Assessment of Peripheral Blood and Bone Marrow Cells Apoptosis Caused by Purine Analogues in Patients with Chronic Lymphocytic Leukemia in Correlation with Parameters of Disease Progression

Abstract

Chronic lymphocytic leukemia (CLL) is a heterogeneous disease with variable clinical course and prognosis. Therefore, the role of prognostic factors is very important, especially for identifying the group of patients who require intensive treatment. The aim of this study was to assess whether the rate of apoptosis caused by purine analogues differs between patients with better or worse prognostic factors. The experiments were preformed in cultures of blood and bone marrow obtained from CLL patients. The cultures were supplemented with cladribine and fludarabine. We determined the percentage of caspase-3-positive cells and the BCL-2/BAX ratio, and subsequently these apoptosis markers were correlated with the expression of ZAP-70 and CD38, lymphocyte counts, lactate dehydrogenase and β₂-microglobulin levels and clinical stage according to the Rai classification. The results showed that bone marrow cells are more sensitive to apoptosis caused by purine analogues than cells derived from blood, supporting the idea that these two compartments have different proliferative statuses. The cells from ZAP-70-positive patients seem to enter apoptosis more readily than those from ZAP-70-negative patients; thus, ZAP-70-positive patients are more likely to benefit from treatment with purine analogues.

The rate of in vitro fludarabine-induced peripheral blood and bone marrow cell apoptosis may predict the chemotherapy outcome in patients with chronic lymphocytic leukemia

“…We detected active caspase-3 expression in malignant CD19+/CD5+ cells in all cell cultures from both peripheral blood and bone marrow after 24 h, similarly to our previously published results [ 12 ]. An increase in the percentage of caspase-3-positive cells was observed after 24 h in control cultures (cells without drugs) relative to the 0-h baseline control.…”

“…This culture medium was supplemented with fludarabine (Schering AG, Germany) at a concentration of 1 μg/ml. We selected such a concentration of fludarabine on the basis of our preliminary experiments in CLL cultures, in which this concentration induced significant number of apoptosis with spontaneous apoptosis that did not exceed 50 % of the total cell culture [ 12 ]. The cells were cultured at 37 °C in a 5 % CO 2 atmosphere, and they were exposed to the drugs for 24 h. Respective cell samples incubated in the absence of any drug for periods of time equivalent to the drug-treated cells were used as a negative control.…”