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CLINICAL OBSERVATIONS, INTERVENTIONS, AND THERAPEUTIC TRIALS

A new morphologic classification system for acute promyelocytic leukemia

distinguishes cases with underlying PLZF/RARA gene rearrangements
Danielle Sainty, Vincenzo Liso, Angelo Cantu`-Rajnoldi, David Head, Marie-Joelle Mozziconacci, Christine Arnoulet, Laurence Benattar,
Susanna Fenu, Marco Mancini, Eliane Duchayne, Francois-Xavier Mahon, Norma Gutierrez, Francoise Birg, Andrea Biondi,
David Grimwade, Marina Lafage-Pochitaloff, Anne Hagemeijer, and Georges Flandrin, on behalf of Groupe Francais dHematologie
Cellulaire, Groupe Francais de Cytogenetique Hematologique, UK Cancer Cytogenetics Group, and BIOMED 1 European
Community-Concerted Action Molecular Cytogenetic Diagnosis in Haematological Malignancies

Acute promyelocytic leukemia (APL) is

typified by the t(15;17) translocation,
which leads to the formation of the PML /
RARA fusion gene and predicts a beneficial response to retinoids. However, approximately 10% of all APL cases lack the
classic t(15;17). This group includes (1)
cases with cryptic PML/RARA gene rearrangements and t(5;17) that leads to the
NPM / RARA fusion gene, which are retinoid-responsive, and (2) cases with t(11;
17)(q23;q21) that are associated with the
PLZF/RARA fusion gene, which are retinoid-resistant. A key issue is how to rapidly distinguish subtypes of APL that demand distinct treatment approaches. To

address this issue, a European workshop

was held in Monza, Italy, during June
1997, and a morphologic, immunophenotypic, cytogenetic, and molecular review
was undertaken in 60 cases of APL lacking t(15;17). This process led to the development of a novel morphologic classification system that takes into account the
major nuclear and cytoplasmic features
of APL. There were no major differences
observed in morphology or immunophenotype between cases with the classic
t(15;17) and those with the cryptic PML /
RARA gene rearrangements. Auer rods
were absent in the t(5;17) case expressing NPM/RARA. Interestingly, this classifi-

cation system distinguished 9 cases with

t(11;17)(q23;q21) and, in addition, successfully identified 2 cases lacking t(11;17), which
were subsequently shown to have underlying PLZF/RARA fusions. The PLZF/RARA
cases were characterized by a predominance of blasts with regular nuclei, an increased number of Pelger-like cells, and by
expression of CD56 in 4 of 6 cases tested.
Use of this classification system, combined
with an analysis for CD56 expression, should
allow early recognition of APL cases requiring tailored molecular investigations. (Blood.
2000;96:1287-1296)
2000 by The American Society of Hematology

Introduction
Acute promyelocytic leukemia (APL) was first recognized for its
particularly poor clinical outcome due to a serious hemorrhagic
syndrome, which is related to disseminated intravascular coagulopathy and abnormal fibrinolysis.1-5 APL is also typified by a unique
differentiation response to retinoids, such as all-trans-retinoic acid
(ATRA), which have transformed clinical practice for this condition.6-9 The initial morphological description of APL emphasized a
specific hypergranular, promyelocyte-like, blast cell component in
the bone marrow. Although the term promyelocytic leukemia has
been widely accepted since 1957,1,2 it is partly misleading because
APL cells are morphologically different from normal promyelocytes. In 1976, the French-American-British (FAB) cooperative
group revised the morphological description of acute myeloid
leukemias (AMLs); the term M3-AML was assigned to hypergranu-

lar promyelocytic leukemia characterized by blast cells with heavy

azurophilic granules, bundles of Auer rods (faggots), and a
reniform or bilobed nucleus.10
Major interest has focused on APL following the identification
of a specific cytogenetic abnormality: t(15;17)(q22;q21) translocation.11 Although the vast majority of M3 cases fit the description of
hypergranular or classical M3, a cytological hypogranular or
microgranular variant form, M3v, has been identified. It is commonly associated with hyperleukocytosis,12 accounts for 15%-20%
of APL cases, and shares the same t(15;17). In M3v-AML, the
majority of blasts have a bilobed, multilobed, or reniform nucleus
and, under usual staining, are devoid of granules or contain only a
few fine azurophilic granules; however, at least a few cells with all
the cytoplasmic features of M3 are present.13 Rare morphological

From the Institut Paoli-Calmettes, Institut National de la Sante et de la Recherche

Medicale (INSERM) U119, IFR 57, and Universite de la Mediterranee, Marseille,
France; Hopital Necker Enfants Malades, Paris, France; Laboratoire dHematologie,
Hopital Purpan, Toulouse, France; Laboratoire dHematologie et dImmunologie,
Hopital Pellegrin, Bordeaux, France; Cattedra di Ematologica i Policlinico di Bari,
Bari, Italy; Dipartimento di Medicina di Laboratorio, Istituti Clinici di Perfezionamento,
Milan, Italy; Dipartimento di Biotecnologie Cellulari ed Ematologia, Universita degli
Studi la Sapienza, Rome, Italy; Centro di Ricerca; M Tettamanti and Ospedale San
Gerardo, Monza, Italy; St Jude Childrens Hospital, Memphis, TN; the Division of
Medical and Molecular Genetics, Guys, Kings and St Thomas School of Medicine,
London, UK; Center for Human Genetics, Leuven, Belgium; and Servicio de
Hematologia, Hospital Universitario de Salamanca, Centro de Investigation del
Cancer, Universidad de Salamanca, Salamanca, Spain.

Supported by grant CT94-1703 from the European Community Biomed

Concerted Action; Molecular Cytogenetic Diagnosis in Haematological
Malignancies by the Comite des Bouches-du-Rhone de la Ligue Nationale
Francaise contre le Cancer; the Leukaemia Research Fund of Great Britain,
UK (D.G.); the Fondazione M. Tettamanti, Associazione Italiana Ricerca sul
Cancro (AIRC), Italy; and MURST (A.B.).
Reprints: Danielle Sainty, Department of Biology, Institut Paoli-Calmettes, 232 bd
Sainte Marguerite, 13009 Marseille, France; e-mail: saintyd@marseille.fnclcc.fr.
The publication costs of this article were defrayed in part by page charge
payment. Therefore, and solely to indicate this fact, this article is hereby
marked advertisement in accordance with 18 U.S.C. section 1734.

Submitted December 9, 1999; accepted April 25, 2000.

A complete list of the contributors appears in the appendix.

