Determination of donor chimerism profiles in blood or bone marrow from patients with allogeneic stem cell transplantation (SCT) is useful for monitoring engraftment or predicting relapse, when specific molecular markers are lacking. CD34+ donor cell chimerism (DCC) analysis in peripheral blood samples from CD34+ acute myeloid leukemia (AML) and myleodysplastic syndrome (MDS) patients proved to be a highly sensitive diagnostic tool that is useful to detect imminent relapse significantly earlier compared to total white blood cell donor cell chimerism monitoring. However, flow-cytometric enrichment of CD34+ cells requires high efforts to human resources and equipment. We present a novel semi-automated CD34+ DCC analysis procedure-employing a magnetic cell-enrichment device, DNA extraction, and short tandem repeat profiling-without the need for flow-cytometric cell sorting. Monitoring 85 patients with AML and MDS over a period of 4 years 24 relapses were detected. Semi-automated peripheral blood CD34+ DCC was diminished below 80 % in all cases of systemic relapse. Significant decrease of the CD34+ DCC value was detected 29-42 days before overt cytological relapse. Our method provides a rapid and sensitive tool for monitoring AML and MDS patients after allogeneic SCT with regard to engraftment and early detection of relapse. Here, we propose a novel semi-automated procedure for CD34+ DCC analysis after allogeneic SCT that is simple, reliable, and therefore applicable in all hematologic laboratories.
Acute myeloid leukemia (AML) is a clonal disease originating from myeloid progenitor cells with a heterogeneous genetic background. High-dose cytarabine is used as the standard consolidation chemotherapy. Oncogenic RAS mutations are frequently observed in AML, and are associated with beneficial response to cytarabine. Why AML-patients with oncogenic RAS benefit most from high-dose cytarabine post-remission therapy is not well understood. Here we used bone marrow cells expressing a conditional MLL-ENL-ER oncogene to investigate the interaction of oncogenic RAS and chemotherapeutic agents. We show that oncogenic RAS synergizes with cytotoxic agents such as cytarabine in activation of DNA damage checkpoints, resulting in a p53-dependent genetic program that reduces clonogenicity and increases myeloid differentiation. Our data can explain the beneficial effects observed for AML patients with oncogenic RAS treated with higher dosages of cytarabine and suggest that induction of p53-dependent differentiation, e.g. by interfering with Mdm2-mediated degradation, may be a rational approach to increase cure rate in response to chemotherapy. The data also support the notion that the therapeutic success of cytotoxic drugs may depend on their ability to promote the differentiation of tumor-initiating cells.
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