ICCS e- Newsletter CSI
Summer 2013
Bioq. Mariela B Monreal
Dra. Marina Narbaitz
Dra. Cecilia Cabral
Division of Hematopathology
FUNDALEU - Buenos Aires - ARGENTINA
Clinical History - Presentation
Previously healthy 63 y/o female, presenting with chills
asthenia , constipation and weight loss of 14 kg
(30 lbs) in one month.
Abdominal ultrasound & CT scan revealed multiple
retroperitoneal and supraclavicular
lymphadenopathies.
Complete blood count & Laboratory results
CBC parameter
WBC
RBC
HGB
HCT
MCV
MCH
MCHC
RDW
PLT
Result
10,40
3,20
8,90
27,00
85,00
28,00
32,80
13,50
309,00
WBC differential
Neutrophils
Lymphocytes
Monocytes
Eosinophils
Others
%
89
7
4
0
0
ESR
LDH
70
667
Units
9
x10 /L
x1012/L
g/dl
%
fl
pg
gm/dl
%
Reference
range
4.6-10.2
4.0-5.48
12.2-16.2
37.7-47.9
80-97
27-31
31.8-35.4
11-14
142-424
50-68
21-30
4-8
2-4
0
mm
UI/l
0-15
260-460
Work – up and Evaluation
Lymph node biopsy was submitted for morphology and flow cytometry, for initial
evaluation.
Flow Cytometry analysis:
Selected files (8-color antibody tubes) are provided for review:
Tubes #1 - 4 tested in Lymph node ; Tube BM : tested in Bone marrow aspirate
Data Acquisition : FACS CANTO II and DIVA software
Data Analysis: Infinicyt (Cytognos)
FITC
PE
PerCP-Cy5
PE Cy7
APC
APC H7
V 450
V500
Tube #1
LAMBDA
KAPPA
CD5
CD19
CD38
CD10
CD20
CD45
Tube #2
CD23
CD7
CD5
CD123
CD2
CD3
HLA DR
CD45
Tube #3
CD4
CD13
CD5
CD123
CD33
CD3
HLA DR
CD45
Tube #4
CD36
CD64
CD34
CD117
IREM
CD14
HLA DR
CD45
Tube BM
CD4
CD7
CD5
CD56
CD2
CD8
CD3
CD45
Tube #1 : B cell markers
FITC
PE
PerCP-Cy5
PE Cy7
APC
APC H7
V 450
V500
LAMBDA
KAPPA
CD5
CD19
CD38
CD10
CD20
CD45
In this first tube , an abnormal population of LARGE CELLS (in grey) is identified . Cells are NEGATIVE for
all B cell markers tested; and also for CD5 and CD10.
Very bright CD45 intensity and dim/neg CD38 exclude plasmacytic differentiation.
Tube #2 : T cell markers / Lymphoplasmacytoid DC
Tube #2
FITC
PE
PerCP-Cy5
PE Cy7
APC
APC H7
V 450
V500
CD23
CD7
CD5
CD123
CD2
CD3
HLA DR
CD45
ABNORMAL POPULATION is shown here in RED
br HLA DR++ and dim/interm CD123+
CD7, CD2 and CD3 are NEGATIVE
LYMPHOPLASMACYTOID DENDRITIC CELLS (DC2)
are CD123++ HLA DR++
1
Tube #3 : Myeloid markers & CD4 CD56 HLA DR
Tube #3
FITC
PE
PerCP-Cy5
PE Cy7
APC
APC H7
V 450
V500
CD4
CD13
CD5
CD123
CD33
CD3
HLA DR
CD45
ABNORMAL CELLS are

br HLA DR+++ and br CD33+++

br CD4++ and dim CD13+

CD56 NEGATIVE (additional tube)
MONOCYTES are distinguished from large abnormal cells,
showing normal FSC/ SSC features ; br CD13++ and dim
CD4 expression
e CSI
Tube #4 : Monocytic maturation. Immature markers
Tube #4
FITC
PE
PerCP-Cy5
PE Cy7
APC
APC H7
V 450
V500
CD36
CD64
CD34
CD117
IREM
CD14
HLA DR
CD45
Bright CD64+ expression allows the
identification of MONOCYTIC LINEAGE.
