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ORIGINAL ARTICLE
Year : 2019  |  Volume : 32  |  Issue : 3  |  Page : 983-990

Effect of urokinase plasminogen activator receptor (CD87) on patients with de-novo acute myeloid leukemia


1 Department of Clinical Pathology, Faculty of Medicine, Menoufia University, Shebin El Kom, Egypt
2 Clinical Oncology Department and Hospital, Faculty of Medicine, Menoufia University, Shebin El Kom, Egypt
3 Department of Clinical Pathology, Shebin El Kom Teaching Hospital, Shebin El Kom, Egypt

Date of Submission24-Mar-2018
Date of Acceptance20-May-2018
Date of Web Publication17-Oct-2019

Correspondence Address:
Arwa Abd El Hamed Hijii
Al Sadat Street, Shebin El Kom, Menoufia 32511
Egypt
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Source of Support: None, Conflict of Interest: None


DOI: 10.4103/mmj.mmj_135_18

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  Abstract 


Objectives
The objective of this study was to study the role and the prognostic value of urokinase-type plasminogen activator receptor (uPAR) (CD87) expression in patients with de-novo acute myeloid leukemia (AML).
Background
The physiological function of the uPAR (CD87) is concerned in various biological processes and signal transduction such as cancer metastasis, angiogenesis, cell migration, and wound healing. The uPAR is expressed in most solid cancer and in several hematological malignancies including myeloproliferative disease, acute leukemia (AML, acute lymphocytic leukemia), and multiple myeloma.
Patients and methods
This study was conducted on 52 newly diagnosed patients with AML and 20 age-matched and sex-matched healthy individuals as a control group. Full history taking, clinical examination, and laboratory investigations were done for all patients. Expression of CD87 was evaluated by flow cytometric analysis.
Results
Positive CD87 expression (CD87+) was significantly higher in patients with AML than in healthy individuals. The highest incidence of CD87+ was found in acute myelomonocytic leukemia and acute monoblastic leukemia (M4/M5). High CD87+ expression in patients with AML predicted poor response to therapy, poor outcome, and shorter overall survival rate.
Conclusion
The high expression of CD87 displays a negative prognostic effect on patients with AML regarding outcome, survival rate, and response to chemotherapy. However, more studies are required to elucidate its role as a target therapy for those patients as its expression is highly restricted to tumor cells.

Keywords: acute myeloid leukemia, flow cytometry, prognosis, urokinase-type plasminogen activator receptor


How to cite this article:
Kandil SH, Ahmedy IA, Soliman GE, El Hamed Hijii AA. Effect of urokinase plasminogen activator receptor (CD87) on patients with de-novo acute myeloid leukemia. Menoufia Med J 2019;32:983-90

How to cite this URL:
Kandil SH, Ahmedy IA, Soliman GE, El Hamed Hijii AA. Effect of urokinase plasminogen activator receptor (CD87) on patients with de-novo acute myeloid leukemia. Menoufia Med J [serial online] 2019 [cited 2019 Nov 14];32:983-90. Available from: http://www.mmj.eg.net/text.asp?2019/32/3/983/268806




  Introduction Top


Acute myeloid leukemia (AML) is a dissimilar neoplastic disorder characterized by unrestrained production of hematopoietic stem cells. The development of AML involves a variety of mechanisms, for example, mutations in the transcriptional factors or mutations in the epigenetic modifiers, aberrant signaling pathways, overexpression of the multiple genes concerning drug resistance, abnormal immune function, and abnormalities in the bone marrow (BM) microenvironment [1].

Approximately 70–80% of patients with de-novo AML achieve an initial complete remission after chemotherapy, but long-term free survival remains as low as 30–50% [2].

The composition of urokinase-type plasminogen activator (uPA) system consists of a proteinase (the uPA) and its receptor [the urokinase-type plasminogen activator receptor (uPAR) (CD87)] [3].

The uPAR performs an important role in plasminogen activation, cell adhesion, and migration – all central aspects of inflammatory processes and numerous disease prognosis [4].

The uPAR is normally expressed on monocytes, eosinophils, neutrophilic granulocytes, skin mast cells, and dendritic cells. However, it is not expressed in CD34+ BM cells, erythrocytes, platelets (PLTs), or resting B lymphocytes and T lymphocyte. However, activated T lymphocytes and natural killer cells express this antigen [5].

Patients with various human cancers showing high levels of uPAR in tumor tissues and plasma are associated with the poor prognosis, increased risk of tumor recurrence, and metastasis [6].

We aimed to study the role of uPAR (CD87) in AML.


  Patients and Methods Top


This study was carried out, under ethical contributions, on patients with AML in the period from January 2016 to January 2018. The laboratory investigations were done in the Clinical Pathology department, Menoufia University. Informed consent was obtained from both patients and controls. The present study involved 52 patients with AML and 20 age-matched and sex-matched healthy individuals as a control group.

