1. Healthcare

Cytogenetic Profile of Minimally Differentiated (FAB MO) Acute Myeloid
Leukemia: Correlation with Clinicobiologic Findings
By Antonio Cuneo, Augustin Ferrant, Jean Louis Michaux, Marc Boogaerts, Hilde Demuynck,
Angeline Van Orshoven, Arnold Criel, Michel Stul, Paola Dal Cin, J e s u s Hernandez, Bernard Chatelain,
Chantal Doyen, Andries Louwagie, Gianluigi Castoldi, Jean-Jacques Cassiman, and Herman Van Den Berghe
Cytogenetic data were studied in 26 patients with de novo
acute myeloid leukemia (AML) with minimal myeloid differentiation, corresponding to the MO subtype of the FrenchAmerican-British classification, in correlation with cytoimmunologic and clinical findings. Clonal abnormalities were
detected in 21 cases (80.7%). 12 of which had a complex
karyotype.Partial or total monosomy 5q and/or7q was
found, either as the sole aberration or in all abnormal metaphases, in 11 patients; in 8 cases, additional chromosome
changes were present, including rearrangements involving
12~12-13 and2p12-15 seen in 3 cases each. Five patients
had trisomy 13 as a possible primary chromosome change;
in 5 cases, nonrecurrent chromsome abnormalitieswere obwith chromosome
served. Comparison ofthesefindings
data from 42 patients with AML-M1 shows that abnormal
karyotypes, complex karyotypes, unbalanced chromosome
changes (-5/5q- and/or -7/7q- and
+l31 were observed
much morefrequently in AML-MO than inAML-M1. Patients
with abnormalities of chromosome 5 and/or 7 frequently
showed trilineage myelodysplasia and low white bloodcell
count. Despite their relatively young age, complete remission was achieved in 4 of 11patients only. Patients with 13
were elderly males with frequent professional exposure t o
myelotoxic agents. Unlike patients with clonal abnormalities, most AML-MO patients with normalkaryotype showed
1% t o 2% peroxidase-positive blast cells at lightmicroscopy
and frequently achieved CR. It is concluded that (1)AML-MO
shows a distinct cytogenetic profile, partially recalling that
of therapy-related AML, (2) different cytogenetic groups of
AML-MO can be idenitified showingcharacteristic clinicobiologic features, and (3)chromosome rearrangements may
partially account for theunfavorable outcome frequently observed in these patients.
0 1995 by The American Society of Hematology.
M
or CD13 myeloid antigens or ultrastructural MPO) as well
as negativecriteria(ie,negativity
for lymphoid antigens)
were put forward to avoid confusing this leukemia with leukemia with stem cell phenotype,' with lymphoblastic leukemias, and with biphenotypic leukemias.'
Little is known about the clinical and biologic significance
of thisnewlyidentifiedsubset
of AML; however,a low
complete remission (CR) rate has recentlybeendescribed
in 15 patientsclassifiedaccording
to the FAB proposals,
suggesting that a poor prognosis may be associated with this
leukemia, possibly because of the convergence of unfavorable cytogenetic and immunologic features.'
In view of the well-established importance of cytogenetic
findings in acuteleukemias,' we analyzedthe cytogenetic
profile in 26 cases of de novo AML-MO, in correlation with
cytologic, immunologic, and clinical features.
INIMALLY DIFFERENTIATED acute myeloid leukemia(AML)was recognizedasadistinctentity
asearly as 1987 by Leeetal,'whodescribedcytologic,
immunologic, and clinical features in 10 patients with morphologically undifferentiated leukemia by light microscopy
and positivity for theultrastructural myeloperoxidase (MPO)
as well as for myeloid antigens.
Later on, a number of studies confirmed that a significant
fraction of leukemias otherwise classified as "undifferentiated" can be shown to be myeloid in nature, when applying
sensitive immunologic and electron microscopy techniques?4
However, because diagnostic criteria in these series were
not uniform, the French-American-British (FAB) Cooperative group recently proposed guidelines for the recognition
of this form of leukemia, now referred to as AML-MO.' The
3% upper cutoff for
M P 0 positivity was setfor its distinction
from AML-M 1, and positive (ie, expression of CD33 and/
+
PATIENTS AND METHODS
From the Institute of Haemutolog>), University of Ferram, Ferrara,Italy; the Department of Haematology,CliniquesUniversitaires St-Luc, Brussels; the Departments of Haematology and ClinicalBiologyand
the Center for Human Genetics,University of
Leuven,Leuven; the Department of Haematology, A.Z. St. Jan,
Brugge; and the Department of Haematology, Cliniques Univer.sitaires de Mont-Godinne, Mont Godinne, Belgium.
Submitted December 7, 1994; accepted Februaty 2, 1995.
Supported in part by MURST fondi (40%)and by Fondi regionali
(60%).
This text presents research results of the Belgian programme on
InteruniversityPoles of attractioninitiatedby the BelgianState,
Prime Minister's Ofice, Science Policy Programming.The scient@
responsibility is assumed by its authors.
Address reprint requests to H. Van Den Berghe, Centerfor
Human
Genetics, Herestraat 49, B-3000 Leuven, Belgium.
The publication costs of this article were defrayedin part by page
chargepayment.Thisarticle
must thereforebeherebymarked
"advertisement" in accordance with 18 U.S.C. section 1734 solely to
indicate this fact.
0 1995 by The American Socier?, of Hemato1og.y.
0006-4971/95/ss12-0035~3.00/0
3688
Patient population. Major criteria for the diagnosis of AML-MO
adopted in this study are the following: ( I ) less than 3% blast cells
stained by light microsopy M P 0 andsudan black-B (SBB); (2)
reactivity with CD33 andor CD13 myeloid antigens; and (3) negativity for lymphoid antigens (the isolated expression of CD7 or
terminal deoxynucleotydil transferase (TdT) did not preclude the
diagnosis of AML-MO). Two patients without expression of myeloid
and lymphoid antigens were classified as AML-MO basedonthe
presence of 1% to 2% positive blast cells with M P 0 and SBB.''
