Aberrant Phenotype of Stem Cells as Assessed by Flow Cytometry Discriminates Between Low and High Risk Myelodysplastic Syndromes and Might Be Instrumental in Predicting Leukemic Evolution.

Abstract 3798

Poster Board III-734

Myelodysplastic syndromes (MDS) can be classified into low risk and high risk categories, with evolution to acute myeloid leukemia (AML) predominantly in the latter cases. In AML, survival of leukemia-initiating cells, often referred to as leukemic stem cells, after chemotherapy is thought to result in minimal residual disease leading to relapse. A larger size of the stem cell compartment in de novo AML is predictive for poor survival [Van Rhenen et al.,Clin Cancer Res 2005,11]. A monoclonal antibody against the cell surface antigen C-type lectin-like molecule-1, CLL-1, together with lineage infidelity markers enables discrimination of normal and malignant stem cells in AML [Van Rhenen et al., Blood 2007, 110; Van Rhenen et al., Leukemia 2007, 21]. Previously, we showed that the size of the total stem cell compartment including both normal and malignant cells, defined as CD45^dimCD34⁺CD38⁻, were significantly more prevalent within the blast cell fraction in high risk MDS as compared to low risk MDS (median 0.77% (n=15) and 0.25% (n=73), respectively, p=0.040) [Westers et al. Blood 2008, 112, abstract 1661]. This might reflect the differences in clinical course in these patients. It could be hypothesized that aberrant marker expression on MDS stem cells may predict leukemic evolution. Therefore, stem cells in MDS bone marrow samples were analyzed by flow cytometry for expression of CLL-1 and aberrant lineage markers. A reliable number of stem cells (>20) could be examined in 22 low risk and 14 high risk MDS patients; patients were classified by WHO2001 as refractory anemia w/o ring sideroblasts (RS; n=6) or refractory cytopenia with multilineage dysplasia w/o RS (n=16) and 14 refractory anemia with excess of blasts type 1 or 2 (n=14). Median CLL-1 expression on the CD34⁺CD38⁻ stem cells was 0.0% (range 0-50, 3 out of 22 clearly (>10%) positive cases) in low risk and 2.2% (range 0-27, 3 out of 14 clearly positive cases) in high risk MDS. For comparison, median CLL-1 expression on stem cells in normal controls was 0.0% (range 0-4.7, not significantly different from MDS cases: p=0.55 and 0.20 as compared to low and high risk categories, respectively). Eleven of the low risk MDS cases were tested for aberrant lineage infidelity marker expression on their myeloid blasts and stem cells. In these cases, myeloid blasts expressed either CD5 or CD7 (n=2 and n=7, respectively; median 33% of blast cells). Median expression of these antigens on stem cells was only 1.2% (range 0-47). There was only one case with high expression (47%); the rest was below 7%. Only four of the high risk cases could be analyzed for lineage infidelity marker expression. In these patients, median 51% of the myeloid blasts expressed either CD5 (n=1), CD7 (n=1) or CD56 (n=2). Median expression of these antigens on stem cells in these high risk MDS cases was 44% (range 3-83); three of four cases had high expression (>40%). Overall, expression of CLL-1 and lineage infidelity markers was higher in high risk MDS as compared to the low risk category (p=0.047 and p=0.036 for CLL-1 and lineage infidelity markers, respectively). Of note, the only low risk case with a high percentage (47%) of aberrant stem cells as assessed by flow cytometry rapidly progressed towards AML. To conclude, CLL-1 is virtually absent on stem cells in low and high risk MDS. Nevertheless, expression of lineage infidelity markers, such as CD5, CD7 and CD56, on CD34⁺CD38⁻ stem cells in some cases strongly suggests that a considerable part of these stem cells was malignant. Aberrant stem cell phenotype as assessed by flow cytometry might discriminate normal from MDS stem cells. This enables further analysis of MDS stem cells and, moreover, makes them amenable for application of targeted therapy. Future analysis might reveal whether the existence of MDS stem cells with an aberrant profile by flow cytometry predicts leukemic evolution.