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BMC Research Notes

Open Access

Persistent polyclonal binucleated B-cell lymphocytosis and MECOM gene amplification

  • Edouard Cornet1, 2Email author,
  • Hossein Mossafa3,
  • Karine Courel3,
  • Jean-François Lesesve4 and
  • Xavier Troussard1, 2
BMC Research Notes20169:138

https://doi.org/10.1186/s13104-015-1742-3

Received: 2 February 2015

Accepted: 25 November 2015

Published: 2 March 2016

Abstract

Background

Persistent Polyclonal Binucleated B-cell Lymphocytosis (PPBL) is characterized by a chronic polyclonal B-cell lymphocytosis with binucleated lymphocytes and a polyclonal increase in serum immunoglobulin-M. Cytogenetic is characterized by the presence of a supernumerary isochromosome +i(3)(q10), premature chromosome condensation and chromosomal instability. Outcome of PPBL patients is mostly benign, but subsequent malignancies could occur. The aim of our study is to provide an update of clinical and cytogenetic characteristics of our large cohort of PPBL patients, to describe subsequent malignancies occurring during the follow-up, and to investigate the role of the long arm of chromosome 3 in PPBL.

Results

We analyzed clinical, biological and cytogenetic characteristics (conventional cytogenetic analysis and fluorescent in situ hybridization) of 150 patients diagnosed with PPBL. We performed high-resolution SNP arrays in 10 PPBL patients, comparing CD19+ versus CD19 lymphoid cells. We describe the cytogenetic characteristics in 150 PPBL patients consisting in the presence of supernumerary isochromosome +i(3)(q10) (59 %) and chromosomal instability (55 %). In CD19+ B-cells, we observed recurrent copy number aberrations of 143 genes with 129 gains (90 %) on 3q and a common minimal amplified genomic region in the MECOM gene. After a median follow-up of 60 months, we observed the occurrence of 12 subsequent malignancies (12 %), 6 solid tumors and 6 Non-Hodgkin’s Lymphomas, and 6 monoclonal gammopathies of undetermined significance (MGUS), requiring a long-term clinical follow-up.

Conclusions

Our clinical and cytogenetic observations lead us to hypothesize that isochromosome 3q, especially MECOM abnormality, could play a key role in PPBL.

Keywords

Persistent Polyclonal Binucleated B-cell LymphocytosisMECOMSNP array

Background

Persistent Polyclonal Binucleated B-cell Lymphocytosis (PPBL) is characterized by a chronic, stable and asymptomatic lymphocytosis with binucleated lymphocytes [1]. Binucleated lymphocytes are not specific for PPBL and can be observed in patients with multiple sclerosis treated by natalizumab [2] or after accidental exposure to ionizing radiation. In the peripheral blood, a polyclonal increase of memory B cells (CD19+, CD5, CD27+, IgM+, IgD+) is usually associated with a polyclonal increase in serum immunoglobulin-M (IgM) [36]. PPBL is characterized by a recurrent supernumerary isochromosome +i(3)(q10), a premature chromosome condensation (PCC) and a chromosomal instability [3, 4]. PPBL evolution is benign in most cases, but non-Hodgkin’s lymphomas and solid tumors (pulmonary blastoma) were previously and rarely described [7, 8]. In this study, we report the follow-up and the cytogenetic characteristics of a large cohort of 150 PPBL patients. We report the occurrence of subsequent malignancies in up to 12 % of patients contrasting with previous studies. Strong association between supernumerary isochromosome 3q, chromosomal instability and PPBL led us to study more extensively the role of the long arm of chromosome 3 using SNP arrays in 10 patients. We observed that the MECOM gene, located on 3q26, was recurrently amplified in B-cells of PPBL patients.

Patients and methods

Patients

PPBL was diagnosed from the persistence during three months of binucleated lymphocytes on a peripheral blood film. Patients were included after written informed consent, in accordance with the Declaration of Helsinki and with institutional guidelines and after approval of the French relevant competent authorities and ethics committees (Committee of Protection of Individuals (CPP), Advisory Committee on the Processing of Information for Medical Research (CCTIRS) and the French National Commission for Data Protection (CNIL)).

Using multiparameter flow cytometry (MFC), B-cells were polyclonal in all cases, based on the expression of CD19 and the absence of a restriction of expression of light chain of immunoglobulin. Blood smears were reviewed in the same laboratory.

Conventional cytogenetic analysis (CCA)

Blood samples were collected on heparin tubes at the time of diagnosis and during the follow-up. All samples were processed in the same laboratory. CCA was performed as previously described [3]. As previously described [9], chromosomal instability was defined as the gain and/or loss of whole chromosomes or chromosomal segments at a higher rate in tumor cell population compared to normal cells.

Fluorescent in situ hybridization (FISH)

FISH was performed in order to detect supernumerary isochromosome +i(3)(q10) in metaphase and interphase cells using alpha-satellite chromosome 3 specific probes and Bcl6 (3q27) specific probes (Vysis™, USA). One hundred metaphases and three hundred interphases cells were analyzed per patient.

