FISH⁺CD34⁺CD38^- Cells Detected In Newly Diagnosed Acute Myeloid Leukemia Patients Can Predict The Clinical Outcome

Acute myeloid leukemia (AML) is a clonal hematologic malignancy arising from a small population of leukemic cells that initiate and propagate the disease. These cells are termed leukemic stem cells (LSCs), which are found within the CD34⁺CD38^- cell compartment. The LICs subpopulation survives chemotherapy and is most probable the cause of minimal residual disease (MRD), which in turn is thought to cause relapse. The aim of this study was to determine the prognostic value of the percentage of LICs measured by fluorescence in situ hybridization (FISH) in flow sorted CD34⁺CD38^- cells (FISH⁺CD34⁺CD38^- population) at diagnosis. Forty-five patients with de novo AML with FISH-detectable cytogenetic abnormalities treated with CHAML 2010 protocol were retrospectively included in this study. The last follow-up was March 31, 2013, and the median follow-up for patients was 12 months (range: 1–27 months). After 24 months, 12 patients (26.7%) had experienced relapse, whereas 7 patients (17.9%) had died due to non-relapse treatment-related complications, 3 from treatment-related complications of transplantation and 4 from infection after chemotherapy. To distinguish normal and leukemic cells within the CD34⁺CD38^- cell compartment, we established a unique protocol for conducting FISH on flow-sorted cells. The FISH positive percentage at diagnosis constituting an average of 2.31% (range: 0.01%-29.4%) of the blast cells and 68.2% (range: 7.8%-89.6%) of the CD34⁺CD38^- cells. According to the LIC level detected by Flow-FISH analysis, patients were categorized into the two groups: low LIC load (＜1%) and high LIC load (≥1%), representing 44.4% (n=20) and 55.6% (n=25) of all patients, respectively. Comparison of clinical and laboratory characteristics showed no significant differences between two groups in age, gender, white blood cell (WBC) count, platelet (PLT) count, blast percentage, FAB subtype distribution, FLT3 mutation status and cytogenetics risk at diagnosis. High LIC load at diagnosis was significantly correlated with increased risk of poor clinical outcome. The 2-year overall survival (OS) for patients with low LIC load was 0.7257[0.4082, 0.8916], compared with 0.1675[0.0323, 0.3947] for the patients with high LIC load (P=0.0004). And 2-year events-free survival (EFS) were 0.6723[0.3262, 0.8687] versus 0.1633[0.0328, 0.3825], respectively (P=0.0029). By multivariate analysis, high LIC load retained prognostic significance for OS and EFS. Furthermore, high LIC load at diagnosis was found to be significantly associated with elevated cumulative incidence of relapse (2-year CIR: 56.7% vs. 18.0%; P = 0.021). In summary, our study established the Flow-FISH protocol as a useful method to distinguish normal and leukemic cells within the CD34⁺CD38^- cell subpopulation. And Flow-FISH might be easily adopted as a powerful tool to predict clinical outcome and help physicians to evaluate criteria for treatment in AML with recurrent cytogenetic abnormalities.