All participants were submitted to the following: clinical history taking and full physical examination. Complete blood count was performed using XT-1800i hematology analyzer (Sysmex, Kobe, Japan), which is based on fluorescent flow technology and hydrodynamic focusing. Peripheral blood film was performed by specialist. The CD87 expression is quantified on the leukemic blast cell by flow cytometric analysis by FACS Calibur flow cytometer in peripheral blood and/or BM aspiration (BD immune cytometry systems; BD Bioscience, San Jose, California, USA). BM film and conventional cytogenetic and/or molecular genetics (by qualitative PCR) were done for patients only.

Blood sampling included venous blood samples (5 ml)/or BM samples (1 ml) aseptically withdrawn on EDTA solution vacutainer tubes from all studied groups.

Sample preparation and staining procedures were as follows: anti-CD34 Fluorescein isothiocyanate (FITC) mouse monoclonal antihuman antibodies (MoAbs) (Immunostep, S.L., Salamanca, Spain) and anti-CD87 Phycoerythrin (PE) MoAbs (Immunostep) were used. Overall, 10 μl of the labeled monoclonal antibodies (MoAbs) were dispensed into all appropriately labeled tubes, and then 100 μl of the sample was added and incubated in the dark for 20 min. Lysing reagent (3.5 ml) (Bio-Rad; Medical Diagnostics GmbH, Dreieich, Germany) was added to each tube and inverted once and then kept for 3 min, followed by centrifugation at 3200 rpm for 3 min. Then 3.5 ml PBS was added and centrifuged again at 3200 rpm for another 3 min.

Flow cytometric analysis of CD87 and CD34 was done as follows: Cell Quest software (BD Biosciences) was used to calculate positive events in the defining gate (CD45 dim population was gated which are the blasts) [Figure 1] compared with the isotype control. CD87 was considered positive when 20% of the blast cells express CD87 [Figure 2]a and [Figure 2]b.
Figure 1: CD45 gating: myeloblasts were shown in red. ECD phycoerythrin-Texas Red conjugate (energy coupled dye), SS, side light scatter.

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Figure 2: (a and b) CD87 expression in patients with AML. AML, acute myeloid leukemia.

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Classification according to prognosis was follows: good prognosis: complete recovery obtained after two cycles or more of the chemotherapy and poor prognosis: residual disease or extramedullary involvement after two cycles or more of the chemotherapy or the patient relapsed or even died during 5 months' therapy.

Statistical analysis

Data collected were tabulated and analyzed by statistical package for the social science (SPSS, version 20; SPSS Inc., Chicago, Illinois, USA) on an IBM personal computer. Level of significance was set at P value equal to 0.05.


  Results Top


The present study involved 52 patients with AML, comprising 26 (50%) male and 26 (50%) female, with age ranged between 19 and 83 years old (mean ± SD, 36.91 ± 15.58 and a median, 40 years), and 20 age-matched and sex-matched healthy individuals as a control group, comprising nine (45%) males and 11 (55%) females with ages range from 18 to 84 years, with a mean of 37.0 ± 17.47 years. We found out no correlation between CD87 and age or sex. The cutoff value of a CD87 expression is considered positive when more than or equal to 20%. Moreover, 20/52 AML cases were CD87+ whereas 32/52 were CD87 expression [Figure 2]a and [Figure 2]b. All control groups were CD87. We revealed that AML group showed statistically significant higher CD87 expression level than the control group (P < 0.001) [Table 1] and [Table 2].
Table 1: Comparison between the two studied groups according to different parameters

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Table 2: Relation between CD87 and demographic data in acute myeloid leukemia cases (n=52)

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Regarding clinical data, we found out that hepatomegaly was found in 12/20 (60%) CD87+ patients and in 9/32 (28.1%) CD87 patients, splenomegaly in 12/20 (60%) CD87+ patients and in 11/32 (34.4%) CD87 patients, lymphadenopathy in 2/20 (10%) CD87+ patients and in 1/32 (3.1%) CD87 patients and central nervous system (CNS) infiltration in 1/20 (5%) CD87+ patients and in 2/32 (6.3%) CD87 patients. Moreover, hepatomegaly shows positive correlation with CD87+ (P = 0.023) whereas splenomegaly, CNS infiltration, and lymphadenopathy did not show any significant statistical relationship between them and CD87 expression [Table 2].

Regarding hematological data, total leukocytic count (76.91 ± 68.48 × 109/l), hemoglobin (7.79 ± 1.53 g/dl), and PLT count (86.40 ± 79.53 × 103/mm 3), did not show any significant statistical relationship correlating them and CD87+ expression [Table 2].

