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 Table of Contents  
ORIGINAL ARTICLE
Year : 2015  |  Volume : 40  |  Issue : 2  |  Page : 99-103

Expression of B-cell activating factor in acute lymphoblastic leukemia patients


1 Department of Clinical and Chemical Pathology, Ain Shams University, Cairo, Egypt
2 Department of Internal Medicine & Clinical Hematology, Ain Shams University, Cairo, Egypt

Date of Submission04-Apr-2015
Date of Acceptance15-May-2015
Date of Web Publication22-Jul-2015

Correspondence Address:
Emad A Abd El-hadi
10 Ahmed Saman Street (from Mostafa El-Nahas Street), Nasr City 11762, Cairo
Egypt
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Source of Support: None, Conflict of Interest: None


DOI: 10.4103/1110-1067.161296

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  Abstract 

Introduction Acute lymphoblastic leukemia (ALL) is a biologically heterogeneous disorder needing determination of morphologic, immunologic, cytogenetic, biochemical, and molecular genetics characterizations of lymphoblasts to establish diagnosis. Microenvironmental cues play critical roles in cancer biology and malignant cells are responsive to multiple extrinsic factors. There is emerging evidence that B-cell activating factor (BAFF) is a critical factor for the growth and survival of both normal and malignant clones of B-cells and can augment tumor cell growth. There is evidence that B-lineage neoplasms have aberrant expression of BAFF.
Objectives The aim of this study was to evaluate serum BAFF levels to assess its prognostic impact on ALL and correlate it with mortality.
Patients and methods The study was conducted at Ain Shams University on 40 ALL pediatric and adult patients, in addition to 20 controls. Patients underwent detailed history and clinical examination, bone marrow examination, and evaluation of serum BAFF levels using enzyme-linked immunosorbent assay. Thereafter, they were assessed for treatment response and patients' mortality on day 28 from induction of chemotherapy and after 6 months, respectively.
Results We found a statistically significant lower BAFF levels among controls compared with the patient group, in which there was no significant difference between pediatric and adult subgroups. High serum BAFF was significantly correlated to patients' poor treatment response but not mortality.
Conclusion High BAFF in our studied patients may be explained by its role in augmenting B-cell tumor growth. In our study, we found that BAFF had no relation with disease outcome. This may be caused by the limited number of patients included. Assessment of BAFF level at the time of diagnosis may be a predictor for response to treatment, as we found a significant relation between BAFF level and response to treatment. This finding recommends that patients with high BAFF level at the time of diagnosis be subjected to intensified course of therapy.

Keywords: Acute lymphoblastic leukemia (ALL), B-cell activating factor (BAFF), expression


How to cite this article:
Abd El Fatah MF, Abd El-hadi EA, Helmy TA. Expression of B-cell activating factor in acute lymphoblastic leukemia patients. Egypt J Haematol 2015;40:99-103

How to cite this URL:
Abd El Fatah MF, Abd El-hadi EA, Helmy TA. Expression of B-cell activating factor in acute lymphoblastic leukemia patients. Egypt J Haematol [serial online] 2015 [cited 2019 Dec 12];40:99-103. Available from: http://www.ehj.eg.net/text.asp?2015/40/2/99/161296


  Introduction Top


Acute lymphoblastic leukemia (ALL) is a malignant proliferation of lymphoid cells blocked at an early stage of differentiation and accounts for three-fourth of all cases of childhood leukemia, with peak prevalence between the ages of 2 and 5 years. ALL is a biologically heterogeneous disorder, and thus morphologic, immunologic, cytogenetic, biochemical [lactate dehydrogenase (LDH) and uric acid], and molecular genetics characterizations of leukemia lymphoblasts are needed to establish the diagnosis, or to exclude other possible causes of bone marrow failure and, finally, to classify ALL subtypes. This heterogeneity reflects the fact that leukemia may develop at any point during the multiple stages of normal lymphoid differentiation [1] .

Increasing evidence indicates that microenvironmental cues play critical roles in cancer biology and that malignant cells are responsive to multiple extrinsic factors from their microenvironment. These stimuli involve both soluble factors and receptor/ligand interactions, which mediate or influence processes such as tumor development, maintenance, drug-resistance, and immune evasion. Studies indicate that the 'leukemia microenvironment' supports ALL cells developing in the bone marrow by providing survival/proliferation signals and by functioning as potential niches for chemotherapy-resistant tumor cells [2] .

