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D. Yehudai et al.
ferentiation by Giemsa stain (Figure 4B). Finally, alovudine decreased global DNA methylation (Figure 4C), in keeping with prior studies showing a reduction in methylation cor- relates with differentiation.25-27
Next, we evaluated alovudine in the 8227 primary AML culture model; 8227 leukemia cells are patient-derived cells that are organized into a hierarchy of stem and bulk cells with the stem cells residing in the CD34+CD38– com-
A
partment.28 Treatment of 8227 cells with alovudine decreased the CD34+CD38– stem cells (Figure 4D).
Inhibition of polymerase gamma but not reductions in oxidative phosphorylation or respiratory chain proteins influence acute myeloid leukemia differentiation
We then explored the mechanism by which alovudine promoted AML differentiation. We tested whether reduc-
B
C
D
Figure 4. Alovudine promotes monocytic differentiation in acute myeloid leukemia. (A) OCI-AML2 and MV4-11 cells were treated with increasing concen- trations of alovudine for 10 days. CD11b expression was assessed by flow cytom- etry. (B) MV4-11 cells were treated with alovudine (200 nM) for 9 days. Morphology was assessed by Giemsa stain (magnification 40X, scale bar=60 μm). (C) OCI-AML2 and MV4-11 cells were treated with alovudine for 6 days. Methylation was assessed by dot-blot assay. (D) 8227 cells were treated with increasing concentrations of alovudine for 6 days. CD34+/CD38– expression was assessed by flow cytometry. For all experiments, *P<0.5, **P<0.01, ***P<0.001, and ****P<0.0001 using Dunnett's multiple comparisons test after one-way ANOVA. Data represent the mean+Standard Deviaiton (SD) from one of three representative experiments, except (C) which represent the mean+SEM of average of 3 (MV4-11) or 4 experiments (OCI-AML2).
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haematologica | 2019; 104(5)