The document discusses acute leukemias of ambiguous lineage, specifically mixed phenotype acute leukemia (MPAL). It defines MPAL as acute leukemias that express antigens from more than one lineage, with at least 20% blasts. The most common combinations are B/myeloid and T/myeloid, seen in 59% and 35% of cases respectively. Diagnosis relies on extensive immunophenotyping using markers like MPO, cCD3, CD19. Patients are typically treated with a corticosteroid trial followed by an AML-like regimen if no response, and stem cell transplant offers the best outcome. The discussion presents examples of typical MPAL immunophenotypes and

1. Acute Leukemias of Ambiguous Origin
Anna Porwit and Marie C. Béné
Am J Clin Pathol September 2015;144:361-376
KEY WORDS : Acute leukemia,
Immunophenotype, Lineage, Myeloid, Lymphoid,
Mixed phenotype acute leukemia, Undifferentiated
acute leukemia
Presenter : Dr.Revathi Krishnamurthy
Moderator : Dr.K.S.Rajasekhar

2. INTRODUCTION

3. • The acute leukemias are a heterogenous group of
neoplasms arising from transformation of
hematopoietic stem cells, usually with a retained
partial capacity of differentiation
• They are characterised by malignant neoplastic
proliferation and accumulation of immature and
non-functional hematopoietic cells in the bone
marrow

4. • The neoplastic cells show :
- Increased proliferation and/or
- Decreased programmed cell death (apoptosis)
• The net effect is expansion of the leukemic clone and
a decrease in normal cells

5. The two major categories of acute leukemias are
classified according to the origin of the cell with the
primary defect
Acute Myeloid
Leukemia[AML]
(Non-
lymphocytic)
Acute Lymphoid
Leukemia
[ALL]
(Lymphoblastic
leukemia)

6. As acute leukemias are a heterogenous group of
neoplasms with differences in clinical course,
prognosis and treatment, with the invent and
application of target-based approach to therapy, their
classification needs to be :
Precise, facilitating non-overlapping
identification of the differing entities, incorporating
all the essential and new information

7. • 1857 : The attempt to classify leukemias was initiated by
Nikolaus Friedreich who categorised leukemias as acute and
chronic
• 1976 : Though the morphological approach to classify
acute leukemias has always been in progress, standard
criteria to distinguish between myeloid and lymphoid acute
leukemias and to subtype them further, based on morphology
and cytochemistry were first laid down by the FAB working
group
EMERGENCE OF FRENCH-AMERICAN-BRITISH
(FAB) CLASSIFICATION

8. Subsequently, with the recognition of new
morphological subsets, the original FAB
classification was modified further
Cut off blast percentage – 30 %

9. PITFALLS OF FAB CLASSIFICATION AND
INTRODUCTION OF WORLD HEALTH
ORGANISATION [WHO] CLASSIFICATION
The FAB classification had major disadvantages as :
• Cursory correlation with clinical outcome
• Poor concordance owing to inter-observer variation
• Failure to incorporate cytogenetic data
• Furthermore, many cytogenetic abnormalities were
identified in the subtypes of leukemias in the latter
half of twentieth century

10. • 1997 : The WHO classification of leukemias
evolved
• Thus, the WHO classification had incorporated
cytogenetic abnormalities and immunology as
principal designating criteria, despite retaining
morphology as the mainstay of the diagnosis
Cut off blast percentage – 20 %

11. Immunophenotype and genetic features have now
become an essential integral part of the definition of
hematopoietic neoplasms

12. 1995 : The European Group for Immunological
Characterising of Acute Leukemia (EGIL)
formulated guidelines for classification of acute
leukemia with biphenotypic marker expression

14. The 2008 WHO classification parameters include not
only morphology and cytochemistry but also :
• Flow cytometry
• Cytogenetics
• Molecular genetic abnormalities
• Clinical findings
WHO CLASSIFICATION

