Abstract
In ALL, current drug efficacy studies focus on reducing leukemia cell burden. However, if drugs have limited effects on LSCs, these cells may expand and eventually cause relapse. The experimental anti-leukemic drug parthenolide (PTL) acts by inhibiting transcription factor nuclear kappa B (NFκB), activating p53 and increasing reactive oxygen species (ROS) in leukemic cells. In the present study we assessed the in vitro effect of PTL on immune-phenotypically defined leukemic stem cells (LSCs) and normal hematopoietic stem cells (HSCs).
Forty ALL samples and 24 normal bone marrow samples were included in this study. ALL samples were sorted into 4 LSC populations (CD34+CD38-CD19+, CD34+CD38+CD19+, CD34+CD38-CD19-, and CD34-CD38-CD19-). Cells were cultured with different PTL concentrations for 24, 48 and 72 hours. Post-culture cell viability was assessed using 7ADD in a flow cytometry-based test. Normal unsorted marrow cells were tested under similar conditions. In addition, colony forming unit assay (CFU) was carried out to assess the effect of PTL on HSCs.
In ALL, LSCs form heterogeneous compartments with different percentages of the four LSC populations. This heterogeneity extended across and within cytogenetically classified groups of cases. There was no significant effect of PTL on the viability of normal cells at concentrations of ≤ 50μΜ; but at 25 μΜ and 10 μΜ after 24 and 72 hours respectively it affected the ability of HSCs to form colonies. For ALL-LSCs PTL was significantly toxic at10 μΜ and most cases with the exception of those with +3 karyotype abnormality and CD34-CD38-CD19- LSCs with t(9;22), showed >50% cell-death at 25μΜ PTL concentration. The CD34+, CD38+, CD19+ LSCs showed significantly higher sensitivity compared to other subpopulations. ALL cases with limited response to PTL expressed significantly higher levels of pNFκBP65 as compared to PTL-sensitive cases.
Overall, CD34+CD38-CD19+ and CD34+CD38-CD19 cells are the predominant LSC populations in B-ALL. PTL has a differential effect on normal HSCs versus ALL cells at lower concentrations in vitro. PTL also has a differential effect on LSC subpopulations. PTL sensitivity/resistance in ALL is related to the level of activated NFκB expression in these cells.