Secondary mutations in t(4;11) leukemia patients
Authors: C Prelle, A Bursen, T Dingermann, R Marschalek
Journal: Leukemia (2012)

MLL rearrangements are a genetic hallmark of acute leukemia patients, which exhibit a particular poor outcome.To date, more than 70 MLL rearrangements have been described at the molecular level. 1 For the most frequently diagnosed MLL rearrangements, for example, AF4-MLL, MLL-AF9, MLL-AF10 or MLL-ENL, it has been shown that these fusion proteins are sufficient for acute leukemia onset in murine model systems. 2 However, these models had a latency time of 4-12 months for the disease phenotype to become overt.This argues in favor of preleukemic clones that carry the fusion genes, but need, in addition, complementing mutations to develop a malignant disease.In case of MLL rearrangements-specifically MLL-AF4-it has been shown that copy number aberrations could not be detected through whole genome high-resolution technologies, 3 favoring the possibility that single nucleotide activating mutations, such as mutated Fms-like tyrosine kinase 3 (FLT3) or RAS, are more important than previously reported. 4-6Therefore, we investigated the incidence of RAS and FLT3 mutations in a large cohort of MLLrearranged leukemia patients (n ¼ 144).This cohort of MLLassociated leukemias was split into three subgroups.The t(4;11) group, which was of major interest includes 21 t(4;11) patients displaying complex genomic rearrangements (42 fusions alleles; 11 pediatric, 10 adult), whereas 79 t(4;11) patients had a balanced translocation (31 pediatric, 48 adult).The control group of 44 patients comprised cases with MLL-AF9 (n ¼ 22; 19 pediatric, 3 adult), MLL-ENL (n ¼ 14; 13 pediatric, 1 adult), MLL-AF10 (n ¼ 4; 4 pediatric), MLL-ELL (n ¼ 2; 2 pediatric) and, finally, 1 case with MLL-AF6 (adult) and one with MLL PTD (adult), respectively.All patient material has been made available by the I-BFM and German multicenter acute lymphoblastic leukemia (ALL) study groups.Genomic DNA of these patients was investigated by amplifying the corresponding genomic regions of the N-RAS and K-RAS genes (exons 2 and 3), and of the FLT3 receptor gene (exons 13-15 and exon 20) and, subsequently, by sequencing the resulting amplimers.FLT3 is predominantly expressed in hematopoietic stem/progenitor cells.In addition, FLT3 is expressed at considerable levels in most clinical samples from acute myeloid leukemia (AML) and B-cell precursor ALL patients.Internal tandem duplication of the juxtamembrane domain of the FLT3 gene is frequently observed in AML.AML patients bearing a particular FLT3 mutation (D835; FLT3-TKD) tend to have poor prognosis, suggesting that FLT3-TKD mutations have an important role in AML. 7Otherwise, it has been published that FLT3-TKD mutations were found in approximately 15% of patients with de novo mixedlineage leukemia (MLL) rearrangements. 5However, these patients were screened by Southern blot experiments, without gaining information about the involved MLL fusion partner.In the cohort of 144 MLL-rearranged leukemia samples we analyzed, none of the investigated patients-regardless of whether diagnosed with balanced, complex t(4;11) rearrangements or other MLL fusions-displayed a genomic FLT3 mutation.This is in line with data reported in the literature that mutant FLT3 is mostly diagnosed in hyperdiploid leukemia cases. 8he reason for the absence of mutant FLT3 receptor genes in MLL-rearranged leukemia might be that a mutant FLT3 receptor gene confers a selective disadvantage for MLL-rearranged cells.Published data demonstrates that MLL-rearranged leukemia cells seem to overexpress wild-type FLT3 receptor and that stimulation