Increased growth inhibition of human chronic myelogenous leukemic cells by a combination of c-myb antisense oligonucleotide and 4-hydroxyperoxycyclophosphamide in vitro

Human chronic myelogenous leukemia (CML) is a unique malignancy in its cellular and molecular phenotypes. High dose therapy followed by stem cell transplantation seems to be one of the most effective treatment modalities for CML. However, allogeneic stem cell transplantation, a curative treatment modality, is limited due to the availability of matched siblings. On the other hand, the autologous stem cell harvests are contaminated with leukemic cells, and therefore a significant reduction of leukemic cells is desired before using the harvest for transplantation. Therefore in the present study, effects of a combination of a suboptimal concentration of 4-hydroxyperoxycyclophosphamide (4HC) and an optimal concentration of c-myb antisense oligonucleotide on the growth of K562 human chronic myelogenous leukemic cells in vitro were determined. The combination significantly (p<0.05) inhibited the growth of K562 cells in vitro when compared to the effects of c-myb oligonucleotide or 4HC alone. The c-myb oligonucleotide alone or in combination with low dose 4HC decreased the expression of c-myb gene as determined by RT-PCR techniques. Cellular uptake and retention of fluoresceinated oligonucleotide in control and treated K562 cells was studied using plain field laser microscopy and flow cytometry. There was an increase in cellular uptake of c-myb oligonucleotide in K562 cells as measured by plain field laser microscopy in the presence of 4HC. The combination of oligonucleotides and 4HC did not significantly decrease the number of hematopoietic stem/progenitor cells from normal hematopoietic stem cell harvests as determined by in vitro colony assays. The combination of low dose 4HC and c-myb antisense oligonucleotides can potentially be applied in CML patients, particularly for purging leukemic cells present in their hematopoietic stem cell harvests.