In this case, mature monocytes:
CD36++ CD14++ and IREM-2 ++.
ABNORMAL CELLS are:
CD64dim+
CD14neg/dim+
CD36neg/dim+
Additional tube demonstrates that MPO is
NEGATIVE
Summary - Immunophenotypic findings (FCM)
Abnormal cells : CD45++ CD33++ HLA DR++ CD123+ CD4++ ,dimCD64+ ,
dim/variable CD14+ , dim/variable CD36+ , IREM neg/dim+
MPOneg CD15neg CD56neg
Lymph node (and BM , see below plots on the right) infiltrated by large /
pathological cells , with phenotypic features that suggests monocytic /
monocytic-related dendritic cell subset.
LYMPH NODE
BM ASPIRATE
LYMPH NODE
Lymph node biopsy demonstrates a diffuse proliferation of pleomorphic large cells.
The nuclei are irregular, often folded, with occassional nucleoli. The cytoplasm is
abundant and eosinophilic.
Reactive eosinophils were observed in the background.
Immunophenotype
CD45 +
MPO -
Histiocytic
markers
CD68 +
(KP-1 Clone,
stains also
myeloid cells)
Unspecific
staining
CD43 +
(Sialoglycoprotein
on the surface of
monocytes)
CD68 +
VIMENTIN +
(PGM-1 Clone)
(Intermediate
filament protein
present in cells of
mesenchymal
origin)
CD163 +
KI67 - 75%
(high
proliferation
index)
BONE MARROW
Bone marrow core biopsy
demonstrates marrow infiltration
by neoplastic cells with similiar
features as observed in lymph
node.
CD45 +
CD43 +
CD163 +
MPO -
Preliminary Diagnosis
LYMPH NODE & BONE MARROW involvement :
Hematologic malignancy with expression of
Mono/Histiocytic markers.
Given this immunophenotypic profile….
Lineage Assignment possibilities
Immunohistochemistry
Flow cytometry:
CD45 + CD43 + CD68 + CD163 +
HLA DR+++, CD33+++,CD123+,CD4+,CD64+w, CD14+w.
MPO - CD34 - CD20 - CD3 -
NG2 -, T y B -, CD13 -, CD1a -, MPO – CD56 --
MYELOID
STEM
CELL
CD14CD11c+
CD1a+
CLA+
LANGHERHANS DC
Monocyte
CD14+
CD11c+
CD68+
CD1aCLA-
MACROPHAGE
INTERSTITIAL
DC
CD14+
CD11c+
CD1aBDCA2+
CD123+
BDCA4+
PLASMOCYTOID
DENDRITIC CELL
(DC2)
CD4+
CD56+
CD123+
CD13+
CD33+
CD14+
CD15+
MPO+/LISOZIMA+
BDCA3+
MYELOID
DENDRITIC CELL
(DC1)
Differential Diagnosis
•
HISTIOCYTIC SARCOMA.
-
Must express two or more monocyte/macrophage lineage antibodies (CD14, CD11c, CD13, CD68,
CD163).
CD4 (cytoplasmic), CD15, CD43, CD45, CD45RO, CD33, lysozyme +/WHO 2008 definition: Express monocyte/macrophage markers. Myeloid lineage antibodies (CD33,
CD13) MUST BE NEGATIVE.
-
•
AML WITH MONOCYTIC DIFFERENTIATION/ MYELOID SARCOMA.
-
> = 20% myeloid blasts, with less than 20% of cells with monocytic differentiation.
AML M4; M5.
-
M4: The PB or BM has more than 20% blast (including promonocytes), neutrophils and their precursors
and monocytes and their precursors.
M5: Myeloid leukaemia in which 80% or more of the leukaemic cells are monocytic lineage including
monoblast, promonocytes and monocytes; a minor neutrophil component, <20%, may be present.
