RUNX1 mutations in cytogenetically normal acute myeloid leukemia are associated with a poor prognosis and up-regulation of lymphoid genes

Abstract

Background: The RUNX1 (AML1) gene is a frequent mutational target in myelodysplastic syndromes and acute myeloid leukemia. Previous studies suggested that RUNX1 mutations may have pathological and prognostic implications.

Design and methods: We screened 93 patients with cytogenetically normal acute myeloid leukemia for RUNX1 mutations by capillary sequencing of genomic DNA. Mutation status was then correlated with clinical data and gene expression profiles.

Results: We found that 15 out of 93 (16.1%) patients with cytogenetically normal acute myeloid leukemia had RUNX1 mutations. Seventy-three patients were enrolled in the AMLCG-99 trial and carried ten RUNX1 mutations (13.7%). Among these 73 patients RUNX1 mutations were significantly associated with older age, male sex, absence of NPM1 mutations and presence of MLL-partial tandem duplications. Moreover, RUNX1-mutated patients had a lower complete remission rate (30% versus 73% P=0.01), lower relapse-free survival rate (3-year relapse-free survival 0% versus 30.4%; P=0.002) and lower overall survival rate (3-year overall survival 0% versus 34.4%; P<0.001) than patients with wild-type RUNX1. RUNX1 mutations remained associated with shorter overall survival in a multivariate model including age and the European Leukemia Net acute myeloid leukemia genetic classification as covariates. Patients with RUNX1 mutations showed a unique gene expression pattern with differential expression of 85 genes. The most prominently up-regulated genes in patients with RUNX1-mutated cytogenetically normal acute myeloid leukemia include lymphoid regulators such as HOP homeobox (HOPX), deoxynucleotidyltransferase (DNTT, terminal) and B-cell linker (BLNK), indicating lineage infidelity.

Conclusions: Our findings firmly establish that RUNX1 mutations are a marker of poor prognosis and provide insights into the pathogenesis of RUNX1 mutation-positive acute myeloid leukemia.

Figure 1.

(A) Overview of mutations in RUNX1. Linear structure of the RUNX1 protein (NP_001745.2) includes the N-terminal RUNT domain and the C-terminal transcriptional activation domain (TAD). Amino acid (aa) changes resulting from mutations found in our cohort of CN-AML patients are detailed. The graph was generated using the software DOG 2.0. (B) Distribution of mutations in RUNX1 and eight additional genes in 93 CN-AML patients. Additional mutations are shown for patients with RUNX1 mutations (n=15) or wild-type RUNX1 (n=78). Seventy-three CN-AML patients were enrolled in the AMLCG-99 clinical trial (left panel). Another 20 CN-AML patients were not homogenously treated (right panel). Genes analyzed for mutations are indicated on the left side.

Figure 2.

Influence of RUNX1 mutations on clinical outcome. Kaplan-Meier estimates for intensively treated CN-AML patients with or without RUNX1 mutations. The censored patient in the RUNX1 mutated group experienced a relapse and was then lost to follow up. (A) The median overall survival in RUNX1 mutated patients was 75 days compared to 442 days for patients with RUNX1 wild-type status. (B) For ELN Intermediate I patients (CN-AML with wild-type CEBPA and wild-type NPM1 and/or FLT3-ITD) median overall survival in RUNX1 mutated patients was 75 days compared to 293 days for patients without this mutation. (C) In elderly AML patients (≥ 60 years) the median overall survival for RUNX1-mutated patients was 86 days and 432 days for patients with wild-type RUNX1.

Figure 3.

Heatmap of genes differentially expressed between RUNX1-mutated and RUNX1-wild-type patients. Each column represents one of 41 CN-AML patients, grouped according to RUNX1 mutation status, and each row represents one of 85 genes that were differentially expressed. Yellow indicates high and blue indicates low gene expression.