BLOOD, 15 AUGUST 2000 VOLUME 96, NUMBER 4

2000 by The American Society of Hematology

1287

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1288

SAINTY et al

forms associated with t(15;17) have been reported. These cases are
characterized by hyperbasophilic microgranular blast cells with
cytoplasmic budding that mimicks micromegakaryocytes14-16 or by
blast cells that exhibit toluidine blue positive basophilic granules17,18 or eosinophilic granules.19 Other cases have been reported
to have M1- or M2-like morphology.15,20
Immunophenotypic studies have shown a distinctive pattern in
APL as compared to other AMLs: positivity for the CD33 and
CD13 myeloid antigens, negativity for HLA-DR, and low frequency of CD34 expression.21-23 Positivity for this latter antigen
and for CD2 and CD19 has been correlated with the M3v form.24,25
Interestingly, rare cases of HLA-DR, CD33 AML, whose
morphology resembles M3v but lacks t(15;17) and expresses the
natural killer (NK)-associated CD56 antigen, have been described.26 Infrequently, however, t(15;17)-associated APL can also
exhibit positivity for this antigen.27
In 1990, t(15;17) was cloned and was found to disrupt a
previously uncharacterized gene, PML, on chromosome 15 and the
gene encoding the retinoic acid receptor (RARA) on chromosome
17. The resultant PML / RARA fusion protein, which retains the
retinoic acid (RA) ligand-binding domain, not only plays a key role
in leukemogenesis, but it is also somewhat paradoxically implicated in mediating the response to retinoids.28 It was originally
thought that all cases of APL were associated with t(15;17),29 but it
is now clear that this chromosomal change is not invariably
present. In some instances, fusion of the PML and RARA genes still
occurs as a result of insertion events or more complex rearrangements,30-31 whereas in others, RARA is fused to a gene other than
PML.28 To date, 3 alternative translocations associated with APL
have been characterized: t(11;17)(q23;q21); t(5;17)(q35;q21); and
t(11;17)(q13;q21). In these translocations, RARA is fused to the
PLZF (promyelocytic leukemia zinc finger),32,33 NPM (nucleophosmin),34-36 and NuMA (nuclear mitotic apparatus)37,38 genes, respectively. Recently, the STAT5b gene located at 17q21 has been
identified as a new RARA partner in a [der(17)] APL case.39
The nature of the fusion partner has a considerable impact on
the characteristics of the disease. PML / RARA-mediated APL is
sensitive to retinoids, which in combination with chemotherapy,
have led to considerable improvements in outcome.9,40,41 Indeed,
this condition is now one of the most prognostically favorable
subtypes of AML.42 Preliminary studies suggest that APL associated with NPM/RARA and NuMA/RARA is also retinoid-responsive.38,43 In marked contrast, PLZF/RARA-associated APL, is
resistant to ATRA in vitro, and patients previously treated with
ATRA as a single-agent therapy had an adverse prognosis.33
Furthermore, distinct from t(15;17)-associated APL, preliminary
studies suggest that patients with the PLZF/RARA fusion gene are
also resistant to arsenic trioxide,44,45 but they could be sensitive to
the combination of ATRA and granulocyte colony-stimulating
factor (G-CSF).46 Clearly, the underlying molecular lesion associated with APL has considerable implications for an optimal
therapeutic approach.
Because initiation of therapy is highly dependent upon a
successful morphological diagnosis, this study sought to determine
whether APL cases lacking t(15;17) could be reliably identified.
This issue was addressed at a European workshop held in Monza,
Italy, during June 1997, whereby a large series of cases with
morphologically suspected APL, but lacking t(15;17), were referred for central morphological, immunophenotypic, cytogenetic,
and molecular review. This process led to the development of a
novel morphological classification system that was found to
reliably distinguish APL cases with the PLZF/RARA fusion gene

BLOOD, 15 AUGUST 2000 VOLUME 96, NUMBER 4

from those with ATRA-responsive disease, including those with

underlying PML / RARA and NPM/RARA gene rearrangements.

Materials and methods

Patient characteristics
The 90 cases referred as APL at diagnosis and lacking the classical t(15;17)
were provided by 42 laboratories from 6 European countries and from
Memphis, TN (see Appendix). For controls, we examined 20 further
cases of APL with the classical t(15;17). The corresponding karyotypes and
molecular data were reviewed concomitantly but separately. Morphological
reviewers were ignorant of the cytogenetic and molecular data and vice
versa. Full details of the molecular and cytogenetic characterization of these
cases are presented by Grimwade et al.31 In a second step, the reviewed data
were combined, and the immunophenotypic data were considered. Unique
patient numbers were assigned to each case and are consistent with those of
Grimwade et al.31
We excluded 23 cases: 4 cases (1 M1-AML and 3 M2-AML cases) were
excluded as a result of morphological review, and furthermore, all were
negative for the PML / RARA transcript by reverse transcriptasepolymerase
chain reaction (RT-PCR); 4 cases were excluded as a result of cytogenetics
because of the presence of a minor clone with the classical t(15;17); and 15
cases were excluded due to insufficient material for adequate molecular
analysis. We retained 67 cases and, on the basis of molecular and
cytogenetic analyses, classified them as cases with or without PML / RARA
gene rearrangements
Cases with a PML / RARA gene rearrangement. The majority of these
49 cases were due to insertion events including 27 cases with documented
formation of PML / RARA and 1 case in which RARA/PML was the sole
fusion gene created. In 14 of 49 workshop cases, the PML / RARA fusion
gene resulted from complex chromosomal rearrangements. In the remaining
7 PML / RARA positive cases, conventional cytogenetic analysis revealed an
isochromosome for the long arm of the derivative chromosome 17,
ider(17)(q10)t(15;17)(q22;q21), [ider(17)]. These cases were subsequently
defined as t(15;17)-positive with an additional abnormality, and they were
retained to determine whether this superimposed cytogenetic abnormality
influenced morphological appearances. This latter group was excluded from
the accompanying molecular study,31 which considered only cases lacking t(15;17).
Cases lacking a PML / RARA gene rearrangement. Of 18 cases
lacking a PML / RARA gene rearrangement, 11 cases possessed PLZF/RARA
rearrangements including 2 new cases lacking t(11;17)(q23;q21); 6 of the
cases with documented t(11;17) were reported previously.26,33,44,46-49 There
were 2 cases with t(5;17): a new case with t(5;17)(q34;q21), expressing
NPM/RARA, and a case with an unbalanced der(5)t(5;17), which has been
partially described previously,50,51 with a proven deletion of 1 RARA
allele.31 In the remaining 5 cases, the RARA gene was not found to
be rearranged.
Morphological analysis
Bone marrow smears were reviewed by a committee of morphologists
(D.S., V.L., A.C.R., D.H., and G.F.) using a Leica-ICG video system
(Rueil-Malmaison, France). Peripheral blood films were reviewed, too; but
according to the FAB classification,10 they were not considered by
themselves as sufficient for diagnosis. Therefore, each case had a minimum
of 20 fields from bone marrow smears registered at a 1000-fold magnification and, according to general agreement, was classified using FAB
criteria.10,13 Furthermore, for each case, a total of 100 bone marrow blast
cells were analyzed by this committee, thereby taking into separate account
the major nuclear and cytoplasmic features considered characteristic of
M3/M3v cells (Figures 1 and 2). The blasts were initially classified
according to the appearance of the nuclear outline, regular (round or oval)
or irregular (reniform, bilobed, or folded). They were then further categorized according to the degree of granularity of the cytoplasm (hypergranular
or hypogranular). Finally, the presence or absence of Auer rods was taken

