1. Art and Diseño

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Significance of the P210 Versus P190 Molecular Abnormalities in Adults With
Philadelphia Chromosome-Positive Acute Leukemia
By Hagop M. Kantarjian, Moshe Talpaz, Kapil Dhingra, Elihu Estey, Michael J. Keating, Stella Ku, Jose Trujillo,
Yang Huh, Sanford Stass, and Razelle Kurzrock
We investigated the significance of p2lO and p190 molecular
abnormalities in 32 adults with Philadelphia chromosome
(Ph)-positive acute leukemia. p210 was detected in 15 patients (47%). p190 in 16 (50%). and both in one (3%). p210
was noted in 11 of 24 patients (46%) with acute lymphocytic
leukemia, and in four of eight patients (50%) with acute
myelogenous or undifferentiated leukemia. Among 29 patients with untreated disease (p210, 14 patients; p190, 15
patients), no significant differences in the two molecularly
distinct groups were observed by pretreatment characteristics including age, degree of organomegaly, anemia, leukocytosis, thrombocytopenia,occurrence of karyotypicabnormalities in addition to Ph, or residual diploid metaphases.
Complete response (CR) rates were also similar. Although
T
HE PHILADELPHIA chromosome (Ph), originally
described as a shortened long arm of chromosome 22,’
is present in more than 90% of patients with chronic
myelogenous leukemia (CML), in 20% to 35% of adults
with acute lymphocytic leukemia (ALL), and in 5% or
fewer children with ALL or patients with acute myelogenous leukemia (AML).2.3 In CML, the Ph abnormality
results from a reciprocal translocation between the long
arms of chromosomes 9 and 22, the t(9;22) (q34;qll)
translocation, which transposes the C-ABL gene from
chromosome 9 to the 5’ half of the BCR gene on chromosome 22, creating a new hybrid gene called BCR-ABL.
Transcription of BCR-ABL results in a hybrid 8.5-kb
messenger RNA (mRNA) that codes for a 210-Kd protein
(p210BCR-ABL;p210)!,5In the 8.5-kb mRNA, either exon b2 or
exon b3 of the major breakpoint cluster region (bcr) is
coupled to C-ABL exon 2 (b3-a2 or b2-a2 junction).
The break on chromosome 22 in acute leukemia may
occur in a different location than in CML, 5’ to the bcr,
within the first intron. The result is a smaller 7.5-kb
BCR-ABL fusion mRNA that encodes a 190-Kd protein
(p190Bc”ABL;p190).6~7
In this mRNA, the first exon of the
BCR gene is spliced to the second exon of the C-ABL gene
(el-a2 junction).
Most children with Ph-positive ALL have the p190 rather
than the ~ 2 1 0 . ~However,
3~
which of the two abnormalities
occurs more frequently in adults with Ph-positive acute
leukemia has not been studied extensively.lo~llIt is also
unknown whether the different molecular abnormalities
are associated with different clinico-laboratory features or
with different outcomes after therapy. These abnormalities
may also diagnostically discriminate patients who present
with a CML lymphoid blastic phase (Ph-positive, bcr
rearranged, p210 disease) from those who have true de
novo ALL (Ph-positive, bcr germline, p190 disease).
In an attempt to answer the above questions, we analyzed
the Ph-associated molecular abnormalities in adults with
Ph-positive acute leukemia.
Blood, Vol78, No 9 (November l), 1991:pp 2411-2418
the remission duration tended to be longer with pl90
(P = .OS), the differences were minor (median duration 29 v
20 weeks) and not paralleled by differences in survival rate. In
10 patients studied by karyotypic analysis in remission, two
of four patients with p190 and two of six patients with p2lO
showed 100% normal metaphases. One of the seven patients
(14%) with p2lO who achieved CR manifesteda morphologic
picture of second chronic-phase chronic myelogenous leukemia lasting for 1 month. We conclude that the molecular
studies in Ph-positiveacute leukemiaare not associated with
significantly different clinico-laboratory,karyotypic, or prognostic implications.
6 1991 by The American Society of Hematology.
MATERIALS AND METHODS
Adults with a diagnosis of Ph-positive acute leukemia referred
between June 1984 and January 1991 were the subjects of this
analysis. Patients were required to have (1) a documented Ph
abnormality by karyotypic analysis,’* (2) a new onset of acute
leukemia presentation without a preceding antecedent hematologic disorder or a CML picture, and (3) 30% or more marrow
blasts. Approval was obtained from the Institutional Review Board
for these studies. Informed consent was provided according to the
Declaration of Helsinki.
