell 1991, 65, 1143–1152. [Google Scholar]
- Rosnet, O; Schiff, C; Pébusque, MJ; Marchetto, S; Tonnelle, C; Toiron, Y; Birg, F; Birnbaum, D. Human FLT3/FLK2 gene: cDNA cloning and expression in hematopoietic cells. Blood 1993, 82, 1110–1119. [Google Scholar]
- Yarden, Y; Ullrich, A. Growth factor receptor tyrosine kinase. Annu. Rev. Biochem 1988, 57, 443–478. [Google Scholar]
- Abu-Duhier, FM; Goodeve, AC; Wilson, GA; Care, RS; Peake, IR; Reilly, JT. Identification of novel FLT-3 Asp835 mutations in adult acute myeloid leukaemia. Br. J. Haematol 2001, 113, 983–988. [Google Scholar]
- Birg, E; Courcoul, M; Rosnet, O; Bardin, F; Pebusque, MJ; Marchetto, S; Tabilio, A; Mannoni, P; Birnbaum, D. Expression of the FMS/KIT-like gene FLT3 in human acute leukemias of the myeloid and lymphoid lineages. Blood 1992, 80, 2584–2593. [Google Scholar]
- Meierhoff, G; Dehmel, U; Gruss, HJ; Rosnet, O; Birnhaum, D; Quentmeier, H; Dirks, W; Drexler, HG. Expression of FLT3 receptor and FLT-3 ligand in human leukemia-lymphoma cell lines. Leukemia 1995, 9, 1368–1372. [Google Scholar]
- Drexler, HG. Expression of FLT3 receptor and response of to FLT3 ligand by leukemic cells. Leukemia 1996, 10, 588–599. [Google Scholar]
- Ihle, JN; Witthuhn, BA; Quelle, FW; Yamamoto, K; Silvennoinen, O. Signaling through the hematopoietic cytokine receptors. Annu. Rev. Immunol 1995, 13, 369–398. [Google Scholar]
- Kelly, LM; Kutok, JL; Williams, IR; Boulton, CL; Amaral, SM; Curley, DP; Ley, TJ; Gilliland, DG. PML/RARalpha and FLT3-ITD induce an APL-like disease in a mouse model. Proc. Natl. Acad. Sci. 2002, 99, 8283–8288. [Google Scholar]
- Kelly, LM; Liu, Q; Kutok, JL; Williams, IR; Boulton, CL; Gilliland, DG. FLT3 internal tandem duplication mutations associated with human acute myeloid leukemias induce myeloproliferative disease in a murine bone marrow transplant model. Blood 2002, 99, 310–318. [Google Scholar]
- Zhao, M; Kiyoi, H; Yamamoto, Y; Ito, M; Towatari, M; Omura, S; Kitamura, T; Ueda, R; Saito, H; Naoe, T. In vivo treatment of mutant FLT3-transformed murine leukemia with a tyrosine kinase inhibitor. Leukemia 2000, 14, 374–378. [Google Scholar]
- Tse, KF; Novelli, E; Civin, CI; Bohmer, FD; Small, D. Inhibition of FLT3-mediated transformation by use of a tyrosine kinase inhibitor. Leukemia 2001, 15, 1001–1010. [Google Scholar]
- Levis, M; Tse, KF; Smith, BD; Garrett, E; Small, D. A FLT3 tyrosine kinase inhibitor is selectively cytotoxic to acute myeloid leukemia blasts harboring FLT3 internal tandem duplication mutations. Blood 2001, 98, 885–887. [Google Scholar]
- Weisberg, E; Boulton, C; Kelly, LM; Manley, P; Fabbro, D; Meyer, T; Gilliland, DG; Griffin, JD. Inhibition of mutant FLT3 receptors in leukemia cells by the small molecule tyrosine kinase inhibitor PKC412. Cancer Cell 2002, 1, 433–443. [Google Scholar]
- O’Farrell, AM; Foran, JM; Fiedler, W; Serve, H; Paquette, RL; Cooper, MA; Yuen, HA; Louie, SG; Kim, H; Nicholas, S; Heinrich, MC; Berdel, WE; Bello, C; Jacobs, M; Scigalla, P; Manning, WC; Kelsey, S; Cherrington, JM. An innovative phase I clinical study demonstrates inhibition of FLT3 phosphorylation by SU11248 in acute myeloid leukemia patients. Clin. Cancer Res 2003, 9, 5465–5476. [Google Scholar]
