Fludarabine ability to down‐regulate Bcl‐2 gene product in CD5⁺ leukaemic B cells: in vitro/in vivo correlations

Abstract

CD5+ B‐chronic lymphocytic leukaemia (B‐CLL) and mantle cell lymphoma (MCL) in leukaemic phase are characterized by defects in cell death induction that primarily involves the Bcl‐2 family of genes. Fludarabine (9‐β‐D‐arabinofuranosyl‐2‐fluoradenine, F‐ara‐A) is a potent inducer of apoptosis in CLL cells. This study aimed to determine whether F‐ara‐A‐induced apoptosis might be related to Bcl‐2 modifications and to evaluate in vitro/in vivo correlations. Peripheral blood lymphocytes from eight B‐CLL and four leukaemic MCL were cultured in the presence of different concentrations of F‐ara‐A ±methylprednisolone (MP). F‐ara‐A down‐regulated the expression of Bcl‐2 in 5/12 cases. mRNA down‐regulation was maximal at 48 h; protein down‐regulation was prominent after 48 h. Both events were dose‐dependent. The amount of apoptosis was significantly higher in the samples treated with F‐ara‐A than in those exposed to MP alone. In the seven remaining cases, no Bcl‐2 down‐regulation was observed after exposure to F‐ara‐A and the degree of F‐ara‐A‐induced apoptosis overlapped that induced by MP. The in vivo outcome after treatment with three to six courses of F‐ara‐A was evaluable in 10 patients: 4/5 cases, whose cells had shown in vitro Bcl‐2 down‐regulation and prominent apoptosis after exposure to F‐ara‐A, had a complete response (CR) and a partial response (PR) was observed in the remaining patient. Of the five patients whose cells had shown no in vitro Bcl‐2 modulation after exposure to F‐ara‐A, two had a PR, but the other three did not show any in vivo clinical response.

Role of caspases and apoptosis-inducing factor (AIF) in cladribine-induced apoptosis of B cell chronic lymphocytic leukemia

“…B-CLL cells expressed significant amounts of Bax and Bcl-2 proteins, which did not significantly change with 2CdA treatment, according to Ref. 35 and it as previously found with Fara-A by most 4,15,36,37 but not all 38 authors. However, Bcl-2 overexpression prevented 2CdA-induced apoptosis in CEM 6 and Jurkat cells.…”

In vitro evaluation of bendamustine induced apoptosis in B-chronic lymphocytic leukemia

“…Recently, it was reported that fludarabine could induce Bcl-2 down-regulation in cultured B-CLL cells and that antisense Bcl-2 oligonucleotides results in the induction of apoptosis in B-CLL cells. 14,37,70 In our study, the Bcl-2 and Bax expression was determined by RNase protection assay and FACS analysis. Using these methods, we found no significant changes (defined as more than 100% for mRNA expression levels or more than 1 log 10 concerning fluorescence intensity Leukemia for FACS analysis).…”

“…11,12 It is yet unknown how the expression of the mem-bers of the Bcl-2 family are regulated and whether they can influence the drug sensitivity of B-CLL cells in vitro. 13,14 Caspase activation, following different stimuli, plays a key role in the apoptotic pathways by a specific cleavage of important cellular essential enzymes. 15,16 This activation mechanism seems to be under the control of dimerizing Bcl-2 family members.…”

Nucleoside analogues: mechanisms of drug resistance and reversal strategies

“…71 Moreover, F-ara-A has been reported to downregulate Bcl-2 mRNA expression, thereby favoring apoptosis. 91 Finally, F-ara-A affects mitochondria but only as a late event, suggesting an indirect mechanism related to the p53-apoptotic pathway. 69 The different effects of F-ara-A and 2-CdA on mitochondria integrity may be related to the fact that 2-CdATP may accumulate more rapidly than F-ara-ATP in the mitochondria because it is converted with higher efficiency by mitochondrial dGK.…”