Forty-one patients with a presumptive diagnosis of de novo AMLMO were selected among approximately 700 newly diagnosed
AMLs, seen at the University Institutes of Hematology in Brussels,
Leuven, Mont-Godinne, Brugge, and in Ferrara since 1984. Fifteen
patients were excluded from this analysis for the following reasons:
( I ) a diagnosis ofAML-M1, of AML-MS,and of biphenotypic
leukemia was thought to be more appropriate at review of cytology
and immunophenotype (7, I , and 2 cases, respectively); and (2)
karyotype not available ( 5 cases).
To compare cytogenetic findings in this cohort of patients and in
a cytologically similar subset of AML, 42 patients with an AMLM1 FAB diagnosis, seen at our institutions during the study period,
were selected for karyotype review. Differences in the distribution
Blood, Vol 85,No 12 (June 15). 1995:pp 3688-3694
CYTOGENETICS OF AML-MO
of clonal abnormalities among different groups were compared using
the x' test.
Morphologic studies. Review of bone marrow (BM) and peripheral blood smears stained by May-Grunwald-Giemsa and by cytochemical reactions including MPO, SBB, and alpha-naphthyl acetate
esterase with and without fluoride inhibition, periodic-acid Schiff
(PAS), and acid phosphatase (AcP)" was performed, and the patients
were classified according to the FAB riter ria.^." Attention was devoted to the presence of dysplastic features of BM cells.13 When all
three lineages were affected, AML with trilineage myelodysplasia
(MDS) was diagnosed as previously reported.I4
Immunologic studies. Cytofluorimetric analysis of the phenotype
ofBM and/or peripheral blood cells was performed as previously
de~cribed,'~
gating primarily on the blast cell population. To minimize nonspecific Fc-receptor binding, all samples were preincubated
with2.5% human AB serum. Nonspecific isotypic mousemonoclonal antibodies (MoAbs) served as negative control for the primary
agent.
The expression of following antigens was tested using commercially available reagents (Ortho Diagnostics, Raritan, NJ;Becton
Dickinson, Mountain View, CA) (1) myeloid, erythroid, and platelet
antigens: CD33, CD13, CD15, CDllb, CD14, glycophorine, CD41,
and/or CD61; and (2) lymphoid antigens: CD7, CD2, CD3, CD5,
CD22, CDlO, and CD19. The expression of the CD34 stem cell
marker was also tested; reactivity for the 17F11 MoAb, recognizing
an epitope of the c-kit protein product (CD1 17)," was assayed since
1993. A polyclonal antiserum was used for TdT assay (Gibco BRL,
Gaithersburg, MD).
A sample was considered positive when 20% ofblast cells showed
fluorescence above control. However, in accordwith previous report~,"~
the
' ~ 10% cutoff was thought to be more appropriate for the
MoAb detecting the c-kit protein product.
Cytogenetic and molecular genetic studies. Cytogenetic analysis
was performed at diagnosis in all patients by a synchronization
technique with methotrexate and bromodeoxyuridine or thymidine.
Metaphases were either R-banded or G-banded. Chromosome aberrations were described according to the International System for
Human Cytogenetic Nornenclat~re.'~
Complex karyotypes were defined by the presence of three or more events of translocation and
nondisjunction in the same clonez0or by the presence of multiple
unrelated clones. The configuration oftheIgand
T-cell receptor
(TCR) genes were analyzed at diagnosis in 15 patients for whom
representative frozen samples were available.
Methods have been detailed elsewhere." After DNA extraction
by standard techniques, digestion with Bgl 11, BamHI, EcoRI, HindIII, and Kpn I, respectively, was performed. DNA fragments were
size-fractionated on 0.7% agarose gels and blotted onto Hybond N+
filters. Hybridization to probes "P-labeled by primer extension was
performed. Ig gene rearrangement was assayed using a heavy chain
joining (JH) region probe (a 3.8-kb BamHI-Hind111 fragment) and
two constant region probes, CK and CA (2.7- and 0.8-kb EcoRI
fragments; from Dr R. Dalla Favera, New York University Medical
Center, New York, NY). TCR genes were analyzed for the 6,y ,
and p chains, respectively. The TCR-6 rearrangement was studied
withthe J6 S16 probe (a 1.5-kb Sac I fragment; from Dr T.H.
Rabbitts, Laboratory of Molecular Biology, MRC, Cambridge, UK).
The TCR-7 gene configuration was investigated withthe Jy 1.3
probe (a 0.8-kb EcoRI-Hind111 fragment; from Dr J. Bolhuis, Dr D.
den Hoed Cancer Centre, Rotterdam, The Netherlands). The TCRp gene was examined with a constant region cDNA probe (a 0.4kb Bgl I1 fragment; from Dr T.W. Mak, Ontario Cancer Institute,
Toronto, Canada).
Clinical dafa. Clinical records were reviewed with particular
reference to the profession, hematologic data at presentation, outcome of induction therapy, presence of an MDS phase after achievement of CR, and cytologic features at relapse. CR was defined by
3689
Table 1. Immunologic Findings in 26 Patients With AML-MO
Immunologic Marker
CD117
CD34
CD33
CD13
CD15
CD1 1b
CD7
TdT
No.
PositiveINo.Tested
316
22/22
21/26
22/25
8/16
11/18
14/26
7/26
% Positive Cells
Median Value (Range)
22 (12-51)
70 (21-83)
62 (25-80)
60 (20-87)
35 (20-52)
54 (22-72)
61 (25-84)
43 (25-75)
the presence of less than 5% BM blast cells with more than 1.5 X
10'L neutrophils and more than 100 X 109Lplatelets and hemoglobin greater than 10 g/dL.
RESULTS
Morphologyandcytochemistry.
All patients classified
as AML-MO showed little or no maturation along the granulocytic lineage. The overall morphologic picture was that of
undifferentiated leukemia, with small- to medium-sized
cells, round nuclei, and open chromatin. One or more
nucleoli were evident, and scanty, moderately basophilic cytoplasm was observed in the majority of cases, whereas heterogeneity of cell size was observed in some cases.
