Characterization of CEBPA mutations and promoter hypermethylation in pediatric acute myeloid leukemia

Abstract

The online version of this article has a Supplementary Appendx. BackgroundDysfunctioning of CCAAT/enhancer binding protein α (C/EBPα) in acute myeloid leukemia can be caused, amongst others, by mutations in the encoding gene (CEBPA) and by promoter hypermethylation. CEBPA-mutated acute myeloid leukemia is associated with a favorable outcome, but this may be restricted to the case of double mutations in CEBPA in adult acute myeloid leukemia. In pediatric acute myeloid leukemia, data on the impact of these mutations are limited to one series, and data on promoter hypermethylation are lacking. Our objective was to investigate the characteristics, gene expression profiles and prognostic impact of the different CEBPA aberrations in pediatric acute myeloid leukemia. Design and MethodsWe screened a large pediatric cohort (n=252) for CEBPA single and double mutations by direct sequencing, and for promoter hypermethylation by methylation-specific polymerase chain reaction. Furthermore, we determined the gene-expression profiles (Affymetrix HGU133 plus 2.0 arrays) of this cohort (n=237). ResultsThirty-four mutations were identified in 20 out of the 252 cases (7.9%), including 14 doublemutant and 6 single-mutant cases. CEBPA double mutations conferred a significantly better 5-year overall survival compared with single mutations (79% versus 25%, respectively; P=0.04), and compared with CEBPA wild-type acute myeloid leukemia excluding core-binding factor cases (47%; P=0.07). Multivariate analysis confirmed that the double mutations were an independent favorable prognostic factor for survival (hazard ratio 0.23, P=0.04). The combination of screening for promoter hypermethylation and gene expression profiling identified five patients with silenced CEBPA, of whom four cases relapsed. All cases characteristically expressed T-lymphoid markers. Moreover, unsupervised clustering of gene expression profiles showed a clustering of CEBPA double-mutant and silenced cases, pointing towards a common hallmark of abrogated C/EBPα-functioning in these acute myeloid leukemias. ConclusionsWe showed the independent favorable outcome of patients with CEBPA double-mutant acute myeloid leukemia in a large pediatric series. This molecular marker may, therefore, improve risk-group stratification in pediatric acute myeloid leukemia. For the first time, CEBPA-silenced cases are suggested to confer a poor outcome in pediatric acute myeloid leukemia, indicating that further investigation of this aberration is needed. Furthermore, clustering of gene expression profiles provided insight into the biological similarities and diversities of the different aberrations in CEBPA in pediatric acute myeloid leukemia.Key words: Pediatric acute myeloid leukemia, CEBPA mutation, promoter hypermethylation, molecular marker, prognostic significance. Citation

Normal karyotype is a poor prognostic factor in myeloid leukemia of Down syndrome: a retrospective, international study

“…Non-DS pediatric NK AML cases are characterized by various abnormalities, including overexpression of specific genes (MN1, BAALC, and ERG), 29 but also single gene mutations such as FLT3-ITD, WT1, NPM1, and CEBPA, [30][31][32] as well as cryptic translocations. 33 We recently showed that the abnormalities mentioned above are absent or rare in (NK-) ML-DS.…”

High GATA2 expression is a poor prognostic marker in pediatric acute myeloid leukemia

“…The independence of prognostic factors was examined by multivariate Cox regression analysis, including in pediatric AML-established prognostic factors such as age, WBC count at diagnosis, favorable karyotype, NPM1 mutation, CEBPA double mutation (CEBPA-DM), and FLT3-ITD. 45 Age and WBC count were included as continuous variables. Favorable karyotype was defined as patients harboring t(15;17)(q21;q22), inv(16)(p13q22), or t(8;21)(q22;q22).…”

Pediatric Acute Myeloid Leukemia

“…Type 1 abnormalities mainly induce proliferation, and consist for instance of mutations in tyrosine kinase receptors such as the FLT3-gene (Zwaan, et al 2003a) or KITmutations (Goemans, et al 2005, Pollard, et al 2010, and type 2 abnormalities induce maturation arrest and mainly result from genetic aberrations in hematopoietic transcription factors, either resulting from translocations, or from mutations in genes such as NPM1, GATA1 and CEBPA. (Ahmed, et al 2004, Hollink, et al 2011, Hollink, et al 2009c Evidence for this model is supported by several factors: 1) AML-specific translocations can already be demonstrated in cord-blood (Wiemels, et al 2002), and may only cause AML several years later, 2) fusion transcripts may be demonstrated using sensitive techniques in patients in long-term clinical remission of AML (Leroy, et al 2005), 3) FLT3 mutations induce a myeloproliferative disorder in mice but lack the maturation arrest typical of full-blown AML (Kelly, et al 2002b), and 4) certain type I and II genetic aberrations cluster together in a non-random fashion. Conventional karyotyping may identify AML-specific abnormalities, which are not only of use in diagnosis and the correct classification of the leukemia, but may also provide prognostic information used for risk-group stratification of pediatric AML.…”

“…(Balgobind, et al 2011a, Marcucci, et al 2011 NPM1 and CEBPA gene mutations confer good clinical outcome, whereas mutations in the FLT3 and WT1-genes confer poor clinical outcome. (Ho, et al 2009, Ho, et al 2010b, Hollink, et al 2011, Hollink, et al 2009a, Hollink, et al 2009c, Zwaan, et al 2003a Figure 2 shows the distribution of type 1 and 2 abnormalities, as identified in >400 cases of pediatric AML. We have arbitrarily included the WT1 mutations as type I aberrations, however, their role in AML still has to be elucidated.…”

“…(Balgobind, et al 2011a) The NPM1 and CEBPA double mutations confer good clinical outcome, allowing risk-stratification with the "good risk" cytogenetic subgroups. (Brown, et al 2007, Ho, et al 2009, Hollink, et al 2011, Hollink, et al 2009c In addition, the following type-1 mutations were identified: FLT3-internal tandem duplications (FLT3-ITD), found in ~30-40% of cases, FLT3-tyrosine kinase domain mutations (FLT3-TKD) in ~2% and N-or K-RAS mutations in ~15-20% of CN-AML cases. (Balgobind, et al 2011a, Goemans, et al 2005 WT1 mutations were found in 20-25% of pediatric CN-AML cases, in approximately half of the cases together with a FLT3-ITD, and in a quarter together with a RAS-mutation.…”