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BLOOD, 15 AUGUST 2000 VOLUME 96, NUMBER 4

NEW MORPHOLOGIC CLASSIFICATION SYSTEM FOR APL

1289

and 2). The classification system also allowed for the presence of a series of
additional morphological changes including basophilic or Chediak-Higashi
granules. The blasts with these latter features were initially assigned to a
category within the 3-parameter classification system (ie, categories 1-12),
then recounted and reassigned to category 13 (basophilic granules) or
category 14 (Chediak granules) (Figures 1 and 2). A category initially
planned for identifying hyperbasophilic cytoplasm was not used because
the hyperbasophilic variant form of APL was absent from our series. Apart
from blasts, a 15th category was defined by Pelger-like maturing cells
(Figures 1 and 2) whose percentage was evaluated on total cells from
granulocytic lineage, including blasts (n 100), and reported on the grid.
In a second step, cytogenetic and molecular data were taken into account,
and a mean value was calculated for each morphological cell category
within each cytogenetic/molecular group (Figure 3).
Figure 1. Classification system used for defining 15 categories of cells. In the
figure, Hyper indicates hypergranular cytoplasm; Hypo, hypogranular or agranular
cytoplasm; , the presence of 1 or 2 Auer rods; , the presence of 3 or more Auer
rods or faggots; Baso G, basophilic granules; Chediak, Chediak-like inclusions; and
Pelger, Pelger-like cells.

into account. Faggots were defined by the presence of at least 3 Auer rods
per cell.
Application of this 3-parameter classification system (nucleus, granularity, and Auer rods) led to the distinction of 12 categories of cells (Figures 1

Figure 2. Representative cells from each cell category of the classification

system. Bone marrow smears were stained with May-Grunwald-Giemsa and
registered at a 1000-fold magnification on a Leica-ICG system database.

Immunophenotype
Immunophenotypic features of 63 of 87 APL cases, including the 20
t(15;17) controls, were available and thus reviewed. This process took into
account surface antigens considered to have a characteristic expression
pattern in APL: HLA-DR, CD34, CD13, and CD33. In some instances, the
CD56 antigen that has been associated with APL was studied. The CD16
data were also available for some CD56 cases. Unfortunately, as limited
data on the CD2, CD15, and CD19 expression were available, these
antigens were not considered. Immunophenotyping was performed by
cytofluorometry using direct immunofluorescence either on bone marrow
samples (58 of 63 cases) after centrifugation through a Ficoll-Hypaque
density gradient (Pharmacia Biotech) or on total peripheral blood (5 of 63
cases) after lysis of red blood cells, in cases where the blast cell count
exceeded 50 109 cells per L (80%-99% blast cells).

Figure 3. Mean percentage of positive cells in each cell category according to

the cytogenetic/molecular groups. Cell categories are noted in Figure 1, and the
mean of positive cells is given as plus or minus the SD. A total of 100 blasts were
analyzed for each case, assigned to categories 1-12, and then recounted and
reassigned to category 13 or 14. Category 15 was defined by Pelger-like maturing
cells whose percentage was evaluated on the total cells, including blasts, from the
granulocytic lineage (n 100).

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1290

SAINTY et al

We used fluorescein isothiocyanate (FITC)- or phycoerythrin (PE)conjugated monoclonal antibodies (mAbs) (Immunotech International,
Marseille, France; Beckman Coulter, High Wycombe, England; and Dako
SA, Trappes, France, and High Wycombe, England). For each sample, a
minimum of 5000 cells were gated on forward and side scatter parameters,
and thus, at least 90% of the analyzed cells were blasts. In all cases,
negative controls included mouse immunoglobulin (Ig) directly conjugated
to FITC or PE. Expression in more than 20% blast cells was required to
define antigen positivity. For the purposes of this study, the typical APL
phenotype was defined as CD34, HLA-DR, CD13, CD33, and
CD56. Immunophenotypic results were compared to white blood cell
(WBC) counts; cases with WBCs greater than 10 109 cells per L were
considered as having hyperleukocytosis.

BLOOD, 15 AUGUST 2000 VOLUME 96, NUMBER 4

Results

0%-75%; mean, 9%). The unique insertion case solely expressing

RARA/PML31 belonged to the M3v category (category 10,
81% cells).
The complex group included a slight excess of M3v cases
compared with the t(15;17) control group (8 M3 and 6 M3v cases,
respectively). Only case 32 had a moderate percentage of blasts
with regular nuclei, whereas other cases had less then 12% of these
cells (categories 1-6; range, 0%-36%; mean, 5%). A slight difference was observed in the ider(17) group. Each of the 7 total cases
was classified as classical M3, as the majority of blasts had
hypergranular cytoplasm. However, the morphological profile of
this group differed from that of the control group with respect to a
higher percentage of category 1 cells (range, 3%-62%; mean,
22%), ie, regular nucleus and hypergranular cytoplasm without
Auer rods (Figure 4). In the ider(17) group, blasts with regular
nuclei (categories 1-6) accounted for a mean value of 26% (range,
3%-69%); 3 cases had more than 33%, whereas other cases had less
than 20% of these cells. In addition, a slight increase in Pelger-like
cells (category 15; range, 0%-17%; mean, 8%), mostly degranulated, was observed (Figure 4).