There is emerging evidence that the tumor necrosis factor super-family member B-cell activating factor (BAFF) (along with its receptors) is a critical factor for the growth and survival of both normal and malignant clones of B-cells [3] . It has been reported that BAFF can augment tumor cell growth of B-cells by either stimulating proliferation, inhibiting apoptosis or protecting malignant cells against drug-induced apoptosis [4] . In accordance with this, there is emerging evidence that B-lineage neoplasms have aberrant expression of BAFF [5] .


  Objectives Top


The aim of this work was to evaluate the circulating levels of BAFF to assess its prognostic impact on ALL and correlate it with patients' outcome.


  Patients and methods Top


This study was conducted at Ain Shams University Hospitals on 40 newly diagnosed patients (24 male and 16 female patients) with ALL with a mean age of 20.23 years (ranging from 3 to 44 years), in addition to 20 healthy participants with matched age and sex as the control group. An informed consent was taken from all participants before participation and enrollment into the study.

Patients were initially assessed with the following:

Full detailed history and clinical examination, complete blood picture, bone marrow aspiration and examination (with immunophenotyping using flow cytometry to confirm ALL diagnosis using CD19, CD20, CD10, CD34, HLADR panel), and evaluation of circulating BAFF levels in serum using the enzyme-linked immunosorbent assay technique.

Thereafter, the patients were assessed on follow-up.

  1. At day 28 from induction, chemotherapy with bone marrow aspirate was carried out to assess treatment response.
  2. After a 6-month duration, patients' clinical outcome and mortality were assessed.


Evaluation of BAFF levels

BAFF level was measured for all enrolled patients at diagnosis and compared with BAFF serum levels of 20 age-matched and sex-matched controls. BAFF was quantitatively measured in serum using Human BAFF ELISA Kit-1 × 96 (Boster Biological Technology Ltd, Pleasanton, California, USA).

The test is based on standard sandwich enzyme-linked immune-sorbent assay technology. Human BAFF-specific polyclonal antibodies were precoated onto 96-well plates. The human specific detection monoclonal antibodies were biotinylated. The test samples and biotinylated detection antibodies were added to the wells subsequently and then followed by washing with PBS or TBS buffer. Avidin-biotin-peroxidase complex was added and unbound conjugates were washed away with PBS buffer. Polymerized horseradish peroxidase (HRP) substrate TMB was used to visualize HRP enzymatic reaction. Tetra methylbenzidine (TMB) was catalyzed by HRP to produce a blue color product that changed into yellow after adding acidic stop solution. The density of yellow is proportional to the human BAFF amount of sample captured in plate.

The reaction was monitored at 450 nm. Quantification of unknown samples is achieved by comparing their absorbance with a reference curve prepared with known standard concentrations.

Statistical methods

The data were coded, entered, and processed on an IBM-PC compatible computer using SPSS (Version 11, SPSS Inc., Chicago, Illinois). The level P value less than 0.05 was considered the cutoff value for significance. Statistical presentation and analysis of the present study was conducted, using the mean, SE, Student's t-test, χ2 -test, and linear correlation coefficient.

Quantitative data were described in the form of mean ± SD and range. Qualitative data were described in the form of number and percentage.

  1. Student's t-test: Unpaired Student's t-test was used to compare two groups (the patient group and the healthy control group) as regards quantitative data.
  2. χ2 -test: The χ2 -test was used to determine the association between variables for categorical data and for comparison between subgroups.
  3. Correlation analysis: Spearman's correlation was used to assess the strength and direction of the linear relationship between two variables.



  Results Top


This study was carried out on 40 newly diagnosed pediatric and adult ALL patients, of whom 24 (60%) were male and 16 (40%) were female, with a male-to-female ratio of 3:2. Their ages ranged from 3 to 44 years, with a median of 20.23 years. The patients were subdivided on the basis of their age into two groups: group I included 20 pediatric ALL patients with age range from 3 to 16 years (with a mean value of 7.95 ± 4.22) and group II included 20 adult ALL patients with age range from 19 to 44 years (with a mean value of 32.5 ± 7.85 years). Twenty healthy age-matched and sex-matched controls were included in the study, of whom 12 were male and eight were female; their ages ranged from 3 to 45 years (with a mean of 18.70 years).

