Date Published: March 23, 2014
Publisher: Hindawi Publishing Corporation
Author(s): Emma Conway O’Brien, Steven Prideaux, Timothy Chevassut.
Acute myeloid leukemia (AML) is a genetically heterogeneous disease. Certain cytogenetic and molecular genetic mutations are recognized to have an impact on prognosis, leading to their inclusion in some prognostic stratification systems. Recently, the advent of high-throughput whole genome or exome sequencing has led to the identification of several novel recurrent mutations in AML, a number of which have been found to involve genes concerned with epigenetic regulation. These genes include in particular DNMT3A, TET2, and IDH1/2, involved with regulation of DNA methylation, and EZH2 and ASXL-1, which are implicated in regulation of histones. However, the precise mechanisms linking these genes to AML pathogenesis have yet to be fully elucidated as has their respective prognostic relevance. As massively parallel DNA sequencing becomes increasingly accessible for patients, there is a need for clarification of the clinical implications of these mutations. This review examines the literature surrounding the biology of these epigenetic modifying genes with regard to leukemogenesis and their clinical and prognostic relevance in AML when mutated.
Acute myeloid leukemia (AML) is a genetically heterogeneous disease characterized by malignant clonal proliferation of immature myeloid cells in the bone marrow, peripheral blood, and occasionally other body tissues [1, 2]. It is the most common acute leukemia in adults and encompasses 15–20% of cases in children . While the disease is most commonly found in individuals over 60 years, AML also occurs in younger people and occasionally may even be present at birth [1, 2]. Environmental factors that increase the risk of developing AML include smoking, benzene exposure, and chemotherapy or radiotherapy treatment [1, 2]. Preceding myelodysplastic syndrome (MDS) or myeloproliferative neoplasm (MPN) may also develop into AML . Although highly variable, the outlook for most AML subtypes is dismal, with an overall 5-year survival rate of approximately 25% . The genetic and epigenetic profile of the malignant cells influences the likelihood of achieving remission and risk of relapse . A greater understanding of the underlying genetic and epigenetic processes may provide insight into the mechanism of leukemogenesis in AML, as well as offering prognostic information and potential therapeutic targets. The prognostic implications of many molecular mutations in AML are well reported . However, the role of mutations in genes with epigenetic function is less clearly understood [6–8]. This literature review, therefore, aims to examine the pathological role and prognostic implications of mutations in epigenetic modifying genes.
Many patients with AML will have cytogenetic aberrations which can be detected through karyotyping or fluorescent in situ hybridization (FISH) [9–11]. Risk stratification—into low, intermediate, or high risk groups—can then be carried out according to the cytogenetic profile of the patient [9, 10]. However, there is variation between different cooperative groups as to the correct stratification of different mutations . Furthermore, nearly half of the patients have cytogenetically normal (CN) AML and are ascribed to the intermediate risk category despite significant heterogeneity . It is clear, therefore, that molecular mutational analysis has the potential to improve prognostication stratification systems. Currently, only a limited selection of genetic mutations is included in widely used prognostic stratification models—in the European LeukemiaNet (ELN) system, for example, NPM1, FLT3-ITD, and CEBPα are the only molecular mutations afforded prognostic significance . The World Health Organization has included a provisional entity in its classification system which includes AML with NPM1 and CEBPα mutations . Nonetheless, mutations which are not included in stratification systems may still impact on prognosis. In addition, increasing access to whole genome or exome mutational analysis techniques is yielding a bewildering array of novel mutations associated with AML. Newly diagnosed AML patients and their doctors are therefore likely to be faced with a complex combination of different mutations, with uncertain clinical significance, on genetic analysis.
For many years, the accepted model of leukemogenesis was the “two-hit hypothesis,” which suggested that two different types of genetic mutation were required for malignant transformation of a myeloid precursor [8, 14]. Class I mutations were thought to lead to uncontrolled cellular proliferation and evasion of apoptosis and included mutations conferring constitutive activity to tyrosine kinases or dysregulation of downstream signaling molecules (in genes such as BCR-ABL, Flt-3, c-KIT, and RAS) [8, 14]. Class II mutations, such as the translocations associated with the core-binding factor (CBF) leukemias, were associated with inhibition of differentiation including key transcription factors, such as CBF and retinoic acid receptor alpha (RARα) [8, 14], and proteins that are involved in transcriptional regulation, such as p300, CBP, MOX, TIF2, and MLL [8, 14].