- Fiedler, W; Mesters, R; Tinnefeld, H; Loges, S; Staib, P; Duhrsen, U; Flasshove, M; Ottmann, OG; Jung, W; Cavalli, F; Kuse, R; Thomalla, J; Serve, H; O’Farrell, AM; Jacobs, M; Brega, NM; Scigalla, P; Hossfeld, DK; Berdel, WE. A phase 2 clinical study of SU5416 in patients with refractory acute myeloid leukemia. Blood 2003, 102, 2763–2767. [Google Scholar]
- DeAngelo, DJ; Stone, RM; Heaney, ML; Nimer, SD; Paquette, RL; Klisovic, RB; Caligiuri, MA; Cooper, MR; Lecerf, JM; Karol, MD; Sheng, S; Holford, N; Curtin, PT; Druker, BJ; Heinrich, MC. Phase 1 clinical results with tandutinib (MLN518), a novel FLT3 antagonist, in patients with acute myelogenous leukemia or high-risk myelodysplastic syndrome: Safety, pharmacokinetics, and pharmacodynamics. Blood 2006, 108, 3674–3681. [Google Scholar]
- Stone, RM; de Angelo, J; Galinsky, I; Estey, E; Klimek, V; Grandin, W; Lebwohl, D; Yap, A; Cohen, P; Fox, E; Neuberg, D; Clark, J; Gilliland, DG; Griffin, JD. PKC 412 FLT3 inhibitor therapy in AML: Results of a phase II trial. Ann. Hematol 2004, 83, 89–90. [Google Scholar]
- Fenski, R; Flesch, K; Serve, S; Mizuki, M; Oelmann, E; Kratz, A; Kienast, J; Leo, R; Schwartz, S; Berdel, WE; Serve, H. Constitutive activation of FLT3 in acute myeloid leukaemia and its consequences for growth of 32D cells. Br. J. Haematol 2000, 108, 322–330. [Google Scholar]
- Kiyoi, H; Towatari, M; Yokota, S; Hamaguchi, M; Ohno, R; Saito, H; Naoe, T. Internal tandem duplication of the FLT3 gene is a novel modality of elongation mutation which causes constitutive activation of the product. Leukemia 1998, 12, 1333–1337. [Google Scholar]
- Yamamoto, Y; Kiyoi, H; Nakano, Y; Suzuki, R; Kodera, Y; Miyawaki, S; Asou, N; Kuriyama, K; Yagasaki, F; Shimazaki, C; Akiyama, H; Saito, K; Nishimura, M; Motoji, T; Shinagawa, K; Takeshita, A; Saito, H; Ueda, R; Ohno, R; Naoe, T. Activating mutation of D835 within the activation loop of FLT3 in human hematologic malignancies. Blood 2001, 97, 2434–2439. [Google Scholar]
- Xu, F; Taki, T; Yang, HW; Hanada, R; Hongo, T; Ohnishi, H; Kobayashi, M; Bessho, F; Yanagisawa, M; Hayashi, Y. Tandem duplication of the FLT3 gene is found in acute lymphoblastic leukaemia as well as acute myeloid leukaemia but not in myelodysplastic syndrome or juvenile chronic myelogenous leukaemia in children. Br. J. Haematol 1999, 105, 155–162. [Google Scholar]
- Abu, D; Goodeve, AC; Wilson, GA; Gari, MA; Peake, IR; Rees, DC; Vandenberghe, EA; Winship, PR; Reilly, JT. FLT3 internal tandem duplication mutations in adult acute myeloid leukaemia define a high-risk group. Br. J. Haematol 2000, 111, 190–195. [Google Scholar]
- Horiike, S; Yokota, S; Nakao, M; Iwai, T; Sasai, Y; Kaneko, H; Taniwaki, M; Kashima, K; Fujii, H; Abe, T; Misawa, S. Tandem duplications of the FLT3 receptor gene are associated with leukemic transformation of myelodysplasia. Leukemia 1997, 11, 1442–1446. [Google Scholar]
