Pediatric Adult Acute Myeloid Leukemia Research Articles

Prognostic significance of WT1 gene expression in pediatric acute myeloid leukemia

The Wilms Tumor gene (WT1) encodes a transcription factor involved in kidney development and malignancy. WT1 expression in a subpopulation of early CD34+ cells has suggested its involvement in hematopoiesis. WT1 is aberrantly expressed in leukemias. High expression of WT1 at diagnosis has been associated with unfavorable prognosis in adult acute myeloid leukemia (AML). The prognostic relevance of WT1 expression in pediatric AML was evaluated in only one study, including 47 patients, which showed that very low levels of WT1 at presentation were associated with an excellent outcome. To test the validity of these findings we measured levels of WT1 in 41 newly diagnosed pediatric AML of the non-M3 FAB subtype. Patients were treated according to an AML-BFM 83-based protocol in a single institution. Mononucleated cells obtained from presentation BM aspirates were cryopreserved and later thawed and used for total RNA extraction and cDNA synthesis. The quantitative assessment of WT1 transcripts was made by real-time PCR (RQ-PCR). WT1 transcripts values were normalized with respect to the number of ABL transcripts. WT1 levels were significantly higher in patients bearing favorable chromosome abnormalities, t(8;21) and inv(16) (P = 0.002). Higher levels of WT1 expression were unexpectedly associated with a higher probability of overall survival by Cox regression analysis (P = 0.002). Multivariate regression analysis could not discriminate between the effects of WT1 and cytogenetics on survival. Higher WT1 expression was associated with favorable cytogenetics subtypes and accordingly with better outcome in children with AML in this study.

Absence of JAK2 V617F Activating Mutations in Children with Acute Megakaryoblastic Leukemia with and without Down Syndrome.

Activating mutations at codon 617 of the Janus-2 tyrosine kinase (JAK2 V617F) have recently been described in hematological malignancies. In adult acute myeloid leukemia (AML), the reported frequencies vary, and JAK2 V617F mutations have mainly been detected in secondary AML following a myeloproliferative disorder. In adult de novo AML, the mutation was less frequent, and detected in 2/11 (18%) acute megakaryoblastic leukemia (FAB M7) samples (Jelinek et al., Blood 2005), and occasionally in other FAB-types.This prompted us to analyze a cohort of pediatric AML FAB M7 samples for this particular mutation. In children, at least 3 different subsets of AML M7 can be identified: infants with AML M7 characterized by t(1;22)(p13;q13), older children with random cytogenetic aberrations, and myeloid leukemia of Down syndrome (DS ML). DS ML is often preceded by transient myeloproliferative disease (TMD), hence we also screened TMD samples to detect whether JAK2 V617F mutations would be involved in clonal evolution from TMD to DS ML. To exclude germ-line mutations in DS, we tested normal mononuclear bone marrow cells (NBMC) from children with DS. These NBMC were obtained from a sternal aspirate from children undergoing cardiac surgery, after informed consent was obtained. Genomic DNA was harvested from leukemic cells, and JAK2 exon 12, including the intron-flanking regions, was amplified and sequenced to screen for the JAK2 V617F mutation. As a positive control for the JAK2 V617F mutation, we used HEL 92.1.7 cells (an erythroleukemic cell line). In a dilution experiment we could still detect the mutation, using direct sequencing, if 10% HEL/JAK2 mutated cells were mixed with 90% wild-type control cells. We tested 49 samples, comprising of 9 NBMC, 11 TMD, 14 DS-ML M7, 11 non-DS AML M7 and 4 relapsed non-DS AML M7 samples (including 2 initial diagnosis-relapse pairs). The median age of the TMD cohort was 3 days, for DS-ML children 1.9 years (range 0.9–3.8 yrs), and for non-DS AML 1.5 years (range 1.2–13.7 yrs). The median white blood cell count for TMD was 25.8x109/l, for DS-ML 13.8x109/l, and for non-DS AML 12.4x109/l. Cytogenetic data were available in 5/11 non-DS AML cases only, which showed no cases with a t(1;22). No JAK2 V617F mutations were detected in any of the clinical samples. We conclude that the role of JAK2 V617F mutations in pediatric DS and non-DS acute megakaryoblastic leukemia is limited at best. However, we were not able to screen the subgroup of non-DS AML cases with t(1;22).

