eCSI case Fall 2014 Andrea M. Sheehan, MD Associate Professor
of Pathology & Immunology Baylor College of Medicine
Slide 2
Clinical History 17 year old boy with a history of B-acute
lymphoblastic leukemia (B-ALL) Expressed CD45(dim), CD19, CD22,
CD38, CD58, CD52, CD34, CD99, and HLA-DR 46,XY,t(4;11)(q21;q23)
Treated according to AALL1131, and did well One year later,
relapses with B-ALL with same immunophenotype Underwent reinduction
chemotherapy
Slide 3
Clinical History Now, three months later, the patient presents
with neutropenic fever CBC shows ParameterResultUnitReference Range
WBC0.05K/ul4.5-13.5 Hgb9.6g/dL13-16 Hct27.7%37-49
Platelet12K/ul150-450 Differential: Lymphocytes54.5%15-55
Monocytes9.1%0-4 Blasts36.4%0 ANC0.0K/ul1.8-80
Slide 4
Clinical History A bone marrow was performed and submitted for
morphology, flow cytometry, and cytogenetics Flow cytometry was
performed using a 6 color leukemia panel on a BD FACSCanto with two
lasers and acquired using BD FACSDiva software Data was analyzed
using FCS Express V4
Slide 5
Bone Marrow Aspirate 40x
Slide 6
Bone Marrow Biopsy 20x
Slide 7
6 Color Panel
Slide 8
Flow Cytometry Large CD45 dim blast population with increased
side scatter Also note smaller CD45 dim population with lower side
scatter
Slide 9
T-cells (Tube 1) Dim CD4 expression Dim CD4 expression is
present on the larger myeloid blast population, but other T cell
markers are negative (CD3, CD2, CD7, CD8). A small population of
normal T cells is present in the background Normal background T
cells
Slide 10
B-cells (Tube 2) The larger myeloid blast population is
negative for B cell markers, but back gating on the CD19+
population shows that smaller CD45 dim/low side scatter population
are immature B cells that are CD10 negative and negative for
surface light chains. Normal T cells are in blue. CD19+ back gate B
cells are CD10 negative
Slide 11
B-cells (Tube 7) CD19+ back gate shows that the B cell
population is composed of aberrant immature B lymphoblasts with
strong expression of CD34 and overexpression of CD58. CD22 is also
positive. The larger myeloid blast population is also positive for
CD58 but negative for B cell markers. Aberrant immature B cells Two
distinct blast populations are present! Those are not just
hematogones!
Slide 12
Myeloid Tube (Tube 3) The larger myeloid blast population is
CD15+, CD38+ with a small subpopulation having CD11b. See that the
smaller B lymphoblast population is CD15 positive (back gate on
CD34) CD42+61 positivity likely represents platelets stuck to the
larger blasts rather than true expression.
Slide 13
Monocytic Tube (Tube 4) Both blast populations are negative for
CD13 and CD117. The larger myeloid blast population shows dim
expression of CD64 with essentially no CD14 or CD16+56. Dim CD64
expression Normal T cells are CD64 negative
Slide 14
B/erythroid (Tube 6) The smaller B lymphoblast population
expresses CD22 and HLA-DR but not CD20. (back gate based on CD22 vs
CD45) There is no increase in CD71. The larger myeloid blast
population expresses heterogenous HLA-DR and dim CD71 but no B cell
markers CD22 back gate Dim CD71 expression In these plots, we
lowered the resolution, making the dots larger. This helps make the
smaller lymphoblast population easier to see
Slide 15
Tube 5 The larger myeloid blast population expresses CD58 and
heterogenous CD33. The smaller B lymphoblast population
overexpresses CD58 but is largely CD33 negative, which is easier to
see on the lymphoblast gate based on CD45 dim/low SSC (no B lineage
specific marker is present in this tube). All events CD45 dim/low
SSC gate
Slide 16
Immunophenotypic Findings Two distinct blast populations #1 97%
- myeloid with monocytic differentiation Positive for CD45
(moderate), increased side scatter, CD33 (heterogenous), CD15,
CD11b (dim, subpopulation), CD64 (dim), CD14 (small subpopulation),
HLA-DR (heterogenous), CD99, CD58, CD38, CD71, CD4 (dim) #2 3% - B
lymphoblast CD45 (moderate), low side scatter, CD19, CD22, CD34,
