High expression of WT1 gene in acute myeloid leukemias with more predominant WT1+17AA isoforms at relapse.
ABSTRACT Real-time quantitative reverse transcriptase polymerase chain reaction method was established for detecting the expression levels of WT1 gene and WT1+17AA isoforms in 226 acute myeloid leukemia (AML) bone marrow (BM) cells. The results showed that WT1 gene was 2-3 logarithms expressed more in AML BM cells at initial diagnosis or relapse than in normal BM cells (p<0.001), with predominant WT1+17AA isoforms expression (the ratio of WT1+17AA/WT1 more than 0.50). Interestingly the ratio of WT1+17AA/WT1 was statistically higher in relapsed AMLs than in initially diagnosed (p=0.01), speculating that WT1+17AA isoforms might participate in AML relapse.
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ABSTRACT: Wilms' tumor gene 1 (WT1) functions including some contradictory effects may be explained by the presence and interactions of its isoforms, however, their evaluation has been so far complicated by several technical problems. We designed unique quantitative PCR systems for direct quantification of the major WT1 isoforms A[EX5-/KTS-], B[+/-], C[-/+] and D[+/+] and verified their sensitivity, specificity and reproducibility in extensive testing. With this method we evaluated WT1 total and isoform expression in 23 normal bone marrow (BM) samples, 73 childhood acute myeloid leukemia (AML), 20 childhood myelodysplastic syndrome (MDS), 9 childhood severe aplastic anemia (SAA), 30 adult AML and 29 adult MDS patients. WT1 isoform patterns showed differences among these samples and clustered them into groups representing the specific diagnoses (P<0.0001). Isoform profiles were independent of total WT1 expression and possess certain common features-overexpression of isoform D and EX5[+] variants. The KTS[+]/KTS[-] ratio was less variable than the EX5[+]/EX5[-] ratio and differed between children and adults (P<0.001); the EX5[+]/EX5[-] ratio varied between diagnoses (AML vs MDS, P<0.001). These findings bring new insights into WT1 isoform function and suggest that the ratio of WT1 isoforms, particularly EX5 variants, is probably crucial for the process of malignant transformation.Leukemia: official journal of the Leukemia Society of America, Leukemia Research Fund, U.K 03/2012; 26(9):2086-95. · 10.16 Impact Factor
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ABSTRACT: Wilms' tumour is a paediatric malignancy of the kidneys and is one of the most common solid childhood cancers. The Wilms' tumour 1 protein (WT1) is a transcription factor that can either activate or repress genes involved in growth, apoptosis and differentiation. It is frequently mutated or aberrantly expressed in Wilms' tumour, where the wild type protein would normally act as a tumour suppressor. Several studies, however, have found that wild type WT1 acts as an oncogene in adult tumours, primarily through the inhibition of apoptosis. The expression of WT1 correlates with the aggressiveness of several adult cancers, and its continued expression following treatment is indicative of a poor outcome.We recently found that the treatment of tumour cell lines with cytotoxic drugs leads to the cleavage of WT1 by the serine protease HtrA2. HtrA2 binds to a specific region of WT1, the suppression domain, and then cleaves WT1 at multiple sites. The HtrA2-mediated proteolysis of WT1 leads to its removal from gene promoter regions and changes in gene expression. Cleavage of WT1 by HtrA2 enhances apoptosis. This event is advantageous to the treatment of adult tumours where WT1 acts as an oncogene. However, when WT1 is acting as a tumour suppressor in paediatric malignancies, proteolysis by HtrA2 would be antagonistic to therapy.Cell cycle (Georgetown, Tex.) 07/2010; 9(13):2508-14. · 5.24 Impact Factor
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ABSTRACT: Early stage detection of cancer is the key to provide a better outcome for therapeutic intervention. Most routine screening tools for cancer detection are largely based on examination of cell morphology, tissue histology and measurement of serum markers, which lack sufficient sensitivity and/or specificity for early detection of cancer. Indeed, most secreted proteins studied as cancer screening biomarkers have low sensitivity and/or low specificity and this could be due to the use of non-sensitive techniques or due the fact that several of these tumor markers are also produced by normal tissues. Altogether, these facts pinpoint to the urgent need for the discovery of innovative tools and novel tumour markers for cancer screening, diagnosis, and prognosis. Recently, scientists and clinicians have shifted to innovative techniques in order to identify and characterize biomarkers that drive the development and progression of cancer, and to discover upstream genes/proteins which could be useful to detect early-stage cancer, predict prognosis, determine therapy efficacy, or to be novel drug targets. This chapter contains three sections that will discuss different but complementary innovative techniques that are being developed for the detection of early biomarkers in cancers. The first technique will focus on the use of Reverse Transcriptase-Polymerase Chain Reaction (RT-PCR) for the quantitation of the Wilms' tumor gene (WT1) mRNA (WT1 Assay). The WT1 mRNA is a relatively new marker of several types of leukemia and myelodysplastic syndrome (MDS). This WT1 assay makes it possible to rapidly assess the effectiveness of treatment and to evaluate the degree of eradication of leukemic cells as well as the continuous assessment of the MDS progression and its evolution to overt acute myeloid leukemia. The second section will highlight the use of Mass Spectrometry (MS) technique as an important analytical tool in clinical proteomics, primarily in the disease-specific discovery, identification and characterization of proteomic biomarkers and patterns. MS-based proteomics is increasingly being used in clinical validation and diagnostic method development. In this section, we will describe the current state of MS in clinical proteomics applied to early detection of cancer biomarkers with a focus on ovarian and breast cancers including both biomarker discovery and clinical diagnosis. In the last section we will focus on the Surface Plasmon Resonance (SPR) technique which is primarily used to detect bimolecular interaction and has recently gained enormous interest and popularity for its versatility and high sensitivity especially when it is coupled to MS.Cancer Biomarkers: Non-Invasive Early Diagnosis and Prognosis, Edited by Debmalya Barh, Angelo Carpi, Mukesh Verma, and Mehmet Gunduz, 10/2013: chapter 4: pages 900; Taylor & Francis/CRC Press., ISBN: 9781466584280