In 9 cases, 1% to 2% positive cells for the MP0 and SBB
stain were detected, whereas only occasional MPO' cells
were observed in the remaining 17 cases. Weak diffuse positivity for the nonspecific esterase stain that was not inhibited
by sodium fluoride was observed in 15 cases; PAS blockpositivity or strong, localized positivity for the AcP was not
observed inany patient, whereas small granular positivity
for the PASstainwas
detected in 10 cases. A moderate
increase (2% to 5% of total cellularity) of morphologically
normal eosinophils in late stage of differentiation was noted
in 4 cases (no. 13, 15, 16, and 26).
Because of overwhelming blast cell infiltrate of the BM,
morphologic features of the residual nonblast cell population
could not be assessed in 10 patients. Dysplastic features of
the nonblast cell population fulfilling criteria for the definition of trilineage MDS were present in 7 cases (no. 1, 4, 8,
10, 11, 14, and 21), whereas dysplastic features were confined to 1 or 2 cell lineages in 2 and 3 patients, respectively.
Immunophenotype. The immunologic profile of our patients is shown in Table l . Positivity for two or more myeloid-associated antigens was detected in22 cases, in the
absence of coordinate expression of lymphoid-associated antigens; whereas, in 2 cases, only one myeloid marker (either
CD13 or CD33) was found to be positive in more than 20%
of blast cells (cases no. 11 and 15). Two patients with SBB
and M P 0 positivity in 1% to 2% blast cells had a stem cell
phenotype with CD34 and HLA-DR positivity with negative
myeloidand lymphoid markers (patients no. 13 and 25).
No patient expressed the CD14, glycophorine, and CD41
antigens normally detected in leukemias with monocytic,
erythroid, or megakaryoblastic differentiation, respectively.
Cytogenetic and molecular genetic jindings.
Results of
chromosome investigations are detailed in Table 2. Clonal
abnormalities were detected at diagnosis in 21 patients
3690
CUNEO ET AL
Table 2. Karyotype and Clinical Outcome in 26 Cases of AML-MO
Patient No.
Karyotype [No. of Cells1
Age (yr)
1
2
3
4
69
39
66
44
5
6
60
73
7
8
Outcome (Duration of CR)
IgH
TCROIyIp
3
3
23
3
G
G
ND
ND
G/G/G
G/G/G
ND
ND
Survival*
45,XX,-7[61/46,XX,[41
45,XX,del(5)(ql3q31),-7[10]
46,XY,de1(5)(q23q32)~31146,XY[131
46,XX,der(2)t(2;?)(p1?4;?),de1(5)(q12q32),-7,
add(l2)(pl2),+mar~l01
47,XY,de1(7)(q21q31),+mar[lOl
45,X,-X.r(2)(p1?5q3?6),+der(3)(q?),-4,-5,+10,
+11,-13[101/46,XX~61
PR
NR
CR (21)
NR
NR
NR
4
3
G
G
GIGIG
GIGIG
33
46,XY,de1(5)(ql4q32),der(6)t(6;?)(p21;?),-7,
der(12)t(7;12)(qll;p12).+mar[8Il46,XY~21
CF (2) ABMT
8
ND
ND
43
50,XY,de1(2)(p15),-5,add~7)(q22)+8,add(2l~~pll~,
NR
1
G
GIGIG
14
R
G/R/G
8
G
GIGIG
ND
ND
ND
ND
+marl,+mar2,+r,+r[lO]
9
37
45,XY,del(l)(q24q42),de1(5)(q2lq32),-8,der(l2)
CR (9)
t(8;121(q21;p13)[251
37
10
CR (2) BMT
45,XY,de1(5)(ql4q32~.der(7~t~7;17)~q22;ql1~,-17~41~145,idem
dei(9)(ql3q33)[61
43
11
12
13
14
15
16
17
61
72
56
73
72
15
60
18
60
19
20
21
22
23
24
25
26
59
66
62
49
26
66
79
45,XY,-7[11145,idem,der~3)(q?)[91
47,XY,+13~61/46,XY~91
48,XY,+13,+13[61/46,XY[121
47,XY,+13~21/49,XY,idem,+9,+11[61/46,XY[41
47,XY,+13[31/47,XY,idem,-7,+mar[71
47,XX,+13[31/49,XY,+8,+10,+13[121
46,XX,t(6;9~~q12;p23~~41146,XX,t~l;19~~q13;p13~~511
46,XX[61
46,XY,der~16~t~16;?~~p12;?~131/44,idem,-5,-14~611
46,XY[11
NR
NR
ND
CR (7)
PR
NR
CR (3) BMT in II CR
NR
45,XY,add~9~~p12~,-12~41146,XY,idem,+8~31146,XY~71
47,XY,+20~31/46,XY~71
45,XX,-21[41/46,XX[81
46,XX[151
46,XY[201
46,XX[18]
46,XX[ 161
ED
CR
CR
CR
CR
CR
CR
46,XX[20]
NA
8
NA
27
3
10
R
RIRIG
<l
5
37
15+
9
96+
NA
NA
G
ND
ND
RIRIG
G
GIGIG
R
ND
GIGIG
GIGIG
5
CR (5)
(4)
(25)
(3) BMT
(2)
(3) BMT
(duration NA)
40+
9
ND
G
G
ND
G
ND
ND
R
GIGIG
GIGIG
ND
GIGG
ND
ND
ND
Abbreviations: CR. complete remission; PR, partial remission; NR, no response; ED, early death; NA, not available; BMT, BM transplantation;
ABMT, autologous BM transplantation; G,germline; R, rearranged; ND, not done.
* Months; +, indicates the patient is alive.