HLA-DR, CD99 (dim), CD58, CD38, CD15
Slide 17
Differential Diagnosis Therapy related myeloid neoplasm with
small clone of persistent B-ALL Lineage switch from a B-lymphoblast
clone to a myeloid clone related to the patients MLL
rearrangement
Slide 18
Cytogenetics
56-63,XY,+X,+Y,+2,+3,t(4;11)(q21;q23),der(4)(t(4;11),
+der(4)t(4;11),+6,+7,+7,del(7)(q22q36)+8,
der(9)t(9;11)(q34;q13)t(4;11)(der(11)t(9;11),+12,+13,+13,+13,+14,
+17,del(17)(p11.1),+18,+19,+21,+22,+mar[cp20] FISH MLL probe -
pattern consistent with MLL gene rearrangement in 200 out of 200
(100%) cells studied, 162 showed additional copy of 3MLL gene and 5
cells showed 2 additional copies of the 3MLL gene Complex karyotype
with presence of t(4;11) clone plus multiple new aberrations
including a secondary translocation with 9q34 and 11q13
Slide 19
Diagnosis Myeloid component represents a lineage switch from
original clone of B-acute lymphoblastic leukemia Bottom line: mixed
phenotype acute leukemia with t(4;11), MLL rearranged, with two
distinct clones present B lymphoblastic (3%) Myeloid (monocytic)
(97%)
Slide 20
Lineage Plasticity Cross lineage expression of antigens is
common in acute leukemias Lymphoid antigen expression in AML CD19
in t(8;21)+ AML CD4 in monoblastic leukemias CD2, CD7 in various
subtypes of AML Myeloid expression in ALL CD13, CD33 in B-ALL CD117
in T-ALL At genetic level, clonal TCR or IgH rearrangements can be
seen in B- ALL, T-ALL, and AML True mixed phenotype acute leukemias
are rare Most common genetic associations t(9;22) or MLL
rearrangements B/myeloid most common phenotype, but T/myeloid, B/T,
or B/T/myeloid can be seen Usually either one blast population with
mixed markers (biphenotypic) or two distinct blast populations
(bilineal)
Slide 21
Lineage Switch in Acute Leukemias True lineage switch is a rare
event Mixed lineage leukemia gene (MLL) rearrangements most common
genetic abnormality in these cases Variety of translocation
partners noted t(4;11), t(9;11), t(11;19), ins(11;4), among others
Also can be seen with t(9;22) BCR-ABL1, especially in setting of
blast crisis of chronic myeloid leukemia Relatively more common in
infantile acute lymphoblastic leukemia MLL rearrangements are seen
in 70-80% of these leukemias May occur early in treatment course
with emergence of aberrant monocytic clones Lineage switch most
commonly B to myeloid Monocytic differentiation is most common Rare
cases of T to myeloid or myeloid to B have been reported Very poor
prognosis when this happens
Slide 22
Lineage Switch in Acute Leukemias Theories as to how lineage
switch occurs Chemotherapy induced selection of a minor subclone
present at diagnosis but not detected Leukemia arises from a
precursor or stem cell with lymphoid and myeloid potential and
changes phenotype under treatment pressure or with the accumulation
of additional genetic abnormalities Gene expression profile studies
of MLL rearranged acute leukemias show patterns in-between ALL and
AML B-macrophage progenitors are present in normal bone marrow
Transdifferentiation from one lineage to another Has been reported
in other settings (follicular lymphoma and follicular dendritic
cell sarcomas)
Slide 23
Follow Up Patient treated on an individualized acute myeloid
leukemia/high risk ALL chemotherapy protocol Patients disease was
refractory to therapy Passed away a few weeks later due to
infectious complications
Slide 24
References Rossi JG et al. Lineage switch in childhood acute
leukemia: an unusual event with a poor outcome. Am J Hematol 2012
87:890-897. Stasik C et al. Infant acute lymphoblastic leukemia
with t(11;16)(q23;p13.3) and lineage switch into acute monoblastic
leukemia Cancer Genetics Cytogenetics 2006 168:146-149. Slamova L
et al. CD2 positive B-cell precursor acute lymphoblastic leukemia
with an early switch to the monocytic lineage Leukemia 2014
28:609-620. Pui C, ed. Childhood Leukemias, 3 rd ed. 2012 Cambridge
University Press.