scholarly journals Detection of internal tandem duplications in the FLT3 gene by different electrophoretic methods

2012 ◽  
Vol 50 (2) ◽  
Author(s):  
Tamás Bubán ◽  
Katalin Koczok ◽  
Róza Földesi ◽  
Gabriella Szabó ◽  
Andrea Sümegi ◽  
...  
Keyword(s):  
Myeloid Leukemia ◽  
Mutant Allele ◽  
Tandem Duplication ◽  
Detection Methods ◽  
Internal Tandem Duplication ◽  
Complex Evaluation ◽  
Analytical Properties ◽  
Genescan Analysis ◽  
Tandem Duplications

AbstractIn acute myeloid leukemia (AML), the internal tandem duplication (ITD) in the juxtamembrane domain of theA complex evaluation of the analytical properties of the three most frequently used detection methods – PCR followed by agarose (AGE), polyacrylamide (PAGE) or capillary electrophoresis (CE) – was performed on 95 DNA samples obtained from 73 AML patients.All the three methods verified the presence of a mutant allele in 20 samples from 18 patients. AGE and PAGE could detect the presence of 1%–2% mutant allele, while the detection limit of CE was 0.28%. However, acceptable reproducibility (inter-assay CV <25%) of the mutant allele rate determination was only achievable above 1.5% mutant/total allele rate. The reproducibility of the ITD size determination by CE was much better, but the ITD size calculated by PeakScanner or GeneScan analysis was 7% lower as compared to values obtained by DNA sequencing. The presence of multiple ITD was over-estimated by PAGE and AGE due to the formation of heteroduplexes.This study suggests the use of PCR+CE in the diagnostics and the follow-up of AML patients. The data further supports the importance of proper analytical evaluation of home-made molecular biological diagnostic tests.

scholarly journals AML engraftment in the NOD/SCID assay reflects the outcome of AML: implications for our understanding of the heterogeneity of AML

Blood ◽  
2006 ◽  
Vol 107 (3) ◽  
pp. 1166-1173 ◽  
Author(s):  
Daniel J. Pearce ◽  
David Taussig ◽  
Kazem Zibara ◽  
Lan-Lan Smith ◽  
Christopher M. Ridler ◽  
...  
Keyword(s):  
Myeloid Leukemia ◽  
Tandem Duplication ◽  
Current Model ◽  
Internal Tandem Duplication ◽  
Similar Frequency ◽  
Nonobese Diabetic ◽  
Significant Difference ◽  
Frequency Source ◽  
Flt3 Internal Tandem Duplication

AbstractThe nonobese diabetic/severe combined immunodeficient (NOD/SCID) assay is the current model for assessment of human normal and leukemic stem cells. We explored why 51% of 59 acute myeloid leukemia (AML) patients were unable to initiate leukemia in NOD/SCID mice. Increasing the cell dose, using more permissive recipients, and alternative tissue sources did not cause AML engraftment in most previously nonengrafting AML samples. Homing of AML cells to the marrow was the same between engrafters and nonengrafters. FLT3 internal tandem duplication (ITD) and nucleophosmin mutations occurred at a similar frequency in engrafters and nonengrafters. The only variable that was related to engraftment ability was the karyotypically defined risk stratification of individual AML cases. Of interest, follow-up of younger patients with intermediate-risk AML revealed a significant difference in overall survival between NOD/SCID engrafting and nonengrafting AMLs. Hence, the ability of AML to engraft in the NOD/SCID assay seems to be an inherent property of AML cells, independent of homing, conditioning, or cell frequency/source, which is directly related to prognosis. Our results suggest an important difference between leukemic initiating cells between engrafting and nonengrafting AML cases that correlates with treatment response.

FLT3 Internal Tandem Duplications in Adults with De Novo Acute Myeloid Leukemia.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 4430-4430
Author(s):  
Sandra G. Xavier ◽  
Rocío Hassan ◽  
Nelma C.D. Clementino ◽  
Daniel G. Tabak ◽  
Nelson Spector ◽  
...  
Keyword(s):  
Myeloid Leukemia ◽  
Tandem Duplication ◽  
De Novo ◽  
Univariate Analysis ◽  
Internal Tandem Duplication ◽  
Hematopoietic Stem ◽  
Proliferation And Differentiation ◽  
Polymerase Chain ◽  
De Novo Aml ◽  
Tandem Duplications

Abstract FLT3 is a receptor tyrosine kinase involved in the proliferation and differentiation of hematopoietic stem cells. Recently, internal tandem duplication (ITD) mutations of the FLT3 gene have been described in patients with AML and associated with a poor prognosis. The aim of this study was to analyze the prevalence of FLT3-ITD in a series of 90 adults with de novo AML and correlate the presence of this mutation with biological characteristics and clinical response. We analyzed diagnostic peripheral blood or bone marrow specimens from 43 women and 47 men, with a median age of 38 years (16–83). Polymerase chain reaction was performed on genomic DNA using previously published primers for exons 11 and 12. An FLT3-ITD was found in 22/89 patients (25%). It was present in 37% (9/24) of the patients with acute promyelocytic leukemia (APL) and in only 20% (13/65) of the patients with non-M3-AML (p=0.07). The FLT3-ITD was not detected in patients with M6 (n=1) and M7-AML (n=3), nor in patients with the AML1-ETO (n=2) or with the CBFb-MYH11 (n=4) fusion genes. The median WBC counts were higher in FLT3-ITD patients than in those without the mutation (37 X 109/L vs. 27 X 109/L, p=0.43). In APL, FLT3-ITD was found in 5 out of 6 patients with the short PML-RARa isoform, but in only 4 out of 18 patients with the non-short isoform (p=0.01). Univariate analysis showed an association between the presence of FLT3-ITD and both a lower complete remission (CR) rate (41% vs. 64%; p=0.05) and a shorter overall survival (14% vs. 34%; p=0.03). However, FLT3-ITD was not associated with the CR rate (p=0.18) or the OS (p=0.07) in the multivariate analysis. The clinical significance of FLT3-ITD in adult AML remains uncertain, and further investigation is clearly warranted.