(80.7%), of which 12 had complex karyotype and 9 had
one or two clonal chromosome changes. Abnormal clones
carrying -515q- andor -717q- were detected in 13 patients; in 8 cases, these abnormalities were associated with
complex karyotypes, including 2p and 12p rearrangements
present in 3 cases each. In 11 cases (no. 1 through 11) -51
5q- andor -717q- were consistently present in all abnormal metaphases, possibly representing the primary chromosome change. Trisomy 13 was present as the sole anomaly
in 2 cases (no. 12 and 13); whereas, in 3 cases, additional
secondary changes were detected (cases no. 14 through 16).
In 5 cases (no. 17 through 21), miscellaneous primary clonal
aberrations were detected. Recurrent clonal abnormalities
in 19of 42 patients with AML-M1 carrying an abnormal
karyotype were -515q- andor -717q- in 4 cases, +8 in 4
cases, 12p abnormalities in 3 cases, and + 11 and 3q21lq26
rearrangements in 2 cases each. Nonrecurring chromosome
changes were observed in the remaining 4 cases.
The observed frequency of recurrent primary chromosome
changes of complex karyotypes and of unbalanced chromosome rearrangements in AML-M0 with respect to AML-M1
is shown in Table 3. Molecular genetic studies showed clonal
rearrangement of the IgH chain gene and of the TCR genes
in 4 of 15 cases and 3 of 15 cases, respectively (see Table
2).In 2 cases (no. 9 and 18), boththeIgHand
the TCR
genes were in a rearranged configuration. There was no rearrangement of the TCR-0 gene in all cases examined.
Clinical features. Hematologic findings at presentation
and correlation of chromosome findings and clinicobiologic
features in patients with AML-M0 are summarized in Tables
4 and 5. The profession wasknown for 20 patients, 7 of
whom (no. l, 2, 6, 7, 13, 14, and 15; 2 truck drivers, 2
factory painters, and 3 farmers) were considered exposed to
petroleum products, organic solvents, or pesticides. Myeloablative chemotherapy was administered to 24 patients:
whereas 1 patient was treated by low-dose cytarabine, and
l patient died before treatment was started. CR was achieved
in 13 of 24 assessable patients, of whom 4 underwent BM
transplantation (3 allogeneic, 1 autologous) in first CR. Median duration of CR in the remaining patients was 6 months.
One patient (no. 16) was succesfully transplanted in second
CR. Overt relapse was preceded by an MDS phase with
pancytopenia and 5% to 10% BM blasts in 2 patients (no.
3 and 7). Cytologic and immunologic data in 10 patients at
CYTOGENETICS
OF AML-MO
3691
Table 3. Observed Frequency of Distribution of Primary
Chromosome Changes, of Complex Karyotype, and of Unbalanced
Reerrangementsin AML-MO Weh Respect to AML-M1
Karyotype
-5/5q- and/or -717q(+l- additional)
+l3 (+l- additional)
Others
Normal
11/26
5/26
5/26
5/26
Complex
1-2abnormalities
Normal
12/26
9/26
5/26
Unbalanced
chromosome
changes
Balanced chromosome
changes
Normal
AML-M1 (No. of
AML-MO (No.
of Casesflotal)
Casesflotal)
P Value
4/42
<.0003
1 l42
14/42
23/42
3/42
<.0001 16/42
23/42
20121
1/26 ,001
5/26
12/42
7/42
23/42
relapse were consistent with a diagnosis of AML-MO in 5
cases; whereas, the features of AML-M1 were observed in
4 cases, and those of AML-M2, in 1 case. Three transplanted
patients (no. 16, 22, and 24) are alive and free of leukemia
at 15, 40, and 96 months; whereas the remaining patients
died at less than 1 to 37 months, with a median overall
survival of 8 months.
DISCUSSION
Diagnosis of AML-MO. Although the formulation of a
generally accepted system of classification of acute leukemias by the FAB group (5,12) has provided a framework of
reference for the identification of various subtypes of AML,
the unequivocal recognition of AML-MO maystill pose some
problems, with particular reference to the possibility of confusing early lymphoblastic leukemia with inappropriate expression of myeloid antigens.
The diagnosis ofAML-MO in our patients fulfilled the
FAB criteria and was supported by immunophenotyping and
by cytochemical features of the blast cell population. Interestingly, SBB appeared to be more sensitive than M P 0 in
this study, and 7 patients who would have otherwise been
classified as AML-MO because of the presence of less than
3% positivity for M P 0 were included among AML-M1 at
cytologic review, showing 3% to 12% SBB-positive cells.
While a pro-B-lymphoid nature of leukemic cells could
be ruled out by negativity for early markers of B-lymphoid
differentiation, such as CD19 and CD10,” the differential
diagnosis with early T-cell acute lymphoblastic leukemia
was not unequivocal for those patients with CD7 positivity.
This problem may be particularly important, especially for
those patients, such as case no. 9, showing a clonally rearranged pattern for the TCR gene.
In the absence of consensus on the diagnostic importance
that should be attributed to lineage-associated antigens,’ we
felt that a diagnosis of AML-MO was appropriate in these
patients, given the absence of coordinate expression of Tlymphoid features (ie, negativity for the AcP stain, for the
TdT as well as for the CD5, CD2, and CD3 molecules).