•
PLASMACYTOID DENDRITIC CELL NEOPLASM/ BLASTIC NK-CELL
LYMPHOMAHEMATODERMIC NEOPLASM.
-
Expression of CD4, CD56 and CD123 antigens with concomitant negativity for other myeloid and
lymphoid associated markers.
• ACUTE MYELOID DENDRITIC CELL LEUKAEMIA.
-
Expression of CD4, CD56 and CD123 antigens with concomitant positivity for myeloid markers
(CD13,CD33, CD64, 7.1, IREM).
ACUTE MYELOID DENDRITIC CELL LEUKAEMIA
• Acute dendritic cell leukemias are very uncommon and have a
lymphoplasmacytoid dendritic cell (DC2) phenotype more often than a
myeloid dendritic cell phenotype (DC1).
• Myeloid dendritic cell leukaemia is exceptional, and it would
differentiate from plasmacytoid dendritic cell leukaemia by the
expression, as well as of CD4, CD56 and CD123, of some myeloid
makers [CD13, CD14, CD15, CD33, myeloperoxidase (MPO),
lysozyme] and specific myeloid dendritic cell antigens (BDCA3)
instead of plasmacytoid dendritic cell antigens (BDCA2, BDCA4).
• Spontaneously occurring acute myeloid dendritic cell leukemia is very
infrequent. It has been estimated, based on the study of 392 cases of
AML, that 0.8% of cases had overt features of a dendritic cell
malignancy.
• Anemia, thrombocytopenia, and blood, marrow, and skin involvement
with dendritic-like blast and more mature appearing dendritic cells are
characteristic findings. Lymph node and spleen enlargement from
leukemic cell infiltration usually is present.
ACUTE MYELOID DENDRITIC CELL LEUKAEMIA
• Blast cells do not display myeloperoxidase or esterases
by cytochemistry.
• The main differential diagnosis of acute myeloid dendritic
cell leukaemia is acute myeloid leukaemia (AML). In fact,
some authors consider myeloid dendritic cell leukaemia
as a morphological variant of AML. AML blast cells
frequently express the dendritic cell-associated marker
CD86, especially among acute monocytic leukaemia
cells. Dendritic cell features can be found in AML cells
after chemotherapy. In addition, using cytokines and
CD40 ligands, a dendritic cell phenotype strikingly similar
to the blast cells of myeloid dendritic cell leukaemia can
be induced from AML blast cells.
References
•
Swerdlow SH, Campo E, Harris NL, et al. WHO Classification of Tumours of Haematopoietic
and Lymphoid Tissues. Lyon, France: (IARC; 2008).
•
Marshall A. Lichtmana, b, George B. Segelc Uncommon phenotypes of acute myelogenous
leukemia: Basophilic, mast cell, eosinophilic, and myeloid dendritic cell subtypes: A review.
Blood Cells Mol Dis.2005 Nov-Dec;35(3):370-83
•
Martín-Martín L, Almeida J, Hernández-Campo PM, Sánchez ML, Lécrevisse Q, Orfao A
Br J Dermatol Immunophenotypical, morphologic, and functional characterization of
maturation-associated plasmacytoid dendritic cell subsets in normal adult human bone
marrow. Transfusion 2009 Aug;49(8):1692-1708
•
M. Ferran, F. Gallardo, A.M. Ferrer,A. Salar, E. Perez-Vila,N. Juanpere, R. Salgado, B. Espinet, A.
Orfao,–L. Florensaand R.M. Pujol Acute myeloid dendritic cell leukaemia with specific
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•
Orfao A. Neoplasias of dentritic cells: are they the counterpart of one or more cell lineages?
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•
Elizabeth L Courville, Yue Wu, Jihen Kourda, Christine G Roth, Jillian Brockmann, Alona
Muzikansky, Amir T Fathi, Laurence de Leval, Attilio Orazi and Robert P Hasserjian
Clinicopathologic analysis of acute myeloid leukemia arising from chronic myelomonocytic
leukemia modern Pathology , (11 January 2013)
•
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