SNP array

SNP arrays were performed using Affymetrix™ Cytogenetics Whole-Genome 2.7M Arrays® (Affymetrix™, USA). All samples were processed in the same laboratory. Patients were selected according to the availability of sufficient fresh cells (diagnosis) or frozen cells (follow-up). Immunomagnetic sorting was performed on whole blood samples or on thawed cells in order to purify CD19+ cells (Miltenyi™ AutoMACS Pro Separator®, Bergisch Gladbach, Germany). The two fractions (CD19+ positive and CD19 negative selection) were kept and the purity was checked to be >95 % by flow cytometry. The DNA was extracted from the two fractions using Gentra Puregene Blood Kit® (Qiagen™, Hilden, Germany). Hybridization of the DNA on chips was performed according the manufacturer’s instructions. Chips were analyzed using Affymetrix™ Chromosome Analysis Suite® (ChASver 1.0.1). Database of annotations was NetAffx Build 30. Quality controls of the chips were set up according Affymetrix™ recommendations (SNP-QC ≥ 1.1 and MAPD (CN-QC) ≤ 0.27). Copy Number Aberrations (CNA) were called according user-defined thresholds (Copy Number (CN) markers >50 and size >25 kb). The Database of Genomic Variants (DGV, http://projects.tcag.ca/variation/) was consulted to determine whether CNA corresponded to genomic variants. Number and size of Copy Number Aberrations (CNAs) were analyzed and compared between patients and between CD19+ and CD19 cells. CNA are called recurrent when at least two patients present the same CNA. Mosaicism phenomenon was detected in case of allele frequencies between disomic and trisomic states.

Results

PPBL was diagnosed in 150 untreated patients, whose main characteristics are described in Table 1. Sixty-nine percent of cases showed an absolute lymphocytosis >4 × 109/L, with a mean percentage of binucleated lymphocytes at 3.9 % (1–40). Median follow-up was 60 months (1–402) and median overall survival was not reached. Eighteen patients (12 %) developed subsequent malignancies, among which nine cases were previously described (non Hodgkin’s lymphomas (NHL) in three cases, solid tumors in two cases and monoclonal gammopathies of undetermined significance (MGUS) in 4 cases) [10]. Among the 18 patients, six patients developed solid tumors with a mean time of occurrence of 87 months (3–156) (4 pulmonary cancers, 1 breast cancer and 1 cervical carcinoma). Twelve patients (8 %) developed hematological malignancies. Six cases of MGUS (IgM) (4 %) and NHL (4 %) occurred with a mean time of 75 months (0–264) and 58 months (0–120), respectively. Four patients developed a diffuse large B-cell lymphoma and 2 patients a splenic marginal zone lymphoma (Table 2 for details). Among these 18 cases, 17 patients were chronic smokers. These data strongly lead us to consider PPBL as a premalignant state requiring a long-term follow-up.
Table 1

Characteristics and follow-up of 150 patients with PPBL

Age (years), Mean (min–max)

40 (18.9–66.2)

  

Sex (M/F)

26 (17 %)/124 (83 %)

  

Tobacco consumption

130/145 (90 %)

  

Clinical presentation

   

 Lymph node(s)

10/108 (9 %)

  

 Splenomegaly

19/106 (18 %)

  

 Hepatomegaly

2/108 (2 %)

  

Hemogram, Mean (min–max)

   

 White blood cells (109/L)

12.8 (7–44.8)

  

 Hemoglobin (g/dL)

13.8 (10.1–16.9)

  

 Platelets (109/L)

228 (83–380)

  

 Lymphocytosis (109/L)

6.5 (2.2–41)

  

 Binucleated Lymphocytes (% of lymphocytes)

3.9 (1–40)

  

IgM (g/L), Mean (min–max)

7.8 (2.17–20)

  

HLA DR7 positive

40/52 (77 %)

  

Multiparameter Flow Cytometry—Mean (min–max)

   

 CD19 (%)

50.4 (7–83)

  

Cytogenetics

 

Diagnosis

Follow-up

 +i(3)(q10) positive by karyotype

 

50/140 (36 %)

20/32 (63 %)

 +i(3)(q10) positive by FISH

 

80/128 (63 %)

24/26 (92 %)

 PCC positive

 

35/140 (25 %)

8/32 (25 %)

 Chromosomal instability

 

76/140 (54 %)

31/32 (97 %)

Subsequent Malignancies

18/150 (12 %)

  

 MGUS

6/150 (4 %)

  

 Non-Hodgkin’s Lymphomas

6/150 (4 %)

  

 Solid tumors

6/150 (4 %)

  

Clinical and biological data were collected from 27 centers. Median follow-up was 60 months (1–402) with unreached median overall survival

