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Characteristics of CD1 l c + C D S + Chronic B-Cell Leukemias and the Identification
of Novel Peripheral Blood B-Cell Subsets With Chronic Lymphoid Leukemia
Immunophenotypes
By Susan B. Wormsley, Stephen M. Baird, Nancy Gadol, Kanti R. Rai, and Robert E. Sobol
Previous studies have indicated that chronic lymphocytic
leukemias (CLL) are characterized by the coexpression of
CD5 and B-cell antigens, while hairy cell leukemias (HCL)
typically express C D l l c'CD5- B-cell immunophenotypes.
In this report we describe the features of B-cell leukemias
with C D l l c+CD5+ immunophenotypes and the identification of novel circulating B-cell subsets defined by the
expression of CD20, CD5, and C D l l c antigens. Morphologic evaluation of 14 CD1lc+CD5+B-cell leukemias showed
that they generally had larger cellular diameters (14 t o 21
pm) and lower nuc1ear:cytoplasm ratios than typical small
lymphocyte CLL. These cases did not exhibit the welldefined nucleoli characteristic of prolymphocytic leukemia
(PLL). The presenting clinical features of C D l l c+CD5+
B-cell leukemias were most consistent with CLL or PLL,
and none of the evaluated cases had pancytopenia, splenomegaly, and cytoplasmic villi characteristic of HCL. Examination of normal peripheral blood (n = 6 ) by three-color
flow cytometry identified four novel B-cell subsets with the
following immunophenotypes (mean percent of total CD20'
B cells ? SEI: CD20+CD5+CDllc+ (8.0 f 1.6); CD20+CD5CD1 IC+
(12.0 f 2.0); CD20fCD5+CD1 IC(35.0 ? 4.9); and
CD2O+CD5-CDll c- (44.0 f 5.0). Our findings suggest that
CDI 1c+CD5+ B-cell leukemias with atypical morphologic
features represent forms of CLL or PLL rather than HCL. In
addition, w e have identified novel subsets of circulating B
cells defined by patterns of CD20, CD5, and C D l l c expression that correspond t o the immunophenotypes of chronic
B-cell leukemias.
0 1990 by The American Society of Hematology.
C
CD 1 I C expression corresponding to the immunophenotypes
of CLL, PLL, and HCL. Novel peripheral blood B-cell
subsets with CD20+CDS+CDllc+,CD2O+CDS+CDlIC-,
CD20+CDS-CDllc+, and CD20+CDYCDl IC- immunophenotypes were identified.
HRONIC LYMPHOCYTIC leukemia (CLL), prolymphocytic leukemia (PLL), and hairy cell leukemia
(HCL) are distinct clinical entities that cah be distinguished
by morphologic, cytochemical, and immunophenotypic
criteria.'-6CLL is generally characterized by small uniform
leukemic cells with weak surface immunoglobulin (sIg)
expression and a CD5' B-cell immunophenotype.' The cells
in PLL are typically larger, with well-defined nucleoli and
bright sIg ~ t a i n i n g . ~PLL
' ~ patients usually have splenomegaly and an aggressive c o ~ r s e .HCL
~ ' ~ is diagnosed by the
triad of pancytopenia, splenomegaly, and the presence of
characteristic leukemic cells with villous cytoplasmic
projection^.^ Most HCL express tartrate-resistant acid phosphatase (TRAP) activity, sIg, and CD1 1c.4-6CDl l c (p150/
95) is a member of a subfamily of heterodimeric integrin
receptors that include the C3b receptor and lymphocyte
function-associated antigen-1 (LFA-l).' CDl I C is characteristically expressed by normal cells of myeloid/monocytoid lineage' and by HCL that typically express
CD20+CDS-CDl IC' immunophenotypes.6s8
As CLL, PLL, and HCL are treated
evaluation of these morphologic, cytochemical, and immunophenotypic characteristics are important in the differential
diagnosis and management of chronic B-cell leukemias.'"
However, it is well-recognized that definitive classification of
a proportion of chronic B-cell leukemias will be problematic
owing to considerable heterogeneity and overlap of the
morphologic and immunophenotypic characteristics of these
disorder^.'.^.^ In this regard, the present study was undertaken to examine the features of chronic B-cell leukemias
with CDl Ic+CDS+B-cell immunophenotypes. Our findings
suggest that chronic B-cell leukemias with atypical morphologic features and CD1 lcfCD5+ immunophenotypes represent forms of CLL or PLL rather than HCL.