Workup at presentation included a history and physical examination; complete blood, differential, and platelet counts; SMA 12,
including hepatic and renal functions; bone marrow (BM) aspiration and biopsy for morphology, cytochemical, and enzymatic stains
(myeloperoxidase, chloroacetate, nonspecific esterase, periodic
acid-Schiff [PAS], terminal deoxynucleotidyl transferase [Tdt]);
imm~nophenotyping’~~’~;
karyotypic analysis”; electron microscopy’s; and molecular studies as described below.
The classification of the acute leukemia was based on the
French-American-British (FAB) criteria and histochemical
In addition to the characteristic morphology, a diagnosis
of AML required 3% or more myeloperoxidase-positiveblasts or
positive monocytic stains. ALL was diagnosed if the blasts were
morphologically lymphoid, myeloperoxidase-negative, and Tdtpositive andlor common acute lymphocytic leukemia antigen
From the Departments of Hematology, Clinical Immunology, and
Biological Therapy, and Laboratory Medicine, M.D. Anderson Cancer
Center, Houston, TX.
Submitted May 6,1991; accepted July 2,1991.
H.M.K. is a Scholar for the Leukemia Society ofAmerica.
Address reprint requests to Hagop M. Kantarjian, MD, Department
of Hematology, Box 61, M.D. Anderson Cancer Center, 1515 Holcombe Blvd, Houston, 7X 77030.
The publication costs of this article were defrayed in part by page
charge payment. Thk article must therefore be hereby marked
“advertisement” in accordance with 18 U.S.C. section 1734 solely to
indicate this fact.
0 I991 by TheAmerican Society of Hematology.
0006-4971191 /78O9-0018$3.00/0
241 1
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2412
(CALLA)-positive. Acute undifferentiated leukemia (AUL) was
diagnosed if the blasts were morphologically undifferentiated and
histochemically negative. The presence of myeloid surface markers
on more than 20% of undifferentiated blasts did not change the
diagnosis from AUL to AML.
Molecular Studies
Several techniques were used to determine whether patients
with Ph-positive acute leukemia produced p190 or p210. To assess
RNA splice junction, the polymerase chain reaction (PCR) and
specific primers to amplify cDNA were used; the product was then
determined with standard Southern blotting and with hybridization
protection assay (HPA) as detailed below.” Patients with a b2-a2
or b3-a2 junction produce p21019;patients with an el-a2 junction
produce ~ 1 9 0 P In
’ ~ 10 patients, the protein product was verified
with the use of specific antisera and the immune complex kinase
assay. In 21 other patients, Southern blotting of DNA was also
performed to determine the genomic configuration.
PCR: Samplepreparation and amplification. Total cellular RNA
was extracted by previously described methods.20RNA from K562
cells, an erythroid blast crisis cell line, and from patient W, a
Ph-positive CML patient, were used as positive controls for mRNA
containing b3-a2’l and b2-a2 junctions, respectively. The ALL-1
cell line (kindly provided by Dr G. Rovera, Wistar Institute,
Philadelphia, PA) was used as a positive control for the el-a2
mRNA transcript encoding ~190.6,’~’
HL-60 (a promyelocytic leukemia cell line) and normal human endometrial RNA were used as
negative controls. One microgram of total RNA from cell lines or
one-tenth of the total RNA from 50 to 200 x 106 white blood cells
(WSCs) from patient samples was used for amplification reactions.
The amplification method and the primers have been previously
described.n Amplification was performed for 40 cycles.
Because contamination has proven to be a significant problem in
some laboratories using this technique, the following precautions
to ensure the accuracy of results were undertaken: (1) the thermal
cycler was kept in a separate laboratory, away from the room where
cell collection, RNA processing, and cDNA synthesis was performed; (2) no amplified samples were allowed to be brought back
into the room where RNA processing was performed; (3) at least
one negative control was run for each equipment; and (4) samples
from each patient were run at least two different times.