MGUS monoclonal gammopathy of undetermined significance

Table 2

Eighteen subsequent malignancies occurred in PPBL patients

Patients

Delay between PPBL and subsequent malignancy’s diagnosis

Type of malignancy

Follow-up

UPN36

38 months

DLBCL

56 months

UPN47

20 months

SMZL

+65 months

UPN57

92 months

DLBCL

99 months

UPN63

Diagnosis of PPBL and lymphoma was concomitant

DLBCL

+13 months

UPN71

77 months

SMZL

+86 months

UPN83

120 months

DLBCL

+131 months

UPN1

264 months

MGUS

+348 months

UPN10

144 months

MGUS

+148 months

UPN157

44 months

MGUS

+47 months

UPN118

Diagnosis of PPBL and MGUS was concomitant

MGUS

+36 months

UPN163

Diagnosis of PPBL and MGUS was concomitant

MGUS

+57 months

UPN105

Diagnosis of PPBL and MGUS was concomitant

MGUS

+42 months

UPN5

96 months

Mammary carcinoma

+272 months

UPN6

3 months

Pulmonary carcinoma

3 months

UPN70

22 months

Pulmonary carcinoma

+22 months

UPN86

132 months

Pulmonary carcinoma

+146 months

UPN160

114 months

Pulmonary carcinoma

112 months

UPN67

156 months

Cervical carcinoma

+181 months

Six patients developed solid tumors (4 pulmonary cancers, 1 breast cancer and 1 cervical carcinoma) and 6 patients hematological malignancies (diffuse large B-cell lymphoma (DLBCL) in 4 cases, splenic marginal zone lymphoma (SMZL) in 2 cases) and 6 patients monoclonal gammopathies of undetermined significance (MGUS) (IgM)

At diagnosis, CCA and FISH were performed in 140 and 128 patients, respectively. During the follow-up, CCA was performed in 32 patients (21 %). CCA and FISH detected no cytogenetic abnormality in 52/140 patients (37 %). Recurrent supernumerary isochromosome +i(3)(q10) was identified in 82/140 patients (59 %). PCC, arising from asynchronous mitotic activity in multinucleated cells, was observed concomitantly with +i(3)(q10) in 30/140 patients (21 %). By CCA, trisomy 8 and del(6q) were also detected either as recurrent abnormalities (2/140 and 5/140, respectively) or as non-recurrent abnormalities (9/140 and 4/140, respectively). Chromosomal instability was observed in 76/140 patients (54 %) and persisted in 31/32 patients (97 %) during follow-up.

To determine whether 3q could be implicated in PPBL pathogenesis, SNP arrays were performed in 10 patients (3 males, 7 females) with +i(3)(q10) in 9/10 patients (Table 3 for details). Written informed consents were obtained from the patients. The comparative analysis of sorted CD19+ and CD19 cells revealed that CNAs were observed predominantly in CD19+ B-cells on 3q (Table 4) with mosaicism phenomenon in 3 patients. Genetic instability was observed in all cases and predominantly in CD19+ B-cells. We observed 143 recurrent CNAs with 129 gains (90 %) on 3q of B-cells (Table 5). Interestingly, we identified with a high frequency (7/9 patients) partial or complete amplification of one particular genomic region located in 3q26.2. The size of this common minimal amplified region was 28 kilobases (85 copy number markers) located in coding region of MECOM gene (Fig. 1). This amplification was not detected in two patients (UPN147 and UPN136). In one of them (UPN147), no +i(3)(q10) was detected by CCA and/or FISH. Unfortunately, due to the mosaicism phenomenon, with less than 20 % of B-cells presenting +i(3)(q10), gain in MECOM gene has not been confirmed yet by other molecular studies, such as quantitative PCR.
Table 3

Characteristics of the 10 patients analyzed by SNP arrays (UPN: Unique Patient Number)

Patient

Karyotype

PCC (%)

FISH +i(3)(q10) (%)

UPN8b

46–47,XX, +i(3)(q10) [3] /46,XX,del(2)(q22), −17, +mar [1] /45, X, −X [1] /46,XX [40]

Absent

Present (6 %)

UPN57a

47,XY, +i(3)(q10) [5] /48,XY, +i(3)(q10), +12 [01]/46,XY,t(14;18)(q32;q22)[01]/47,XY,t(11;14)(q13;q32), +mar [01]/46,XY,add(3)(p26) [1] /47,XY, +22[01]/49,XY, +i(3)(q10), +8, +mar[01]/46,XY [39]

Absent

Present (7 %)

UPN136a

46,XX [48]/PCC [2]

Present (4 %)

Present (4 %)

UPN71c

47,XX, +X,del(6)(q15q26)[01]/46,XX,del(6)(q15q26),der(6)t(6;6)(q21;q23)[08]/46,XX,del(1)(q12),der(14)t(1;14)(p22;q32)[02]/46,XX[09]

Absent

Present (2 %)

UPN127b

47,XY, +i(3)(q10) [3] /46,X,der(Y)t(Y;?)(q12;?) [3] /46,XY [12]

Absent

Present (12 %)

UPN138a

46,XX,del(6)(q21q24) [6] /46,XX,der(8)t(3;8)(q11;q11),der(17)t(17;?)(p11;?) [2] /46,XX,del(17)(p11) [2] /46,XX,t(1;6)(q24;q21) [1] /46,XX,der(14)t(14;?)(p25;?) [1] /46,XX,dup(3)(p13p26) [1] /46,XX,der(4)t(4;?)(p16;?) [1] /46,XX [26]

Absent

Present (11 %)

UPN99a

47,XX, +18 [2] /47,XX, +3 [1] /46,XX [37]/PCC [1]

Present (2 %)

Present (4 %)

UPN147a

46,XX [50]

Absent

Absent

UPN73a

46,XX [50]

Absent

Present (1.4 %)

UPN105a

46,XY [46]/46,XY [cp 4]

Absent

Present (3 %)

Depending on the quality of extracted DNA, we performed DNA arrays on aboth CD19+ and CD19 cells in 7 patients, bCD19+ cells in 2 patients and cCD19 cells in 1 patient. CCA and/or FISH detected +i(3)(q10) in 9/10 patients