We also examined normal peripheral blood B cells for the
expression of CD20, CD5, and C D l l c antigens by threecolor flow cytometry. Consistent with the view that the
immunophenotypes of B-cell leukemias reflect normal stages
of B-cell differentiation or activation, our analyses identified
normal circulating B cells with patterns of CD20, CD5, and
Blood, VOI 76, NO 1 (July 1). 1990: pp 123-130
MATERIALS AND METHODS
Patients. In a consecutive series of 119 CD5+ B-cell leukemias
referred for immunophenotype determinatons, 26 (22%) were found
to express greater than 20% CDl IC+leukemic cells. Wright-Giemsa
stained peripheral blood and/or bone marrow films were reviewed by
the authors in 14 CDllc'CD5+ cases. The percentages of small
lymphocytes, large lymphocytes, and prolymphocytes were scored
for each of these patients using criteria adapted from Melo et al.'
Briefly, small lymphocytes were defined by cellular diameters less
than 14 pm, very condensed nuclear chromatin, the absence of a
nucleolus, and a high nuc1ear:cytoplasm ratio. Large lymphocytes
were identified by cellular diameters greater than 14 pm, very to
moderately condensed chromatin, absent or poorly defined nucleolus, and an intermediate to low nuc1ear:cytoplasmratio. Prolymphocytes had cellular diameters usually greater than 14 pm, very to
moderately condensed chromatin, the presence of a large prominent
From the Cytometrics. Inc, Division of Specialty Laboratories,
Inc; Departments of Pathology and the Cancer Center, University of
California; Mercy Hospital Medical Center; Veterans Administration Medical Center, San Diego; Becton Dickinson, Mountain View,
CA; and Long Island Jewish Medical Center and Albert Einstein
College of Medicine. New Hyde Park, NY.
Submitted November I O , 1989; accepted March 6, 1990.
Supported in part by Specialty Laboratories. Inc, the University
of California, San Diego Cancer Center, the Veterans Administration, the Aaron Diamond Foundation, the Helena Rubinstein
Foundation. and Mercy Hospital.
Address reprint requests to Susan B. Wormsley, Cytometrics.
Inc. 11575 Sorrento Valley Rd, Suite 202. San Diego, CA 92121.
The publication costs of this article were defrayed in part by page
charge payment. This article must therefore be hereby marked
"advertisement" in accordance with 18 U.S.C.section I734 solely to
indicate this fact.
0 1990 by The American Society of Hematology.
0006-4971/90/7601-001I %3.00/0
123
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124
WORMSLEY ET AL
nucleolus, and intermediate to low nuc1ear:cytoplasm ratios. In
selected cases, evaluation of TRAP activity was performed as
described previously." Presenting clinical features were obtained by
retrospective review of patient medical records.
Immunophenotype Determinations
Leukemic cells. Immunophenotype determinations were performed with leukemic cells isolated from heparinized peripheral
blood and/or bone marrow by standard Ficoll-Hypaque density
gradient centrifugation.' I Previously described indirect immunofluorescence and flow cytometric methods" were used with the panel of
monoclonal antibodies (MoAbs) listed in Table 1. Monocytes were
depleted from study specimens by adherence to plastic after culture,
and their exclusion was confirmed by light scatter analyses and by
the absence of staining with a pan-monocyte reactive MoAb,
LeuM3, as described.".25Cell sIg light and heavy chain phenotyping
was performed by indirect immunofluorescence methods".25 using
affinity purified goat anti-human Ig heavy chain (IgM, IgD, IgG)
and light chain ( K or A) antisera and fluorescein-conjugated swine
anti-goat Ig (Tago, Inc, Burlingame, CA). Flow cytometric analyses
were performed as previously described"," with either a Cytofluorograf 50H and 2150 computer system (Ortho Diagnostic Systems,
Boston, MA) or an Epics Profile (Coulter Electronics, Inc, Hialeah,
FL). Dead cells were eliminated from analyses based on their uptake
of propidium iodide.26 The criterion for marker positivity was
expression by at least 20% of the leukemic cells. sIg expression of the
leukemic cells was interpreted as "weak" if the mean intensity
fluorescence (MIF) of the positive cells was 5 5 0 and "bright" if >50
(linear acquisition, 0 to 200 channel range).
