DNMT3A mutants provide proliferating advantage with augmentation of self-renewal activity in the pathogenesis of AML in KMT2A-PTD-positive leukemic cells

Rabindranath Bera,Yun-Shien Lee,Gang Huang,Ying-Jung Huang,Lee-Yung Shih,Ming-Chun Chiu

doi:10.1038/s41389-020-0191-6

Abstract

Acute myeloid leukemia (AML) with partial tandem duplication of histone-lysine N-methyltransferase 2A (KMT2A-PTD) is a subtype of AML and is associated with adverse survival, yet the molecular pathogenesis of KMT2A-PTD is not fully understood. DNA methyltransferase 3A (DNMT3A) is mutated in various myeloid neoplasms including AML, especially at the Arg882. Recently, it has been found that DNMT3A mutations frequently coexisted with KMT2A-PTD and are associated with inferior outcomes. We aimed to understand the biological role of DNMT3A mutation in KMT2A-PTD-positive cells. Herein, we found that overexpression of DNMT3A mutants (MT) in KMT2A-PTD-positive EOL-1 cells augmented cell proliferation and clonogenicity. Serial colony replating assays indicated that DNMT3A-MT increased the self-renewal ability of Kmt2a-PTD-expressing mouse bone marrow cells with immature morphology. At 10 months post bone marrow transplantation, mice with the combined Kmt2a-PTD and DNMT3A-MT showed hepatosplenomegaly and leukocytosis with a shorter latency compared to control and DNMT3A-wild-type. Gene expression microarray analyses of bone marrow samples from human AML with KMT2A-PTD/DNMT3A-MT showed a stem cell signature and myeloid hematopoietic lineage with dysregulation of HOXB gene expression. In addition, human bone marrow AML cells carrying KMT2A-PTD/DNMT3A-MT showed abnormal growth and augmented self-renewal activity in primary cell culture. The present study provides information underlying the pathogenic role of DNMT3A-MT with KMT2A-PTD in proliferating advantage with augmentation of self-renewal activity in human leukemia, which may help to better understand the disease and to design better therapy for AML patients with these mutations.

Highlights

Acute myeloid leukemia (AML) is a heterogeneous malignant hematopoietic disorder, which is classified by distinct morphologies, cytogenetics and molecular subtypes[1]
Transduced EOL-1 cells treated with all-trans retinoic acid (ATRA) or Na-butyrate induced morphological change with modest differentiation; very little effect was observed in EOL-1 cells treated with 500 nM suberanilohydroxamic acid (SAHA) (Supplemental Fig. S2a, b)
From gene expression microarray data analyses, we found that several genes including the HOXB cluster were upregulated in KMT2A-PTD AML with DNA methyltransferase 3A (DNMT3A) mutations compared to DNMT3A-WT

Summary

Introduction

Acute myeloid leukemia (AML) is a heterogeneous malignant hematopoietic disorder, which is classified by distinct morphologies, cytogenetics and molecular subtypes[1]. The histone-lysine N-methyltransferase 2A (KMT2A) gene, located on chromosome (11q23), has. Our previous study found that 6% of de novo AML patients had KMT2A-PTD mutation, which was associated with poor outcomes[9]. Using a large cohort of 98 de novo AML patients with KMT2APTD, we reported that 90.8% of patients had at least one additional gene mutation including FLT3-ITD (44.9%), DNMT3A (32.7%), RUNX1 (23.5%) and TET2 (18.4%) mutations[12]. We observed a high frequency of coexistence of DNMT3A mutations with KMT2A-PTD and a poor outcome in patients carrying both mutations[12]. The biological functions of DNMT3A mutation in KMT2A-PTD-positive leukemia cells have not been studied before

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