MLL rearrangements are induced by low doses of etoposide in human fetal hematopoietic stem cells

During fetal development, the liver serves as the primary hematopoietic organ in which hematopoietic stem cells (HSC) comprise a large proportion of hepatic cell populations. Because HSC are capable of initiating long-term hematopoiesis, injury to these cells during pregnancy may play a role in the development of hematopoietic disorders manifested after birth. Of interest is the role of genetic injury to fetal HSC in the etiology of the infant acute leukemias, which are characterized by chromosomal rearrangements in the 11q23 region involving the mixed lineage leukemia (MLL) gene. These gene fusions also occur in leukemias in adults following chemotherapy with etoposide and other inhibitors of DNA topoisomerase II. We used etoposide as a model compound to determine the sensitivity of human fetal HSC to DNA damage and to determine whether we could induce MLL rearrangements in cultured human fetal HSC. Exposure of HSC to etoposide resulted in a dose-dependent loss of viability, with effects observed at low nanomolar concentrations. DNA strand breaks were observed on exposure to 140 nM etoposide, and higher etoposide concentrations stimulated an increase in early lymphoid populations and elicited G(2)/M cell cycle arrest. Immunophenotyping of MLL translocations revealed a significant increase in positive flow cytometry events at low etoposide concentrations and were consistent with MLL recombination. MLL translocations were confirmed using fluorescent in situ hybridization. In vitro inhibition of DNA topoisomerase II was observed at >= 25 mu M etoposide, but was not evident at lower etoposide concentrations associated with DNA damage. Our data indicate that low acute doses of etoposide can cause DNA strand breaks and chromosomal rearrangements involving MLL in human fetal HSC. Ultimately, such injury may have ramifications with regards to transplacental exposures to environmental chemicals linked to the etiology of infant acute leukemias.