In selected cases, two-color immunofluorescence staining was
performed to determine whether the C D l IC antigen was coexpressed
by sIg leukemic B cells. The cells were incubated with C D l I C and
the appropriate phycoerythrin (PE)-conjugated goat anti-human
light chain antisera, washed, and then incubated with fluorescein
isothiocyanate (F1TC)-conjugated goat anti-mouse Ig antisera.
Isotype-matched myeloma protein and the opposite phycoerythrinconjugated goat anti-human Ig light chain antisera, respectively,
were used as substitution controls. Two-color flow cytometric analysis to detect double-staining cells was performed as previously
described.' '
The relationship between antigen expression and cellular light
scatter characteristics of the leukemic cells was studied by arbitrarily dividing the leukemia cells into two populations of lower and
higher forward and right angle light scatter as described.,' For the
+
eight cases that were evaluated, the mean percentages % S E of the
leukemia cells in the low and high light scatter populations were
67 t 7.7 and 33 i 7.7, respectively. The two light scatter gated
populations were then analyzed separately for antigen expression.
Differences in the mean percentages of antigen positive cells between
the leukemic cell populations with lower and higher forward and
right angle light scatter characteristics were evaluated for statistical
significance by the paired Student's t-test.28
Normal peripheral blood lymphocytes. To determine whether
normal B cells expressed patterns of CD20, CD5, and C D l l c
antigens corresponding to the immunophenotypes of chronic B-cell
leukemias, three-color flow cytometric analyses were performed as
described29with enriched B lymphocytes obtained from the peripheral blood of six normal adults. Briefly, mononuclear cells were
separated by Ficoll-Hypaque gradient centrifugation and the monocytes were depleted with iron filings (lymphocyte separator reagent;
Technicon, Tarrytown, NY). To remove T lymphocytes, the mononuclear cells were rosetted with sheep erythrocytes treated with a 1%
solution of 2-aminoethyleso-thiouronium (AET) in 50% fetal bovine
serum. The rosetted cells were centrifuged over cold Ficoll-Hypaque
and the interface B cells were collected as de~cribed.~'
The enriched
B cells were then stained with FITC-conjugated CD20, PEconjugated C D l I C and biotin-conjugated CD5 followed by streptavidin-conjugated allophycocyanin (APC). Substitution controls incorporated FITC-IgG,, PE-IgG,,, and biotin-IgG,, murine myeloma
proteins with single and combined test MoAbs as described.29
Analysis of three-color fluorescence staining was performed using a
FACS 440 (Becton Dickinson, Mountain View, CA) equipped with
an argon-ion laser at 488 nm and a helium-neon laser at 633 nm.29
RESULTS
CDl lc+CDS+B-Cell Leukemia
Immunophenotype studies. The results of immunophenotyping studies in patients with C D l lc+CD5+ B-cell leukemia are summarized in Table 2. All cases were positive for
CD20 (Leul6), CD19 (Leul2), and CD24(BA-1) antigens.
The interleukin-2 (IL-2) receptor (CD25) was expressed in
over half of the cases tested (12 of 19), and most cases
evaluated (1 1 of 13) were positive for the B-cell activation
antigen CD23. With respect to other monocyte/myeloid
associated antigens, 44% (1 1 of 25) were My4 and none
expressed LeuM3 positivity. None of the cases expressed the
Table 1. MoAbs Used for lmmunophenotype Determinations
Antibody
Cluster
Designation
LeuM5
CDllc
Leu 1
Leu16
Leu12
BA- 1
IL-2R
6532
My4
LeuM3
CALLA
FMCJ
Leu5
Leu3a
Leu2a
T10
CD5
CD20
CD19
C024
CD25
CD23
CD14
CDlO
CD2
CD4
CD8
CD38
Molecular Weight
Predominant Reactivity
of Antigen (daltons)
Reference
Macrophage, myeloid monocyte cells
Pan T cell
Mature B cells
Pan B cell
B cells. mature granulocytes
IL-2 receptor
B-cell activation (germinal centers)
Monocytes, B cells
Monocytes
Common acute lymphoblastic leukemia antigen (CALLA)
B-cell subset
E-rosette receptor
Helper/inducer T cells
Suppressor/cytotoxic T cells
Thymocytes, myeloid progenitor, 8 cells (terminally differentiated), plasma cells
150.000; 95,000
65,000-6 7,000
35,000
45,000
45,000; 55,000
55,000
45,000
55,000
6
12
13
14
15
16
17
18
19
100,000
20
50,000
65,000
32,000-43,000
45,000
21
22
23
24
23
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125
CD1 1ctCD5+ CHRONIC B-CELL LEUKEMIAS
Table 2. Antigen Expression in CD11ctCD5+ 6-Cell Leukemia
NO.