Morphological analysis of cases with PML / RARA

rearrangements

Morphological analysis of cases lacking PML / RARA

rearrangements

Control cases with the classical t(15;17). The mean percentage of

each cell category in each cytogenetic/molecular group is reported
in Figure 3. In the control group (n 20) with classical t(15;17),
the majority of blasts had an irregular nucleus and hypergranular
cytoplasm, which either lacked Auer rods (category 7; range,
3%-90%; mean, 51%) or contained faggots (category 9; range,
0%-41%; mean, 10%). These 2 categories correspond to the
classical M3 form of the FAB classification,10 which accounted for
16 of 20 control cases. Another peak was noted with 18% of the
cells falling within category 10, ie, irregular nucleus, poorly
granular cytoplasm, and no Auer rods. These cells therefore
corresponded to the characteristic cells of the M3v form of the FAB
classification.13 These cells were not a minor component of the
classical form of M3, but all cells belonged to the M3v subtype,
which accounted for the remaining 4 cases. Indeed, each of these
M3v cases had a majority of blast cells classified in category 10
(range, 73%-90%) and very few blasts belonging to category 7
(range, 3%-25%) or category 9 (range, 0%-8%).
Within the whole control group, Auer rods were seen in each
M3 and M3v case; faggots were absent in 4 cases (2 M3 and 2 M3v
cases); only a minority of blasts with regular nuclei were observed
(categories 1-6; range, 0%-27%; mean, 9%); no basophilic granules (category 13) were noted; and Chediak-like inclusions were
seen in only 1 case in a few blast cells (category 14; 1%).
Pelger-like cells were rarely observed (category 15; range, 0%10%; mean, 3%) and were not hypogranular; dysplastic features
were not observed in the rare cells from the erythroid and
megakaryocyte lineages. Using this classification system, M3 and
M3v cases can be readily distinguished; M3 cases are characterized
by a majority of blasts of categories 7-9; while in M3v, the majority
of blasts belong to categories 10-12.
Cases lacking the classical t(15;17) but with PML / RARA
rearrangements. No major morphological difference was seen
between the control group and the PML / RARA groups (Figure 3).
In the insertion group, the proportion of M3 and M3v cases (22 M3
and 6 M3v cases) and the morphological profile were similar to the
t(15;17) control group. Only 2 cases classified as M3 had more than
33% blasts with regular nuclei (cases 8 and 26), and other cases had
less than 27% of these cells (overall: categories 1-6; range,

Cases with PLZF/RARA rearrangements. Comparison of the

morphological features of t(15;17) controls with the PLZF/RARA
group (n 11) revealed clear differences (Figures 3 and 5). Only 2
cases, 44 and 46, could be readily classified as APL. Case 44 had
62% blasts with irregular nuclei; most of them had hypergranular
cytoplasm (53% blast cells belonged to categories 7-9) and could
be classified using FAB criteria as M3. In case 46, 80% basophilic
granular cells (category 13) were observed, which corresponded to
the basophilic M3-like form,17 but the majority (77%) of blasts
corresponded to category 1 (ie, regular nucleus and hypergranular
cytoplasm without Auer rods) (Figure 5). Other cases were
reminiscent of M3 according to their granularity and sometimes by
the presence of faggots. However, the majority of blasts had a
regular round or oval nucleus that was more suggestive of FAB
type M2, but these cases were retained as APL because of the
presence of hypergranular cells. Indeed, in the whole PLZF/RARA
group, the majority of blasts had (1) a regular nucleus (categories
1-6; range, 38%-100%; mean, 88%) and (2) an abundant cytoplasm
with either coarse granules (category 1; range, 29%-83%; mean,
56%; Figure 5, case 43A, B) or, less frequently, with fine or no

Statistical analysis
The Fisher exact test was used to test for associations between the
cytogenetic/molecular subgroup, M3/M3v morphology, hyperleukocytosis,
and immunophenotype.

Figure 4. PML / RARA cases of the ider(17) group. The majority of blasts with (A,
B) regular nuclei and (B) a Pelger-like cell.

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NEW MORPHOLOGIC CLASSIFICATION SYSTEM FOR APL

1291

more copious and less basophilic cytoplasm, appeared more

mature, and thus resembled cells that were blocked at a promyelocyte-myelocyte stage (Figure 5, cases 43A, 44, 50A, 51, and 52B).
Our classification system revealed an increased number of Pelgerlike cells (category 15; range, 0%-22%; mean, 10%) which were
hypogranular and seen in all cases except case 49 (Figures 3 and 5,
cases 46, 50A, 51, and 52C). Such appearances were reminiscent of
a myelodysplastic syndrome (MDS); however, in contrast to MDS,
the erythroid and megakaryocyte lineages were absent or very
reduced, impairing qualitative analysis, and the majority of cells
appeared blocked at the promyelocytic-myelocytic stage.
Two cases lacking t(11;17) had morphological features typical
of this group; they were subsequently shown to have underlying
PLZF/RARA rearrangements.31 Case 50 was associated with
del(11)(q23) terminal deletion with a breakpoint at 11q23 and case
52 with a normal karyotype. The latter case was shown to have a
submicroscopic insertion of RARA into band 11q23, thereby
leading to the expression of PLZF/RARA as the sole fusion
transcript. Both cases had a similar morphology (Figure 5, cases
50A, B and 52A-C): blasts with regular nuclei and either hypergranular cytoplasm (category 1; 57% and 52%, respectively) or
hypogranular cytoplasm (category 4; 43% and 48%, respectively)
associated with a chromatin pattern similar to that observed in
cases with documented t(11;17). In these 2 cases, Pelger-like cells
were also noted (category 15; 8% and 2%, respectively) (Figure 5,
cases 50A and 52C).
Cases with t(5;17). There were no morphological similarities
between the 2 APL workshop cases (54 and 55) associated with
t(5;17), and they were subsequently shown to be molecularly
different.31 In case 54 (a 9-year old child), t(5;17)(q34;q21) led to
the formation of NPM/RARA and RARA/NPM fusion genes. In case
55, with an unbalanced der(5)t(5;17), molecular analysis revealed
no evidence of an NPM/RARA rearrangement, and 1 RARA allele
was found to be deleted.31 The morphology of case 54 fitted with
M3, and the majority of blasts (97%) were hypergranular, with an
irregular or less frequently regular nuclear outline (category 7,
82%; category 1, 15%). In contrast with the t(15;17) control group,
no Auer rods were seen (Figures 3 and 6A).
The unbalanced der(5)t(5;17) case 55 had hyperleukocytosis
(Table 1) and a very peculiar morphology: the high percentage of
blasts with hypergranular cytoplasm and the presence of faggots
were reminiscent of M3, but the predominance of blasts with