Epigenetic regulation refers to modulation of genetic transcription and expression which does not alter the genetic code . Epigenetic modifications can be transient or physiologically irreversible and play key roles in developmental patterning in the embryo [7, 26]. Following embryogenesis, epigenetic changes continue throughout an organism’s life . The two main mechanisms of epigenetic regulation in the cell are posttranslational histone modifications (see Figure 4), discussed later, and DNA methylation and hydroxymethylation, discussed below [6, 7, 24, 27].
It is evident that methylation patterns play a role in altering expression of genes crucial to leukemogenesis (see Figures 2 and 3). Figueroa et al. carried out DNA methylation profiling of 344 AML samples and found that subjects could be separated into 16 subclasses according to methylation signatures . These subclasses often reflected cytogenetic or molecular subgroups: PML-RARα, CBFβ-MYH11, and RUNX1-RUNX1T1 (AML1-ETO) were each associated with specific methylation signatures. Specific genetic lesions were enriched in further eight groups, while the remaining five groups did not appear to be associated with particular mutations. The finding that AML cases could be separated according to methylation signature, with some clusters highly enriched in specific mutations (t(8;21), inv(16), t(15;17), and 11q23), has been observed in a number of studies [24, 30, 31]. Figueroa et al. found that clinical outcomes could be predicted according to DNA methylation cluster, including the groups without specific mutations . Moreover, cases in clusters enriched for a particular mutation, but not bearing it themselves, shared the prognostic implications of the group as a whole. This was seen in 9 patients classified into one of the CBF leukemia clusters . The groups that were not associated with particular mutations may be reflecting a shared but as yet unknown genetic lesion, or there may be a number of mutations which result in the same epigenetic profile. It is apparent, therefore, that epigenetic changes in leukemic cells occur in a specific and distinct manner—methylation patterns may vary more between subclasses of AML than between AML and controls—and appear to be responsive to overlying genetic mutations .
Histone tail modifications play a key role in epigenetic modulation of gene expression and may include methylation, acetylation, phosphorylation, ADP-ribosylation, and ubiquitination (see Figure 4) [27, 72]. Mechanisms of aberrant histone modification in AML include mutations in genes concerned with polycomb group complexes (PcG), widely considered to be the “bridge” between histone modification and DNA methylation [72, 73]. PcGs maintain stable and heritable transcriptional repression in specific target genes . PcGs are related to body patterning, stem cell renewal, and they also may have pathogenic roles to play in oncogenesis [72, 73]. Genes coding for components of the PcG may be amplified or overexpressed, or the PcG may be “ectopically recruited” to nontarget genes in cancer development . Mutations have been detected in a number of PcG components in myeloid disorders, with some, unexpectedly, conferring a loss of function [27, 73–76].
Recent DNA sequencing studies have facilitated the identification of a hitherto unrecognized class of genetic mutations in AML—mutations in epigenetic modifying genes (see Table 1). The occurrence of mutations in epigenetic modifiers in AML highlights the inadequacy of the “two-hit model” as a mechanistic explanation of leukemogenesis. Mutations in genes concerned with regulation of the epigenome potentially offer a valuable insight into the process of leukemogenesis. These mutations also contribute to the existing body of knowledge that aids risk stratification of AML through molecular and cytogenetic analysis of leukemic cells. Mutations in genes such as TET2, DNMT3A, and ASXL-1 may be associated with a poor prognosis and as such may represent a novel subset of high risk AML which requires more aggressive treatment. The prognostic implications of IDH 1 and 2, and EZH2 mutations are unclear. There is considerable debate about the prognostic implications of various genetic mutations in AML, in part due to the fact that direct comparison between studies is difficult, if not impossible. Patient cohorts frequently vary according to age, type and intensity of therapy, and inclusion of different AML subtypes (e.g., all AML compared with CN-AML). Studies may also vary in their methodology, such as in differences in the subgroup analysis performed or the proportion of patients selected for analysis, which if low (e.g., Marcucci et al. and Ribeiro et al. only analysed 18% and 13% of their cohort resp.) [35, 44] has the potential to introduce an element of selection bias.