It is noteworthy that, although the finding of inappropriate
rearrangement of the IgH chain and TCR6 or y gene is not
surprising in AML,23 especially in those patients with blast
cell immaturity and TdT p o ~ i t i v i t ythe
, ~ ~incidence of such
genetic events in our series is not dissimilar as compared
with previous studies of unselected AML case^.'^.'^
Clinical and cytologic follow-up in this series yields the
following observations confirming the “nonlymphoid nature” of these leukemias: (1) the presence of trilineage MDS
in 7 of 16 evaluable cases; ( 2 )an MDSphase preceding overt
relapse in 2 cases achieving CR; and (3) a more differentiated
myeloid phenotype at relapse in 5 of 10 cases, in the absence
of lineage switch. These features would be unusual in lymphoblastic leukemias because trilineage MDS has been described in approximately 15% de novo AML14 and, more
frequently, in erythroleukemia and megakaryoblastic leukemia~.~~,~~
Immunologic findings in AML-MO document the consistent expression of the CD34 stem cell marker in association
with CD13 and/or CD33 and with other myeloid associated
antigens, such as C D l l b and CD15. However, no patient
was found with isolated expression of CD15 or CD1 lb, thus
confirming the notion that CD 13 and CD33 are to be considered the most sensitive and reliable markers for the immunologic diagnosis of AML with minimal myeloid differentiation,2S.29
Finally, attention should be drawn to the fact that the antiCD1 17 MoAb 17F11, recognizing an epitope of c-kit protein
product, functioning as a receptor for the stem cell factor
(c-kit ligand),I6 was found to be reactive withmorethan
10% leukemic cells in 50% of our cases (3 of 6). The distribution of the CD117 antigen in different FAB subtypes of
AML is still controversial, some investigators having found
100% positive cases in AML-MO and AML-M1 and others
having reported on a 14.9% positivity in children AMLM1.I8 Obviously, more cases need to be studied to clarify
the clinicobiologic significance of CD1 17 positivity in AML.
Cytogenetic proJle of AML-MO: Comparison with unselected AML cases and with AML-MI. A preliminary methodological problem in the definition of the cytogenetic profile ofAML-MO was represented by the selection ofan
appropriate control group serving as reference for comparative analysis. Data for comparison of AML-MO with unse-
Table 4. Clinical Findings in 26 Patients With AML-MO
Age in years
WBC ( ~ 1 0 7 ~ )
Pits ( ~ 1 0 9 1 ~ )
Hb (g/dL)
%BM blasts
%CR
Duration of CR (mo)”
Survival (mo)
60 ( 15-79)
4.9 (1.2-64)
57 (24-550)
8.7 (4.1-12.1)
91 (49-99)
54
6 (2-25)
8 (<1-96+)
Values shown are given as the median with range in parentheses,
and indicates that the patient is alive.
Abbreviations: WBC, white blood cellcount; Plts, platelets; Hb, hemoglobin.
* Excluding patients transplanted in CR.
+
3692
CUNEO ET AL
Table 5. Correlation of Cytogenetic Findings, Cytologic Features, and Clinical Outcome in 26 Patients With AML-MO
Primary Clonal
Abnormality
-5/5q+l3
WBC Count
No. of
Cases
and/or -7Dq-
miscellaneous
normal
11
(15-73)
72 5
(59-66)
60 5
5
Age (yrl’
(XlOS/LI*
(33-73)
44
% BM
Blasts’
3.6 (1.2-29)
79 (49-93)
11.8 (5.7-64) 95 (74-97)
4.6 (1.8-61) 95 (63-97)
6.6 (1.6-16) 01492 (68-99)
49 (26-79)
1%-2% SBB/
M P 0 +/Total
311 1
1l5 515
1 l5 415
415 415
TMDSflotal
CR (YesiTotal)
CD7 or TdT’TTotal
517
1I2
1l3
411 1
215
314
414
811 1
Abbreviation: TMDS, trilineage MDS (assessable in 14 of 26 patients).
* Values shown are given as the median with the range inparentheses.
lected AML cases were derived from the report of the Sixth
higher than the 21% derived from 357 de novo AML cases
International Workshop on Chromosomes in Leukemia
with abnormal karyotype reported at the SIWCL.
(SIWCL).30In addition, we elected to include into this study
Interestingly, a 66% and 72.4% frequency of 5q andor
original cytogenetic data from a cohort of patients affected
7q aberrations was found in twostudies of 39 and 58 therapywith de novo AML without maturation, corresponding to the
related AML (t-AML) cases with abnormal karyotype.”.’3
M1 type of the FAB classification, to be able to compare
These findings show that some cytogenetic features in our
chromosome findings in these cytologically similar forms of
patients with AML-MO recall those typically found in a subleukemia, the distinction of which may not be immediate on
set of t-AML and may suggest that similar leukemogenic
cytologic grounds and may appear somewhat artificial from
mechanism may frequently be operative in these forms of
a biologic point of view.
leukemia.
According to our data, de novo AML-MO stands out as a
The cytogenetic profile of AML-MO shows similar differdisease entity with a higher percentage of abnormal karyoences even when compared with the cytologically closest
types (80.7%) as compared with unselected cases of AML
form of leukemia, namely AML-M1. In addition, analysis
reported at the SIWCL, where upto46% cytogenetically
of our data shows that trisomy 13 and aberrations of2p
normal cases were found. In addition, the type of chromoare more frequently observed in AML-MO, that 3q2Uq26
some changes in AML-MO differs significantly with respect
rearrangements may be confined to AML-M1, and that the
to the general cytogenetic profile of AML, a conclusion reinobserved frequency of primary chromosome changes in
forced by a literature review of previously reported cases
AML-MO differs significantly with respect to AML-M1 . Bewith AML-MO diagnosed according to the FAB proposals
sides abnormalities of5q and 7q, complex karyotypes and
(see Table 6).
unbalanced chromosome changes leading to gain or loss of
None of the chromosome translocations associated with
genetic material, normally regarded as characteristic chrowell-defined cytologic subsets of AML9 was encountered in
mosome changes in t-AML,33appeared more frequently in
this series, nor was any previously unrecognized translocaAML-MO than in AML-M1. In this respect, it is interesting
tion found as a recurrent chromosome change in our patients
to note that a viewis emerging that balanced reciprocal
with AML-MO. Notably, 1lq23 rearrangements, possibly astranslocations and unbalanced chromosome abnormalities
sociated with stem cell involvement in acute l e ~ k e m i a , ’ ~ . ~ ’may contribute differently to malignant transformation, the
were not observed in this series. Chromosome changes, such
former type possibly resulting from exposure to agents taras 5q and 7q abnormalities, normally found in many FAB
geting DNA topoisomerase I1 and the latter type being charsubtypes of AML were detected in 13 of 21 (61.9%) AMLacteristically associated with genetic damage following “in
MO cases with abnormal karyotype, a figure significantly
vivo” and “in vitro” exposure to mutagens, such as alkylat-
Table 6. Cytogenetic and Clinical Findings in AML-MO:
Data From the Literature
No. of
Karyotype
Cases
Median Age in
Years (Range)”
12 (32-59)
50
-5/5q- and/or
(29-70)
62.5
10
-7i7q+8
(54-68)
62 7
+ l 9 (45-56)
4
+l3
3
(61-67)
62
2
61-70
deI(3)
2
42-51
t(12;13)(p13;q14)
Normal
Cr (Yes/
Total)*
Reference
46 417
44, 8, 1 , 4,
016
213
111
213
012
NR
1 , 4, 8, 46
1 , 5,
8, 46
5, 8, 43, 45
8,39,40,43
8
47
Only those cases with clonal aberrations observed in at least
2
patients are shown. del(3p) was associated in both cases with -5 or
-7; + l 9 was associated with + l 3 in 1 case.
