Role of Epigenetics in Chronic Myeloid Leukemia

Katerina Machova Polakova 0 1 Jitka Koblihova 0 1 Tomas Stopka 0 1 0 T. Stopka Institute of Pathophysiology, First Faculty of Medicine, Charles University in Prague , U Nemocnice 5, Prague 2 128 53, Czech Republic 1 K. Machova Polakova ( The efficacy of therapeutic modalities in chronic myeloid leukemia (CML) depends on both genetic and epigenetic mechanisms. This review focuses on epigenetic mechanisms involved in the pathogenesis of CML and in resistance of tumor cells to tyrosine kinase inhibitors leading to the leukemic clone escape and propagation. Regulatory events at the levels of gene regulation by transcription factors and microRNAs are discussed in the context of CML pathogenesis and therapeutic modalities. - It has been over half a century since a specific chromosomal aberration the Philadelphia chromosome (Ph chromosome), was found in chronic myeloid leukemia (CML) [1], but still many questions regarding molecular mechanisms of pathogenesis in CML remain unanswered. The CML genetics provided detailed analyses of the Ph chromosome that is a result of a reciprocal translocation t(9;22)(q34;q11) accompanied by breaks on long arms of chromosome 9 (9q34) proximally from the ABL gene and on long arms of chromosome 22(q11) with BCR (Breakpoint Cluster Region) [2]. The hybrid gene BCR-ABL product translates a chimeric protein with strong and constitutive tyrosine kinase activity that phosphorylates target proteins to facilitate expansion of hematopoietic stem and progenitor cells. The end of the last century was marked by significant progress in CML treatment following the introduction of the tyrosine kinase inhibitor (TKI) imatinib (IM) that binds to the kinase domain (KD) of BCR-ABL and inhibits its tyrosine kinase activity [3]. Despite the high efficiency of imatinib therapy, still approximately 30 % of patients develop resistance to imatinib resulting in first line therapy failure. Imatinib resistance due to the mutations in KD of BCRABL can be bypassed by 2nd line TKIs such as dasatinib or nilotinib [4]. Resistance to the 2nd line therapy also develops and not surprisingly is also associated with specific KD mutations [5]. Genetic mechanisms at the level of KD sequence integrity must be important; but other mechanisms of primary or acquired resistance to TKI are also now being studied including additional clonal aberrations, BCR-ABL overexpression, and TKI bioavailability. However, apart from BCR-ABL, there are other genetic or epigenetic alterations that are still unknown as they may contribute to CML stem cells survival during a long term TKI therapy that successfully inhibits the BCR-ABL activity but is not curative. One can imagine that TKI therapy may in the future benefit from being combined with other agents in order to achieve deep and long term molecular responses. Abnormal epigenetic regulation of the expression of CML-associated genes may play a critical role in its pathogenesis and in the mechanisms modulating therapeutic responsiveness. Epigenetics is thought to involve well recognized regulatory mechanisms of gene expression such as DNA methylation or covalent post-translational modifications of histone core proteins that lead to changes in the chromatin accessibility for mRNA transcription regulation [6, 7]. As well as nuclear events, other mechanisms including non-coding RNA-mediated (by microRNAs, miRs) specific mRNA silencing at the levels of translation and RNA stability are also considered to be very powerful epigenetic mediators to modulate CML expression profiles and phenotypic outcomes. MicroRNAs are able to control hundreds of mRNAs and thus they control broad physiological and pathological features including tumor aggressiveness. Unlike mRNAs, microRNAs are stable and therefore can be routinely quantitated and potentially may also serve as disease biomarkers. This review summarizes the role of epigenetics in CML, and focuses on DNA methylation and histone modification as well as post-transcriptional effects of microRNAs in the pathogenesis of CML from diagnosis and throughout treatment. DNA Methylation in CML The methylation of CpG islands is an active enzymatic and transcription-inhibiting control mechanism that balances the levels of gene expression that is frequently dysregulated in hematological malignancies. A large number of genes (mostly tumor suppressors) are inactivated by hypermethylation of CpG islands mainly in the promoter regions while some genes (such as oncogenes) are hypomethylated. This phenomenon has been documented to play a critical role in both solid tumors and leukemias [8]. ABL1 (v-abl Abelson murine leukemia viral oncogene homolog 1) methylation at its Pa promoter represents a likely marker of CML pathogenesis [9, 10]. The frequency of methylation in chronic phase (CP) CML however ranges from 26 % [11] to 77 % [12], 78 % [10] and 81 % [13 ]. Sun et al. [14] confirmed the high incidence of Pa methylation in CP bone marrow (BM) samples in contrast to normal BM. They observed copies of ABL1 promoter Pa to be methylated 2060 % in BM from 7 CP CML patients at diagnosis. No Pa methylation was detected in normal BMs or colonies derived from them. On the other hand, most colonies from CP CML patients were methylated at the Pa. The authors suggested that ABL1 Pa methylation was an early marker of CML in BM. Asimakopoulos et al. [9] demonstrated that in accelerated phase (AP) CML, methylation is likely to be an allele-specific process, since each progenitor cell carries both methylated and unmethylated alleles. This paragraph documents the promising but also quite highly disputed significance of the ABL1 hypermethylation in CML. One of the most frequently studied genes in leukemias is the cell cycle regulating gene p15 (CDKN2B). Cyclindependent kinase (CDK) inhibitor p15 (p15Ink4b) together with CDK4 or CDK6 are known to negatively regulate cyclin D transcription leading to inhibition of cell cycle progression. Abnormal hypermethylation of p15 gene regions has been associated with the disease progression in myelodysplastic syndrome (MDS) [15, 16] and with the poor outcome in acute myelogenous leukemia (AML) [17]. The clinical importance of p15 methylation in AML patients is not conclusive [18, 19]. Similarly, the significance of p15 methylation in CML patients is not fully understood as the p15 promoter in CML patients is hypomethylated [20, 21], while others observed p15 hypermethylation in 18 % and 24 % of patient samples, respectively [12, 22]. To conclude, p15 methylation in CML patients requires additional work to fully understand its clinical relevance. Ras association domain-containing protein 1 (RASSF1A) is another candidate gene for DNA methylation. Avramouli et al. [23] however did not observe in 41 CML patients in different stages of the disease any methylation of RASSF1A that is normally involved in cell cycle control. In contrast, RASSF1A was methylated in the CML-derived K562 cell line. Methyl (...truncated)