The incidence of CD87+ cells in AML French–American–British (FAB) subtypes was in the following descending order: M4 (12/20, 60%) more than M5 (4/20, 20%) more than M1 (2/20, 10%) more than M2 (1/20, 5%) more than M3 (1/20, 5%) more than M7 (0/20, 0%). In addition, there was a positive correlation between both CD87+ and M4 (P = 0.002) and M5 (P = 0.018), whereas there was a negative correlation between CD87 and M2 (P = 0.005) [Table 2].

Regarding immunophenotypic markers, CD34 was positive in 6/49 cases, whereas CD117 was positive in 19/38 cases. There was no significant relationship between the expression of CD87+ and both CD34 (P = 0.813) or CD117 (P = 0.304) [Table 2]. The favorable cytogenetics abnormalities were found in 46/49 patients: five patients t(8:21), three patients inv (16) and eight patients t(15:17) and 30 patients normal karyotype. The favorable cytogenetics cases were further divided into 16/18 (88.9%) CD87+ patients and 30/31 (96.8%) CD87− patients, meanwhile the unfavorable cytogenetics were found in 3/49, one patient t (9:11) and two patients t(9:22). The unfavorable cytogenetics cases were further divided into 2/18 (11.1%) CD87+ cases and 1/31 (3.2%) CD87 negative cases. Thus, there were no significant relationship between CD87 and the cytogenetic information [Table 2].

In the present study, we also correlated CD87 expression in 52 patients with the outcome of patients with AML after follow-up for 6 months following induction chemotherapy. Regarding the CD87+ cases (20), 1/20 (5.0%) was alive and 16/20 (80.0%) died whereas 3/20 (15%) were lost to follow-up. However, we depicted among the CD87 cases (32) that 12/32 (37.5%) were alive and 18/32 (56.3%) died whereas 2/32 (6.3%) were lost to follow-up. Consequently, CD87+ patients had a poor outcome than CD87 patients (P = 0.016) [Table 2]. Furthermore, our study revealed that CD87+ patients have shorter overall survival (mean, 1.54 ± 1.40 and median, 1.0) than patients with CD87 (mean, 3.09 ± 2.31 and median, 2.0) [Table 2].

In our study, 21 patients achieved complete remission whereas 26 patients did not achieve it, and five were lost to follow-up. Achievement of complete remission was correlated with all variable in the study and showed the following relation with FAB subtypes (M2 > M3, M4 > M1), CD87+ and overall survival [Table 3] and [Table 4]. We revealed from our multivariate analysis that CD87 was the most independent factor affecting the achievement of complete remission [Table 5].
Table 3: Relation between complete remission achievement and different parameters in acute myeloid leukemia group (n=47)

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Table 4: Relation between complete remission achievement and different parameters in acute myeloid leukemia group (N=47)

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Table 5: Multivariate analysis logistic regression for complete remission achievement

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  Discussion Top


AML is a heterogeneous disease due to associated molecular defect with variable clinical outcomes. Cytogenetic analysis indicates which patients may have favorable risk disease. However, the 5-year survival in this category is only ∼60% [7]. The uPAR (CD87) is found in many normal human body tissues and several different tumor cells. It affects malignancy development through adhesion, relocation, chemotaxis, and proteolysis and also invasion mechanisms [8]. The uPAR expression is strictly linked to cellular migration through its capacity to promote pericellular proteolysis in response to uPA binding [9] Many studies had elucidated the role of CD87 in solid cancers such as endometrial, ovarian, cervical, colon, non-small-cell lung cancer, breast, and stomach [2], but few of them had addressed its role in hematological malignancies [10]. We aimed in our work to shed light on the role and the prognostic value of flow cytometric expression of CD87 in newly discovered patients with AML.

The study included 52 patients with de-novo AML and 20 age-matched and sex-matched healthy control group. Diagnosis of AML was based on blasts immunophenotyping, FAB recommendation, and cytogenetic studies. The patients were followed up for 6 months after induction chemotherapy.

Our study revealed that CD87 expression was statistically higher in patients with AML than the control group (P < 0.001). Aref et al. [11] and Lanza et al. [12] stated that the CD87 is expressed on normal monocyte, neutrophils, and myeloid precursors but not on CD34+ cells. Furthermore, Kwak et al. [13] clarified that uPAR induces cellular migration and activation of intracellular signaling pathways through mechanisms that are independent of proteolysis.

Regarding demographic data, we demonstrated no significance between age, sex, and CD87, like other studies such as Atfy et al. [10], Lanza et al. [12], Rigolin et al. [14], and Sawai et al. [15].