Abbreviation: NR, not reported.
* Data not available for all cases.
ing
Finally,isit noteworthy that aberrations of
5q and 7q and deletions or translocations of 12p are also
very common in e r y t h r ~ l e u k e m i a , ~which
~ ~ ~ ‘is~usually
~~
regarded as a stem cell disorder with multiple cell-lineage
involvement. In the absence of reliable markers of early
erythroid differentiation, the theoretical possibility should be
considered that some leukemias fulfilling the FAB criteria
for AML-MO may in fact represent proliferations of immature erythroid precursors.
Correlation of chromosome jindings with clinicobiologic
features. Further insights into the significance of chromosome findings in AML-MO maybe derived from the observation that, in this study, 11 cases had -5/5q- andor -7/7qas a possible primary chromosome change, 5 cases had + 13,
5 patients had different nonrecurring primary chromosome
aberrations, and 5 patients had a normal karyotype.
As shown in Table 5, analysis of clinicobiologic findings
in these cytogenetic groups ofAML-MO, yields important
3693
CYTOGENETICS OF AML-MO
observations, some of which deserve particular attention.
Those patients carrying aberrations of the long arms of chromosome 5 and/or 7 share with t-AML the primary chromosome change, the frequent presence of a complex karyotype,
and, possibly, some additional aberrations such as rearrangements involving l 2 ~ . ~
In*addition, they may frequently show abnormalities of the short arm of chromosome
2, the presence of which was only detected sporadically in
other forms of de novo or t-AML.39Interestingly, a possible
exposure to myelotoxic agents in the workplace could be
documented in 4 cases.
Other hematologic features in these patients, such as the
presence of trilineage MDS in frequent association with a
relatively low white blood cell count and low percentage of
BM blasts also recaIl the clinicobiologic picture commonly
observed in t-AML.40Most importantly, although patients in
this cytogenetic subset of AML-MO were generally young,
they infrequently achieved CR under conventional myeloablative chemotherapy.
Trisomy 13 may identify a group of AML-MO preferentially affecting elderly males with frequent professional exposure to myelotoxic agents. A moderate increase ofBM
eosinophils, an unusual finding in other patients with AMLMO, was observed in 3 of 5 cases, whereas the presence of
trilineage MDS could only be assessed in 2 patients because
of almost complete BM replacement byblast cells in the
remaining 3 cases. All cases expressed either TdT or CD7
(3 cases and 2 cases, respectively).
Although + l 3 has been reported in a wide spectrum of
myeloid neoplasias;’ our data seem to support the existence
of a strong association between this numerical aberration
and AML-MO, because + 13 has only been found in 10 patients with other FAB subtypes of AML seen at our institutions during the study period. An association of + 13 with
cell immaturity in acute leukemia was previously described
by Sreekantaiah et al,42whereas a more heterogeneous cytologic picture was described by Dohner et a14’ in 8 cases
collected in a multicenter study of 621 patients. It is noteworthy that 3 of 36 AML-MO collected in a literature review
(see Table 6) had trisomy 13 as the primary change.
Little is known about the prognostic implication of + l 3
in AML, although a relatively low CR rate was noted at
review of 21 published case^.^' However, this finding must
be weighed against the advanced age of most patients with
+13. Not unexpectedly, 2 patients with + l 3 achieving CR
in this series were less than 60.
Finally, unlike most karyotypically abnormal AML-MO,
those patients with normal karyotype frequently showed positivity for “myeloid” cytochemistry in 1% to 2% cells and
achieved CR under conventional chemotherapy. As shown
in Table 3, a normal karyotype is more frequently encountered in AML-M1, thus suggesting that those cases of AMLMO with few MPO+ blast cells may not be dissimilar, on
cytogenetic grounds, from AML-MI, the distinction of these
FAB subtypes in such cases being only represented by the
arbitrary 3% cutoff for SBBMPO positivity. In conclusion,
these findings confirm that the recognition of AML-MO is
essential for a complete cytologic classification of AML.
These findings also seem to indicate that the identification
of AML-MO as a distinct entity may be justified on cytoge-
netic and clinicobiologic grounds and indicate that chromosome changings may partially account for the unfavorable
outcome usually associated with this leukemia.