15. MAJOR DIFFERENCES BETWEEN
2001 AND 2008 WHO
CLASSIFICATIONS

17. WHO 2001 WHO 2008

18. Identification of cell lineage
The first step in classifying acute leukemia is to
differentiate ALL from AML, usually by identifying
the lineage of the blasts

19. Morphology
It is the most important since some neoplasms have
characteristic or diagnostic cell features

20. Nucleus
Size Position Shape Chromatin Cytoplasm
Myeloblast 10 -18 µ Eccentric or
central
(3-5
nucleoli)
Round or
oval
Very fine
meshwork
( Lacy )
Light blue
Lymphoblast 10 – 18 µ Eccentric or
central
(1 or more
nucleoli)
Round or
oval
Moderately
coarse
particles,
stippled
Scant
amount,
usually
granular
Myeloblast Lymphoblast

21. • However, differentiation of ALL from AML
(minimally differentiated or AML without
maturation) may not be possible by morphological
examination of Romanowsky stained smears alone
• Thus, when morphological examination cannot
differentiate between the blasts, cytochemistry may
be helpful

22. Cytochemistry
Myeloperoxidase and Sudan Black-B :
• To differentiate myeloid and lymphoid blasts
• Myeloblasts are positive
• Lymphoblasts are negative
• In ALL, up to 3% positivity in blasts is acceptable
to accommodate residual normal myeloblasts in the
sample

23. Esterases :
• To distinguish monoblasts from myeloblasts
• Myeloblasts – Positive for specific esterases
• Monoblasts – Positive for non-specific esterases
Non-specific esterase

24. Periodic Acid Schiff :
• Helpful in diagnosing ALL, acute erythroid and
megakaryoblastic leukemias
• Lymphoblasts – Block-like PAS positivity
• Erythroblasts in erythroid leukemia – Positive
• Megakaryoblasts – Fine granular PAS positivity

25. Terminal Deoxynucleotidyl Transferase : (TdT)
• It is an intracellular enzyme helpful in identifying
cellular subtypes
• DNA polymerase found in cell nuclei
• This enzyme is not present in normal mature
lymphocytes but can be found in 65% of the total
thymic population of lymphocytes with TdT
positive cells localised in the cortex

26. Can also be found in :
• Very early B cells
• Very early myeloblasts (occasionally)
Value in ALL : To differentiate early
precursor lymphoblasts from more mature
cells

27. Immunophenotyping
• It is the identification of antigens using detection
antibodies
• Antibodies are utilised because they bind
specifically to antigens and can be labelled with
fluorochromes to provide a sensitive and specific
detection method

28. • Most flow cytometric immunophenotyping studies
involve detecting cell surface antigens
• Intracytoplasmic and intranuclear antigens can be
detected following permeabilisation of cell
membrane with detergent and/ or alcohol

29. • Immunophenotyping allows a more precise
characterisation of acute leukemias by detecting
lineage-associated markers expressed by the
leukemic cells
• It has become an indispensable part of the
integrated hematopathological diagnostics of
leukemia
• This methodology relies on the use of an antibody
panel, to be chosen among the more than 300
clusters of differentiation (CD)

30. ALL :
Immunophenotyping is essential for :
• The diagnosis of ALL
• Separation of T- and B-ALL

31. B lineage :
CD markers
Commitment • CD19
• Cytoplasmic CD22
Immaturity • CD10
Others • CD20
• Cytoplasmic
CD79a

32. T lineage :
• CD1a
• CD2
• CD3
• CD4
• CD5
• CD7
• CD8

33. AML :
• CD34
• HLA-DR
• CD117
• CD13
• CD33
• CD15

34. Acute Leukemias of Ambiguous Lineage :
• However, some cases of acute leukemia do not fit
neatly into the categories ALL and AML
• It can be impossible to assign lineage because :
Blasts express very few
antigens
Mixed Phenotype
Acute Leukemia
Blasts express antigens from
more than one lineage
Acute
undifferentiated
Leukemia