- Kiyoi, H; Naoe, T; Nakano, Y; Yokota, S; Minami, S; Miyawaki, S; Asou, N; Kuriyama, K; Jinnai, I; Shimazaki, C; Akiyama, H; Saito, K; Oh, H; Motoji, T; Omoto, E; Saito, H; Ohno, R; Ueda, R. Prognostic implication of FLT3 and N-RAS gene mutations in acute myeloid leukemia. Blood 1999, 93, 3074–3080. [Google Scholar]
- Yokota, S; Kiyoi, H; Nakao, M; Iwai, T; Misawa, S; Okuda, T; Sonoda, Y; Abe, T; Kahsima, K; Matsuo, Y; Naoe, T. Internal tandem duplication of the FLT3 gene is preferentially seen in acute myeloid leukemia and myelodysplastic syndrome among various hematological malignancies. A study on a large series of patients and cell lines. Leukemia 1997, 11, 1605–1609. [Google Scholar]
- Wang, L; Lin, D; Zhang, X; Chen, S; Wang, M; Wang, J. Analysis of FLT3 internal tandem duplication and D835 mutations in Chinese acute leukemia patients. Leuk. Res 2005, 29, 1393–1398. [Google Scholar]
- Auewarakul, CU; Sritana, N; Limwongse, C; Thongnoppakhun, W; Yenchitsomanus, PT. Mutations of the FLT3 gene in adult acute myeloid leukemia: determination of incidence and identification of a novel mutation in a Thai population. Cancer Genet. Cytogenet 2005, 162, 127–134. [Google Scholar]
- Thiede, C; Steudel, C; Mohr, B; Schaich, M; Schäkel, U; Platzbecker, U; Wermke, M; Bornhäuser, M; Ritter, M; Neubauer, A; Ehninger, G; Illmer, T. Analysis of FLT3 activating mutations in 979 patients with acute myelogenous leukemia: association with FAB subtypes and identification of subgroups with poor prognosis. Blood 2002, 99, 4326–4335. [Google Scholar]
- Ganguly, A. An update on conformation sensitive gel electrophoresis. Hum. Mutat 2002, 19, 334–342. [Google Scholar]
- Williams, IJ; Abuzenadah, A; Winship, PR; Preston, FE; Dolan, G; Wright, J; Peake, IR; Goodeve, AC. Precise carrier diagnosis in families with haemophilia A: Use of conformational sensitive gel electrophoresis for mutation screening and polymorphism analysis. Thromb. Haemost 1998, 79, 723–726. [Google Scholar]
- Goodeve, AC; Williams, I; Bray, GL; Peake, IR. Relationship between factor VIII mutation type and inhibitor development in a cohort of previously untreated patients treated with recombinant factor VIII (Recombinate). Recombinate PUP Study Group. Thromb. Haemost 2000, 83, 844–848. [Google Scholar]
- Hashemi Soteh, M; Peake, IR; Marsden, L; Anson, J; Batlle, J; Meyer, D; Fressinaud, E; Mazurier, C; Goudemand, J; Eikenboom, J; Goodeve, A. MCMDM-1VWD Study Group. Mutational analysis of the von Willebrand factor gene in type 1 von Willebrand disease using conformation sensitive gel electrophoresis: a comparison of fluorescent and manual techniques. Haematologica 2007, 92, 550–553. [Google Scholar]
- Hinks, JL; Winship, PR; Makris, M; Preston, FE; Peake, IR; Goodeve, AC. Rapid method for haemophilia B mutation detection using conformation sensitive gel electrophoresis. Br. J. Haematol 1999, 104, 915–918. [Google Scholar]
- Gari, MA; Abuzenadah, AM; Chaudhary, AG; Al-Qahtani, MH; Al-Says, FM; Dmanhouri, G. Pilot study of DNA extraction from archival unstained bone marrow slides: comparison of three rapid methods. Af J. Biotech 2006, 5, 532–535. [Google Scholar]
