Increased Expression Level of Noxa in Peripheral Versus Lymph Node Chronic Lymphocytic Leukemia Cells Is Correlated with Survival Capacity.

Abstract

Background: The gradual accumulation of chronic lymphocytic leukemia (B-CLL) cells is presumed to derive from proliferation centers in lymph nodes and bone marrow. To what extent these cells possess the purported anti-apoptotic phenotype of peripheral B-CLL cells is unknown. Recently, we have described that in B-CLL samples from peripheral blood, aberrant apoptosis gene expression was not limited to protective changes but also included increased levels of pro-apoptotic BH3-only member Noxa. The functional consequence of this finding is not known, nor whether this aberrant apoptosis gene profile is also present in CLL proliferation centers.Aim: To perform a functional comparison of apoptosis gene profiles from peripheral blood (PB) B-CLL versus lymph node (LN) proliferation centers.Methods: Immunofluorescence microscopy, RT-Multiplex-Ligation-dependent Probe Amplification (RT-MLPA), Western Blot, Transduction, RNA interference.Results: PB samples (>90% CD5/CD19/CD23+ B-CLL cells) from 16 B-CLL patients and LN samples from 9 B-CLL patients were included in this study. LN samples contained over 90% CD5/CD19/CD23+ lymphocytes with Ki67+ cells either scattered throughout the LN or in follicle-like structures. RNA samples were subjected to the RT-MLPA procedure which monitors expression of 34 apoptosis genes. Apart from expected differences in survivin and Bcl-xL, the most prominent distinction with PB B-CLL cells was the generally low levels of Noxa in LN samples. [Display omitted] A reduction in Noxa RNA and protein levels could also be obtained by in vitro stimulation of PB B-CLL with CD40. Direct manipulation of Noxa protein levels was achieved by proteasome inhibition in B-CLL cells and via RNAi in model cell lines. In all these instances the viability of the cells was inversely correlated with Noxa levels.Conclusions: These data indicate that spontaneous apoptosis of PB B-CLL cells in vitro is linked with high Noxa levels. We propose that suppression of Noxa in the LN contributes to the persistence of B-CLL, and that therapeutic targeting of Noxa might be beneficial.