No.
Cases
Antigen
Tested
Cases
Positive (%)
CDllc
CD5
CD20
CD19
CD24
slst
CD23
CD25
My4
CD38
LeuM3
CDlO
FMC7
CD2
CD4
CD8
26
26
26
26
25
26
13
19
25
25
25
25
11
26
26
26
26 ( 100)
26 ( 100)
26 ( 100)
26 ( 100)
25 (100)
26 ( 100)
11 (85)
12 (63)
11 (44)
6 (24)
0 (0)
0 (0)
0 (0)
0 (0)
0 (0)
0 (0)
% Antigen Positive Cells.
Median
41
94
89
89
88
56
39
47
73
37
+ SE
+3
+3
f3
k 3
k 4
f5
f6
& 7
t 8
t 6
Range
26-89
30-99
42-96
47-98
26-98
20-9 1
23-70
21-87
20-94
30-65
*For antigen positive cases.
tBased on monotypic Ig light chain expression.
common acute lymphoblastic leukemia antigen (CD10). Of
11 cases tested, none expressed the B-cell differentiation
antigen defined by MoAb FMC7. None of the cases were
judged to express the T-cell associated antigens CD2 (LeuS),
CD4 (Leu3a), or CD8 (Leu2a).
Two groups of C D l l c + C D 5 + B-cell leukemia could be
distinguished based on the intensity of sIg staining. Fifteen
patients had weak ( M I F less than 50) sIg expression (mean
M I F SE = 30 * 3) and 11 patients had bright sIg staining (mean M I F * SE = 111 * 13). Expression of CD38
(T10) was restricted to the bright sIg+ group. Analysis of Ig
heavy chain expression was performed in 25 patients. The
most frequent heavy chain phenotype was IgM+IgD+(n = 8,
32%). An IgM+IgD+IgG+phenotype was observed in four
patients (16%). Some cases expressed only one type of heavy
chain: IgM+ (n = 1, 4%); IgD+ (n = 6, 24%); or IgG+
(n = 1, 4%). Five cases (20%) lacked detectable surface
IgM, IgD, or IgG heavy chains. All cases expressed monotypic light chains. Seventy-three percent were K positive and
27% were X positive.
Double-label immunofluorescence staining of leukemic
cells demonstrated coexpression of C D l I C and monoclonal
light chain sIg in the two cases that were evaluated. Representative histograms depicting C D l I C expression in CD5+
B-cell leukemia and cytograms reflecting coexpression of sIg
and CD1 I C are shown in Figs 1 and 2, respectively.
Analysis of leukemic cell light scatter characteristics and
antigen expression are summarized in Table 3. Leukemic cell
populations with higher forward and right angle light scatter
had a significantly greater mean percentage of C D l l c + cells
than those with lower light scatter properties (79% v 39%,
P < .001). With respect to the expression of other markers,
there were no significant differences in the mean percentages
of cells expressing My4, LeuM3, CD20, or CD5 antigens
A
B
6REEN FL
CDllc
682 POSITIVE
Fig 1. Histograms A, 6, and C depict representative CD5. CD20, and C D l l c expression in
C D l l c'CD5' chronic 6-cell leukemia. Dotted line
histograms represent background staining with
isotype-matched negative control myeloma proteins. Solid lines represent cells stained with CD5
(A). CD20 (61. and C D l l c (C) specific antibodies.
FL, fluorescence.
6REEN FL
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126
WORMSLEY ET AL
B
C IGGzn
IGGzB CONTROL
Fig 2. (A) Two-color immunofluorescencecontour plots of leukemic cells demonstrating coexpression of K light chain slg and C D l l c. No doublestaining cells were observed with antibody
substitution controls: 16) anti-A and isotypematched negative control myeloma protein: IC)
anti-r and isotype-matched negative control myeloma protein: ID) anti-A and anti-CD1 IC
(LeuM5).
FL, fluoroscence.
CONTROL
GREEN FL
between leukemic cell populations with higher and lower
light scatter characteristics.