REFERENCES
1. Lee W, Pollak A, Leavitt RD, Testa JR, Schiffer CA: Mini-
mally differentiated acute nonlymphocytic leukemia: A distinct entity. Blood 70:1400, 1987
2. Matutes E, Pombo de Oliveira M, Foroni L, Morilla R, Catovsky D: The role of ultrastructural cytochemistry and monoclonal
antibodies in clarifying the nature of undifferentiated cells in acute
leukaemia. Br J Haematol 69:205, 1988
3. Campos L, Guyotat D, Archimbaud E, Devaux Y, Treille D,
Larese A, Maupas J, Gentilhomme 0, Ehrsam A, Fiere D: Surface
marker expression in adult acute myeloid leukaemia: correlations
with initial characteristics, morphology and response to therapy. Br
J Haematol 72:161, 1989
4. LeMaistre A, Childs CA, Hirsch-Ginsberg C, Reuben J, Cork
A, Trujillo JM, Andersson B, McCredie KB, Freireich E, Stass SA:
Heterogeneity in acute undifferentiated leukemia. HematolPathol
2:79, 1988
5. Bennett JM, Catovsky D, Daniel MT. Flandrin G, Galton DAG,
Gralnick HR, Sultan C: Proposals for the recognition of minimally
differentiated acute myeloid leukaemia. Br J Haematol 78:325, 1991
6. Brito-Babapulle F, Pullon H, Layton DM, Etches A, Huxtable
A, Mangi M, Bellingham AJ, Mufti GJ: Clinicopathological features
of acute undifferentiated leukaemia with a stem cell phenotype. Br
J Haematol 76:210, 1990
7. Catovsky D, Matutes E, Buccheri V, Shetty V, Hanslip J,
Yoshida N, Morilla R: A classification of acute leukaemia for the
1990s. Ann Hematol 62:16, 1991
8. Stasi R, Del Poeta G, Venditti A, Masi M, Stipa E, Dentamaro
T, Cox C, Dallapiccola B, Papa G: Analysis of treatment failure in
patients with minimally differentiated acute myeloid leukemia
(AML-MO). Blood 8311619, 1994
9. Le Beau MM,Rowley JD: Cytogenetics, in Williams WJ, Beutler E, Erslev AJ, Lichtman MA (eds): Hematology (ed 4). New
York, NY, McGraw-Hill, 1990, p 78
IO. Seshi B, Kashyap A, Bennett JM: Acute myeloid leukemia
withunusual phenotype: Myeloperoxidase (+), CD13 (-), CD14
(-) and CD33 (-). Br J Haematol 81:374, 1992
11. Shibata A, Bennett JM, Castoldi CL. Catovsky D, Flandrin
G, Jaffe ES, Katayama I, Nanba K, Schmalz1 F, Yam LT, and
The International Committee for Standardization in Haematology
(ICSH): Recommended methods for cytologic procedures in hematology. Clin Lab Haematol 7:55, 1985
12. Bennett JM, Catovsky D, Daniel MT. Flandrin G, Galton
DAG, Gralnick HR, Sultan C: Proposed revised criteria for tha classification of acute myeloid leukemia: A report of the French-Amencan-British Cooperative group. Ann Intern Med 103:626, 1985
13. Bennett JM, Catovsky D, Daniel MT, Flandrin G, Galton
DAG, Gralnick HR, Sultan C, andthe French-American-British
(FAB) Co-operative group: Proposals for the classification ofthe
myelodysplastic syndromes. Br J Haematol 51:189, 1982
14. Brito-Babapulle F, Catovsky D, Galton DAG: Clinical and
laboratory features of de novo acute myeloid leukemia with trilineage myelodysplasia. Br J Haematol 66:445, 1987
15. Cuneo A, Michaux JL, Ferrant A, Van Hove L, Bosly A, Stul
M, Dal Cin P, Vandenberghe E, Cassiman JJ, Negrini M, Piva N,
Castoldi GL, Van Den Berghe H: Correlation of cytogenetic patterns
and clinicobiological features in adult acute myeloid leukemia expressing lymphoid markers. Blood 79:720, 1992
16. Lerner NB, Nocka KH, Cole SR, QiuF, Strife A, Ashman
LK, Besmer P: Monoclonal antibody YB5.B8 identifies the human
c-kit protein product. Blood 77: 1876, 1991
3694
17. Reuss-Borst MA, Buhring HJ, Schmidt H, Muller CA: AML:
Immunophenotypic heterogeneity and prognostic significance of ckit expression. Leukemia 8:258, 1994
18. Smith FO, Broudy VC, Zsebo KM, Lampkin BC, Buckley
CV, Buckley JD,OpieT, Woods WG, Denmann Hammond G,
Bernstein ID: Cell surface expression of c-kit receptors by childhood
acute myeloid leukemia blasts is not of prognostic value: A report
from the Childrens Cancer Group. Blood 842347, 1994
19. Mitelman F (ed): ISCN: Guidelines for Cancer Cytogenetics.
Supplement to an International System for Human Cytogenetic Nomenclature. Basel, Switzerland, Karger, 1991
20. Sandberg AA: Cytogenetic features of special hematologic
disorders, including some aspects of acute leukemia, in Sandberg
AA (ed): The Chromosomes in Human Cancer and Leukemia (ed
2). New York, NY, Elsevier, 1990, p 374
21. Kerim S, Stul M, Mecucci C, Vandenberghe E, Cuneo A,
Dal Cin P, Michaux JL, Louwagie A, Cassiman JJ, Van Den Berghe
H: Rearrangements of immunoglobulin and TCR genes in lymphoid
blast crisis of Ph+ chronic myeloid leukemia. Br J Haematol74:414,
1990
22. Foon KA, Todd RF 111: Immunologic classification of leukemia and lymphoma. Blood 68: I , 1986
23. Gale RP, Ben-Bassat I: Hybrid acute leukemia. Br J Haematol
65:261, 1987
24. Foa R, Casorati G, Guibellino MC, Basso G, Schiro K, Pizzolo G, Lauria F, Lefranc MP, Rabbitts TH, Migone N: Rearrangements of immunoglobulin and T-cell receptor beta and
gamma genes are associated with terminal deoxynucleotidyl transferase expression in acute myeloid leukemia. J Exp Med 165:879,
1987
25. Oster W, Konig K, Ludwig WD, Ganser A, Lindemann A,
Mertelsmann R, Hemnann F: Incidence of lineage promiscuity in
acute myeloblastic leukemia: Diagnostic implications of immunoglobulin and T-cell receptor gene rearrangement analysis and immunological phenotyping. Leuk Res 125387, 1988
26. Knowles DM, Dalla Favera R, Pelicci PG: T-cell receptor
beta-chain gene rearrangements. Lancet 2:159, 1985
27. Cuneo A, Van Orshoven A, Michaux JL, Boogaerts M, Louwagie A, Doyen C, Dal Cin P, Fagioli F, Castoldi GL, Van Den
Berghe H: Morphologic, immunologic and cytogenetic studies in
erythroleukaemia: Evidence for multilineage involvement and identification of two distinct cytogenetic-clinicopathological types. Br J
Haematol 75:346, 1990
28. Cuneo A, Mecucci C, Kerim S, Vandenberghe E, Dal Cin P,
Van Orshoven A, Rodhain J, Bosly A, Michaux JL, Martiat P.