Figure 5. PLZF/RARA cases. Case 43: Blasts have regular nuclei with (A)
hypergranular cytoplasm and (B) faggots of Auer rods. Case 44: Blasts have irregular
nuclei with (A) hypergranular cytoplasm and (B) Chediak-like granules. Case 45:
Blasts have regular nuclei and poorly granular cytoplasm. Case 46: Blasts with
regular nuclei, basophilic granules, and a Pelger-like cell. Case 50: Blasts with
regular nuclei and either (A) hypogranular or (B) hypergranular cytoplasm and (A) a
Pelger-like cell. Case 51: Majority of blasts have regular nuclei and a Pelger-like cell.
Case 52: Blasts with regular nuclei and either (A) hypogranular or (B) hypergranular
cytoplasm with myelocyte-like features and (C) a Pelger-like cell.

granules (category 4; range, 3%-67%; mean, 28%; Figure 5, cases

45, 50A, 51, and 52A). Only 2 cases, 43 and 53, exhibited faggots
(category 3; 15% and 18%, respectively; Figure 5, case 43B). Case
44 was the sole case with Chediak-like granules (category 14; 12%)
(Figure 5, case 44B).
Noticeably, in 5 of 11 PLZF/RARA cases, the majority of blasts
had a more condensed nuclear chromatin pattern. Compared with
blasts from the t(15;17) control group, the PLZF/RARA blasts had

Figure 6. Chromosome transcription t(5;17) cases. (A) The NPM/RARA case:

Majority of blasts have irregular nuclei and hypergranular cytoplasm without Auer
rods. (B) The der(5)t(5;17) case: Blasts with regular nuclei, hypergranular cytoplasm
with Chediak-like granules, and Pelger-like cells.

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SAINTY et al

Table 1. WBC count and immunophenotypic characteristics of APL cases

Groups (cases)

WBC count, 109

cells per L

HLA-DR (%)

CD34 (%)

CD13 (%)

CD33 (%)

CD56 (%)

PML / RARA (50/69)*

t(15;17) control group (20/20)
Range
Median
Insertions (16/28)
Range
Median
Complex (10/14)
Range

(20/20)

0.5-86.1
1.7
(27/28)
4.4

0.7-60

Median

3.1

ider(17) (4/7)

(7/7)

(13/14)

(12/15)

(19/20)

8-98
79

(15/16)

(16/20)

(14/14)

0-98

30 -100

85

91

(6/10)

(9/9)

(8/9)

0-84

0-91

30-94

20-97

55

60

53

(3/4)

Median

2.8

0
(7/7)

Range

2-69.5

0-13

Median

9.9

(4/4)

0-3
1

(7/7)

0-3
0

(4/4)

(3/4)

46-97

0-85

69

(7/7)

(6/7)

(20/20)
0-8

74

(9/10)

0-98

0-74

0-43

0.8-10.8
(11/11)

(19/20)
0-46

0-25

Range

PLZF/RARA (7/11)

0.2-257
(13/14)

(19/20)
0-37

(4/5)
0-30
1

(2/2)
0-2
1

(1/1)
0
0

(4/6)

77-100

0-100

0-100

96

93

70

t(5;17) (2/2)

NPM/RARA (1/1)

17

97

der (5) t(5;17)

43.1

13

83

(5/5)

(4/4)

(3/4)

(3/4)

RARA not rearranged (4/5)

Range
Median

3.3-84

0-7

11.3

(3/4)

0-22

1-98

1-99

82

89

(2/2)
1-9
5

The positivity of each immunophenotype is given on the left, the indicated characteristics (number of cases and percentages) on the right.
* Indicates the number of cases with available phenotype/total number of cases.
Indicates the number of cases with available WBC/total number of cases.
Indicates the number of positive () or negative () cases/number of tested cases, the cut-off is more than 20% positive cells.
Indicates the percentage of positive cells.
P .002, comparing CD34 positivity between complex cases and noncomplex cases in the PML / RARA group; CD34 positivity among complex cases reflected an
association with M3v morphology.
P .001, comparing CD56 positivity between PLZF/RARA and PML / RARA cases; P .0008, comparing CD56 positivity between PLZF/RARA and all non-PLZF/RARA
cases.

regular nuclei (categories 1-6, 99%) was more suggestive of M2.

About half of the blasts possessed hypergranular cytoplasm
(category 1, 43%) and half were hypogranular (category 4, 46%). A
few faggots were seen (category 3, 8%; category 6, 2%). In
addition, Pelger-like cells were numerous (category 15, 26%), and
a significant proportion of blasts contained Chediak-like granules
(category 14, 13%) (Figure 6B). This profile (Figure 3) was thus
reminiscent of PLZF/RARA-associated APL; however, an underlying PLZF/RARA fusion gene was excluded by molecular analysis.50
APL cases lacking evidence of a RARA rearrangement. In 5
workshop cases, molecular analysis revealed no evidence of an
underlying RARA rearrangement.31 Two cases (59 and 60) could be
classified as M3 (Figure 7A) because they exhibited a predominance of blasts with irregular nuclei. The remaining cases (56, 57,

and 58) were difficult to classify as M3 or M3v according to FAB

criteria as they presented with a majority of blasts with regular
nuclear outline (categories 1-6, more than 67% in each case). But
they were nonetheless retained as APL because of the presence of
hypergranular blasts (Figure 7B,C). All cases in the whole group
possessed more than 36% blasts with regular nuclei (categories
1-6; range, 37%-93%; mean, 66%). This profile (Figure 3) was
close to that of PLZF/RARA APL. In contrast to the typical
features observed in the latter subtype of APL, case 56 exhibited a
high number of blasts with faggots (category 3, 55%) (Figure 7C),
and Pelger-like cells were rarely observed (category 15; range,
0%-13%; mean, 3%). The ATRA responsiveness of cases lacking
RARA rearrangements could not be formally established due to the
lack of material available for in vitro differentiation assays.
Nevertheless, in case 58, there was no observed response after 14
days of ATRA treatment. Sensitivity to retinoids could not be
determined in the remaining patients because 3 patients received
ATRA in combination with chemotherapy, and 1 patient died prior
to therapy.
Immunophenotypic analysis of cases with PML / RARA
rearrangements

Figure 7. Cases lacking RARA rearrangements. (A) Case 60: The majority of
blasts have irregular nuclei, hypergranular cytoplasm, and faggots. (B) Case 57: The
blasts have regular nuclei and hypergranular cytoplasm. (C) Case 56: Blasts with the
same nuclear features and a high number of faggots.