HPA. We have recently described the application of a new
rapid method for detecting amplified BCR-ABL CDNA.’’.~’This
method uses acridinium ester-labeled probes (Genprobe, Inc, San
Diego, CA) with high chemoluminescent properties. In the presence of hydrolysis buffer, the rate of hydrolysis of free probe is
much faster than that of hybridized probe. Therefore, separation of
hybridized probe on a solid support, as in Southem blotting, is
unnecessary. HPA is performed in solution, and allows for reliable
detection of transcripts within 30 minutes after amplification. HPA
results correlated with those of Southern blotting of amplified
product in all 60 samples from patients with Ph-positive CML,
acute leukemia, Ph-negative leukemia, or normal volunteers.”
Acridinium ester-labeled oligonucleotides complementary to the
BCR-Al3L junction sequences were synthesized by Genpr~be.~’
The chemical labeling of the DNA probes with acridinium ester
was achieved by reacting alkylamine linker-arms, which were
introduced during DNA synthesis, and an N-hydroxysuccinimide
ester of a methyl acridinium phenyl ester. The b2-a2 probe was a
28mer with 22 bases from bcr exon 2; the b3-a2 probe was a 25mer
with 11 bases from bcr exon 3. These probes were used to detect
transcripts encoding p210. The el-a2 probe was a 26mer with 17
bases from the first exon of the BCR gene and was used to detect
transcripts coding for p190. The method for application of this
technique was previously reported.’* A chemiluminescence read-
KANTARJIAN ET AL
ing, expressed in relative light units (RLUs), indicates whether a
sample is positive or negative.
Southem blotting of amplified products. To verify the results
obtained by HPA, conventional Southern blotting and hybridization of amplified samples were performed. Ten microliters of the
amplified product was run on 3% Nusieve/l% Seakem (FMC,
Rockland, ME) composite gels, transferred overnight to Genescreen Plus membrane (New England Nuclear, Boston, MA), and
baked at 80°C for 2 hours. Oligonucleotide probes complementary
to the junctional sequences” were 5’ end-labeled with ”P and
hybridization was performed overnight using hybridization buffer
with (for b3-a2 and el-a2) or without (for b2-a2 probe) formamide.
The membranes were washed as recommended by the manufacturer and exposed to Kodak XAR film (Eastman Kodak Co,
Rochester, NY) for 3 to 48 hours. The b3-a2 and b2-a2 probes
detect 200-bp and 125-bp amplification products, respectively,
while the el-a2 probe detects a 307-bp product.
Immune complex kinase assay. p210 and p190 can be detected
in vitro by exploiting their tyrosine phosphokinase enzymatic
activity in the immune complex kinase a s ~ a y ? The
~ , ~ antiserum
used for this assay was anti-ABL389-403,I9 a rabbit polyclonal
serum made against the predicted hydrophilic domains of v-ABL.”
The immune complex kinase assay was performed on 2 x lo7
low-density cells separated by Ficoll-Hypaque. In the case of fresh
samples, K562 cells (p210-positive CML blast crisis cell line) and
ALL-1 cells (pl90-positive, Ph-positive ALL cell lines) were used
as a positive control. To ensure that the 210- and 190-Kd bands on
the gel represented proteins recognized by the anti-ABL serum,
rather than background phosphorylation, alternate samples were
incubated with anti-ABL serum and blocking cognate ~eptide.’~.’~
DNA analysis for bcr rearrangement. Ten micrograms of DNA
were digested with restriction endonucleases in conditions recommended by the supplier (Boehringer Manheim, Indianapolis, IN),
electrophoresed on 0.7% agarose gel, blotted, and hybridized,
using a slight modification of the previously described procedure.M
The probes were labeled by oligo primer extension to a specific
activity to 2.5 x lo7 dpm/kg of DNA. After hybridization, filters
were washed once for 15 minutes at 60”C, twice for 15 minutes each
at room temperature, and once for 60 minutes at 52°C with a
solution of 0.1X SSC (1X SSC = 0.15 mol/L sodium chloride +
0.015 mol/L sodium citrate) and 0.1% sodium dodecyl sulfate,
dried, and autoradiographed. The probes were a 3’ genomic, 1.2-kb
HindIIIIBgl I1 bcr probe (bcr[PR-11) with DNA digest with BamHI
and Bgl 11, and a large “universal” bcr probe (Phl-bcr/3) encompassing the entire 5.8-kb bcr region with DNA digested with Xba
and Bgl 11. These probes detect all breakpoints resulting in p210.
They do not detect the proximal breakpoint resulting in p190.