PCC premature chromosome condensation

Table 4

Repartition of CNAs observed in CD19 and CD19+ cells. CD19+ cells presented twice as many CNAs as CD19 [83 CNAs (12–218) versus 42 (3–184)]

CD19−

CNAs—Total (gains/losses)

Chromosome

UPN73

UPN71

UPN57

UPN136

UPN138

UPN99

UPN147

UPN105

Mean

1

2 (2/0)

0 (0/0)

4 (1/3)

1 (0/1)

3 (3/0)

1 (0/1)

0 (0/0)

17 (0/17)

3.5 (0.7/2.8)

2

3 (1/2)

1 (1/0)

0 (0/0)

2 (0/2)

2 (2/0)

0 (0/0)

0 (0/0)

24 (0/24)

4.0 (0.5/3.5)

3

2 (1/1)

0 (0/0)

3 (3/0)

0 (0/0)

0 (0/0)

0 (0/0)

0 (0/0)

8 (1/7)

1.6 (0.6/1.0)

3p

0 (0/0)

0 (0/0)

0 (0/0)

0 (0/0)

0 (0/0)

0 (0/0)

0 (0/0)

0 (0/0)

0.0 (0.0/0.0)

3q

2 (1/1)

0 (0/0)

3 (3/0)

0 (0/0)

0 (0/0)

0 (0/0)

0 (0/0)

8 (1/7)

1.6 (0.6/1.0)

4

0 (0/0)

0 (0/0)

0 (0/0)

2 (1/1)

0 (0/0)

0 (0/0)

1 (1/0)

30 (1/29)

4.1 (0.4/3.7)

5

0 (0/0)

1 (0/1)

1 (0/1)

0 (0/0)

1 (1/0)

0 (0/0)

0 (0/0)

15 (0/15)

2.3 (0.1/2.2)

6

0 (0/0)

0 (0/0)

0 (0/0)

0 (0/0)

1 (1/0)

0 (0/0)

0 (0/0)

11 (0/11)

1.5 (0.1/1.4)

7

0 (0/0)

0 (0/0)

0 (0/0)

0 (0/0)

1 (1/0)

0 (0/0)

1 (1/0)

12 (0/12)

1.8 (0.3/1.5)

8

1 (0/1)

0 (0/0)

0 (0/0)

1 (1/0)

3 (3/0)

0 (0/0)

0 (0/0)

6 (0/6)

1.4 (0.5/0.9)

9

0 (0/0)

4 (0/4)

3 (0/3)

0 (0/0)

4 (2/2)

1 (1/0)

0 (0/0)

7 (0/7)

2.4 (0.4/2.0)

10

1 (1/0)

1 (0/1)

1 (0/1)

0 (0/0)

1 (1/0)

0 (0/0)

1 (1/0)

7 (0/7)

1.5 (0.4/1.1)

11

0 (0/0)

2 (1/1)

0 (0/0)

0 (0/0)

2 (2/0)

0 (0/0)

1 (0/1)

10 (0/10)

1.9 (0.4/1.5)

12

2 (2/0)

0 (0/0)

1 (0/1)

0 (0/0)

1 (1/0)

0 (0/0)

0 (0/0)

8 (1/7)

1.5 (0.5/1.0)

13

0 (0/0)

0 (0/0)

2 (0/2)

0 (0/0)

1 (0/1)

0 (0/0)

0 (0/0)

8 (1/7)

1.4 (0.1/1.3)

14

0 (0/0)

0 (0/0)

0 (0/0)

0 (0/0)

0 (0/0)

0 (0/0)

0 (0/0)

6 (1/5)

0.8 (0.1/0.7)

15

2 (2/0)

1 (1/0)

1 (0/1)

2 (0/2)

2 (1/1)

0 (0/0)

1 (1/0)

4 (1/3)

1.6 (0.9/0.7)

16

1 (1/0)

0 (0/0)

0 (0/0)

1 (1/0)

3 (1/2)

0 (0/0)

0 (0/0)

1 (0/1)

0.8 (0.4/0.4)

17

0 (0/0)

0 (0/0)

1 (0/1)

0 (0/0)

1 (1/0)

0 (0/0)

0 (0/0)

4 (2/2)

0.8 (0.4/0.4)

18

0 (0/0)

0 (0/0)

0 (0/0)

0 (0/0)

1 (1/0)

0 (0/0)

0 (0/0)

1 (0/1)

0.2 (0.2/0.1)

19

0 (0/0)

1 (0/1)

0 (0/0)

0 (0/0)

0 (0/0)

1 (0/1)

1 (0/1)

1 (0/1)

0.5 (0.0/0.5)

20

1 (1/0)

0 (0/0)

0 (0/0)

0 (0/0)

1 (1/0)

0 (0/0)

0 (0/0)

0 (0/0)

0.3 (0.3/0.0)

21

1 (0/1)

1 (0/1)

1 (0/1)

0 (0/0)

0 (0/0)

0 (0/0)

0 (0/0)

1 (0/1)

0.5 (0.0/0.5)

22

0 (0/0)

0 (0/0)

0 (0/0)

0 (0/0)

0 (0/0)

0 (0/0)

1 (1/0)

1 (1/0)

0.3 (0.3/0.0)

X

1 (1/0)

7 (6/1)

13 (13/0)

1 (0/1)

25 (24/1)

0 (0/0)

2 (1/1)