Details of the presenting WBC count and immunophenotype of the

t(15;17) control group and the cases with PML / RARA rearrangements defined by molecular methods are presented in Table 1. No
significant differences were observed between the immunophenotypic profile or the WBC count of the molecularly defined group
and the t(15;17) controls. Overall, there were 69 cases with
PML / RARA rearrangements (53 M3 and 16 M3v cases). The

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BLOOD, 15 AUGUST 2000 VOLUME 96, NUMBER 4

median WBC count was low for the whole group (median,
3.1 109 cells per L; range, 0.2-257 109 cells per L), reflecting a
preponderance of M3 cases (median, 2.4 109 cells per L; range,
0.5-97 109 cells per L), whereas the median WBC count of the
M3v cases was significantly higher (median, 32.9 109 cells per
L; range, 0.2-257 109 cells per L; P .0004).
In 50 cases it was possible to correlate immunophenotype with
morphological features. There were no differences observed between M3 and M3v in the expression pattern of HLA-DR (M3:
median, 3.5%; range, 0%-84%; and M3v: median, 0%; range,
0%-13%); CD13 (M3: median, 82%; range, 0%-98%; and M3v:
median, 70%; range, 30%-97%); or CD33 (M3: median, 78%;
range, 0%-100%; and M3v: median, 83%; range, 5%-98%).
However, expression of CD34 was correlated with M3v morphology (M3: median, 0%; range, 0%-74%; and M3v: median, 23%;
range, 0%-91%; P .002) and with hyperleukocytosis (CD34:
median WBC, 29 109 cells per L; range, 0.8-133 109 cells per
L; and CD34: median WBC, 1.9 109 cells per L; range, 0.5257 109 cells per L; P .023). CD56 expression was found to be
rare in APL with PML / RARA rearrangements, with only 1 positive
case among 28 cases tested. The PML breakpoint was determined
in 41 cases including 13 controls; 23 cases had a 3-bcr1-2
breakpoint (19 M3 and 4 M3v cases), and 18 cases had a 5-bcr3
breakpoint (11 M3 and 7M3v cases; P .16).
Immunophenotypic analysis of cases lacking PML / RARA
rearrangements

Cases with PLZF/RARA rearrangements. In this group, 5 of 11

cases had hyperleukocytosis, and the median WBC count was
moderate (9.9 109 cells per L). Immunophenotyping performed
in 7 of 11 cases showed that the blasts were typically HLA-DR,
CD34, CD13, and CD33 (Table 1). Interestingly, the CD56
antigen was found to be strongly expressed in 4 of 6 tested cases
(case 47, 100%; case 49, 53%; case 51, 90%; and case 52, 86%)
and weakly in one case (case 50, 16%), with a median of 70%. In 2
cases strongly expressing CD56, CD16 expression was negative in
case 47 and positive in case 51.
Cases with t(5;17). Cases 54 and 55 had hyperleukocytosis.
With the exception of the lack of CD13 expression, their immunophenotype was typical of APL (Table 1), with negativity for
CD56 expression.
APL cases lacking evidence of a RARA rearrangement. In
this group, 3 of 5 cases had hyperleukocytosis, and the median
WBC count was higher (11.3 109 cells per L). Immunophenotyping performed in 4 of 5 cases revealed no major differences from
t(15;17) APL (Table 1). CD34 was weakly expressed in case 60,
which had hyperleukocytosis, and CD56 was negative in the 2
cases tested (56 and 58).

Discussion
We performed morphological review of 90 cases referred as APL
and lacking the classical t(15;17); using the FAB criteria, we were
able to formally exclude 4 cases (1 M1 and 3 M2 cases) and to
classify the majority of cases as either M3 or M3v. Regarding
morphology, WBC count, or immunophenotype, there was no
major difference seen between the t(15;17) control group and the
cases lacking t(15;17) with molecular evidence of PML / RARA
rearrangements. All cases with ider(17) possessed M3 morphology,
which is in accordance with 9 previously published cases.52,53 If

NEW MORPHOLOGIC CLASSIFICATION SYSTEM FOR APL

1293

this association is confirmed in a larger series, it could be related to

duplication of the RARA/PML gene and/or loss of the p53 gene on
the ider(17). In the whole PML/RARA group, hyperleukocytosis
was associated with M3v morphology and CD34 expression, in
accordance with previous studies.24,25
In contrast to the PML / RARA cases, all cases with PLZF/RARA
rearrangements were reminiscent of M3 with regard to the hypergranular cytoplasm. Thus they were retained in this study, but in 10
of 11 cases, the majority of blasts lacked the bilobed or folded
nuclei characteristic of M3 or M3v. The chromatin pattern was also
more condensed than typically observed in AML blasts, mimicking
a block at the promyelocytic-myelocytic stage, and in addition,
hypogranular Pelger-like cells were observed in the majority.
Hence, these cases possessed morphological features associated
with FAB types M2 and M3 as well as MDS. Previous reports of
t(11;17)-associated APL indicate a wide range of morphological
appearances. In a review of 6 cases,33 common morphologic
features distinct from M3 or M3v were found: the blasts were more
granular than typical M2-AML cases, but less granular than usually
observed in M3-AML (hypergranular). Furthermore, the authors
focused on the: (1) loss of the usual reddish color shift of M3
granules, (2) absence of the obscured nuclear outline typically
observed in M3, (3) absence of faggot cells, (4) loss of the bilobed
grooved nucleus, and (5) presence of fine granules reminiscent of
M3v-AML. We agree with regard to the rarity of Auer rods and
faggot cells and the loss of the bilobed grooved nucleus.33
Furthermore, our classification system demonstrates that this latter
feature (regular nuclear outline) is the key criterion in distinguishing this subtype of APL; in addition, we have identified a number of
other features of value including a more condensed chromatin
pattern in blasts and the presence of Pelger-like cells.
Identification of these features allowed us to focus molecular
analysis on 2 cases lacking t(11;17), which were subsequently
found to harbor cryptic PLZF/RARA rearrangements. These 2 cases
had a similar morphology that was very typical of the PLZF/RARA
group. One case has been proven to express solely PLZF-RARA,31
suggesting that this transcript could be responsible for the peculiar
morphology of this APL subtype. Indeed, this case had all the
characteristic features observed in cases with t(11;17) in which
both PLZF-RARA and reciprocal RARA/PLZF transcripts were
detectable, namely blast cells with regular nuclei, a maturation
block at the promyelocyte/myelocyte stage, the absence of Auer
rods, and the presence of Pelger-like cells (Figure 5, case 52A-C).
This finding would indicate that there are some differences between
t(11;17) APL arising in man and recent transgenic mouse models,
which have suggested that RARA/PLZF influences morphological
appearances and have implied that both fusion transcripts are
required for the development of APL.55
The present study has shown that the presence of a regular
nuclear outline is a key feature of t(11;17)-associated APL, and we
suggest that the FAB classification could be extended to include a
new subtype of M3, M3r, which is characterized by regular
nuclei. As all PLZF-RARA cases had more than 37% blasts with
regular nuclei, and the majority of PML / RARA cases had less than
30% such cells (6 of 69 cases had more than 30%), we propose a
30% threshold for defining M3r-APL, keeping all the cytoplasmic
FAB criteria necessary for APL diagnosis. However, it is also clear
that the 5 of 5 cases lacking RARA rearrangements, 3 of 7 cases
with ider(17), and the sole der(5)t(5;17) case could be classified as
M3r. Moreover, in common with cases with PLZF/RARA rearrangements, the latter 2 groups demonstrated an increased number of
Pelger-like cells; this feature could be related to the loss of