35. Biphenotypic Acute Leukemias : (BAL)
• The first attempt to standardise the diagnostics of BAL
was provided by the EGIL group in 1995
• Fine morphologists had repeatedly recognised
leukemia cases in which there seemed to be 2 different
clones among the proliferating cells
• Later, observations of aberrant co-expressions of
immunophenotypic markers considered to have an
association with B, T, or myeloid lineages added to the
complexity, even in morphologically homogenous
proliferations

36. Thus, to describe these rare leukemias, the acronym
BAL was used, encompassing :
• Proliferations exhibiting 2 different clones :
Bilineal/Biclonal leukemias
• Cases with co-expression of markers from 2 or
sometimes 3 lineages on blast cells : Biphenotypic
leukemias

37. Scoring system proposed by EGIL :

38. • A major change in the 2008 revision of the WHO
classification was to simplify the definition of BAL
• A group of experts reviewed available data from the
literature and defined the new entity of Mixed
phenotype acute leukemia

39. Mixed Phenotype Acute Leukemia (MPAL)
• In this new proposal, only markers considered most
specifically lineage associated have been retained
for lineage assignment
• MPAL represent less than 5% of acute leukemia
• Display myeloid morphology in 60% of cases

40. ARTICLE PROPER

41. What is the Definition of Acute Leukemia of
Ambiguous Lineage ?
• In the 2008 World Health Organisation (WHO)
classification,acute leukemias of ambiguous lineage
were defined as leukemias that show no clear
evidence of differentiation along a single lineage
• The recognition of acute leukemia of ambiguous
lineage requires extensive multiparametric flow
cytometry (FCM) immunophenotyping disclosing
the specific features of these diseases

42. Extensive immunophenotyping
Acute
undifferentiated
leukemia
Mixed
Phenotype
Acute Leukemia
Others,
including
Natural Killer
With
t(9;22)
With
t(n;11q23)
Not otherwise
specified

43. • In extremely rare cases of acute undifferentiated
leukemia (AUL) :
Although markers of all lineages have been
investigated, no significant expression is detected on
blast cells except for :
- CD34 and/or
- HLA-DR
• A second, slightly more frequent occurrence is that
of blast cells labelling with several monoclonal
antibodies, recognising antigens normally expressed
on different lineages

44. Biphenotypic Acute
Leukemias (BALs)
Mixed-Phenotype Acute
Leukemia (MPAL)

45. Mixed Phenotype Acute Leukemias
t (9;22)/BCR-
ABL1
t (v;11q23)/MLL
Not otherwise
specified
According to the presence of Philadelphia chromosome

46. All other unusual immunophenotypes, including early
natural killer (NK) leukemias, constitute the
remainder of leukemias of ambiguous origin

47. Introduction to MPAL :
• The morphological features of the blasts in most
cases of MPAL are mostly uninformative
• Therefore the diagnosis of MPAL relies exclusively
on immunophenotypic features

48. • MPAL cells appear most often as morphologically
undifferentiated
• Sometimes the neoplastic cells may display more
lymphoblastic or myeloblastic cytology

49. • In some cases, two types of blasts with a distinctive
size and morphology point to the extremely rare
occurrence of bilineal or biclonal leukemia
• These neoplasms can also be recognised by FCM
analysis as two cell subsets with different
immunophenotypes

50. Several markers specific for :
• Myelo/monocytic lineage
• B- and T- lymphoid lineages must be tested
excluding a restrictive strategy of quick orientation
followed by selected lineage-specific markers

51. At least two key cytoplasmic (c)markers must be
investigated :
Myeloperoxidase
(MPO)
Myeloid lineage
cCD3
T lineage

52. • In the 2008 WHO classification, the name of these
leukemias and their immunophenotypic definition
were changed
• MPO stands as the most robust marker that will
identify the myeloid engagement

