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 Table of Contents  
ORIGINAL ARTICLE
Year : 2016  |  Volume : 41  |  Issue : 3  |  Page : 128-131

The prognostic role of chemokine ligand-3 in chronic lymphocytic leukemia


1 Department of Clinical and Chemical Pathology, Faculty of Medicine, Benha University, Banha, Egypt
2 Department of Clinical and Chemical Pathology, Faculty of Medicine, Ain Shams University Hospital, Cairo, Egypt
3 Department of Clinical Pathology, Faculty of Medicine, Benha University, Banha, Egypt

Date of Submission30-Apr-2016
Date of Acceptance07-May-2016
Date of Web Publication27-Dec-2016

Correspondence Address:
Yasmin N El-Sakhawy
Department of Clinical and Chemical Pathology, Faculty of Medicine, Ain Shams University, Cairo
Egypt
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Source of Support: None, Conflict of Interest: None


DOI: 10.4103/1110-1067.196196

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  Abstract 

Introduction Chemokines are responsible for dissemination and survival of many malignant tumors including chronic lymphocytic leukemia (CLL). Chemokine ligand-3 (CCL3), previously known as macrophage inflammatory protein-1a, has been found to be secreted from CLL cells in response to B-cell receptor activation, enhancing the interaction between CLL cells and the leukemia microenvironment. In the present study, we measured CCL3 serum levels by using the enzyme-linked immunosorbent assay in 40 CLL patients and examined CCL3 levels for associations with established prognostic markers. A significant increase in the serum level of CCL3 in CLL patients (mean=147.2±62.3?pg/ml) was found in comparison with normal healthy controls (8.2±23.7?pg/ml); in addition, this increase in the CLL serum levels was significantly correlated with established prognostic factors as CD38 and the advanced stages of Rai and Binet staging system.
Conclusion CCL3 serum level could be a valuable and independent prognostic marker and should be useful in risk assessment in patients with CLL, and it may provide insights into creating a new therapeutic modality.

Keywords: cd38, chemokine ligand-3, chronic lymphocytic leukemia


How to cite this article:
Tolbah FM, Kamal HM, El-behisy MM, El-Sakhawy YN, Abd-El Latif S. The prognostic role of chemokine ligand-3 in chronic lymphocytic leukemia. Egypt J Haematol 2016;41:128-31

How to cite this URL:
Tolbah FM, Kamal HM, El-behisy MM, El-Sakhawy YN, Abd-El Latif S. The prognostic role of chemokine ligand-3 in chronic lymphocytic leukemia. Egypt J Haematol [serial online] 2016 [cited 2019 Dec 10];41:128-31. Available from: http://www.ehj.eg.net/text.asp?2016/41/3/128/196196


  Introduction Top


Chronic lymphocytic leukemia (B-CLL) is characterized by the accumulation of monomorphic CD5 positive small round B-lymphocytes in peripheral blood, bone marrow (BM), spleen, and/or lymph nodes [1]. Within the BM and secondary lymphoid tissue, CLL cells engage in complex interaction with stromal cell matrix and leukocytes. These are collectively referred to as the microenvironment, and are thought to promote CLL survival and proliferation [2] by displaying signs of B-cell receptor (BCR) activation [2],[3].

The chemokine ligand-3 (CCL3) − previously called macrophage inflammatory protein-1α − is a chemokine of the CC subfamily and inducible in a number of hematopoietic cells, particularly in those involved in adaptive immune responses (macrophages, dendritic cells, B and T lymphocytes) [4].

By analyzing the impact of the microenvironment on CLL gene expression in vitro, it was found that CLL cells upregulate and secrete CCL3 in response to BCR stimulation, inducing trafficking and homing of accessory cells to the malignant B cells and thus promoting the survival of these malignant cells [2]. This BCR induction of CCL3 was also found to be sensitive to the inhibition of BCR signaling using a spleen tyrosine kinase inhibitor [2],[5]. The importance of CCL3 level has been lately highlightened as a biomarker for BCR activation of CLL cells and for response assessment after therapies targeting the BCR-signaling pathway [6].

The aim of the present study was to evaluate the clinical significance of CCL3 in CLL and its value as a prognostic factor for CLL patients, in relationship with other well-established prognostic factors, mainly the CD38 expression and CLL stage of the disease.


  Patients and methods Top


The present study was carried out on 40 adult patients presenting with CLL. A written consent was signed by all patients. They were selected from the Hematology/Oncology Clinics of Ain Shams and Benha University Hospitals during the period from August 2014 to July 2015. Their ages ranged from 35 to 80 years, with a mean age of 57.8±8.8 years. They were 25 males and 15 females, with a male to female ratio of 1.7 : 1. In addition, 40 age and sex-matched healthy individuals were included in the study as a control group. They were 21 males and 19 females, with a male to female ratio of 1.1 : 1.