1 (0/1)

6.2 (5.6/0.6)

Y

1 (1/0)

1 (1/0)

2 (0/2)

0 (0/0)

2 (2/0)

0 (0/0)

0 (0/0)

1 (0/1)

0.9 (0.5/0.4)

Total

18 (13/5)

20 (10/10)

33 (18/15)

10 (3/7)

55 (48/7)

3 (1/2)

9 (6/3)

184 (9/175)

41.5 (13.5/28)

CD19+

CNAs—total (gains/losses)

Chromosome

UPN73

UPN8

UPN57

UPN136

UPN127

UPN138

UPN99

UPN147

UPN105

Mean

1

20 (19/1)

0 (0/0)

0 (0/0)

4 (0/4)

0 (0/0)

0 (0/0)

5 (4/1)

8 (1/7)

0 (0/0)

5.0 (3.6/1.4)

2

22 (22/0)

2 (1/1)

0 (0/0)

2 (0/2)

0 (0/0)

1 (0/1)

3 (3/0)

12 (1/11)

0 (0/0)

5.7 (3.9/1.8)

3

20 (19/1)

4 (4/0)

26 (25/1)

0 (0/0)

46 (46/0)

1 (1/0)

2 (2/0)

5 (0/5)

113 (113/0)

25.0 (24.2/0.8)

3p

11 (11/0)

0 (0/0)

1 (0/1)

0 (0/0)

0 (0/0)

0 (0/0)

0 (0/0)

0 (0/0)

0 (0/0)

1.8 (1.7/0.1)

3q

9 (8/1)

4 (4/0)

25 (25/0)

0 (0/0)

46 (46/0)

1 (1/0)

2 (2/0)

5 (0/5)

113 (113/0)

23.2 (22.6/0.7)

4

9 (9/0)

0 (0/0)

2 (0/2)

5 (1/4)

0 (0/0)

1 (1/0)

0 (0/0)

10 (1/9)

0 (0/0)

4.0 (2.3/1.7)

5

14 (13/1)

0 (0/0)

0 (0/0)

2 (0/2)

0 (0/0)

0 (0/0)

0 (0/0)

5 (0/5)

1 (0/1)

3.4 (2.4/1.0)

6

10 (10/0)

1 (1/0)

0 (0/0)

1 (0/1)

0 (0/0)

1 (1/0)

1 (1/0)

5 (1/4)

0 (0/0)

2.9 (2.3/0.6)

7

12 (12/0)

1 (0/1)

0 (0/0)

1 (0/1)

1 (0/1)

0 (0/0)

1 (1/0)

6 (1/5)

0 (0/0)

2.4 (1.6/0.8)

8

13 (11/2)

0 (0/0)

0 (0/0)

0 (0/0)

0 (0/0)

1 (0/1)

1 (1/0)

2 (0/2)

0 (0/0)

2.3 (1.8/0.5)

9

14 (14/0)

0 (0/0)

0 (0/0)

4 (0/4)

0 (0/0)

5 (0/5)

1 (1/0)

4 (1/3)

0 (0/0)

3.8 (2.4/1.4)

10

8 (8/0)

0 (0/0)

0 (0/0)

3 (0/3)

0 (0/0)

0 (0/0)

0 (0/0)

0 (0/0)

0 (0/0)

1.6 (1.2/0.4)

11

9 (9/0)

0 (0/0)

0 (0/0)

2 (0/2)

0 (0/0)

0 (0/0)

1 (1/0)

7 (0/7)

0 (0/0)

2.8 (1.8/1.0)

12

5 (5/0)

1 (0/1)

2 (0/2)

0 (0/0)

1 (1/0)

0 (0/0)

1 (1/0)

8 (1/7)

0 (0/0)

2.7 (1.6/1.1)

13

4 (4/0)

0 (0/0)

1 (0/1)

2 (0/2)

0 (0/0)

1 (0/1)

0 (0/0)

2 (0/2)

1 (1/0)

1.3 (0.7/0.6)

14

3 (3/0)

0 (0/0)

1 (0/1)

0 (0/0)

0 (0/0)

0 (0/0)

2 (2/0)

0 (0/0)

0 (0/0)

1.1 (1.0/0.1)

15

5 (5/0)

0 (0/0)

1 (1/0)

1 (0/1)

2 (0/2)

1 (0/1)

0 (0/0)

2 (1/1)

1 (1/0)

1.8 (1.2/0.6)

16

2 (2/0)

0 (0/0)

0 (0/0)

2 (0/2)

1 (1/0)

2 (0/2)

0 (0/0)

1 (1/0)

1 (0/1)

1.6 (1.0/0.6)

17

4 (4/0)

0 (0/0)

0 (0/0)

1 (0/1)

1 (1/0)

0 (0/0)

0 (0/0)

1 (1/0)

0 (0/0)

0.9 (0.8/0.1)

18

5 (5/0)

1 (1/0)

0 (0/0)

0 (0/0)

0 (0/0)

3 (3/0)

1 (1/0)

1 (0/1)

0 (0/0)

1.9 (1.8/0.1)

19

1 (1/0)

0 (0/0)

0 (0/0)

0 (0/0)

1 (0/1)

0 (0/0)

0 (0/0)

1 (0/1)

0 (0/0)

0.4 (0.2/0.2)

20

5 (5/0)

0 (0/0)