53. An alternative, for MPAL with a strong monocytic
differentiation, is represented by cases in which blasts
will be positive for :
• Nonspecific esterase and/or
Express at least two of the following antigens:
• CD14
• CD11c
• CD36
• CD64
• Cytoplasmic Lysozyme

54. Enagagement of blast cells in the B-lymphoid
lineage will rely on the expression of surface
CD19
If Bright
Expression of one
other B-lineage-
associated
marker
If weak
Expression of at
least two other B-
lineage markers
Other B-lineage associated markers : cCD79a , surface or
cCD22, or CD10

55. • Assessment of T-lineage engagement relies on the
demonstration of cCD3 expression
• The fluorochrome used - should yield strong
fluorescence, for instance phycoerythrin or
allophycocyanin
• The immunophenotypic strategy to explore these
cells involves the use of two monoclonal antibodies
directed to CD3 (usually anti-ϵ chain) conjugated to
different fluorochromes

56. Cells are incubated without manipulation, ideally
using the whole bone marrow (BM)
Permeabilizing agents will allow both :
- Lysis of erythrocytes
- Intracytoplasmic labelling with the
second anti-CD3 antibody
Analysis by FCM :
 Normal residual T-lymphocytes : Costained
with both antibodies
 T-lineage blast cells lacking surface CD3 :
Stained with the second anti-CD3 only

57. What possible combinations of MPAL occur and
what are their frequencies ?
All possible combinations of MPAL can be observed,
including :
 B/myeloid
 T/myeloid
 B/T
 B/T/myeloid

58. In a series of 100 cases of MPAL published by the
European Group for the Immunological
Characterisation of Leukemias (EGIL) :
B/Myeloid
59%
T/Myeloid
35%
B/T
4%
B/T/Myeloid
2%

59. How are patients with MPAL managed clinically ?
• The clinical management of MPAL cases is
problematic
Initial course of corticosteroids
Acute lymphoblastic leukemia-like approach

60. In the absence of response to corticosteroids
AML – like approach with alkylating agents
• However, patients who fared well in the literature
were clearly those who could benefit from allogenic
stem cell transplantation

61. • It is important to identify MPAL cases and not
misdiagnose these tumours as ALL or AML by
using immunophenotypic panels that are not
sufficiently comprehensive
• Indeed, some refractory cases of acute leukemia,
with poor response to therapy, may represent
undetected cases of MPAL that were incorrectly
assigned to a single lineage

62. • Using an FCM intracytoplasmic orientation panel
combining MPO, cCD3 , CD19 (possibly with
cCD79a , cCD22, or CD10) could be a good
approach to detect such cases early in the process of
immunophenotyping a neoplasm
• It is important to point out that
immunohistochemistry (IHC) on BM biopsy
sections also can be useful to confirm B, T, or
myeloid differentiation by using antibodies specific
for PAX5, CD3, and MPO and/or lysozyme,
respectively

63. Immuohistochemical analysis may be of great help in
cases with two or more different populations with
specific distributions in BM or other tissue biopsy
specimens since it allows direct visualisation of cell
location

64. DISUSSION OF WORKSHOP
CASES

65. • There were 32 cases included in this workshop
session
• The following discussion describes :
- The characteristics of MPAL cases
- Technical issues involved in MPAL diagnosis
- Groups of cases that can be linked by some
common characteristics or diagnostic issues

66. TYPICAL EXAMPLES OF MPAL

67. MPAL B / Myeloid :
MGG MPO

68. Bone marrow biopsy specimen
CD79 positive
MPO positive

69. CD 19
Moderate to strong in 5 cases,
(10,208,285,323,328)
weak/partial in all other cases
B cell
lineage
cCD79a, CD22, and/or PAX5
CD 10 Positive in three cases
(10,208,214)
• Most blast cells- MPO
positive in 4 cases(10,76,285,323)
• Other cases showed only
subpopulations of MPO-
positive cells
MPO

