Molecular alterations and clonal evolution in acute myeloid leukemia : prognostic and therapeutic implications

Acute myeloid leukemia (AML) is a clonal disorder of hematopoietic stem cells characterized by inhibition of differentiation, subsequent accumulation of incomplete matured cells at various stages and reduced production of healthy hematopoietic elements. Despite significant progress, the outcome of AML is variable and often suboptimal. Many patients will ultimately relapse and approximately 60% of patients will succumb to their disease. Deep understanding of the leukemogenesis and progression of AML and searching for new prognosis markers, risk stratification as well as therapeutic targets are of great importance to scientists, clinicians and of course future patients.

In the first paper, we conducted a study on 48 de novo AML patients and found downregulation of miR-370 expression as a frequent event. Ectopic expression of miR-370 in two AML cell lines led to cell growth arrest and senescence, while depletion of miR-370 expression enhanced the proliferation of those leukemic cells. Mechanistically, miR-370 targeted the transcription factor FoxM1, a well-established oncogenic factor promoting cell cycle progression. The treatment of AML cells with 5-aza-2’-deoxycytidine (5-aza-CdR), a DNA methylation inhibitor, led to the up-regulation of miR-370 expression, which indicates epigenetic silencing of miR-370 in leukemogenesis. In conclusion, miR-370 acts as a tumor suppressor in AML by targeting FoxM1.

Acute promyelocytic leukemia (APL) is a distinct subtype of AML characterized by the balanced reciprocal translocation t(15;17)(q22;q12-21) that encodes a fusion protein PMLRARα. In study II, we determined the clonal evolution scenario in an APL patient who presented the same disease after 17 years to distinguish between a very late relapse and newly developed de novo APL. The patient APL cells carried the identical PML-RARa fusion gene between two occasions, however, exhibited significant other genetic alterations. FLT3ITD and FLT3D835 mutations were observed in the first and second APL cells, respectively. Thus, the patient experienced a very true late relapse of the disease. The data also suggest that PMLRARa fusion-mediated APL development needs a second oncogenic event (FLT3 mutations in the present case).

DNA methyltransferase inhibitors such as 5-azacytidine (5-AZA) have been used for the treatment of AML and other malignancies. In study III, we identified that 5-AZA induced DNA damage, telomere dysfunction and telomerase inhibition in AML cells, which was coupled with cellular apoptosis. Telomerase over-expression significantly attenuated 5-AZA-mediated DNA damage, telomere dysfunction and apoptosis of AML cells. Collectively, 5-AZA-mediated telomere dysfunction contributes to its anti-cancer activity.

Somatic mutation of FMS-like tyrosine kinase 3 (FLT3) occurs in 30% of AML, with the majority of mutations exhibiting internal tandem duplication (ITD) in the juxtamembrane domain to drive leukemogenesis. In study IV, we observed that, in FLT3ITD-harboring primary cells from AML patients and AML cell lines, FLT3 inhibitor PKC412 down-regulated telomerase (TERT) gene expression and telomerase activity in a MYC-dependent manner. This effect was required for its optimal anti-AML efficacy. Ectopic expression of TERT significantly attenuated the apoptotic effect of PKC412 on AML cells. Mechanistically, TERT enhanced the activity of FLT3 downstream effectors or alternative tyrosine kinase receptor signaling pathways through which PKC412 effect was attenuated. In conclusion, FLT3ITD regulates TERT expression via a MYC-dependent manner, and TERT plays an important role in FLT3 inhibitor-mediated anti-AML efficacy.

List of scientific papers

I. Zhang X, Zeng J, Zhou M, Li B, Zhang Y, Huang T, Wang L, Jia J, Chen C. The tumor suppressive role of miRNA-370 by targeting FoxM1 in acute myeloid leukemia. Mol Cancer. 2012; 11:56.
https://doi.org/10.1186/1476-4598-11-56

II. Zhang X, Zhang Q, Dahlström J, Tran AN, Yang B, Gu Z, Ghaderi M, Porwit A, Jia J, Derolf Å, Xu D, Björkholm M. Genomic analysis of the clonal origin and evolution of acute promyelocytic leukemia in a unique patient with a very late (17 years) relapse. Leukemia. 2014; 28, 1751-1754
https://doi.org/10.1038/leu.2014.113

III. Zhang X, Li B, Jonge N, Björkholm M and Xu D. The DNA methylation inhibitor induces telomere dysfunction and apoptosis of leukemia cells that is attenuated by telomerase over-expression. [Manuscript]

IV. Zhang X, Li B, Dahlström J, Tran AN, Jia J, Björkholm M, Xu D. Functional interplay between FLT3-ITD and telomerase reverse transcriptase (hTERT) in acute myeloid leukemia (AML). [Manuscript]