Prognostic significance of quantitative analysis of <i>WT1</i> gene transcripts by competitive reverse transcription polymerase chain reaction in acute leukaemia

Garg, Mamta; Moore, Helen M.; Tobal, Khalid; Yin, John

doi:10.1046/j.1365-2141.2003.04552.x

Cited by 74 publications

(81 citation statements)

References 29 publications

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“…The RQ-PCR measurement of WT1 levels is especially useful as a high percentage of patients have high levels at diagnosis and relapse, including those patients who have no specific marker for detection of MRD. 8 Our results are also in accordance with those for MRD quantitation in BM harvests used for autografting in ALL, where higher MRD levels detected by PCR were associated with a worse RFS. 13 It could be argued that higher WT1 levels may merely reflect higher CD34 counts in BM harvests.…”

Section: Discussionsupporting

confidence: 80%

“…Extraction of RNA and cDNA synthesis was as previously described. 8,10 Quantitation of WT1 transcript was performed using real-time-PCR. The housekeeping gene ABL was used as the control gene for these quantifications.…”

Section: Sample Preparation and Quantitation Of Wt1 Transcriptsmentioning

confidence: 99%

“…Equal degradation rates for ABL and WT1 transcripts have previously been described. 8 Amplification reactions were carried out using the ABI Prism 7700 Sequence Detector System. In all, 2 ml cDNA was added to each reaction (25 ml) and a 40-cycle amplification was then performed.…”

Section: Sample Preparation and Quantitation Of Wt1 Transcriptsmentioning

confidence: 99%

“…This gene is overexpressed in over 90% of leukaemia cells and several studies have shown that quantitation of WT1 transcripts for patients in remission has prognostic value. [5][6][7][8] This study was undertaken to measure MRD levels in BM harvests used for autografting in AML, through WT1 quantitation, and to correlate these results with clinical outcome. …”

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confidence: 99%

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Elevated levels of WT1 transcripts in bone marrow harvests are associated with a high relapse risk in patients autografted for acute myeloid leukaemia

Osborne

Frost

Tobal

et al. 2005

Bone Marrow Transplant

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Summary:Relapse postautograft in acute myeloid leukaemia (AML), may in part arise from leukaemia cells present in the bone marrow (BM) inoculum, and the level of minimal residual disease (MRD) in BM harvests used for autografting may therefore be clinically important.We have used the WT1 transcript as a marker of MRD, which was quantitated by RQ-PCR, in the BM harvests of 24 patients receiving an ABMT for AML. ABL was used as a control gene with WT1 level being normalised to 10 5 copies of ABL per sample. Median WT1 level was 651 copies (range ¼ 113-32 700) for the 13 patients with relapse-free survival (RFS) of less than 5 years, and 174 (range ¼ 0-1900) for patients with RFS of over 5 years postautograft (Po0.04). The RFS was 10.5 months for patients with WT1 level of 42000 copies (n ¼ 5), and has not yet been reached for patients with WT1 levelo2000 (n ¼ 21), at a median follow-up of 92 months (Po0.05). We show that elevated levels of MRD in BM harvests are associated with a higher relapse risk in patients autografted for AML. Autologous bone marrow transplantation (ABMT), as consolidation therapy, can prolong disease-free survival (RFS) for patients with acute myeloid leukaemia (AML) in first CR. 1-3 However, relapse remains the main cause of treatment failure, and may in part arise from leukaemia cells present in the bone marrow (BM) inoculum. 4 Thus, the level of minimal residual disease (MRD) in BM harvests may be relevant in determining the risk of relapse. Methods of detection of MRD with increased sensitivity, such as RQ-PCR are now available and quantitation of WT1 is useful in patients who lack specific fusion transcripts. This gene is overexpressed in over 90% of leukaemia cells and several studies have shown that quantitation of WT1 transcripts for patients in remission has prognostic value. [5][6][7][8] This study was undertaken to measure MRD levels in BM harvests used for autografting in AML, through WT1 quantitation, and to correlate these results with clinical outcome. Materials and methods PatientsBM harvest samples were available from 24 patients who had received an ABMT as consolidation treatment for AML between 1987 and 2000 (Table 1). There were 14 female and 10 male patients, with a median age of 42.5 years (range 16-55). Cytogenetics analysis at diagnosis showed two patients to be in a poor prognostic and 17 in a standard prognostic group using UK MRC criteria. 9 Five patients were in a favourable prognostic group (two with t(15;17), two with inv16, and one with t(8:21)) because the autograft was performed before risk stratification by cytogenetic analysis came into practice. Induction and consolidation treatments were given as per UK MRC 10 and 12 trials and responding patients were autografted after two courses of consolidation treatment. All patients were in morphological and cytogenetic remission at the time of BM harvest and also at autografting. A total of 18 patients were in 1st CR, five in 2nd and one in 3rd CR. Conditioning consisted either of standard Cy/TBI, Bu/Cy, or high-d...