As regards demographic data, hemoglobin concentration of patients ranged from 4.8 to 12.5 g/dl, with a mean of 8.03 g/dl ± 2.01. Their platelet count ranged from 6.5 to 220 × 10 9 /l, with a mean of 72 × 10 9 /l ± 72.46. Their total leukocytic count ranged from 0.1 to 350, with a mean of 33.48 × 10 9 /l ± 75.08. Their peripheral blood blasts ranged from 15 to 65%, with a mean of 38.50 ± 13.31. The bone marrow blast ranged from 46 to 97%, with a mean of 80.90% ± 10.98, and LDH concentration ranged from 550 to 1250 (IU/l), with a mean of 812.98 IU/l ± 195.68 ([Table 1]).
Table 1 Laboratory data of the studied B-ALL patients


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Comparison between the pediatric and adult B-ALL patient groups showed significant difference as regards sex, splenomegaly, and percentage of bone marrow-blasts, which was seen in 18 male adult patients (90%) and 14 female patients (70%) in the pediatric group. Notable splenomegaly was seen in 40% of adult patients compared with 10% of pediatric cases. In contrast, the pediatric group showed higher bone marrow-blasts% compared with the adult group (mean ± SD 86.7 ± 6.25 and 75.1 ± 11.72, respectively); otherwise, no significant difference was noted between the two groups as regards lymphadenopathy, hepatomegaly, hemoglobin and platelets levels, total leukocytic count, LDH, and peripheral blood blasts (%) ([Table 2]).
Table 2 Comparison between pediatric and adult B-ALL patients as regards clinical and laboratory data


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On comparing BAFF levels among patients and controls, there was statistically significant difference, being lower in the control group (range 200-320 pg/ml; mean 270 ± 34.18) compared with the patient group (range 550-5000 pg/ml; mean 1606.50 ± 1002.15 pg/ml) ([Table 3]).
Table 3 Comparison between BAFF levels in the control and B-ALL patient groups


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On comparing the pediatric and adult patient groups as regards BAFF level, there was insignificant difference between the two groups. In contrast, on comparing the two groups as regards response to treatment at day 28, there was statistically significant difference [16 (80%) patients of the pediatric group vs. eight (40%) patients of the adult group were in remission]. Moreover, there was statistically significant difference as regards patient outcome [i.e. in the pediatric group 20 (100%) patients survived, whereas in the adult group 10 (50%) patients survived]. On comparing all patients all together at day 28 for treatment response, 24 (60%) out of 40 patients were in remission, whereas 16 (40%) did not remit with statistically significant difference. In contrast, as regards mortality (i.e. patient outcome), 30 (75%) were alive out of 40 patients, whereas 10 (25%) patients died, with statistically nonsignificant difference ([Table 4]).
Table 4 Comparison between pediatric and adult B-ALL as regards response to treatment and patient outcome


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There was statistically significant correlation between all patients' BAFF level as regards treatment response (i.e. high levels of BAFF were associated with poor response to treatment and no remission). In contrast, as regards patient outcome, there was no statistically significant correlation between all patients' BAFF levels and patient outcome ([Table 5]).
Table 5 Correlation between BAFF in all B-ALL patients with their response to treatment and mortality


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  Discussion Top


Although significant progress has been made in the treatment of ALL, the prognosis following relapse is still poor. High-risk patients have been offered more intensive treatment at the time of diagnosis to improve leukemia-free survival [6] . However, intensive therapeutic strategies are also associated with increased treatment-related morbidity and mortality. Therefore, accurate assignment of relapse risk at diagnosis is essential for the optimal treatment of ALL [7] .

Indeed, the overall majority of patients experiencing disease relapse cannot be adequately assessed for their chance of experiencing relapse at diagnosis [8] . In recent years, the use of uniform criteria (including age, sex, race, leukemic burden, white blood cell count at presentation, serum immunoglobulins, early response to therapy, cytogenetics, central nervous system status, immunophenotyping, drug-resistance profiles, and minimal residual disease) to assign risk-based therapy for patients with ALL has been advocated. Assessment of these factors is mandatory for therapeutic assignment [9] .