70. Cases 208, 328, 214 represented the most common
cytogenetic variant of B/myeloid MPAL, associated
with t(9;22)(q34;q11.2)/BCR-ABL1

71. 3 cases
Highly hyperdiploid and/or
Near-tetraploid karyotype
1 case Hypodiploidy
• Highly hyperdiploid and/or near-tetraploid
karyotypes are rare in AML and ALL
• Have been reported previously as common in MPAL
and BAL classified according to the EGIL system

72. • One of the near-tetraploid cases (case 285) carried also
the t(12;17)(p13;q11.2)
• The t(12;17)(p13;q11)  described as a rare but
recurrent abnormality, predominantly observed in
B-ALL cases that usually have an early pre-B
immunophenotype and showing aberrant expression
of the myeloid antigens CD13 and/or CD33

73. • The t(12;17) is less common in AML, and reported
cases often showed aberrant expression of CD19
• Case 285 is the first case with this translocation in
which the WHO criteria for MPAL are fulfilled
(CD19+, cCD79a+, MPO+, CD33+, CD34,HLA- DR+,CD38+,and with
partial expression of CD13 & CD15)

74. • Another chromosomal abnormality hitherto not
described in MPAL, inv(3)(q21q26.2) was detected
in case 283
• Inv(3)(q21q26.2) or t(3;3)(q21;q26.2) displays an
inversion or homologous reciprocal translocation
that leads to juxtaposition of the ecotropic viral
integration site 1(EVI1) gene with the ribophorin
1(RPN1) gene

75. • AML cases carrying this abnormality are classified
as one of the categories of AML with recurrent
cytogenetic abnormalities in the WHO classification
• Most commonly have a myeloid or myelomonocytic
immunophenotype and poor prognosis
• An MPAL phenotype has not been reported
previously in a leukemia with inv(3) or t(3;3)

76. Flow cytometry immunophenotypic findings in case 208

77. MPAL T / Myeloid
Bone marrow aspirate
MGG
Bone marrow biopsy
H&E

78. Lymph node sections
Positivity for cytoplasmic CD3
Positivity for MPO

79. Most T/Myeloid MPALs were characterised by two
population of blasts :
T-precursor
immunophenotype
(usually co-
expressing some
myeloid markers
but negative for
MPO)
Fulfilling MPAL
criteria

80. 2 cases
Myeloid lineage showed a
monocytic differentiation pattern
2 cases
Different immunophenotypes
were observed in a lymph node
biopsy specimen and the BM

81. In one case (200) :
• BM : Immunophenotype suggestive of AML with
monocytic differentiation
• Lymph node : Two aberrant populations were
detected
1. cCD3 positivity
2. Myelomonocytic immunophenotype
• Cytogenetics : Two different clonal populations
1.46,XY,der(12)t(11;12)(q21;p11.2)
2.45,XY,dic(12;18)(p11.2;p11.2)

82. In one case (402) :
• Lymph node biopsy : T-precursor
immunophenotype
• BM : Population of blasts fulfilling the MPAL
criteria

83. Flow cytometry immunophenotypic findings in case 279

84. MPAL B / T
• MPAL cases that express both B- and T- lineage
markers are rare
• Three examples (312,359,390) were submitted to the
workshop

85. MPAL B/T
Bone marrow aspirate
MGG
Bone marrow biopsy
H&E

86. Flow cytometry immunophenotypic findings in case 305

87. In two cases :
• A population of blasts : CD 19 (strong)
cCD3
• One of these cases had a normal karyotype and one
was hyperdiploid
B-cell lineage
1.CD 22
2.CD 79a
T-cell lineage
1.CD 7
2.No other T-
cell associated
marker

88. Third case :
• Trisomy 11
• Two populations of blasts :
- Immunophenotype of B-precursor ALL (EGIL B-II)
- T-ALL with early T-cell precursor
immunophenotype (EGIL T-I)