Moreover, at the clinical level, we revealed a significant correlation between CD87+ and hepatomegaly and no correlation with splenomegaly, lymphadenopathy, or CNS infiltration, which agree with Nassar et al. [16] However, Atfy et al. [10] reported that high level of CD87+ in patients with AML was an indicator of poorly differentiated disease and associated with mucocutaneous infiltration, organomegaly (hepatosplenomegaly, lymph node enlargement), bleeding complication, CNS infiltration, and more aggressive course of the disease.

Regarding hematological parameters, we did not observe any statistical correlation between hemoglobin, PLT, total leukocytic count, blast cells, and CD87+ expression, and these were in accordance with Nassar et al. [16] However, Erkut et al. [2] observed a positive correlation between CD87 levels and the number of circulating white blood cells. Therefore, they suggested that the production of CD87 is related to blast cells in the peripheral circulation.

In the present study, the incidence of CD87+ cells in AML FAB subtypes was in the following descending order: M5more than M4 more than M1 more than M2 more than M3 more than M7. In addition, CD87+ was highly expressed in M4 (P = 0.002) and M5 (P = 0.018), whereas low expressed in M2 (P = 0.005). These findings were in accordance with Aref et al. [11], Nassar et al. [16], and Nadir et al. [17]. Additionally, Atfy et al. [10] suggested that CD87 expression was higher in M4 and M5 than the poorly differentiated M0 subtype of AML. Lanza et al. [12] added that various expression of CD87 depends on the cell lineage concerned (granulocytic or monocytic) and the level of maturation on AML blasts. Knapp et al. [18] showed that CD87 was highly expressed in monoblastic AML (FAB M4 and M5), and in most positive cases, it was co-expressed with CD14. Moreover, M4 and M5 are characterized by an elevated rate of extramedullary dissemination that is partially mediated by high expression of CD87, which bind uPA and start the conversion of plasminogen to plasmin which leads to proteolytic cleavage of endothelium-associated adhesion molecules and then extravasion of the cells through the endothelium [10].

Koch and his colleagues reported that immature hematopoietic cells do not express CD87 including CD34+ progenitor cells, and less differentiated BM cells. However, CD87 is found on the mature monocytes but not found on the red blood cells, erythroblasts, or PLTs [19]. In the present study, we correlated CD87+ with CD34+ and CD117+ in patients with AML which revealed no statistical relationship between them. Moreover, Atfy et al. [10] suggested that the co-expression of both CD34+ and CD87+ was related with a shorter overall survival rate (P = 0.05). In our study, favorable cytogenetics were found in 16/18 (88.9%) CD87+ cases and in 30/31 (96.8%) CD87 cases. On the contrary, unfavorable cytogenetics were found in 2/18 (11.1%) CD87+ cases and in 1/31 (3.2%) CD87 cases. However, no relationship between CD87 and cytogenetic information was found. These findings are in contrast to Atfy et al. [10] who depicted that 81.8% of patients with AML were associated with cytogenetic alterations when CD87+ was highly expressed. Besides Lanza et al. [12] reported that 93% of patients with AML with highly expressed CD87+ had cytogenetic abnormalities and added that high CD87+ expression and complex karyotype were often associated with M5 AML, and this helps to identify the abnormal variant of patients with AML with the poor prognosis.

Regarding disease status, we observed significantly shorter overall survival in CD87+ than CD87 patients. This agrees with Atfy et al. [10] who stated that high CD87 expression had shorter overall survival more than low CD87 expression (P = 0.0188).

In addition, our data revealed that complete remission achievement was inversely related to CD87+ (P = 0.001). Furthermore, from our multivariate analysis, we revealed that CD87 is considered the only independent prognostic factor predicting chemotherapy response in patients with AML. Similarly, Lanza et al. [12] and Graf et al. [20] observed that patients with AML with the marked increase of CD87+ receptors relapsed after using the induction consolidation chemotherapy. Additionally, Guo et al. [21] observed that CD87+ expression was related to pathogenesis and treatment response of AML; its high level was associated with decreased chemosensitivity, and its level on the multidrug resistant K562/ADM cell surface was increased compared with that on the sensitive K562 cell surface. Moreover, Gutova et al. [22] observed that patients with small cell lung cancer with CD87+ were more resistant to the chemotherapeutic drug than CD87 patients.

In conclusion, CD87 is an attractive goal for cancer treatment not only because of its several functional roles related to tumor progression but also the expression of CD87 is restricted quite tightly to the tumor tissue and is infrequently expressed in the neighboring normal quiet tissue.


  Conclusion Top


In conclusion, the high expression of CD87 displays a negative prognostic effect on patients with AML patients regarding outcome, survival rate and response to chemotherapy. However, more studies are required to elucidate its role as a targeted therapy for those patients as its expression is highly restricted to tumor cells.

Financial support and sponsorship

Nil.

Conflicts of interest

There are no conflicts of interest.



 
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