Zhi-Jian Zou

Downregulated Dicer expression predicts poor prognosis in chronic lymphocytic leukemia

Chronic lymphocytic leukemia (CLL) is the most common leukemia in the western world. Alterations in microRNAs (miRNAs) expression have been proposed to play a role in CLL pathogenesis. Dicer and Drosha are the main regulators of miRNA biogenesis, and deregulation of their expression has been indicated as a possible cause of miRNA alterations observed in various cancers. To investigate the role of Dicer and Drosha in CLL, we assessed the expression of Dicer and Drosha and their correlation with other prognostic factors, including Binet stages, immunoglobulin heavy chain variable gene (IGHV) mutation status, TP53 mutation status, ZAP‐70 protein and CD38 expression level in 165 CLL patients by using real‐time polymerase chain reaction methods. Patients with unmutated IGHV genes had significantly lower expression of Dicer than patients with IGHV mutations. The lower expression level of Dicer was also significantly associated with higher level of CD38 and ZAP‐70, and more aggressive Binet stage. We also analyzed Dicer expression in different cytogenetic subgroups. Lower Dicer level was found in patients with unfavorable cytogenetic aberrations (deletion in 17p13 or 11q22.3) in contrast to higher level in good risk cytogenetics (deletion in 13q14 as the sole abnormality). Furthermore, the lower expression of Dicer in CLL shows a strong association with shorter overall survival (OS) (P = 0.0046) as well as with reduced treatment free survival (TFS) (P = 0.0006). By contrast, no differences in the expression of Drosha among these groups of patients were observed. Our data suggest that Dicer expression may play an important role in the progression and prognosis of CLL. (Cancer Sci 2012; 103: 875–881)

Expression levels of Lyn, Syk, PLCγ2 and ERK in patients with chronic lymphocytic leukemia, and higher levels of Lyn are associated with a shorter treatment-free survival

Abstract There is now strong evidence that B-cell receptor (BCR) signaling plays a major role in the development of chronic lymphocytic leukemia (CLL). The purpose of this study was to investigate the expression levels of some of the molecules involved in the signaling cascade originating from the BCR (Syk, Lyn, PLCγ2, ERK) and analyze possible correlations of mRNA levels with biological/clinical features. Our study population consisted of 92 consecutive patients with newly diagnosed CLL. Several genes of BCR signaling (Lyn, Syk, PLCγ2 and ERK) and ZAP-70 were measured by quantitative polymerase chain reaction (qPCR). The signaling molecules of BCR were strongly associated with each other, and ZAP-70 correlated well with Lyn, Syk, PLCγ2 and ERK. Associations between treatment response and Lyn, Syk, PLCγ2 and ERK were not found. Moreover, a higher level of Lyn mRNA was associated with a shorter treatment-free survival (TFS) (univariate analysis only; multivariate Cox analysis showed that only ZAP-70 and Binet stage were independent prognostic factors and associated with TFS). Though Lyn was not an independent prognostic factor, it still might be a new therapy target of CLL. BCR signaling provides perspectives for future development of an exciting new class of kinase inhibitors.

Richter transformation in 16 of 149 Chinese patients with chronic lymphocytic leukemia.

Abstract No confirmed risk factors are known to predict Richter syndrome (RS), and the value of clinical prognosticators conventionally applied to chronic lymphocytic leukemia (CLL) is not firmly established in this setting. The objective of this study was to present evidence for RS in Chinese patients with CLL and risk factors for CLL transformation to Richter syndrome. With a median follow-up of 43 months from CLL diagnosis in 149 Chinese patients, 16 (10.7%) patients progressed to diffuse large B-cell lymphoma (DLBCL). According to correlation analysis, a high level of lactate dehydrogenase (LDH) and CD38 positivity were found to be independent predictors of transformation to RS. Survival analysis showed that presence of RS, advanced Binet stage, high level of LDH, high level of β2-microglobulin, high concentration of thymidine kinase (TK), ZAP-70 and CD38 expression, unmutated immunoglobulin heavy chain variable (IGHV) gene status and del(17p13) were adverse factors in determining overall survival (OS). Only del(17p13) was strongly associated with survival by multivariate Cox regression analysis. Median OS after transformation was 16 months (95% confidence interval, 5.6–26.4 months). The results support that RS is associated with a poor outcome, and a policy of close monitoring and careful biopsy is needed in patients carrying transformation risk factors.