0 (0/0)

0 (0/0)

0 (0/0)

0 (0/0)

2 (2/0)

1 (1/0)

0 (0/0)

1.2 (1.2/0.0)

21

1 (0/1)

0 (0/0)

0 (0/0)

1 (0/1)

0 (0/0)

1 (0/1)

0 (0/0)

2 (0/2)

0 (0/0)

0.6 (0.0/0.6)

22

0 (0/0)

0 (0/0)

0 (0/0)

0 (0/0)

0 (0/0)

0 (0/0)

0 (0/0)

1 (1/0)

0 (0/0)

0.3 (0.3/0.0)

X

28 (27/1)

1 (0/1)

1 (1/0)

7 (5/2)

2 (2/0)

5 (4/1)

1 (1/0)

5 (3/2)

0 (0/0)

7.0 (6.2/0.2)

Y

4 (4/0)

1 (1/0)

0 (0/0)

6 (6/0)

0 (0/0)

14 (14/0)

2 (2/0)

0 (0/0)

0 (0/0)

3.0 (3.0/0.0)

Total

218 (211/7)

12 (8/4)

34 (27/7)

44 (12/32)

55 (51/4)

37 (24/13)

24 (23/1)

89 (15/74)

117 (115/2)

82.7 (66.6/16.1)

28 % of CNAs (0–97 %) were located on 3q in CD19+ cells compared to 5 % (0–11 %) in CD19 cells (data not shown)

Table 5

Recurrent Copy Number Aberrations (CNA) in CD19+ B-cells. 143 CNA had been observed

Chr

Cytoregion

Recurrence

Recurrence including mosaicism

CN state

Gene

Minimal common size (kbp)

Genic region: total (T)

Exonic (E)

Intronic (I)

