Cellular Reprogramming Erases Aberrant DNA Methylation and the Malignant Phenotype in Chronic Myeloid Leukemia

Chronic Myeloid Leukemia (CML) is a myeloproliferative disorder characterized by the presence of the Philadelphia chromosome deriving from the genetic translocation t(9;22)(q34;q11.2) that encodes for the BCR-ABL fusion gene. BCR-ABL is a constitutively active tyrosine kinase believed to be the primary genetic event driving CML development (Daley and Baltimore, 1988; Huettner et al., 2000). Tyrosine Kinase Inhibitors (TKI) targeting the BCR-ABL kinase have revolutionized CML therapy, however, they fail to fully eradicate the disease due to the presence of a drug-resistant stem cell pool that sustains continued growth of the malignant cells. Indeed, discontinuation of TKI results in relapse and/or disease progression.

It has been shown that the emergence of leukemic clones resistant to TKI and responsible for CML evolution is correlated with aberrant DNA methylation (Machova Polakova et al., 2013). DNA methylation is a key epigenetic signature implicated in regulation of gene expression (Robertson, 2001), that occurs predominantly within CpG dinucleotides. CpG-rich regions (namely CpG islands) are frequently located within promoter regions (~70%) of human protein-coding genes (Illingworth et al., 2010). Methylation of CpG-rich promoters negatively correlates with gene expression levels and it is considered an important regulatory mechanism for long-term gene silencing (Herman and Baylin, 2003). Although numerous studies have established a link between aberrant promoter DNA methylation and cancer (Costello et al., 2000; Feinberg et al., 2006), the impact of DNA methylation in CML is still poorly understood (Yamazaki et al., 2012), particularly due to the lack of the specific animal models.

In this study we tested the functional relevance of aberrant methylome in CML development and investigated the possibility that BCR-ABL oncogene triggers DNA methylation changes leading to an aggressive leukemic phenotype. To this end, we have combined cellular reprogramming (Amabile and Meissner, 2009; Carette et al., 2010; Kumano et al., 2012; Miyoshi et al., 2010; Takahashi et al., 2007), due to its ability to erase tissue-specific DNA methylation and to re-establish an embryonic stem-like DNA methylation state (Mikkelsen et al., 2008), with a previously developed BCR-ABL inducible murine model (Koschmieder et al., 2005). Using this approach, we demonstrate that the presence of a single genetic aberration is sufficient to trigger DNA methylation changes and leads to an aggressive leukemic phenotype. We show that by resetting the normal DNA methylation profile of primary human CML cells through cellular reprogramming, we are able to re-establish normal myeloid differentiation, despite the persistence of the native genetic lesion. Finally, by combining a comprehensive genome-scale methylation analysis with cellular reprogramming of leukemic cells, we elucidate the sequential events driving leukemogenesis and reveal the reciprocal interplay of genetic and epigenetic mechanisms during malignant transformation.

In conclusion, these results dissect the role of DNA methylation alterations in CML development and warrant an application of demethylating agents such as 5-azacytidine as adjuvant treatment in therapeutic approaches to CML.