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1294

BLOOD, 15 AUGUST 2000 VOLUME 96, NUMBER 4

SAINTY et al

chromosome 17p material and/or the p53 gene, as reported in some

myelodysplastic syndromes.56,57
In a series of 25 APL cases, Neame et al15 identified 3
PML / RARA M2-like cases, where the majority of blasts were
normal-appearing differentiated promyelocytes without evidence
of folding and lobulation of the nucleus. In our classification
system, these would be classified as M3r; a further category of cells
(blasts with hyperbasophilic cytoplasm) could be added in order to
classify the hyperbasophilic M3 described by McKenna et al.14 Our
series did not include the NUMA/RARA or STAT5b/RARA rearrangements. The previously described NUMA/RARA case37,38 had hypergranular blasts, with irregular nuclei and Pelger-like cells, and is
therefore reminiscent of M3 morphology. The case expressing
STAT5b/RARA39 was classified as M1-AML, with only a minority
of blast cells evocative of M3v.58
In situations in which the morphology is similar to that
associated with the PLZF/RARA rearrangement, the present study
suggests that immunophenotype analysis could be helpful. Indeed,
the NK-associated CD56 antigen was positive in 4 of 6 PLZF/
RARA cases tested. Moreover, in a case lacking t(11;17), the CD56
positivity, as well as the evocative morphology, prompted further
molecular analysis, which lead to identification of a cryptic
PLZF/RARA rearrangement. Positivity for CD56 and myeloid
antigens, with negativity for HLA-DR and CD34, has also been
associated with the NK/myeloid leukemia identified by Scott et
al.26 This study, which included 1 case with a PLZF/RARA
rearrangement, highlighted the presence of a particular morphology in two-thirds of cases, whereby blasts were characterized by a
deeply invaginated nuclear membrane and fine azurophilic granulations in the cytoplasm. This appearance is clearly distinct from that
observed in PLZF/RARA-associated APL; indeed, such CD56
cases identified in the present study were characterized by a regular
nuclear membrane and hypergranular cytoplasm. CD56 positivity
was also recently described in a new t(11;17)(q23;q21) case, which
was classified as M3v but had a poorly lobulated nucleus.59 The
detection of CD56 in t(11;17)-associated APL led to the suggestion
that this form of AML could originate from a more immature
progenitor than t(15;17) APL, which is common to myeloid and
NK cell lineages.33 However, as CD56 corresponds to the N-CAM
molecule whose gene is located at band 11q23,60 one cannot
exclude an aberrant expression related to a gene-position effect at
the chromosomal breakpoint.
In 12 cases that lacked a PLZF/RARA rearrangement, the
morphologic appearances were reminiscent of the PLZF/RARA

group; these included 6 PML / RARA cases (2/28 insertions; 1/14

complex; 3/7) ider[17]; the case with der(5)t(5;17); and 5/5 cases
lacking RARA rearrangements. The CD56 antigen was tested in 5
cases: 1 PML / RARA insertion, 1 ider(17), 1 der(5)t(5;17), and 2
cases without RARA rearrangement. In each case the antigen was
negative, which highlights the potential value of this antigen for
distinguishing subtypes of APL, although this should be confirmed
in a larger number of patients. Moreover, with the exception of the
PLZF/RARA group, expression of CD56 in APL was found to be
rare; it was detected in only 1 of 28 PML / RARA cases analyzed
(P .001). This is in accordance with data from the literature;
indeed, only 14 cases of PML / RARA APL expressing CD56 have
been reported to date including 1 M2-like case.15,27
We report a new t(5;17) case expressing NPM/RARA, which, in
common with the 2 previous reported cases,34,36 occurred in a
pediatric patient. Whereas the cases described by Corey et al34 and
us could be classified as M3 and presented with hyperleukocytosis,
the remaining case36 had an M3v morphology. Auer rods were not
observed in any of these cases, a feature very unusual in APL. The
immunophenotype of our t(5;17) case was typical of APL (CD13
but CD33) and, in contrast to the case published by Corey et al,
was CD56.
Finally, in a few APL cases, rearrangements of RARA were not
found31; most of them presented with hyperleukocytosis and a
morphological profile close to the PLZF/RARA group, but CD56
was negative in the 2 cases tested. The identification of such cases
suggests that alternative pathways could mediate the differentiation
block that characterizes APL, although it remains a possibility that
RARA could still be involved by mutation or epigenetic mechanisms. Such events are currently under investigation.
In conclusion, no major morphological or immunophenotypic
difference exists between APL harboring the classical t(15;17) and
PML / RARA cases lacking t(15;17). Interestingly, our study highlights a specific morphological pattern for the PLZF/RARA group,
which was associated with the CD56 expression. These features
should alert one to the possibility of an underlying PLZF/RARA
rearrangement that can be cryptic. Distinguishing APL cases with
the PLZF/RARA fusion from other forms of the disease is critical
because they are resistant to arsenic trioxide and ATRA as a
single-agent therapy. The present study underlines the importance
of combining morphologic, immunophenotypic, cytogenetic, and
molecular studies to distinguish patients who could benefit from
tailored therapeutic strategies.

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Appendix
The Monza Workshop comprised 41 cytogenetic laboratories
representing 6 European countries and 1 American laboratory
from St Jude Childrens Hospital, Memphis, TN. The participants are listed below; participating cities or the names of
participating centers and the number of cases supplied are given
in parentheses. The workshop was organized by Dr Andrea
Biondi. The European communitysponsored concerted action,

Molecular and Cytogenetic Diagnosis in Hematological Malignancies, was headed by Prof Anne Hagemeijer. *Indicates a
member of the Monza working party; indicates a member of the
working committee.
A. Hagemeijer,* H. Van Den Berghe, F. Speleman, L. Michaux,
H. Vranckx, Ph. Martiat, A. Van Orshoven, J. Rhodin, J. M. Scheiff,
A. Criel, G. Verhoef, D. Selleslag, J. L. Michaux, A. Delannoy, P.