- Bennett, JM; Catovsky, D; Daniel, MT; Flandrin, G; Galton, DA; Gralnick, HR; Sultan, C. Proposal for the classification of the acute leukaemias. French American British (FAB) cooperative group. Br. J. Haematol 1976, 33, 451–458. [Google Scholar]
- Grimwade, D; Walker, H; Oliver, F; Wheatley, K; Harrison, C; Harrison, G; Rees, J; Hann, J; Hann, I; Stevens, R; Burnett, A; Goldstone, A. The importance of diagnostic cytogenetics on outcome in AML: Analysis of 1,612 patients entered into the MRC AML 10 trial. The Medical research Council Adult and Children's Leukaemia Working Parties. Blood 1998, 92, 2322–2333. [Google Scholar]
Case no. | Age (years) | Sex | FAB subtype | Karyotype | Mutations |
---|---|---|---|---|---|
5 | 73 | F | M3 | 46;XY, t(15;17) | ITD |
7 | 32 | M | M3 | 46;XY, t(15;17) | ITD |
9 | 48 | F | M3 | 46;XY, t(15;17) | ITD |
12 | 51 | F | M2 | 46;XY, t(8;21) | ITD |
18 | 24 | F | M3 | 46;XY, t(15;17) | ITD |
20 | 62 | F | M3 | 46;XY, t(15;17) | ITD |
23 | 32 | M | M4 | 46;XY, inv(16) | ITD |
25 | 33 | M | M4 | 46;XY, inv(16) | ITD |
28 | 38 | M | M3 | 46;XY, t(15;17) | ITD |
29 | 46 | F | M2 | 46;XY, t(8;21) | ITD |
41 | 48 | M | M3 | 46;XY, t(15;17) | ITD |
44 | 62 | F | M3 | 46;XY, t(15;17) | ITD |
49 | 64 | F | M4 | 46;XY, inv(16) | ITD |
53 | 43 | F | M2 | 46;XY, t(8;21) | ITD |
58 | 49 | M | M3 | 46;XY, t(15;17) | ITD |
64 | 48 | M | M4 | 46;XY, inv(16) | Asp 835 Tyr |
69 | 47 | M | M4 | 46;XY, inv(16) | Asp 835 Tyr |
80 | 34 | M | M4 | 46;XY, inv(16) | Asp 835 Tyr |
83 | 39 | M | M2 | 46;XY, t(8;21) | Asp 835 Tyr |
91 | 37 | M | M2 | 46;XY, t(8;21) | Asp 835 Tyr |
96 | 38 | F | M6 | 46;XY | Asp 835 Tyr |
108 | 35 | F | M5 | 46;XY | Asp 835 Tyr |
116 | 31 | F | M1 | 46;XY | Asp 835 Tyr |
118 | 33 | F | M4 | 46;XY | Asp 835 Tyr |
121 | 25 | M | M2 | 46;XY, inv(16) | Asp 835 Tyr |
128 | 45 | F | M6 | 46;XY | Asp 835 Tyr |
Primer sequences | Fragment size (bp) | ||
---|---|---|---|
F | 5′-TCTGTTTCATCGCTGAGTGAC-3′ | ||
ITD | 396 | ||
R | 5′-AGCCTTGAAACATGGCAAAC-3′ | ||
F | 5′-CCAGGAACGTGCTTGTCA-3′ | ||
Exon 20 | 352 | ||
R | 5′-TCAAAAATGCACCACAGTGAG-3′ |
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Gari, M.; Abuzenadah, A.; Chaudhary, A.; Al-Qahtani, M.; Banni, H.; Ahmad, W.; Al-Sayes, F.; Lary, S.; Damanhouri, G. Detection of FLT3 Oncogene Mutations in Acute Myeloid Leukemia Using Conformation Sensitive Gel Electrophoresis. Int. J. Mol. Sci. 2008, 9, 2194-2204. https://doi.org/10.3390/ijms9112194
Gari M, Abuzenadah A, Chaudhary A, Al-Qahtani M, Banni H, Ahmad W, Al-Sayes F, Lary S, Damanhouri G. Detection of FLT3 Oncogene Mutations in Acute Myeloid Leukemia Using Conformation Sensitive Gel Electrophoresis. International Journal of Molecular Sciences. 2008; 9(11):2194-2204. https://doi.org/10.3390/ijms9112194
Chicago/Turabian StyleGari, Mamdooh, Adel Abuzenadah, Adeel Chaudhary, Mohammed Al-Qahtani, Huda Banni, Waseem Ahmad, Fatin Al-Sayes, Sahira Lary, and Ghazi Damanhouri. 2008. "Detection of FLT3 Oncogene Mutations in Acute Myeloid Leukemia Using Conformation Sensitive Gel Electrophoresis" International Journal of Molecular Sciences 9, no. 11: 2194-2204. https://doi.org/10.3390/ijms9112194