S‐MDM4 mRNA overexpression indicates a poor prognosis and marks a potential therapeutic target in chronic lymphocytic leukemia

The purpose of the present study was to investigate the prognostic significance of murine double minute 4 (MDM4) in chronic lymphocytic leukemia (CLL) and to characterize the role of MDM4 in the p53 pathway. Full‐length MDM4 (FL‐MDM4), a splicing variant of MDM4 (S‐MDM4) and murine double minute 2 (MDM2) mRNA expressions were detected by quantitative PCR in 140 Chinese patients with CLL, and primary CLL cells were treated in vitro with either fludarabine or Nutlin‐3 to explore the interaction between p53 status and MDM4 or MDM2 expression. A marked increase of FL‐MDM4 and S‐MDM4 expressions were observed in the CLL patients with p53 aberrations (deletion and/or mutation) (P = 0.024, P < 0.001). A high level of S‐MDM4 mRNA expression was associated with short treatment free survival (TFS) (P = 0.004). FL‐MDM4 expression was significantly decreased after fludarabine treatment (P = 0.001) but increased after Nutlin‐3 treatment (P = 0.008) of primary CLL cells without p53 aberrations. Both S‐MDM4 and MDM2 expressions were significantly increased after fludarabine treatment of CLL cells without p53 aberrations (P = 0.013 and P = 0.030). MDM2 overexpression also occurred in CLL cells with p53 wild type after Nutlin‐3 treatment (P = 0.018). FL‐MDM4 and S‐MDM4 overexpression are indicators of p53 aberrations in CLL patients, suggesting that those patients have a poor prognosis. FL‐MDM4 inhibitory effects on p53 can be removed by MDM2‐p53 and saved by Nutlin‐3.

Low expression level of phosphatase and tensin homolog deleted on chromosome ten predicts poor prognosis in chronic lymphocytic leukemia

Abstract Phosphatase and tensin homolog deleted on chromosome ten (PTEN) is one of the best-studied tumor suppressor genes which can promote cell proliferation and contribute to tumorigenesis. This study aimed to investigate the PTEN mRNA (Pm) expression level in patients with chronic lymphocytic leukemia (CLL) and healthy controls, and its correlation with prognostic factors. Quantitative polymerase chain reaction (qPCR) was used to detect Pm expression. Compared to controls, patients with CLL presented a lower expression level of Pm (p < 0.001). In univariate analysis, the expression level of Pm was significantly decreased in patients with Binet C (p < 0.001) and higher level of β2-microglobulin (β2-MG) (p = 0.036), lactate dehydrogenase (LDH) (p = 0.019) and ZAP-70 (p = 0.008). Higher Pm expression level was found in favorable cytogenetic aberrations (p = 0.016) and the group without p53 aberration (p = 0.005). Multivariate analysis showed that advanced Binet stage (p = 0.027) and p53 aberration (p = 0.007) were associated with a low PTEN expression level. Survival analysis showed that low expression of PTEN was associated with shorter time to first treatment (TTFT) (p = 0.040). These results indicate that PTEN might be a new prognostic marker in patients with CLL.

High levels of CD20 expression predict good prognosis in chronic lymphocytic leukemia

Heterogeneity of CD20 expression exists in chronic lymphocytic leukemia (CLL), therefore, we explored the prognostic significance of CD20 expression in Chinese patients with CLL. Multiparameter flow cytometry was used to detect the expression of CD20 in CD5+CD19+ cells. In 172 CLL patients, the median expression percent of CD20 was 97.82% (range, 0–100), and the median mean fluorescence intensity (MFI) of CD20 in CLL cells was 731.45 (range, 0.00–9071.90). The percentage of CD20+ cells in the patient group with mutated variable region of immunoglobulin genes (IGHV) was higher than in the non‐mutant IGHV group (mean, 92.1% vs 80.4%, P < 0.001). There were no differences in the MFI of CD20+ cells in all prognostic factor groups. Representation of the data using a receiver operating characteristic plot reflected separation between the two IGHV groups, with an area under the curve of 0.661 (95% confidence interval, 0.569–0.753). At the cut‐off value of 60.3% for percentage of CD20, the sensitivity and specificity were 90.00% and 38.46%, respectively. Patients whose percentage of CD20 antigen was above 60.3% had longer treatment‐free survival (hazard ratio, 0.452; 95% confidence interval, 0.232–0.884, P = 0.020). Percentage and MFI of CD20 were the variables not associated with treatment‐free survival by multivariate Cox regression analysis (P < 0.05). High level of CD20 expression in de novo CLL appears to be associated with a good prognosis.