Cytology and cytochemistry. Peripheral blood and/or
bone marrow smears from 14 patients were available for
review. The morphologic characteristics of these cases are
summarized in Table 4. In 11 cases, the majority of leukemic
cells had diameters greater than 14 pm. Three patients (CB,
BW, AS) had lower percentages of large lymphocyte type
leukemic cells, and these cases tended to have lower percentages of C D l IC+ cells. The nucleus was round and centrally
located in the cells from 12 patients, while two cases (NH,
FL) had eccentrically placed nuclei. The majority of leukemic cells had clumped to moderately dispersed nuclear
chromatin patterns. In most cases ( 1 1 of 14), nucleoli were
observed in less than 5% of the leukemic cell population. In
Table 4. Morphologic Characteristics of CDl IC+
CD5' B-Cell
Leukemias
% of Leukemic Cells
Small
Large
Cytoplasmic
Patient Lymphocyte Lymphocyte Prolymphocyte
Villi
C D l IC'
Table 3. Antigen Expression in Leukemia Cell Populations With
Low and High Light Scatter Characteristics
% Antigen Positive Cells
(Mean SE,n = 8 )
*
Antigen
Low Light
Scatter
Population
High Light
Scatter
Pooulation
P Value.
~
CD1 IC
My4
LeuM3
CD20
CD5
39
40
+ 3.5
* 8.7
1 k 1.8
80 t 7.8
81
?
4.0
79
42
2
82
83
t 5.5
t
7.8
k 3.0
k 6.1
k 4.0
<.001
.12
.27
.35
.33
Combined forward and right angle light scatter.
'Comparison of mean percentages of antigen positive cells between
low and high light scatter leukemia populations by paired t-test.
ET
RR
NH
BD
CB
BW
AS
IS
SP
MS
FL
RG
MM
26
9
20
20
ao
90
75
20
49
29
4
30
35
73
91
80
70
10
10
20
70
40
68
91
67
60
1
10
0
0
0
0
0
10
0
0
0
10
11
3
5
3
5
0
0
0
0
0
0
5
0
0
64
46
36
89
31
20
37
36
68
42
40
36
40
Criteria used for morphologic evaluations were adapted from Melo et
a1.3
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CD1 lc'CD5'
127
CHRONIC 8-CELL LEUKEMIAS
three patients (BD, IS, SP), the percentages of leukemic cells
with well-defined nucleoli were slightly higher (10% to 11%).
Three patients (CB, AS, MM) had small subsets of leukemic
cells (range 5% to 10%) with cytoplasmic villi. Cytochemical
stains for TRAP activity were negative in 4 of 4 cases tested.
Clinical features. The presenting clinical characteristics
of 14 cases of CDllc+CD5+ B-cell leukemia are summarized in Table 5. The median age at presentation was 63
years and the majority of patients were male (10 of 14). The
median white blood cell count was 20.4 x 103/mm3(range
6.1 to 230.0). None of the three patients with a subset of
leukemic cells with villous cytoplasmic projections (CB, AS,
MM) had splenomegaly and pancytopenia characteristic of
HCL. Two patients (MS, TW) with bright sIg+ phenotypes
had high leukocyte counts, splenomegaly, and no lymphadenopathy characteristic of PLL. Overall, the majority of
patients had lymphadenopathy (64%) at presentation, while
splenomegaly (36%), anemia (43%), neutropenia (14%), and
thrombocytopenia (21%) were less frequently observed.
Normal Peripheral Blood Lymphocytes
To determine whether normal circulating B cells have
patterns of CD20, CD5, and CDl I C expression corresponding to the immunophenotypes of chronic B-cell leukemias,
enriched peripheral blood B-cell specimens from six healthy
donors were evaluated by three-color flow cytometry. The
following subsets of normal circulating B cells were identified:
C D 2 0 + C D S + C D l l c + , CD20CCD5-CD1I C + , CD20+
CDS+CDl IC-, and CD20+CDS-CDllc-. Table 6 lists the
mean percentages of total CD20' B cells represented by each
of these subsets. Figure 3 depicts representative three-color
immunofluorescence cytograms of these normal B-cell subpopulations.