All patients were subjected to the following: full history taking and clinical examination, complete blood count, and immunophenotyping of the BM or whole peripheral blood using a Coulter EPICS-XL flow cytometer with system II software (USA). The following panel of monoclonal antibodies was used: CD5/CD19, CD20, CD79b, CD23, s-IgM, CD10, CD103, CD123, CD38, FMC7, κ and λ light chain − all obtained from Coulter Beckman (Florida, USA). Positivity threshold was defined as expression of more than or equal to 20% of the cells for the marker. Analysis for CCL3 serum level was carried out using an enzyme-linked immunosorbent assay on Cusabio Kits (SB-E04662h), according to the manufacturer’s instructions (R&D systems).

Statistical analysis

Statistical analyses of data in the present study were carried out using the computer program SPSS, version 16 (SPSS Inc., Chicago, Illinois, USA). Student’s t-test was used to compare between the means of two groups of numerical (parametric) data and analysis of variance for more than two groups of numerical (parametric) data. For continuous nonparametric data, the Mann–Whitney U-test was used. Pearson correlation coefficient (r) test was used to correlate different parameters. The sensitivity and specificity were examined at different cutoff points using the receiver operating characteristic curve analysis to determine the best cutoff point as well as the diagnostic power of each test.


  Results Top


The present study was carried out on 40 adult patients presenting with CLL ([Table 1]).
Table 1 Clinical and laboratory data of cases

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There was a highly significant elevation in CCL3 in the patient group compared with the control group ([Table 2]). Regarding the relation of CCL3 levels to the different laboratory prognostic markers, there was a highly significant negative correlation between CCL3 level and hemoglobin level and platelet count. In addition, there was a highly significant correlation between CCL3 level from one side and Rai staging, Binet staging, and CD38 level from the other side ([Table 3]). There was also a highly significant difference in CCL3 level between different Rai and Binet stages, with high levels in stages III and IV and Binet stage C, respectively ([Table 4]).
Table 2 Comparison of cases and control regarding CCL3 serum level

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Table 3 Correlation between CCL3 and different variables

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Table 4 Comparison of CCL3 serum level in Rai and Binet stages

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However, no correlation was found between CCL3 level and age, sex, total leukocytic count (TLC), and absolute lymphocytic count.

In addition, CCL3 was found to have high sensitivity (95%) and specificity (95%) in CLL patients compared with CD38 ([Table 5] and [Figure 1]).
Table 5 Area under the curve of ROC curves of CCL3 and CD38

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Figure 1 ROC curve comparing between CCL3 & CD38.

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  Discussion Top


BCR signaling is a central pathogenetic mechanism in CLL. Most CLL cells express both IgM and IgD receptors. Increased responsiveness to IgM stimulation correlates with the expression of immunoglobulin heavy chain variable region (IGHV)-unmutated immunoglobulin receptors (U-CLL) and with poor disease outcome. Recently it was revealed that IgM stimulation induces a long-lived signaling response, increased CLL-cell survival, and CCL3 secretion [7]. New therapeutic agents that interfere with BCR signaling, such as spleen tyrosine kinase, are emerging and show promising results in patients with CLL and other B-cell malignancies [8].

In agreement with many studies [6][9], the present study showed that patients with CLL had higher levels of CCL3 than did healthy controls. Yan et al. [9] reported that CCL3 was the most effective in distinguishing patients of CLL from healthy individuals. Burger et al. [10] recently demonstrated that CLL B cells produce high levels of CCL3 and its related chemokine CCL4 in a coculture with CD68+ nurselike cells after BCR stimulation.

The present study revealed a nonsignificant negative correlation between CCL3 serum levels and TLC, and this was in contrast to Sivina et al. [6], who revealed that if the TLC was less than 100 000/μl, there would be a nonsignificant correlation between them, whereas there was a high significant correlation if TLC was greater than 100 000/μl.

This study revealed a highly significant negative correlation between CCL3 and both hemoglobin concentration and platelet count, a finding that was consistent with those of other studies [6],[7]. However, the clinical staging systems developed by Rai and Binet remain the standard methods for risk assessment in CLL [6]. In the present study, the increased concentration of CCL3 in CLL patients was found to correlate with these classical prognostic staging systems (Rai and Binet). CCL3 level was significantly higher in patients with advanced Rai (stage III and IV) and Binet (stage C) staging compared with an early-staged disease. These finding were compared with those reported by Sivina and colleagues in 2011, who, in 2014, measured the CCL3 in serial samples collected over a period of time (ranging from 5 months to 12 years) in untreated CLL patients, concluding that CCL3 levels presumably reflect the dynamic changes of CLL pathogenesis over time, strengthening the relevance of CCL3 chemokine production in CLL [6],[7].