FLT3 internal tandem duplication during myelodysplastic syndrome follow-up: a marker of transformation to acute myeloid leukemia

2008 ◽  
Vol 183 (2) ◽  
pp. 89-93 ◽  
Author(s):  
Ronald Feitosa Pinheiro ◽  
Eloisa de Sá Moreira ◽  
Maria Regina Régis Silva ◽  
Fernando Lopes Alberto ◽  
Maria de Lourdes L.F. Chauffaille
Keyword(s):  
Acute Myeloid Leukemia ◽  
Myelodysplastic Syndrome ◽  
Myeloid Leukemia ◽  
Tandem Duplication ◽  
Internal Tandem Duplication ◽  
Flt3 Internal Tandem Duplication ◽  
Acute Myeloid

scholarly journals Size of FLT3 internal tandem duplication has prognostic significance in patients with acute myeloid leukemia

Blood ◽  
2006 ◽  
Vol 107 (9) ◽  
pp. 3724-3726 ◽  
Author(s):  
Derek L. Stirewalt ◽  
Kenneth J. Kopecky ◽  
Soheil Meshinchi ◽  
Julia H. Engel ◽  
Era L. Pogosova-Agadjanyan ◽  
...  
Keyword(s):  
Acute Myeloid Leukemia ◽  
Myeloid Leukemia ◽  
Tandem Duplication ◽  
Prognostic Significance ◽  
Juxtamembrane Domain ◽  
Internal Tandem Duplication ◽  
Southwest Oncology Group ◽  
Flt3 Internal Tandem Duplication ◽  
Tandem Duplications ◽  
Acute Myeloid

FLT3 internal tandem duplications (FLT3/ITDs) in the juxtamembrane domain are found in approximately 25% of acute myeloid leukemia (AML) patients, ranging in size from 3 to hundreds of nucleotides. We examined whether the sizes of FLT3/ITDs were associated with clinical outcomes in 151 AML patients enrolled in Southwest Oncology Group studies: S9333 and S9500. FLT3/ITDs were identified in 32% of patients (median ITD size = 39 nucleotides; range, 15-153 nucleotides). The CR rates were 35%, 67%, and 52% for patients with large (≥ 40), small (< 40), and no ITDs, respectively (P = .19). Increasing ITD size was associated with decreasing OS (estimated 5-year OS: large = 13%, small = 26%, and no ITD = 21%, P = .072) and RFS (estimated 5-year RFS: large = 13%, small = 27%, and no ITD = 34%, P = .017). These studies suggest that ITD size may have prognostic significance.

FMS-Like Tyrosine Kinase 3 Internal Tandem Duplication and the Patterns of Its Gene Sequence in 207 Chinese Patients With De Novo Acute Myeloid Leukemia

2012 ◽  
Vol 136 (1) ◽  
pp. 84-89 ◽  
Author(s):  
Ling Zhong ◽  
Yong Qian Jia ◽  
Wen Tong Meng ◽  
Xun Ni
Keyword(s):  
Acute Myeloid Leukemia ◽  
Tyrosine Kinase ◽  
Myeloid Leukemia ◽  
Tandem Duplication ◽  
De Novo ◽  
Chinese Patients ◽  
Wild Type ◽  
Internal Tandem Duplication ◽  
Tandem Duplications ◽  
Acute Myeloid

Context.—Constitutive activation of the FMS-like tyrosine kinase 3 (FLT3) receptor tyrosine kinase by internal tandem duplication (ITD) has been researched in patients with de novo acute myeloid leukemia (AML). Objective.—To study the patterns of FLT3-ITD in Chinese patients with AML. Design.—A total of 207 patients with de novo AML were enrolled in the study. Genomic DNA was extracted from peripheral blood and polymerase chain reaction was performed. GeneScan was used to analyze the mutant to wild-type ratio. The sequencing of mutated genes was performed to confirm the mutation types and exclude false positives. Results.—A total of 42 cases (20.3%) were associated with mutations. FLT3-ITD was found equally in AML subtypes M1 to M6. The level of the ITD allele was heterogeneous. GeneScan showed that the mutant to wild-type ratio ranged from 0.03 to 3.78 (median, 0.43). Patients with a high ratio had significantly lower cancer remission rates and shorter survival. They also showed distinct clinical features including higher white blood cell counts and higher CD7 and CD56 expression. The length of the duplicated fragment was 26 to 57 bp (median, 43 bp). Twenty-two cases (52%) had simple tandem duplications, while 20 other cases (48%) had an extra interval of 12 to 30 bp before the tandem duplications. A hexanucleotide consisting of GAAAAG was found exclusively in the intervals. Patients with this GAAAAG interval showed better survival. The ITD to wild-type ratio, gene pattern, and CD7 expression status appear to be independent prognostic indices for patients with AML. Conclusion.—Detection of FLT3 mutation is fast, easy, and inexpensive. The mutant to wild-type ratio is helpful for performing detailed risk stratification. DNA sequence analysis is more precise for confirming and evaluating the mutation pattern.

Sign in / Sign up

Export Citation Format

Share Document