89. MPAL B / T/ Myeloid
Case 268 was the only case of acute leukemia that
expressed markers of all three lineages fulfilling
criteria for B/T/myeloid MPAL

90. • The patient was a 58 year old man with two
separate leukemic cell populations
• Corresponded to
AML
• Positive for : CD7,
CD4 (dim),
CD11b, CD13,
CD33, CD34,
CD117, MPO,
HLA-DR
• Corresponded to
MPAL with B/T
immunophenotype
• Positive for : TdT,
CD19, CD20,
CD10, cCD79a,
CD22, PAX5,
CD33 (dim),
cCD3, CD5(dim)
• Negative for :
MPO

91. • Conventional cytogenetics showed a normal
karyotype and BCR-ABL1 was negative

92. CD 117 +
PAX 5 –
AML
CD 117 -
PAX 5 +
MPAL

93. Higher magnification of areas outlined in A and B respectively

94. • Another case fulfilling the criteria for T/Myeloid
MPAL and also showing weak CD19 expression
was case 356
• The patient was a 14 year old girl in whom both a
lymph node biopsy specimen and BM showed blasts
positive for cCD3 and MPO with weak expression
of CD19 and CD79a
• However, other B-cell markers were not found and
the B-lineage differentiation were not fulfilled

95. MLL Rearrangement and Immunophenotype
Switch
• MPAL with MLL rearrangement is considered a
separate entity in the WHO classification
• Frequency of approximately 10% of adult and 12%
to 18% paediatric MPAL cases
• Most cases of MPAL with MLL rearrangement are
B/myeloid

96. • There were 4 examples of MPAL associated with
MLL rearrangement
• 1 case illustrated a rare occurrence of T/Myeloid
MPAL
• The other three cases demonstrated an
immunophenotype switch, previously reported in
MLL rearranged leukemias

97. • MLL rearrangement juxtaposes the amino-terminus
of the histone methyltransferase MLL with a variety
of different fusion partners
• To date, more than 70 fusion partners of the MLL
gene have been characterised
• Deregulation of HOX seems to be one of the most
important factors in MLL-induced leukemogenesis

98. Since HOX expression is high in stem cells and early
precursors and needs to be downregulated for
maturation, a continuous ectopic HOX expression
will create a basis for the development of abnormal
preleukemic precursors that may progress to various
forms of acute leukemia

99. • In 2 cases (180,370) submitted to this session:
- Primary diagnosis AML with t(9;11)(p22;q23)
- Relapse  ALL/lymphoma with an EGIL B-I
immunophenotype (CD10 negative)
• In the third patient (392), a therapy-related myeloid
neoplasm presented as a t(4;11)(q21;q23) B-
ALL/lymphoma with an EGIL B-I immunophenotype,
whereas :
- AML with monocytic differentiation -First relapse
- B/myeloid MPAL - Second relapse

100. The MLL-A4 fusion has been reported in therapy-
related ALL, after treatment with either
topoisomerase II inhibitors or alkylating agents, and
seems to be associated with a worse clinical outcome

101. What are some of the important technical and
interpretation issues in the diagnosis of MPAL ?
MPO
• Previous French-American-British group
guidelines, based on cytochemistry, used 3% of
MPO-positive blasts in BM smears as sufficient to
call a leukemia MPO positive
• A threshold of 10% expression has been used by
the EGIL group

102. • However, discordant cases are found when both
methods are compared, leading to the conclusion that
the 10% threshold may be conservative but not very
sensitive
• Thus, MPO expression in leukemic blasts has to be
compared with internal negative and positive
controls
• In a study comparing MPO expression detected by
FCM and cytochemistry in cases of AML and ALL, a
13% threshold was found to be relevant using an
isotype control as a background reference
(sensitivity-95.1%, specificity- 91.7%)