CNA reported in DGV

1

p33

2

2

Loss

FAF1

31

I

No

1

p32.2

2

2

Gain

C1orf168

50.2

E/I

No

2

p23.2

2

2

Gain

ALK

62

E/I

Yes

2

q21.2–q21.3

2

2

Gain

MGAT5

72

E/I

No

3

p24.2

2

2

Gain

THRB

55.7

I

No

3

q11.2

2

3

Gain

LOC255025

50

E/I

No

3

q12.2

2

3

Gain

ABI3BP

143

E/I

Yes

3

q13.13

2

3

Gain

DZIP3

10.6

I

No

3

q13.31

2

3

Gain

ZBTB20

39

I

No

3

q13.31

2

3

Gain

GAP43

399

T

No

3

q13.31

2

3

Gain

LSAMP

53

I

No

3

q13.33

2

3

Gain

TMEM39A

176

T

No

3

q13.33

2

3

Gain

KTELC1

176

T

No

3

q13.33

2

3

Gain

C3orf1

176

T

No

3

q13.33

2

3

Gain

CD80

176

T

No

3

q13.33

2

3

Gain

ADPRH

176

T

No

3

q21.1

2

3

Gain

HSPBAP1

101

E/I

No

3

q21.1

2

4

Gain

DIRC2

101

T

No

3

q21.1

2

4

Gain

LOC100129550

101

T

Yes

3

q21.1

2

4

Gain

SEC22A

114

T

No

3

q21.1

2

4

Gain

PTPLB

125

T

No

3

q21.1

3

4

Gain

MYLK

70

E/I

No

3

q21.1

2

4

Gain

CCDC14

121

E/I

Yes

3

q21.2

2

4

Gain

KALRN

169

T

No

3

q21.2

2

4

Gain

UMPS

169

T

No

3

q21.2

2

4

Gain

ZNF148

164

E/I

Yes

3

q21.2

2

4

Gain

ALDH1L1

171

E/I

No

3

q21.3

3

4

Gain

TXNRD3IT1

299

E/I

No

3

q21.3

3

4

Gain

CHCHD6

299

E/I

No

3

q21.3

2

4

Gain

KLHDC6

95

T

No

3

q21.3

2

4

Gain

RUVBL1

211

E/I

Yes

3

q21.3

2

4

Gain

EEFSEC

211

E/I

Yes

3

q21.3

2

4

Gain

GATA2

76

E/I

Yes

3

q21.3

3

4

Gain

LOC90246

76

T

Yes

3

q21.3

2

4

Gain

C3orf27

120.7

T

Yes

3

q21.3

2

4

Gain

TMCC1

268

T

No

3

q21.3

2

4

Gain

COL6A4P2

131

T

Yes

3

q22.1

2

4

Gain

MRPL3

62

E/I

Yes

3

q22.1

2

4

Gain

SNORA58

62

T

Yes

3

q22.1

3

5

Gain

CPNE4

46

I

Yes

3

q22.1

2

4

Gain

CPNE4

155

E/I

Yes

3

q22.1

2

4

Gain

TMEM108

120

I

Yes

3

q22.1

2

4

Gain

TOPBP1

69

E/I

No

3

q22.1

2

4

Gain

RYK

225

T

No

3

q22.1

2

4

Gain

ANAPC13

197

T

Yes

3

q22.1

2

4

Gain

CEP63

197

T

Yes

3

q22.2

2

4

Gain

EPHB1

144

E/I

No

3

q22.2

2

4

Gain

PPP2R3A

85

E/I

No

3

q22.3

2

4

Gain

SOX14

925

T

No

3

q22.3

3

4

Gain

CLDN18

121

T

Yes

3

q22.3

2

4

Gain

ARMC8

77

E/I

Yes

3

q22.3

2

4

Gain

TXNDC6

77

E/I

Yes

3

q22.3

2

4

Gain

ESYT3

202.7

E/I

No

3

q22.3

2

4

Gain

CEP70

202.7

T

No

3

q22.3

2

4

Gain

FAIM

202.7

T

No

3

q22.3

2

4

Gain

PIK3CB

202.7

E/I

No

3

q22.3

3

4

Gain

LOC729627

193

T

No

3

q22.3

3

4

Gain

LOC389151

193

T

No

3

q22.3

3

4

Gain

FLJ46210

193

T

No

3

q22.3

3

4

Gain

BPESC1

193

T

No

3

q22.3

2

4

Gain

PISRT1

319

T

No

3

q23

2

4

Gain

MRPS22

89

E/I

No

3

q23

2

4

Gain

COPB2

89

T

No

3

q23

3

4

Gain

NMNAT3

277.8

E/I

No

3

q23

4

5

Gain

CLSTN2

46

I

Yes

3

q23

2

4

Gain

TRIM42

443

T

Yes

3

q23

2

4

Gain

SLC25A36

443

T

Yes

3

q24

2

4

Gain

SLC9A9

138

E/I

Yes

3

q24

2

4

Gain

PLSCR4

47

E/I

No

3

q24

2

4

Gain

PLSCR5

69

T

No

3

q24

2

4

Gain

AGTR1

194

E/I

No

3

q25.1

2

4

Gain

P2RY13

74

E/I

No

3

q25.1

2

4

Gain

MED12L

74

E/I

No

3

q25.1

2

4

Gain

P2RY13

74

T

No

3

q25.2

2

4

Gain

SGEF

364

E/I

Yes

3

q25.2–q25.31

3

4

Gain

MME

87.4

E/I

No

3

q25.32

2

4

Gain

VEPH1

78

E/I

Yes

3

q25.32

2

4

Gain

C3orf55

78

E/I

No

3

q25.32

3

4

Gain

MLF1

50

E/I

No

3

q26.1

2

4

Gain

C3orf57

120.6

E/I

No

3

q26.1

2

4

Gain

OTOL1

120.6

T

No

3

q26.1

3

4

Gain

SI

747

T

No

3

q26.1

3

4

Gain

BCHE

329

E/I

No

3

q26.1

2

4

Gain

ZBBX

307

T

No

3

q26.2

6

7

Gain

MDS1

28

E/I

No

3

q26.2

2

4

Gain

TERC

59

T

Yes

3

q26.2

2

4

Gain

ARPM1

59

T

Yes

3

q26.2

2

4

Gain

MYNN

59

T

Yes

3

q26.2

2

4

Gain

LRRC34

59

E/I

Yes

3

q26.2

3

5

Gain

TNIK

31

E/I

No

3

q26.31

2

4

Gain

NLGN1

125

I

Yes

3

q26.31

2

4

Gain

NLGN1

64

E/I

No

3

q26.31

2

4

Gain

NAALADL2

113

E/I

Yes

3

q26.32

2

4

Gain

TBL1XR1

60

E/I

No

3

q26.32

2

4

Gain

KCNMB2

121

E/I

No

3

q26.33

2

4

Gain

USP13

59

E/I

No

3

q26.33

2

4

Gain

PEX5L

81

E/I

No

3

q26.33

2

4

Gain

CCDC39

118

E/I

Yes

3

q27.1

2

4

Gain

YEATS2

112

E/I

No

3

q27.1

2

4

Gain

MAP6D1

112

T

No

3

q27.1

2

4

Gain

PARL

112

E/I

No

3

q27.2

2

4

Gain

VPS8

218

E/I

No

3

q27.2

2

4

Gain

ETV5

157

T

No

3

q27.2

2

4

Gain

DGKG

157

E/I

No

3

q27.3

2

4

Gain

CRYGS

110

E/I

No

3

q27.3

2

4

Gain

TBCCD1

110

T

No

3

q27.3

2

4

Gain

DNAJB11

110

T

No

3

q27.3

2

4

Gain

AHSG

110

T

Yes

3

q27.3

2

4

Gain

FETUB

110

E/I

Yes

3

q27.3

2

4

Gain

ST6GAL1

46

E/I

Yes

3

q27.3

2

4

Gain

MASP1

428

E/I

No

3

q27.3

3

4

Gain

RTP4

148

T

No

3

q27.3

2

4

Gain

SST

428

T

No

3

q27.3

2

4

Gain

FLJ42393

191

T

Yes

3

q28

3

4

Gain

LPP

191

E/I

Yes

3

q28

2

4

Gain

TP63

142

E/I

No

3

q28

2

4

Gain

CLDN1

203

T

No

3

q28

2

4

Gain

CLDN16

203

T

No

3

q28

2

4

Gain

TMEM207

203

T

No

3

q29

2

4

Gain

C3orf59

396

E/I

No

3

q29

2

4

Gain

MGC2889

396

T

Yes

3

q29

2

4

Gain

HRASLS

396

T

Yes

3

q29

2

4

Gain

ATP13A5

396

E/I

No

3

