RUNX1 Mutations Are the Most Frequent Leukemia Associated Mutations in Congenital Neutropenia Patients

Abstract 7

Congenital neutropenia is a rare inborn heritable disease with a 20% risk to develop acute myeloid leukemia (AML). One of the most frequently mutated genes in de novo as well as in secondary AML is RUNX1. The aim of our study was to investigate whether patients with congenital neutropenia who developed secondary AML (CN/AML) acquire mutations within the RUNX1 gene prior to the development of AML and whether RUNX1 mutations might be involved in leukemogenesis. Mutation analysis was performed using a sensitive next-generation amplicon deep-sequencing assay (454 Life Sciences, Branford, CT) with minimum coverage of at least 200-fold and confirmed by ABI Sanger sequencing. Using these methods we tested 22 patients with CN/AML for the presence or absence of RUNX1 mutations. Fourteen of the 22 patients (63.6%) demonstrated mutations within the RUNX1 gene. The mutation load was between 8 % and 78 % as assessed by the deep-sequencing read counts. Most of the mutations were detected within the Runt homology/DNA binding domain. Four patients have even two RUNX1 mutations. The RUNX1 mutations consisted of 11 missense mutations, 2 nonsense mutations, 4 frameshift mutations, and one ins/del mutation. Of the 14 patients with RUNX1 mutations 12 were tested positive for inherited congenital neutropenia causing mutations: 10 ELANE mutations, one HAX1 mutation, and one G6PT (SLC37A4) mutation. Cooperating mutations were detected in 8 patients, 7 with CSF3R mutations, one with FLT3-ITD mutation. Cytogenetic analysis of the 14 patients with RUNX1 mutations revealed that 4 patients had monosomy 7 and 3 patients trisomy 21. Sequential analysis of the time prior to overt AML revealed one patient harboring RUNX1 mutation already at birth and a second patient had a RUNX1 mutation 6 months prior to AML. The time course of acquisition of RUNX1 mutations in the remaining 12 patients is presently under investigation. The incidence of RUNX1 and CSF3R mutations in at least 7 patients clearly suggests that these two mutations cooperate in causing leukemia. In conclusion, these data show that patients suffering from congenital neutropenia frequently acquire mutations within the RUNX1 gene prior to development of leukemia.

We thank the Investigators of the Severe Chronic Neutropenia International Registry for supplying us with patient material and clinical data.