Expression level of DEK in chronic lymphocytic leukemia is regulated by fludarabine and Nutlin-3 depending on p53 status

Human oncogene DEK has been shown to be upregulated in a number of neoplasms. The purpose of this study was to investigate DEK expression level in chronic lymphocytic leukemia (CLL), analyze the correlation between DEK expression and CLL prognostic markers, and characterize the role of DEK in the response to either chemotherapeutic drugs or nongenotoxic activators of the p53 pathway. DEK mRNA was evaluated by real-time quantitative reverse transcriptase-polymerase chain reaction (qPCR), and primary CLL samples were treated in vitro with either fludarabine or Nutlin-3 to explore the interaction of p53 status and DEK mRNA expression. The median expression levels of DEK mRNA were 6.792 × 10−2 (1.438 × 10−2−3.201 × 10−1) in 65 patients with CLL. A marked increase of DEK mRNA expression was observed in the CLL patients with unmutated immunoglobulin heavy chain variable (IGHV) gene (p = 0.025), CD38-positive (p = 0.047), del(17p13) (p = 0.006). Both fludarabine and Nutlin-3 significantly downregulated DEK in the primary CLL cells which were with normal function of p53, or without deletion or mutation of p53 (p = 0.042, p = 0.038; p = 0.021, p = 0.017; p = 0.037, p = 0.017). However, the downregulation of DEK was not observed in the primary CLL cells which were with dysfunction of p53, or with deletion or mutation of p53 (p = 0.834, p = 0.477; p = 0.111, p = 0.378; p = 0.263, p = 0.378). These data show that DEK might be applied for the assessment of prognosis in patients with CLL, and fludarabine and Nutlin-3 regulate DEK expression depended on p53 status.

High expression of cyclic nucleotide phosphodiesterase 7B mRNA predicts poor prognosis in mantle cell lymphoma

In order to determine the relationship between expression of cyclic nucleotide phosphodiesterase isoform 7B (PDE7B) and mantle cell lymphoma (MCL) progression, PDE7B mRNA expression was measured by qRT-PCR in 21 untreated MCL patients with bone marrow involvement and 20 healthy donors. The expression level of PDE7B was markedly higher in MCL patients compared with normal controls (P=0.001), and the higher level of PDE7B expression was associated with unfavorable cytogenetic characteristics in MCL. It was showed that higher expression of PDE7B might be a novel unfavorable prognostic indicator in MCL, which possess important clinical significance.

The BH3-only protein Puma plays an essential role in p53-mediated apoptosis of chronic lymphocytic leukemia cells.

Abstract The purpose of this study was to explore the characteristics and functions of BH3-only proteins Puma, Noxa and Bim in the prognosis, therapy and drug resistance of chronic lymphocytic leukemia (CLL). Puma, Noxa and Bim mRNAs were evaluated by real-time quantitative reverse transcriptase-polymerase chain reaction, and correlations between their expression levels and CLL prognostic markers were analyzed. Primary CLL samples were treated in vitro with fludarabine to investigate the role of Puma, Noxa and Bim in the response to chemotherapeutic drugs which act through activation of the p53 pathway. We found that a low expression level of Puma was associated with some markers of poor prognosis. However, the level of Noxa or Bim was not different in patients with CLL with variant clinical features and prognostic factors. Puma expression was up-regulated after fludarabine treatment in primary CLL cells, but there was no significant difference for Noxa and Bim. Up-regulation of Puma occurred only in CLL cells with functional p53. CLL cells with p53 abnormalities were deficient in the activation of Puma by chemotherapeutics. These results suggest that a lack of Puma induction may contribute to the development of resistance to anticancer agents in CLL.

Polyclonal Antibody Targeting SOX11 Cannot Differentiate Mantle Cell Lymphoma From B-Cell Non-Hodgkin Lymphomas

OBJECTIVES To determine whether SOX11 is a diagnostic marker of mantle cell lymphoma (MCL). METHODS We analyzed SOX11 expression in 349 B-cell non-Hodgkin lymphomas (B-NHLs) via immunohistochemistry. RESULTS Nuclear staining of SOX11 was observed in 54 (93.1%) of 58 MCLs. We noticed that SOX11 protein was also expressed on the nuclei in 8 (21.6%) of 37 B-lymphoblastic lymphomas, 45 (32.6%) of 138 diffuse large B-cell lymphomas, 15 (44.1%) of 34 follicular lymphomas, 8 (30.8%) of 26 Burkitt lymphomas, 2 (10.0%) of 20 chronic lymphocytic leukemia/small cell lymphomas, and 3 (18.8%) of 16 marginal zone lymphomas. CONCLUSIONS Although the positive rate of SOX11 expression in MCL was significantly higher than other B-NHLs (P < .001), polyclonal antibody targeting SOX11 is not able to identify MCL from B-NHLs because the nuclear staining of SOX11 was widely positive in B-NHLs.