Impact of malignant stem cell burden on therapy outcome in newly diagnosed chronic myeloid leukemia patients

Abstract

Chronic myeloid leukemia (CML) stem cells appear resistant to tyrosine kinase inhibitors (TKIs) in vitro, but their impact and drug sensitivity in vivo has not been systematically assessed. We prospectively analyzed the proportion of Philadelphia chromosome-positive leukemic stem cells (LSCs, Ph+CD34+CD38-) and progenitor cells (LPCs, Ph+CD34+CD38+) from 46 newly diagnosed CML patients both at the diagnosis and during imatinib or dasatinib therapy (ClinicalTrials.gov NCT00852566). At diagnosis, the proportion of LSCs varied markedly (1-100%) between individual patients with a significantly lower median value as compared with LPCs (79% vs 96%, respectively, P=0.0001). The LSC burden correlated with leukocyte count, spleen size, hemoglobin and blast percentage. A low initial LSC percentage was associated with less therapy-related hematological toxicity and superior cytogenetic and molecular responses. After initiation of TKI therapy, the LPCs and LSCs rapidly decreased in both therapy groups, but at 3 months time point the median LPC level was significantly lower in dasatinib group compared with imatinib patients (0.05% vs 0.68%, P=0.032). These data detail for the first time the prognostic significance of the LSC burden at diagnosis and show that in contrast to in vitro data, TKI therapy rapidly eradicates the majority of LSCs in patients.

Leukemic Stem Cell Quantification in Newly Diagnosed Patients With Chronic Myeloid Leukemia Predicts Response to Nilotinib Therapy

“…3B). A similar tendency toward a better response in patients with lower LSC burden at diagnosis was observed at later time points (6,12,15,18, and 21 months), although statistical significance was only reached at 15 months (P ¼ 0.034, at other time points P values were between 0.05 and 0.08). FISH analysis also predicted milestones; all patients not achieving MR 3 at 12 or 18 months had a high LSC burden at diagnosis (>80% Ph þ cells; Fig.…”

“…À stem cell compartment at diagnosis, as assessed by multiparameter flow cytometry (MPFC) or stem and progenitor cell sorting followed by FISH, not only predicts for later response, but also for hematologic toxicity during TKI treatment (14,15). Reliable and early response prediction is of great value as it enables personalized tailoring of therapy before progression to advanced phases in patients who are deemed to respond poorly.…”

“…The protocol for sorting and FISH has been described previously (15). In short, CD34 þ cells were enriched with paramagnetic beads (Miltenyi Biotech), stained with CD34 and CD38 antibodies (BD Biosciences) after which the primitive cell fractions were sorted by a flow cytometer (FACSAria; BD Biosciences)…”

Single-cell molecular analysis defines therapy response and immunophenotype of stem cell subpopulations in CML

“…To discriminate between leukemic and normal cells, 2 primer assays targeting BCR-ABL were included. By analyzing a combination of cell lines expressing a variety of BCR-ABL transcripts and the LSC population of a CML patient with known frequency of Ph 1 cells, 30 we confirmed a high specificity and sensitivity of the BCR-ABL primers at the single-cell level and a close to absolute correlation between the proportion of Ph 1 cells and the fraction of BCR-ABL-expressing primary LSCs (supplemental Figure 1). Additionally, we performed a number of control experiments to confirm that any effects on the data from pretreatment of cells (eg, cryopreservation and FACS), messenger RNA (mRNA) preamplification, and technical noise is minimal and significantly lower than observed cell-to-cell heterogeneity (supplemental Figure 2).…”

Inhibition of Ras-Mediated Signaling Pathways in CML Stem Cells

“…As soon as it became clear that BCR-ABL1 transcripts could be detected in HSCs, the existence of a pre-CML state before the chronic phase was firmly proved over the recent years. Oncofusion gene is expressed at low levels in pre-LSCs, which is increased in parallel with CML development, so that the progress of the phase transformation is vital for LSC formation [4]. By considering this issue, identification of CML using detection techniques for BCR-ABL1 chimaera seems to be impossible in the initial stages of disease.…”