WT1 protein expression in slowly proliferating myeloid leukemic cell lines is scarce throughout the cell cycle with a minimum in G0/G1 phase.

Wilms' tumor gene 1 (WT1) is overexpressed in various hematological malignancies and has been proposed as a target for minimal residual disease (MRD) detection and for immunotherapy. Although WT1 is known as a key molecule for tumor cell proliferation, the expression pattern of WT1 in leukemic cells in dependency of proliferation has not yet been investigated. Furthermore, WT1 expression was mostly studied by reverse transcriptase PCR and the expression of WT1 protein has not been extensively studied. Here, we analyzed WT1 protein expression in the human myeloid leukemia cell lines K562 and HL-60 by indirect immunofluorescence and flow cytometry. Both cell lines exhibited varying nuclear WT1 immunoreactivity pointing to a cell cycle-dependent and/or proliferation-dependent WT1 expression. In rapidly proliferating cells high levels of WT1 protein were detected by flow cytometry. A reduced proliferation rate was associated with a low WT1 protein expression and an accumulation of cells in G(0)/G(1) phase. During G(0)/G(1) phase cells expressed WT1 at a lower level than in S or G(2)/M phase. Moreover, WT1 expression was diminished in all cell cycle phases in slowly proliferating cells. We conclude that WT1 protein expression is dependent on the cell cycle phase as well as on the proliferation rate. This finding might be relevant for MRD studies and immunotherapeutic strategies targeting WT1.

WT1 protein expression in childhood acute leukemia.

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 = 15 and T-ALL, n = 10) and acute myeloid leukemia (n = 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.