Chromosomal translocation 8;21 is present in 10–15% of patients with acute myeloid leukemia and results in the expression of the AML1-ETO (AE) fusion protein. AE expression by itself is not sufficient for malignant transformation of bone marrow progenitor/stem cells, but the nature of the cooperating genetic alterations are unknown. It is unclear whether AE facilitates acquisition of these synergizing events. We have previously shown that retroviral transduction of primary human CD34+ cells with AE causes a dramatic expansion of CD34+ progenitor cells that retain the ability to differentiate into myeloid, lymphoid, and erythroid lineages and engraft NOD/SCID mice. Consistent with mouse studies, these transgenic human cells fail to cause disease in immunodeficient mice or become transformed in vitro. These cells serve as a highly relevant pre-leukemia model for t(8;21) leukemogenesis. Here we demonstrate that AE downregulates genes involved in multiple DNA repair pathways, an effect that is evident as early as two days after retroviral transduction of human CD34+ cells. The human AE cells show delayed repair of induced DNA damage as shown by kinetics of a comet assay. During in vitro culture, AE cells show spontaneous accumulation of DNA damage, as demonstrated by γH2AX staining. Using a transgenic mutation frequency assay based on the lacZ reporter gene, we show that the in vivo mutation frequency of murine BM cells was increased approximately two fold by AE expression. Possibly as a result of this chronic DNA damage, AE cells have increased p53 protein expression and an upregulation of multiple p53 response genes. This results in increased basal apoptosis and enhanced sensitivity to DNA damaging agents compared to control cells. Intriguingly, microarray data indicate that t(8;21) patient samples also exhibit increased expression of p53 response genes when compared to other AML patient samples. Inhibition of the p53 pathway by shRNA interferes with the G1 cell cycle checkpoint and increases the resistance of AE cells to DNA damage. We thus speculate that AML1-ETO may indeed facilitate accumulation of additional genetic alterations by suppressing endogenous DNA repair, but that this activity also renders the cells more susceptible to stress. It is possible that the superior outcome of t(8;21) patients is partly due to an activated p53 pathway, and that loss of the p53 response pathway in disease progression will consequently negatively impact overall outcome. We are currently investigating whether a “p53 activation” profile can be used as a prognostic tool in AML.
Disclosure: No relevant conflicts of interest to declare.