From www.bloodjournal.org by guest on March 19, 2016. For personal use only.
1296

SAINTY et al

Meeus, Z. Berneman (Belgium: Leuven, 6; Brugge, 3; Brussels, 3;

Jolimont, 1; Antwerp, 1); N. Dastugue, E. Duchayne, F. RigalHuguet, A. Huynh (Toulouse, France: 7); D. Head,* W. Miller Jr,
P. B. Brett, J. Bickers, and E. Magenis (Memphis, TN; New York,
NY; Sacramento, CA; New Orleans, LA; Portland, OR: 6); M.
Lafage-Pochitaloff,* M. J. Mozziconacci,* F. Fernandez, A.
Restoin, J. Gabert, F. Birg,* D. Sainty,* C. Arnoulet, V. J. Bardou,
A. M. Stoppa, D. Coso, D. Blaise (Institut Paoli-Calmettes,
Marseilles, France: 6); F. Mugneret, B. Favre,* M. A. Collonge,
P. M. Carli, F. Bailly, M. Manadie, E. Racadot, D. Caillot, A.
Rozenbaum, B. Salles (Dijon, France: 5); D. Belotti, S. Tosi, A.
Biondi,* G. Cazzaniga,* A. Cantu`-Rajnoldi,* G. Rossi, S. Cortelazzo, G. Gaipa, P. Airo, G. Borleri, A. Rambaldi, G. Dotti (Monza,
Italy: 5); J. H. Jansen,* B. van der Reijden,* A. Hagemeijer,*
H. J. C. M. Holdrinet (The Netherlands: Erasmus University,
Rotterdam, 4; Breda, 1); I. Radford-Weiss,* E. Delabesse, G.
Flandrin,* L. Benattar, F. Valensi, L. Hoang, B. Varet, E.
Macintyre (Paris Necker, France: 3); M. Mancini,* D. Diverio, F.
Lo Coco, M. Nanni, S. Fenu, C. Nervi, G. Alimena (University La
Sapienza, Rome, Italy: 3); C. Charrin, D. Treille-Ritouet, I. Tigaud,
X. Thomas (Lyon, France: 3); F. Pasquali,* L. Seghezzi, E.
Maserati, R. Invernizzi, G. Basso, R. Maccario, A. Gabbas, F.
Locatelli, G. Bergamashi (Italy: Pavia, 2; Nuoro, 1); J. M.
Hernandez, M. Gonzalez, A. Orfao, N. C. Gutierrez* (Hospital
Universitario, Salamanca, Spain: 2); R. Berger,* M. Busson-Le
Coniat, M. T. Daniel (Paris, Saint-Louis, France: 2); J. van den
Akker, C. Perot, D. Bories, J. M. Pignon, M. F. Portno, C.
Cordonnier (Paris, St Antoine and Henri Mondor, France: 2); V.
Eclache* (Bobigny, France: 2); S. Raynaud, I. Sudaka (Nice,
France: 2); A. Aventin, M. Espadaler,* M. Carrasco (Sant Pau
Hospital, Barcelona, Spain: 2); M. Neat,* S. M. Kelsey, A. C.
Newland (St Bartholomews and Royal London Hospitals, London,

BLOOD, 15 AUGUST 2000 VOLUME 96, NUMBER 4

England: 2); J. Waters, M. Pomfret, D Milligan (Heartlands

Hospital, Birmingham, England: 1); H. Walker, F. Oliver, S.
Langabeer, A. H. Goldstone, (University College Hospital, London, England: 1); P. Chipping (North Staffordshire Hospital, North
Staffordshire, England: 1); P. Revell (Stafford District General
Hospital, Stafford, England: 1); J. Ropner (Gloucester Royal
Infirmary, Gloucester, England: 1); D. Stevenson, D. Culligan
(Aberdeen Royal Infirmary, Aberdeen, Scotland: 1); K. Rodgers, J.
Emmerson, G. Abrahamson, D. Samson (Charing Cross Hospital,
London, England: 1); P. Gorman, D. Sheer (ICRF, London,
England); D. Grimwade,* K. Howe,* E. Solomon (Guys, Kings
& St Thomas School of Medicine, London, England); C. Harrison,
L. Secker-Walker (Royal Free Hospital, London, England); E.
Vandenberghe, G. Wilson, A. Watmore (Royal Hallamshire &
Sheffield Childrens Hospitals, Sheffield, England); S. Sobolewski,
A.V. Kiorkian (Pilgrim Hospital, Boston, UK: 1); C. Mecucci, C.
Liberatore, C. Matteucci,* R. La Starza, Fr Grignani, P. G. Pelicci*
(Perugia, Italy: 1); C. Bilhou-Nabera, Ph. Bernard, M. Micheau, A.
Notz-Carre`re, F. X. Mahon, J. Reiffers (Bordeaux, France: 1); F.
Viguie, S. Ramond, D. Bouscary (Hotel-Dieu and Cochin, Paris,
France: 1); M. J. Gregoire, Ph. Jonveaux, A. Guerci, J. Buisine
(Nancy, France: 1); F. Brizard, A. Brizard, A. Sadoun (Poitiers,
France: 1); F. Uetwiller, F. Maloisel, M. Mark (Strasbourg, France:
1); P. Talmant, H. Avet-Loiseau, R. Garand (Nantes, France: 1); S.
Taviaux, M. Dupont, J. Taib, F. Bernard, N. Sarran, G. Margueritte
(Montpellier, France: 1); D. Leroux, M. Callanan, M. F. Sotto
(Grenoble, France: 1); R. Casalone, M. G. Biotti, G. Pinotti
(Varese, Italy: 1); C. Chomienne (Paris Saint-Louis, France) for
RT-PCR experiments on 4 cases; M. Lessard, A. Hery, (Brest,
France) and I. Chudoba (Altlussheim, Germany) for M-FISH and
m-band analyses on 3 cases; and V. Liso* (Bari, Italy).

From www.bloodjournal.org by guest on March 19, 2016. For personal use only.

2000 96: 1287-1296

A new morphologic classification system for acute promyelocytic

leukemia distinguishes cases with underlying PLZF/RARA gene
rearrangements
Danielle Sainty, Vincenzo Liso, Angelo Cant-Rajnoldi, David Head, Marie-Jolle Mozziconacci,
Christine Arnoulet, Laurence Benattar, Susanna Fenu, Marco Mancini, Eliane Duchayne,
Franois-Xavier Mahon, Norma Gutierrez, Franoise Birg, Andrea Biondi, David Grimwade, Marina
Lafage-Pochitaloff, Anne Hagemeijer and Georges Flandrin

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