DISCUSSION
We have examined the features of chronic B-cell leukemias with an unusual immunophenotype characterized by
the expression of C D l l c and CD5 antigens. In our study,
Table 6. Normal Peripheral Blood E-Cell Subsets Identified by
Three-Color Immunofluorescence Analysis of CD20. CD5. and
CD1IC
Expression
E-Cell Subset
Mean Percentage (+SEI of
Total CD20' B Cells (n = 6)
CD20+CD5+CDllc'
CD20+CD5-CD 1 1c+
CD20+CD5+CDlICCD20'CD5-D 1 1 c-
8.0 1.6
12.0 2 . 0
35.0 t 4.9
44.0 5 . 0
*
*
*
CD1 lc+CD5+B-cell leukemias were generally found to have
larger cellular diameters, more abundant cytoplasm, and
lower nuc1ear:cytoplasm ratios than typical CLL. Consistent
with these observations, CD1 I C expression was associated
with leukemic cells having higher forward and right angle
light scatter properties. In contrast to leukemic cells characteristic of PLL, CD1 lc+CD5+B-cell leukemias did not have
prominent nucleoli. None of the cases examined in our series
had TRAP activity or a preponderance of cells with cytoplasmic villi. Clinically, most patients had features of CLL at
presentation, and none of the evaluated cases had splenomegaly, pancytopenia, and cytoplasmic villi characteristic of
HCL. Two patients with bright sIg+ phenotypes presented
with high leukocyte counts, splenomegaly, and minimal
lymphadenopathy characteristic of PLL. Overall, our observations suggest that CD1 lc+CD5+ B-cell leukemias with
atypical morphologic features represent forms of CLL or
PLL rather than HCL. Prospective studies of CDl lc+CD5+
B-cell leukemias, examining their clinical course and response to therapy, will be required to extend our evaluation of
morphologic, immunophenotypic, and presenting clinic features. These additional studies should also determine whether
CD1 lc+CDS+B-cell leukemias represent a clinically significant subgroup of chronic B-lymphoid neoplasms.
Our CD1 lc+CDS+ B-cell leukemias possessed several
differences and shared some similarities with previously
reported "variants" of CLL, PLL, HCL, and B-cell lymphomas. In contrast to splenic lymphomas with circulating
Table 5. Presenting Clinical Features of CD1lc+CD5+E-Cell Leukemias
Pt
Age
RR
ES
76
71
67
50
67
70
64
77
44
NU
ED
C8
8W
AS
IS
SP
WH
CR
MS
TW
MM
Sex
WEC/mm3 x lo3
Lymphadenopathy
Splenomegaly
F
19.4
37.0
17.3
83.4
8.6
14.4
58.4
21.4
44.5
10.3
14.4
230.0
204.9
6.1
-
+
-
M
M
F
F
F
82
M
M
M
M
71
52
52
45
M
M
M
M
+
+
+
+
+
+
i
+
-
+
+,
Neutropenia
Thrombocytopenia
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
+
+
-
-
-
-
-
-
-
-
+
+
+
+
+
+
+
+
+
+
Anemia
-
-
-
-
+
+
+
-
-
-
-
Summary
Diagnosis
CLL
CLL
CLL
CLL
CLL
CLL
CLL
CLL
CLL
CLL
CLL
PLL/CLL*
PLL/CLL*
CLL
Abbreviations: Pt, patient: WBC, white blood cell count;
present: -, absent.
*Patients had leukemic cells with bright slg staining and PLL clinical features, but lacked the well-defined nucleolus charactertistic of PLL cells.
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WORMSLEY ET AL
128
Fig 3. Representative three-color immunofluorescence analysis of 6-cell enriched normal peripheral blood lymphocytes labeled with
anti-CDPO-FITC, anti-CDSbiotin, and anti-CD11 c-PE followed by streptavidin-conjugated APC. The cytogram quadrants were set using
appropriate negative controls. Subsets of normal CD20+ 6 cells corresponding to the immunophenotypes of chronic ‘B-lymphoid
malignancieswere detected: yellow, CDPO’CDS’CDl 1c’; green, CD20fCD6-CD1 1c’; pink, CD20+CD5‘CDl 1c-; blue. CD20’CDS-CDll c-.
The mean percentages (n = 6) of normal CD20f 6-cell subsets with these immunophenotypes are listed in Table 6.
villous lymphocytes3’ and variant HCL,32our cases expressed
CD5 and the majority of leukemic cells did not have villous
cytoplasmic projections. Unlike “prolymphocytoid” transformation of CLL,3 C D l lc+CD5+ leukemias were not characterized by the presence of cells with prominent nucleoli.