103. • If residual normal lymphocytes were used as
reference, a threshold of 28% had to be applied,
yielding an improved 97.4% sensitivity and 96.1%
specificity in distinguishing between ALL and AML
• The WHO criteria for MPAL do not indicate any
lower limit for MPO expression in leukemic blasts,
but it seems reasonable to use published thresholds
• Since MPAL cases often show more than one
population of blasts, MPO could be present in only a
minor population, which has to be identified as the
myeloid component

104. • Another issue is that AML with minimal
differentiation and no MPO expression could be
involved in MPAL
• Therefore, as stated in the WHO criteria, when there
are two or more distinct populations of leukemic
cells, one with co-expression of a number of
myeloid markers, no MPO and no lymphoid
markers also can be accepted to define the myeloid
component of MPAL

105. • Rarely, cases of otherwise typical B-ALL /
lymphoma can express MPO by FCM or IHC or
least often by cytochemistry
• These cases are best classified and treated as B-
ALL / lymphoma

106. • In contrast, most MPAL cases show expression of
other myeloid-associated markers
• Also characterised by the presence of two or more
subpopulations of blasts with slightly different
immunophenotypes
• In some cases of MPAL, MPO can be expressed
only in a small subset of blasts, with or without
lymphoid markers (biphenotypic or bilineal
presentation)

107. Cytoplasmic CD3
• Cytoplasmic CD3 (cCD3) is considered the most
specific marker for the T-cell lineage
• It has been used to diagnose T-cell malignancies
since the 1980s
• FCM immunophenotyping methods to detect cCD3
were established in the 1990s when
permeabilization reagents became available

108. • Expression of cCD3 in leukemic blasts has to be
determined in comparison with internal negative
and positive controls
Negative controls
• B cells
• Monocytes
• Granulocyes
Positive control
• Normal T
lymphocytes

109. • At least a fraction of blasts should express cCD3 at the
level of normal T cells, and weak expression in a minor
fraction of blasts is insufficient to diagnose MPAL
• Many AML cases show aberrant expression of other T-
cell associated markers, such as:
• Or NK-cell-associated markers such as CD 56
CD 2
CD 4
CD 5
CD 7

110. • Thus, the correct interpretation of cCD3 staining is
important for final diagnosis
• If a BM biopsy specimen is available, the
expression of cCD3 may be confirmed by IHC

111. CD19 and other B-cell markers
• The WHO classification provides no definite
threshold for the size of the population expressing
CD19 that is necessary to define B-lineage
involvement
• In two cases of bilineal B/myeloid MPAL (cases 206,365),
only a minor CD19+ populations were detected
• Since B-cell lineage was confirmed by other
markers, criteria were met for the diagnosis of MPAL

112. The intensity of CD19 expression by blasts should be
compared with that expected by normal B cells and, if
comparable, considered bright

113. • It has to be stressed that in cases in which :
- The whole blast cell population : strongly
expresses CD 19
- Immunophenotype : consistent with B-
ALL/lymphoma
Presence of MPO expression as a sole aberrant
marker should not dissuade therapy using a B-ALL
regimen, and patient prognosis does not seem to
differ from other patients with B-ALL

114. What criteria are helpful in the differential
diagnosis of AUL v/s AML with minimal
differentiation ?
Five cases submitted to this session (cases 199,250,261,349,396)
illustrated classification difficulties between AUL and
AML with minimal differentiation

115. True AUL is very rare, and these neoplasms :
• Should not express markers of B, T, or myeloid
lineage
• Should not fulfil criteria for B- or T- ALL , AML with
minimal differentiation, or MPAL
• Nevertheless, they can be positive for:
CD 34
CD 38
HLA-DR
TdT

116. • There may be cases that cannot be adequately
classified due to insufficient immunophenotyping
data or discordant expression of various markers,
rendering definitive classification impossible
• The latter cases should be designated as acute
unclassifiable leukemia, which is different from
AUL

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119. Thank you !