The leukemic cell is phenotypically different from any normal cell, but the degree of difference is extremely variable. There is emerging evidence that the tumor necrosis factor super-family member BAFF, along with its receptors, is a critical factor for the growth and survival of both normal and malignant clones of B-cells [3] . It has been reported that BAFF can augment tumor cell growth of B-cells by either stimulating proliferation, inhibiting apoptosis or protecting malignant cells against drug-induced apoptosis [4] . In accordance with this, there is emerging evidence that B-lineage neoplasms have aberrant expression of BAFF [5] .

The present study aimed at determining the circulating levels of BAFF to assess its prognostic impact on ALL and correlate it with patients' outcome in 40 newly diagnosed ALL patients. In our study we found that response to treatment of the pediatric group is better than the response to treatment in the adult group, with a statistically significant difference in response to treatment between the pediatric and adult groups. This is in agreement with the study conducted by Hunger and colleagues, who confirmed that survival trends in ALL trials conducted by the Children's Oncology Group (COG) between 1990 and 2005 predict the 10-year survival of children with ALL who entered onto COG trials between 2006 and 2010 and will approach or exceed 90%. They also believe that it is extremely unlikely that there will be a significant increase in deaths beyond 5 years for patients diagnosed between 2000 and 2005 [10] .

As regards BAFF results in our study, it showed high levels in pediatric and adult patients, with a mean ± SD of 1606.50 ± 1002.15 pg/ml and a range of 550-5000 pg/ml, which was high compared with the control group (range: 250-320; mean ± SD: 18.70 ± 13.58). Similar results were obtained by Bienertova-Vasku et al. [3] , who confirmed that BAFF has high levels in acute leukemia, with a mean ± SD of 7764 ± 6329. This may be explained by the fact that BAFF can augment tumor cell growth of B-cells by either stimulating proliferation, inhibiting apoptosis or protecting malignant cells against drug-induced apoptosis.

In contrast, BAFF in chronic lymphocytic leukemia behaved in a different way. Haiat et al. [11] compared the level of serum BAFF in the serum of B-chronic lymphocytic leukemia patients versus normal controls and found significantly lower levels of serum BAFF in the sera of B-chronic lymphocytic leukemia patients (920 ± 54 ng/ml) compared with normal controls (1183 ± 76 ng/ml) (P = 0.0334), suggesting that the circulating BAFF might be overconsumed by leukemic cells.

In our study we found that BAFF had no relation with prognostic factors and disease outcome. This may be caused by the limited number of patients included in the study. Moreover, our study included pediatric and adult ALL patients who had different prognosis, as pediatric ALL patients had better prognosis compared with adult ALL patients. Assessment of BAFF level at the time of diagnosis may be a predictor for response to treatment as we found a significant relation between BAFF level and response to treatment. This finding recommends that patients with high BAFF level at the time of diagnosis should be subjected to intensified course of therapy. Therefore, Bienertova-Vaskua et al. [3] categorized patients with high BAFF level as high-risk ALL patients (especially if BAFF level was more than 10 000 pg/ml).

The majority of ALL cases are of B-cell lineage. Heisterkamp and Parameswaran had previously discovered that these leukemia cells have a protein on their surface called BAFF-receptor (BAFF-R). It was known that this BAFF-R was present on mature B-cells, but finding it on pre-B-cells was surprising and also presented a therapeutic target for selectively killing the pre-B-ALL cells. The investigators made use of the fact that a protein called BAFF specifically binds to the BAFF-R protein and then is allowed entry into the cell. They tested a toxin-BAFF fusion protein. Using a 'Trojan horse' approach, the investigators showed that when ALL cells were exposed to the BAFF-toxin, the ALL cells bound to the BAFF-toxin, transported it inside the cell, and were then killed. The BAFF-R is only present on certain blood-forming cells and so the BAFF-toxin is not expected to harm any other cells, making it much less toxic compared with standard chemotherapy [12] .