While sharing some cytologic features with our cases, the
recently described monocytoid B-cell lymphomas do not
express CD5 or CD24 and they are rarely associated with a
leukemic p h a ~ e . Den
~ ~ Ottolander
.~~
et ai” have described a
form of CLL termed “lymphoplasmacytoid leukemia” characterized by eccentrically placed nuclei, moderately condensed chromatin, inconspicuous nucleoli, and moderately
abundant cytoplasm. Approximately half of these lymphoplasmacytoid leukemias expressed CD5 and C D l 1b; however,
they were not tested for C D l I C expression and the clinical
characteristics of these cases were not r e p ~ r t e d . ’Some
~
of
our cases, particularly those with eccentrically placed nuclei,
appeared to express some morphologic and immunophenotypic similarities with this subset of CLL.
With respect to the expression of other hematopoietic
markers, none of the CD1 lc’CD5’ B-cell leukemias tested
were FMC7 +, an antigen commonly expressed by HCL and
PLL.8.2’.36
All of our cases were CD24+, an antigen typically
expressed by CLL that is infrequently or weakly expressed
by HCL.5.35A subset of our cases were My4 positive, another
monocyte-associated antigen frequently expressed by CLL.37
Our cases were divided between those with “weak” and
“bright” sIg expression. Over one half (63%) of the cases
studied were CD25’ (IL-2 receptor), while the vast majority
expressed the B-cell activation antigen CD23 (B532). These
latter observations suggest that CD1 I C expression may be
associated with activated CD5+ malignant B cells. This view
is supported by previous studies describing the induction in
CLL cells of C D l I C expression by agents that promote
lymphoid activation and differentiati~n.~”.~’
It is presently
unknown whether the spectrum of C D l IC’ leukemia immunophenotypes identified in this study reflect normal stages of
B-cell activation/differentiation or aberrant C D l I C eXpreSsion associated with the leukemic process.
Consistent with the hypothesis that the immunopheno-
types of some B-cell neoplasms correspond to normal stages
of B-cell differentiation and activation, our three-color flow
cytometric analyses identified novel circulating B-cell subsets that expressed patterns of CD20, CD5, and C D l IC
antigens corresponding to the immunophenotypes of chronic
B-cell leukemias and lymphomas. The most common peripheral blood B-cell subpopulation had a CD20CCD5-CD1 ICimmunophenotype, and this subset represented nearly half of
the circulating CD20’ B cells. Our studies also showed that
normal circulating CD5+ B cells are comprised of CD1 I C +
and C D l IC- subsets. In our study, the mean percentage of
normal CD5’ B cells (approximately 40%) was higher than
the values reported in earlier studies (approximately 20% to
30% CD5’ B cell^).^'.^^ The higher value observed in our
study may be explained in part by our use of more sensitive
indirect staining methods, compared with the direct staining
techniques used by others.43 In addition, as some normal
subjects with greater than 50% CD5+ B cells have been
r e p ~ r t e d , ~ ’the
. ~ ~inclusion of individuals with high normal
CD5’ B-cell levels also may have contributed to the higher
mean percentage of CD5’ B cells observed in our investigation. Our analyses also identified normal peripheral blood B
cells with the immunophenotype of HCL (CD20’CDl IC+
CD5-). These CDllc+CDS- B cells may correspond to the
recently described subset of normal B cells defined by the
expression of a novel HCL associated antigen (B-ly 7).44
These circulating C D l lc’CD5- B cells may also represent
the peripheral blood counterpart of lymph node C D l lc+CD5B cells presumed to be the cellular origin of monocytoid
B-cell lymphoma.33Further studies of these previously unrecognized peripheral blood B-cell subsets will be required to
integrate their immunophenotypes into the pathways of
normal B-cell differentiation/activation and to determine
whether they have distinct functional characteristics.
ACKNOWLEDGMENT
We thank Drs Frederick Davey and Paul Kurtin for reviewing the
manuscript, and Bonnie Kats for assistance with manuscript preparation.
From www.bloodjournal.org by guest on June 18, 2017. For personal use only.
CD1 lC'CD5'
129
CHRONIC B-CELL LEUKEMIAS
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1990 76: 123-130
Characteristics of CD11c+CD5+ chronic B-cell leukemias and the
identification of novel peripheral blood B-cell subsets with chronic
lymphoid leukemia immunophenotypes [see comments]
SB Wormsley, SM Baird, N Gadol, KR Rai and RE Sobol
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