In addition, there was a highly significant positive correlation between CCL3 plasma level and CD38 status (P=0.001). This finding was also reported by Sivina et al. [6] in their study, where CCL3 level was significantly higher on the CD38+ subset compared with CD38 counterparts. Concomitantly, Zucchetto and colleagues demonstrated that CD38+CD49d+ CLL cells selectively and aberrantly express CCL3 through interaction with CD38 and CD31 expressed on stromal cells in the BM microenvironment. CCL3 subsequently induces recruitment of macrophage-lineage cells into the leukemia niche. The recruited cells produce inflammatory factors including tumor necrosis factor-α that eventually activate stromal cells to express vascular cell adhesion molecule-1, a ligand for CD49d that delivers prosurvival signals to CLL cells through its interaction with CD49d. Thus, CCL3 can promote establishment of the leukemia niche, which is essential for CLL-cell survival [11].

In the present study, the receiver operating characteristic curve demonstrated that specificity and sensitivity of CCL3 were more than those of CD38. However, a detailed analysis of a larger number of sequential samples in correlation to clinical and other prognostic factors needs to be performed to better understand the value of CCL3 in disease activity.


  Conclusion Top


The present study concluded the validity of measurement of CCL3 serum levels for prognostication in CLL. The simplicity of the assay using the enzyme-linked immune sorbent assay technique, the significance of the prognostic information, the relationship to BCR signaling, and the emerging therapies related to this pathway indicate that CCL3 will become a highly useful prognostic marker in CLL and potentially other B-cell malignancies.

Financial support and sponsorship

Nil.

Conflicts of interest

There are no conflicts of interest.

 
  References Top

1.
Goldin LR, Slager SL, Caporaso NE. Familial chronic lymphocytic leukemia. Curr Opin Hematol 2010; 17:350–355.  Back to cited text no. 1
    
2.
Burger JA, Grubben JG. The microenviroment in chronic lymphocytic leukemia (CLL) and other B cell malignancies: insight into disease biology and new targeted therapies. Semin Cancer Biol 2014; 24:71–81.  Back to cited text no. 2
    
3.
Herishanu Y, Pérez-Galán P, Liu D, Biancotto A, Pittaluga S, Vire B et al. The lymph node microenvironment promotes B-cell receptor signaling, NF-kappaB activation, and tumor proliferation in chronic lymphocytic leukemia. Blood 2011; 117:563–574.  Back to cited text no. 3
    
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Rusak M, Eljaszewicz A, Bołkun Ł, Łuksza E, Łapuć I, Piszcz J et al. Prognostic significance of PD-1 expression on peripheral blood CD4+ T cells in patients with newly diagnosed chronic lymphocytic leukemia. Pol Arch Med Wewn 2015; 125:553–559.  Back to cited text no. 4
    
5.
Langerbeins P, Busch R, Anheier N, Dürig J, Bergmann M, Goebeler ME et al. Poor efficacy and tolerability of R-CHOP in relapsed/refractory chronic lymphocytic leukemia and Richter transformation. Am J Hematol 2014; 89:239–243.  Back to cited text no. 5
    
6.
Sivina M, Hartmann E, Kipps TJ, Rassenti L, Krupnik D, Lerner S et al. CCL3 (MIP-1α) plasma levels and the risk for disease progression in chronic lymphocytic leukemia. Blood 2011; 117:1662–1669.  Back to cited text no. 6
    
7.
Sivina M, Werner L, Rassenti L, Wierda WG, Keating MJ, O’Brien S et al. Dynamics changes in CCL3 and CCL4 plasma chemokine levels in patients with chronic lymhocytic leukemia (CLL) managed with observation. Blood 2014; 124:21.  Back to cited text no. 7
    
8.
Friedberg JW, Sharman J, Sweetenham J, Johnston PB, Vose JM, Lacasce A et al. Inhibition of Syk with fostamatinib disodium has significant clinical activity in non-Hodgkin lymphoma and chronic lymphocytic leukemia. Blood 2010; 115:2578–2585.  Back to cited text no. 8
    
9.
Yan XJ, Dozmorov I, Li W, Yancopoulos S, Sison C, Centola M et al. Identification of outcome-correlated cytokine clusters in chronic lymphocytic leukemia. Blood 2011; 118:5201–5210.  Back to cited text no. 9
    
10.
Burger JA, Quiroga MP, Hartmann E, Bürkle A, Wierda WG, Keating MJ, Rosenwald A. High-level expression of the T-cell chemokines CCL3 and CCL4 by chronic lymphocytic leukemia B cells in nurse like cell cocultures and after BCR stimulation. Blood 2009; 113:3050–3058.  Back to cited text no. 10
    
11.
Zucchetto A, Benedetti D, Tripodo C, Bomben R, Dal Bo M, Marconi D et al. CD38/CD31, the CCL3 and CCL4 chemokines, and CD49d/vascular cell adhesion molecule-1 are interchained by sequential events sustaining chronic lymphocytic leukemia cell survival. Cancer Res 2009; 69:4001–4009.  Back to cited text no. 11
    


    Figures

  [Figure 1]
 
 
    Tables

  [Table 1], [Table 2], [Table 3], [Table 4], [Table 5]



 

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