q29

2

4

Gain

ATP13A4

158

E/I

Yes

3

q29

2

4

Gain

OPA1

158

T

Yes

3

q29

2

4

Gain

GP5

97

E/I

No

3

q29

2

4

Gain

ATP13A3

97

T

Yes

3

q29

2

4

Gain

TM4SF19

87

E/I

Yes

3

q29

2

4

Gain

UBXN7

87

E/I

Yes

3

q29

2

4

Gain

DLG1

240

E/I

Yes

3

q29

2

4

Gain

FYTTD1

50

T

Yes

3

q29

2

4

Gain

LRCH3

50

E/I

Yes

3

q29

2

4

Gain

RPL35A

91

E/I

Yes

3

q29

2

4

Gain

IQCG

91

E/I

Yes

3

q29

2

4

Gain

LMLN

91

T

Yes

4

q13.3

2

2

Gain

SLC4A4

46

E/I

No

11

p15.1

2

2

Gain

NELL1

43

I

No

14

q13.1

2

2

Gain

NPAS3

43

I

Yes

16

p11.1

2

2

Gain

LOC283914

277

T

Yes

21

p11.2–p11.1

2

3

Loss

TPTE

107

T

Yes

X

p22.33

3

3

Gain

DHRSX

31

E/I

Yes

X

q12

2

2

Gain

EDA2R

91

E/I

Yes

Y

q11.21

2

2

Gain

USP9Y

60

E/I

No

129 gains concerned the long arm of chromosome 3 (3q). 123 gains concerned gene coding regions. 75 CNA did not include previously reported polymorphism (Database of Genomic Variants, DGV). Gain of one exon of MDS1 (part of MECOM gene) was recurrently observed in 7 patients (including mosaicism phenomenon)

Chr chromosome, Recurrence number of patients with the same CNA, CN state Copy Number state, gain or loss

Fig. 1

Schematic representation of region 3q26.2 corresponding to MECOM gene in CD19+ B-cells of 9 patients. We observed a common minimal amplified region of 28 kilobases (85 copy number markers) in 7 patients. This amplification is observed in all the CD19+ B-cells in 6 patients (copy number at 3, CN 3) and in a part of CD19+ B-cells in 1 patient (copy number between 2 and 3 revealing a mosaicism phenomenon)

Discussion

Similar to the described link between aneuploidy, genetic instability and the development of human cancers [11, 12], supernumerary isochromosome 3q could be the cause of chromosomal instability observed in PPBL. Transfer of isochromosome 3q into myoblast cell line caused abnormal cytokinesis, centrosome amplification, aneuploidy and abolished G1 arrest following DNA damage. These observations might be related to an increasing expression of ATR gene located on 3q [13]. Moreover, isochromosome 3q has been implicated in the progression of cervical carcinomas, where cells exhibiting either tetrasomy or aneusomy for chromosomes 3 and 17 increased significantly with disease progression [1316]. Supernumerary isochromosome 3q could explain binucleated lymphocytes and chromosomal instability observed in PPBL. MECOM abnormalities, particularly the overexpression of EVI1, have been described in the pathogenesis of myeloid neoplasm such as acute myeloid leukemia and myelodysplastic syndrome, especially concerning cell-cycle disorders [1721]. Furthermore, as observed by Stein et al., EVI1 activation could lead to genetic instability [22]. Even if it has never been observed in lymphoid neoplasm, the potential implication of MECOM in PPBL has to be elucidated.

The link between PPBL and subsequent malignancies remains unclear and the role of tobacco is probably dominant. Majority of our patients (17/18) with subsequent malignancies were chronic smokers. We reported recently a detailed description of 2 heavy smokers patients with subsequent malignancies, UPN57 and UPN71 [23]. Tobacco use is a recognized risk factor in the development of solid tumor, such as pulmonary cancer, and also lymphoma [24]. Therefore, in PPBL, where tobacco consumption is frequent (90 % of our cohort of 150 patients), smoking could represent a confounding factor in interpreting the link between PPBL and subsequent malignancies.

Isochromosome 3q has been described in cell-cycle deregulation, chromosomal instability and progression of cervical cancers. Our cytogenetic and clinical observations lead us to hypothesize that isochromosome 3q in B-cells plays a key role in the physiopathology and evolution of PPBL. Although isochromosome 3q has not been yet identified in tumor cells of subsequent malignancies [23], it could be implicated in chromosomal and genomic instability. This genomic instability could be part of a multi-step process leading to the emergence of a malignant B lymphoproliferation. MECOM gene could be a good candidate to explain these observations and remains to be explored.

Availability of data and materials

All raw data are available from the authors upon request.

Declarations

Authors’ contributions

HM and XT designed the study. EC and XT wrote the manuscript. EC and HM analyzed SNP arrays data. KC performed SNP arrays. HM analyzed conventional cytogenetic and FISH. EC, JFL and XT examined the patients and collected clinical/biological data. All authors read and approved the final manuscript.

Acknowledgements

We acknowledge all the members of the Groupe Francophone d’Hématologie Cellulaire (GFHC) who participated in this study and followed up PPBL patients.

Competing interests

The authors have nothing to disclose.

Open AccessThis article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.

Authors’ Affiliations

(1)
Laboratory of Hematology, Caen University Hospital
(2)
University of Caen, Medical School
(3)
Laboratoire Cerba, Department of Genetic
(4)
Laboratory of Hematology, Nancy University Hospital

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