Gene expression profiles based on ZAP70 mRNA level.
Stamatopoulos B, Haibe-Kains B, Meuleman N, Bron D, Martiat P and Lagneaux L
Background: Gene expression
profile is a powerful tool to better understand the biology, the
clinical outcome and the molecular mechanism implicated in chronic
lymphocytic leukemia (CLL). This disease presents an extremely variable
clinical course with overall survival times ranging from months to
decades. Therefore a plethora of prognostic factors which classified
patients in poor or good behaviour have been investigated.
Zeta-associated protein 70 (ZAP70) is one of the most promising
prognostic factors to predict CLL evolution. Furthermore, we previously
described a quantitative real-time PCR (qPCR) method to measure
ZAP70 and demonstrated its prognostic power (Stamatopoulos et al, Clin.
Aims: In this study, we
compared gene expression profile of patients expressing high versus low
ZAP70 mRNA level in order to find genes not only associated with
prognosis but also with cell biology. We also confirmed some microRNA
differentially expressed between these two groups and linked them to
treatment-free survival (TFS) and overall survival (OS).
Methods: ZAP70 was evaluated by
qPCR in a cohort of 108 patients ; two groups of 7 patients were
chosen in the top-20 of patients expressing high and low level of ZAP70
mRNA and their gene expression profiles were compared using Affymetrix
technology. Selected genes were verified by qPCR in an extended patient
cohort (n=85) with a median follow-up of 72 months. Adhesion/migratory
capacities into a stromal microenvironment or in response to
conditioned medium were also evaluated. Finally, we investigated some
microRNA differential expression by qPCR in a cohort of 61 patients
with a median follow-up of 74 months.
Results: 43 probe sets were
differentially expressed with a FDR<10%, 135 with a P<0.001 and
932 with a P<0.05. Several of these genes were TFS and/or OS
significant predictors: PDE8A and FCRL family genes were downregulated
in ZAP70+ patients and can predict TFS and OS; ITGA4 mRNA was
upregulated in ZAP70+ patients and can significantly predict OS.
Moreover pathway analysis reveals an over-representation of
adhesion/migration genes. CXCR4/SDF1-alpha pathway was one of them. We
observed a downregulation of CXCR4 in stromal-adherent cells only in
ZAP70+ patients indicating that only these patient cells can respond to
microenvironment stimulus. Furthermore, ZAP70+ patient cells can
significantly better adhere to fibronectin. MicroRNA comparison
confirmed the differential expression of miR-29c and miR-223 and we
showed for the first time that these two microRNA had a TFS and OS
individual prognostic power.
Conclusions: This study
identifies new prognostic factors (genes and microRNA) and shows the
better adhesion/migratory capacities of ZAP70+ cells in their
microenvironment explaining their better survival and the
aggressiveness of the disease.