  Conclusion Top


We concluded that BAFF analysis at diagnosis and at day 28, in addition to other prognostic markers, may contribute to improvement in the management of ALL patients. Moreover, high BAFF level in ALL patients indicates its role in ALL pathogenesis. Although BAFF has no relation with prognosis, it shows significant relation with response to treatment and it could be used as a predictor for response to treatment.

From the data mentioned above, we recommend that further studies be conducted on a wide scale of pediatric or adult ALL patients for accurate assessment of prognostic value of BAFF expression in pediatric ALL and/or adult ALL. In addition, we recommend further assessment of patients for high, low, and intermediate risk in correlation with BAFF levels.


  Acknowledgements Top


Conflicts of interest

There are no conflicts of interest.

 
  References Top

1.
Pidala J, Djulbegovic B, Anasetti C, Kharfan-Dabaja M, Kumar A. Allogeneic hematopoietic cell transplantation for adult acute lymphoblastic leukemia (ALL) in first complete remission. Cochrane Database Syst Rev 2011; 5 :CD008818.  Back to cited text no. 1
    
2.
Maia S, Pelletier M, Ding J, Hsu YM, Sallan SE, Rao SP, et al. Aberrant expression of functional BAFF-system receptors by malignant B-cell precursors impacts leukemia cell survival. PLoS One 2011; 6 :e20787.  Back to cited text no. 2
    
3.
Bienertova-Vasku J, Bienert P, Kodytkova D, Zlamal F, Tomandl J, Tomandlova M, et al. BAFF Levels are elevated in pediatric patients with acute lymphoblastic leukaemia compared to other B-lineage neoplasms. J Hematol 2012; 1 :20-22.  Back to cited text no. 3
    
4.
Lied GA, Berstad A. Functional and clinical aspects of the B-cell-activating factor (BAFF): a narrative review. Scand J Immunol 2011; 73 :1-7.  Back to cited text no. 4
    
5.
He B, Chadburn A, Jou E, Schattner EJ, Knowles DM, Cerutti A. Lymphoma B cells evade apoptosis through the TNF family members BAFF/BLyS and APRIL. J Immunol 2004; 172 :3268-3279.  Back to cited text no. 5
    
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Forestier E, Izraeli S, Beverloo B, Haas O, Pession A, Michalová K, et al. Cytogenetic features of acute lymphoblastic and myeloid leukemias in pediatric patients with Down syndrome: an iBFM-SG study. Blood 2008; 111 :1575-1583.  Back to cited text no. 6
    
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Caglar K, Varan A, Akyuz C, Rondelli R. Second neoplasms in pediatric patients treated for cancer: a center′s 30-year experience, J Pediatr Hematol Oncol 2006; 28 :374-378.  Back to cited text no. 7
    
8.
Khan NI, Cisterne A, Devidas M, Larson R. Expression of CD44, but not CD44v6, predicts relapse in children with B cell progenitor acute lymphoblastic leukemia lacking adverse or favorable genetics. Lymphoma 2008; 49 :710-718.  Back to cited text no. 8
    
9.
Mi JQ, Wang X, Yao Y, Lu HJ, Jiang XX, Zhou JF, et al. Newly diagnosed acute lymphoblastic leukemia in China (II): prognosis related to genetic abnormalities in a series of 1091 cases. Leukemia 2012; 26 :1507-1516.  Back to cited text no. 9
    
10.
Hunger SP, Lu X, Devidas M, Camitta BM, Gaynon PS, Winick NJ, et al. Improved survival for children and adolescents with acute lymphoblastic leukemia between 1990 and 2005: a report from the children′s oncology group. J Clin Oncol 2012; 30 :1663-1669.  Back to cited text no. 10
    
11.
Haiat F, Billard C, Quiney C, Ajchenbaum FC, Kolb J. Role of BAFF and APRIL in human B-cell chronic lymphocytic leukemia. Immunology 2006; 118 :281-292.  Back to cited text no. 11
    
12.
Heisterkamp N , Parameswaran R. Saban Research Institute of Children′s Hospital Los Angeles: Effector-mediated eradication pf precursor B-ALL with a novel Fc-engineered monoclonal antibody targeting the BAFF-R; Mol Cancer Ther 2014; 13 :1567-1577.  Back to cited text no. 12
    



 
 
    Tables

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



 

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Abstract
Introduction
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Patients and methods
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