T(9;11)(p22;p15) with NUP98-LEDGF fusion gene in pediatric acute myeloid leukemia

The rare t(9;11)(p22;p15) translocation is associated with adult acute myeloid leukemia (AML) with immature forms. We report a novel fusion of the NUP98 and LEDGF genes in a pediatric AML with intermediate characteristics between M2-M3 French-American-British (FAB) subtypes exhibiting the same chromosomal rearrangement. Fluorescence in situ hybridization (FISH) and reverse transcriptase-PCR (RT-PCR) studies identified the chimeric transcript product of in-frame fusion of NUP98 exon 8 to LEDGF exon 4.

High Expression Levels of Iaps Predict Poor Overall Survival in Childhood De Novo Acute Myeloid Leukemia.

Apoptosis-related proteins are important molecules for predicting chemotherapy response and prognosis in adult acute myeloid leukemia (AML). However, data on the expression and prognostic impact of these molecules in childhood AML are rare. Using flow cytometry and western blot analysis, we therefore investigated 45 leukemic cell samples of children with de novo AML enrolled and treated within the German AML-BFM93 study for the expression of apoptosis-regulating proteins (CD95, Bcl-2, Bax, Bcl-xL, Procaspase-3, XIAP, cIAP-1, Survivin). XIAP (p<0.002) but no other apoptosis regulators showed maturation-dependent expression differences as determined by FAB morphology with the highest expression levels observed within the immature M0/1 subtypes. XIAP (p<0.01) and Bcl-xL (p<0.01) expression was lower in patients with favorable than intermediate/poor cytogenetics. After a mean follow-up of 34 months, a shorter overall survival was associated with high expression levels of XIAP {30 (n=10) vs. 41 months (n=34); p<0.05} and Survivin {27 (n=10) vs. 41 months (n=34); p<0.05}. We conclude that apoptosis-related molecules are associated with maturation stage, cytogenetic risk groups and therapy outcome in childhood de novo AML. The observed association of XIAP with immature FAB types, intermediate/poor cytogenetics and poor overall survival should be confirmed within prospective pediatric AML trials.

High expression levels of x-linked inhibitor of apoptosis protein and survivin correlate with poor overall survival in childhood de novo acute myeloid leukemia.

Apoptosis-related proteins are important molecules for predicting chemotherapy response and prognosis in adult acute myeloid leukemia (AML). However, data on the expression and prognostic impact of these molecules in childhood AML are rare. Using flow cytometry and Western blot analysis, we, therefore, investigated 45 leukemic cell samples from children with de novo AML enrolled and treated within the German AML-BFM93 study for the expression of apoptosis-regulating proteins [CD95, Bcl-2, Bax, Bcl-xL, procaspase-3, X-linked inhibitor of apoptosis protein (XIAP), cellular inhibitor of apoptosis protein-1 (cIAP-1), survivin]. XIAP (P < 0.002) but no other apoptosis regulators showed maturation-dependent expression differences as determined by French-American-British (FAB) morphology with the highest expression levels observed within the immature M0/1 subtypes. XIAP (P < 0.01) and Bcl-xL (P < 0.01) expression was lower in patients with favorable rather than intermediate/poor cytogenetics. After a mean follow-up of 34 months, a shorter overall survival was associated with high expression levels of XIAP [30 (n = 10) versus 41 months (n = 34); P < 0.05] and survivin [27 (n = 10) versus 41 months (n = 34); P < 0.05]. We conclude that apoptosis-related molecules are associated with maturation stage, cytogenetic risk groups, and therapy outcome in childhood de novo AML. The observed association of XIAP with immature FAB types, intermediate/poor cytogenetics, and poor overall survival should be confirmed within prospective pediatric AML trials.