Uncontrolled cell proliferation is one of the hallmarks of cancer and the transition from the G1 to S phase is the most commonly reported cell cycle abnormality in tumors. It has been shown that the oncogenic activity of G1 cyclin E (CCNE) can be amplified by generating hyperactive low molecular weight forms (LMW) through elastase-mediated proteolytic processing. Neutrophil elastase (NE) and proteinase 3 (PR3) are 2 proteases that are aberrantly expressed in breast cancer cells and seem to be involved in cell proliferation. In this study, we evaluated the effect of the expression of these 2 proteases in addition to 2 potential intracellular targets of NE (CCNE1 and CCNE2) on clinical outcome in a population of 205 primary breast cancer patients. By univariate analysis, CCNE1, CCNE2, estrogen receptor and grade significantly predicted relapse free interval (RFI). NE and PR3 did not achieve statistical significance. In a multivariate analysis, elevated CCNE2 [hazard ratio (HR) 2.10, p 5 0.008] predicted shorter RFI. In subgroup analyses of the tamoxifen-only treated patients, high CCNE1 levels predicted treatment resistance, while high levels of CCNE2 were associated with poor RFI in untreated patients. Investigation of the relationship between CCNE1, CCNE2 and NE did not show any impact on RFI. To conclude, this study was the first to evaluate these markers at the mRNA level by RT-PCR in a series of primary breast cancer patients, and our results confirmed the impact of high CCNE levels on clinical outcome in systemically untreated and of CCNE1 in tamoxifen-only treated early breast cancer patients. ' 2006 Wiley-Liss, Inc.Key words: breast cancer; elastase; proteinase 3; cyclin E; mRNA Uncontrolled cell proliferation is one of the hallmarks of cancer and the transition from the G1 to S phase is the most commonly reported cell cycle abnormality in tumors. 1 Cyclin E (CCNE), a G1 cyclin which is essential for S-phase entry, has a profound role in oncogenesis, 2,3 and its overexpression has been repeatedly linked with poor patient outcome in breast cancer (reviewed in Ref. 4) and with endocrine therapy failure. 5 Recent studies identified low molecular weight isoforms (LMW) of CCNE, generated specifically in tumors by elastase-mediated proteolytic processing, 6 which were shown to be hyperactive compared to full-length CCNE and associated with poor clinical outcome in stage I to III breast cancer patients. 7 Recently, it has been shown that high levels of neutrophil elastase (NE) protein in breast cancer tissue extracts were associated with poor clinical outcome and with endocrine treatment failure in patients with advanced breast cancer disease. 8,9 This prognostic value was supposedly linked with the essential role of NE in the degradation of the extracellular matrix and the metastasis process. 10,11 But since NE may be aberrantly produced by breast cancer cells themselves and because NE has been implicated in the intracellular cleavage of CCNE in its LMW forms, its prognostic value may be linked to its property ...

The online version of this article contains a supplementary appendix.Background Zeta-associated protein 70 (ZAP70) is a widely recognized prognostic factor in chronic lymphocytic leukemia, but mechanisms by which its higher expression leads to a poor outcome must still be fully explained. Design and MethodsIn an attempt to unveil unfavorable cellular properties linked to high ZAP70 expression, we used gene expression profiling to identify genes associated with disparities in B cells from chronic lymphocytic leukemia patients expressing high versus low ZAP70 mRNA, measured by quantitative real-time PCR. Two groups of 7 patients were compared, selected on the basis of either high or low ZAP70 mRNA expression. ResultsTwenty-seven genes were differentially expressed with an FDR<10%, and several genes were significant predictors of treatment-free survival (TFS) and/or overall survival; PDE8A and FCRL family genes (down-regulated in ZAP70 + patients) could predict TFS and overall survival; ITGA4 mRNA (up-regulated in ZAP70 + patients) could significantly predict overall survival. Importantly, gene set enrichment analysis revealed overrepresentation of adhesion/migration genes. We therefore investigated in vitro adhesion/migration capacity of chronic lymphocytic leukemia cells into a stromal microenvironment or in response to conditioned medium. We showed that ZAP70 + cells had better adhesion/migration capacities and only ZAP70 + patient cells responded to microenvironment contact by CXCR4 downregulation. ConclusionsWe concluded that several prognostic factors are the reflection of microenvironment interactions and that the increased adhesion/migratory capacity of ZAP70 + cells in their microenvironment can explain their better survival and thus the aggressiveness of the disease.Key words: chronic lymphocytic leukemia, ZAP70, microarrays, microenvironment, prognosis.Citation: Stamatopoulos B, Haibe-Kains B, Equeter C, Meuleman N, Sorée A, De Bruyn C, Hanosset D, Bron D, Martiat P, and Lagneaux L. Gene expression profiling reveals differences in microenvironment interaction between patients with chronic lymphocytic leukemia expressing high versus low ZAP70 mRNA. Haematologica 2009; 94:790-799. doi:10.3324/haematol.2008 This is an open-access paper.Gene expression profiling reveals differences in microenvironment interaction between patients with chronic lymphocytic leukemia expressing high versus low ZAP70 mRNA

Autologous transplantation using peripheral blood stem cells (PBSCs) collected after chemotherapy, followed by growth factor administration (ASCT), is increasingly used in the treatment of non‐Hodgkin's lymphoma (NHL). However, quantitative data regarding contaminating malignant cells in the harvests are still scarce. We prospectively investigated 37 diffuse large‐cell lymphomas (DLCLs) in complete remission (CR) that were treated according to multicentric protocols at our centre. DNA was extracted from the diagnostic lymph node. The complementarity‐determining region (CDR) III was sequenced and a patient‐specific oligomer synthesized. Contamination was evaluated semiquantitatively by polymerase chain reaction (PCR) and was confirmed by a limiting dilution analysis. PBSCs collected at regeneration after administration of granulocyte colony‐stimulating factor (G‐CSF), steady‐state bone marrow (BM) and peripheral blood samples at CR were compared. DNA was available in 37 patients, from which 22 rearrangements could be sequenced. Patients (n = 15) who had both the required follow‐up samples and a suitable clonal marker were investigated. In two cases, the patient‐specific PCR assay set up at diagnosis later gave false‐negative results in samples in which clonal DNA was still detectable by other sets of primers. PBSC contamination was highly variable: 7 out of 15 patients showed a PBSC/BM ratio of NHL cells greater than 1 log, whereas 8 out of 15 patients showed no difference and could vary from one apheresis to another. Eight ASCTs were performed, five of which used highly contaminated PBSCs: four patients relapsed early, three with disseminated lymphoma. Thus, 50% of DLCLs in CR seem to mobilize significantly malignant cells at regeneration under G‐CSF. Considering the higher numbers of cells reinfused, this translates into a much higher number of lymphoma cells reinfused when compared with autologous bone marrow transplantation (ABMT). However, their clonogenic potential remains unknown and, despite concerning observations in certain cases, it is still unclear whether this has an impact upon the outcome of ASCT.