Boogaerts M, Carli MG, Castoldi GL, Van Den Berghe H: Multipotent stem cell involvement in megakaryoblastic leukemia: Cytological and cytogenetic evidence in 15 patients. Blood 74:1781, 1989
29. Van’t Veer MB: The diagnosis of acute leukemia with undifferentiated or minimally differentiated blasts. Ann Hematol 64: 161,
1992
30. Arthur DC, Berger R, Golomb HM, Swansbury GJ, Reeves
BR, Alimena G, Van den Berghe H, Bloomfield CD, de la Chapelle
A, Dewald GW, Garson OM, Hagemeijer A, Kaneko Y, Mitelman
F, Pierre RV, Ruutu T, Sakurai M, Lawler SD, Rowley JD from the
Sixth International Workshop on Chromosomes in Leukemia: The
clinical significance of karyotype in acute myelogenous leukemia.
Cancer Genet Cytogenet 40:203, 1989
3 l , Castoldi GL, Cuneo A, Tomasi P: Phenotype-related chromosome aberrations and stem cell involvement in acute myeloid leukemia. Haematologica 74525, 1989
32. Iurlo A, Mecucci C, Van Onhoven A, Michaux JL, Boogaerts
CUNEO ETAL
M, Noens L, Body A, Louwagie A, Van Den Berghe H: Cytogenetic
and clinical investigations in 76 cases with therapy-related leukemia
and myelodysplastic syndrome. Cancer Genet Cytogenet 43:227, 1989
33. Pedersen-Bjergaard J, Philip P, Olesen Larsen S, Jensen G,
Byrsting K: Chromosome aberrations and prognostic factors in therapy-related myelodysplasia and acute nonlymphocytic leukemia.
Blood 76:1083, 1990
34. Pedersen-Bjergaard J, Rowley JD: The balanced and the unbalanced chromosome aberrations of acute myeloid leukemia may
develop in different ways and may contribute differently to malignant transformation. Blood 83:2780, 1994
35. Shulman LN: The biology of alkylating-agent cellular injury.
Hematol Oncol Clin North Am 7:325, 1993
36. Olopade 01, Thangavelu M, Larson RA, Mick R, KowalVern A, Schumacher HK, Le Beau MM, Vardiman JW, Rowley JD:
Clinical, morphologic and cytogenetic characteristics of 26 patients
with acute erythroblastic leukemia. Blood 80:2873, 1992
37. Kowal-Vem A, Cotelingam J, Schumacher HR: The prognostic significance of proerythroblasts in acute erythroleukemia. Am J
Clin Pathol 98:32, 1992
38. United Kingdom Cancer Cytogenetic Group (UKCCG):
Translocations involving 12p in acute myeloid leukemia: Associationwith prior myelodysplasia and exposure to mutagenic agents.
Genes Chromosom Cancer S:252, 1992
39. Dewald GW, Schad CR, Lilla VC, Jalal SM: Frequency and
photographs of HGMl1 chromosome anomalies in bone marrow
samples from 3966 patients with malignant hematologic neoplasms.
Cancer Genet Cytogenet 68:60, 1993
40. Le Beau MM, Albain KS, Larson R A , Vardiman JW, Davis
EM, Blough RR, Golomb HM, Rowley JD: Clinical and cytogenetic
correlations in 63 patients with therapy-related myelodysplastic syndromes and acute nonlymphocytic leukemia: Further evidence for
characteristic abnormalities of chromosomes number S and 7. Clin
Oncol 4:325, 1986
41. Baer MR, Bloomfield CD: Trisomy 13 in acute leukemia.
Leuk Lymphoma 7.1, 1992
42. Sreekantaiah C, Baer MR. Morgan S, Isaacs ID, Miller KB,
Sandberg AA: Trisomy/tetrasomy 13 in seven cases of acute leukemia. Leukemia 4:78 1, 1990
43. Dohner H, Arthur DC, Ball ED, Sobol RE, Davey FR, Lawrence D, Gordon L, Patil SR, Surana RB, Testa JR, Verma RS.
Schiffer CA, Wurster-Hill D, Bloomfield CD: Trisomy 13: A new
recurring chromosome abnormality in acute leukemia. Blood
76:1614, 1990
44. Yokose N, Ogata K, Ito T, Miyake K,An E, Inokuchi K,
Tamada T, Gomi S, Tanabe Y, Ohki I, Kuwabwd T, Hasegawa
S, Shinohara T, DanK, Nomura T: Chemotherapy for minimally
differentiated acute myeloid leukemia (AML-MO). Ann Hematol
66:67, 1993
45. Johansson B, Billstrom R, Mauritzson N. Mitelman F: Trisomy 19 as the sole chromosomal anomaly in hematologic neoplasms: Cancer Genet Cytogenet 74:62, 1994
46. Venditti A, Del Poeta G, Stasi R, Masi M, Bruno A, Buccisano F. Cox C, Coppetelli U, Aronica G, Simone MD, Papa G,
Tribalto S, Amadori S: Minimally differentiated acute myeloid leukemia (AML-MO): Cytochemical, immunophenotypic and cytogenetic analysis of 19 cases. Br J Haematol 88:784, 1994
47. Tosi S, Stilgenbauer S, Giudici G, Capalbo S , Specchia G ,
Lis0 V, Castagna S, Ianzi E, Lichter P, Biondi A: Reciprocal translocation t(12; 13)(p13;q14) in acute nonlymphoblastic leukemia: Report and cytogenetic analysis of two cases. Cancer Genet Cytogenet
77: 106, I994