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Section: Discussionsupporting

confidence: 80%

Section: Sample Preparation and Quantitation Of Wt1 Transcriptsmentioning

confidence: 99%

Section: Sample Preparation and Quantitation Of Wt1 Transcriptsmentioning

confidence: 99%

mentioning

confidence: 99%

See 2 more Smart Citations

Elevated levels of WT1 transcripts in bone marrow harvests are associated with a high relapse risk in patients autografted for acute myeloid leukaemia

Osborne

Frost

Tobal

et al. 2005

Bone Marrow Transplant

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“…On the contrary, a greater agreement was found among groups that have used WT1 levels as a marker of minimal residual disease (MRD) in AML remission BMs (less than 5% of blast cells). In particular, WT1 expression has been shown to predict disease progression in AML patients treated with conventional chemotherapy (8)(9)(10)(15)(16)(17) and patients undergone allogeneic stem cell transplantation (allo-SCT) (18)(19)(20)(21)(22). Furthermore, when WT1 expression was compared with widely used techniques in monitoring MRD such as multiparameter flow cytometry (MFC) (23) or specific molecular targets such as fusion genes transcripts (PMLRARa, AML-ETO1, and CBFb-MYH11), comparable sensitivities were found in predicting the relapse in AML.…”

Section: Introductionmentioning

confidence: 99%

Wilms’ Tumor Gene (WT1) Expression and Minimal Residual Disease in Acute Myeloid Leukemia

et al. 2016

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Copyright: The Authors.Licence: This open access article is licensed under Creative Commons Attribution 4.0 International (CC BY 4.0). http://creativecommons.org/licenses/by/4.0/ Users are allowed to share (copy and redistribute the material in any medium or format) and adapt (remix, transform, and build upon the material for any purpose, even commercially), as long as the authors and the publisher are explicitly identified and properly acknowledged as the original source. AbstractThe identification of minimal residual disease (MRD) has led to substantial improvements in early recognition of the recurrence of acute myeloid leukemia (AML). Flow cytometry (FC), real-time quantitative polymerase chain reaction (RQ-PCR) and fluorescence in situ hybridization are useful methods for the detection of MRD in AML patients although molecular monitoring of leukemia-specific rearranged (RUNX1-RUNX1T1 and CBFB-MYH11) or mutated genetic (NPM1, CEBPA) sequences represents the most sensitive methodology. Rossi et al. 274Besides, more than 50% of all AML patients lack one of these specific sequences, so it is crucial to identify molecular targets applicable for the majority of patients. WT1 is overexpressed at the mRNA level in 80-90% of AML cases at diagnosis in both peripheral blood and bone marrow, and is detectable in a consistent low range in normal donors. These features have led to its adoption for MRD detection using RQ-PCR. A European LeukemiaNet Study found the magnitude of WT1 log reduction after induction chemotherapy to be an independent predictor of relapse. Other studies showed a poorer outcome in patients having WT1 levels above reference thresholds at specific time points. WT1 expression was compared with other modalities of MRD assessment, such as RQ-PCR of specific fusion genes and FC, but no differences in terms of predictive value emerged. Finally, some authors translated the use of WT1 in the clinic giving donor lymphocytes infusions to patients with increasing WT1-mRNA levels after allogeneic stem cell transplantation and obtaining an improvement of survival in this subset. Data collected on WT1 expression over the past years provided evidence for the use of this molecular marker to stratify high-risk AML patients. It can also be used as a marker for early interventional therapy, but further studies are needed to demonstrate it.

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WT1 protein expression in childhood acute leukemia

et al. 2007

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In patients with acute leukemia, Wilms' tumor gene 1 (WT1) has been used as a target for the detection of minimal residual disease (MRD) by PCR techniques. The expression of WT1 protein, however, has not been extensively studied. To determine the relation between expression of WT1 transcripts and of the encoded protein, we examined leukemic cell lines and primary childhood leukemia samples using both real-time quantitative PCR (RQ-PCR) and flow cytometry. WT1 protein was highly expressed in the leukemic cell lines K562, HL-60, PLB 985, KG-1a and CEM. By contrast, 40 primary samples of acute lymphoblastic leukemia (ALL; B-ALL, n 5 15 and T-ALL, n 5 10) and acute myeloid leukemia (n 5 15) expressed low levels of WT1 protein. RQ-PCR detected WT1 transcript levels in the same range as reported in earlier studies in childhood acute leukemia. The results of this study indicate the following: (i) there are considerable discrepancies between WT1 transcripts and protein expression; (ii) WT1 is not a suitable marker for flow cytometric MRD detection in childhood acute leukemia. Am. J. Hematol. 83:382-386, 2008. V

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Prognostic significance of quantitative analysis of WT1 gene transcripts by competitive reverse transcription polymerase chain reaction in acute leukaemia

Cited by 74 publications

References 29 publications

Elevated levels of WT1 transcripts in bone marrow harvests are associated with a high relapse risk in patients autografted for acute myeloid leukaemia

Elevated levels of WT1 transcripts in bone marrow harvests are associated with a high relapse risk in patients autografted for acute myeloid leukaemia

Wilms’ Tumor Gene (WT1) Expression and Minimal Residual Disease in Acute Myeloid Leukemia

WT1 protein expression in childhood acute leukemia

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