Therapy
Patients with AML or AUL received cytosine arabinoside
(ara-C) in conventional or high-dose schedules, alone or in
combination with amsacrine or daunorubicin. One patient with
AML (patient 24) and one with AUL (patient 27) were treated
with anti-ALL therapy (Table 1).
Patients with ALL were treated with vincristine-adriamycindexamethasone (VAD)followed by long-term maintenance therapy using alternations of different chemotherapy combinations for
consolidation, maintenance, and intensification as previously described.x One patient with ALL (patient 7) received amsacrine,
vincristine, ara-C, and prednisone (AMSA-OM) (Table 1).
Statistical Considerations
Differences in the distribution of variables among different
groups were compared using the x’ test. Complete remission (CR)
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2413
PH-POSITIVEACUTE LEUKEMIA; MOLECULAR STUDIES
Table 1. Patient Characteristics, Response to Therapy, and Outcome
Patient
No.
Age
(yr)/Sex
FAB
Diagnosis
Molecular
Abnormality
Newly diagnosed
ALL
1
53lM
ALL
2
58/M
ALL
3
71/M
ALL
4
321F
ALL
5
511F
p210
p210
p210
p210
p210
bcr
Rearrangement1
Site
Karyotype
+/3'
Ph
+/3'
+/5'
+/5'
Ph other
Ph
Double Ph
other
Double Ph,
+8
Ph, -7
Ph, -7
Ph, -7
+I? Ph + other
+
+
6
62/F
ALL
p210
+ 13'
7
8
9
56lF
24lM
471M
ALL
ALL
ALL
p210
p210
p210
ND
ND
ND
10
591M
ALL
p210
+I? Double Ph
11
12
601M
341F
ALL
ALL
p210
p190
ND
13
57lF
14
631F
711M
15
16
27lF
17
46/F
18
73/M
491M
19
20
671M
38lM
21
22
631M
50lM
23
24
45lF
25
65/F
26
56lF
27
34/F
28
30lM
29
39/F
30
43lM
Refractory
31
41/M
32
531F
ALL
ALL
ALL
ALL
ALL
ALL
ALL
ALL
ALL
ALL
AML
AML
AML
AUL
AUL
AUL
AUL
ALL
p190
p190
p190
p190
p190
pl90
p190
pl90
p190
pl90
p210
p190
p190
p210
p210
pl90
p190
p210 p190
+
+/5'
AML
ALL
p210
p190
+I?
I%)
ND
ND
ND
ND
ND
ND
ND
-
-
Duration
Therapy
CR
CR
CR
CR
CR
33
23
5+
20
9
44+
38
8+
100
VAD
CR
21
40
100
29
100
AMSA-OAP
VAD
VAD
CR
PR*
PR*
0
0
t
VAD
Res
0
40
100
VAD
VAD
Res*
CR
16+
100
46
75
71
100
+
CR
CR
CR
CR
PR
Res
Res*
Res
Res
Res
IE
CR
CR
ID
+
Res*
CR
CR
CR
0
0
0
1+
35
0
0
51
20
30
Res
Res
0
0
100
100
100
25
100
100
100
100
100
100
90
100
VAD
VAD
VAD
VAD
VAD
VAD
VAD
VAD
VAD
VAD
DNR + HDAC
VAD
HDAC
AMSA HDAC
VAD
HDAC
AMSA HDAC
VAD
Ph
Ph
100
42
ADOAP
VAD
+
+
+
+
+ other
Survival
(wk)
(wk)
VAD
VAD
VAD
VAD
VAD
Ph
Double Ph
+ other
Ph, -7
Ph other
Ph other
Ph
Ph, -7
Ph
Ph
Ph, -7
Ph
Double Ph
Ph other
Ph other
Ph other
Ph other
Ph, -7
Ph, +8
Ph, -7
Ph
+
+
Response
92
72
26
85
100
+ other
-
CR
Abnormal
Metaphases
44
1
0
1+
22
29
30
0
0
0
0
64
37
50
50
31 (Died in second
CR Post-ABMT)
18
+
42
22 (Died in CR
post-A BMT)
3+
65
50
68
11+
6+
45 +
42
24
14
12
3+
58
2
35
63
55
133
34
30
Abbreviations: Res, resistant; IE, inevaluable; ID, induction death; ABMT, allogeneic BM transplantation; ND, not done; DNR, daunorubicin;
HDAC, high-dose ara-C; AMSA, amsacrine; OAP, vincristine, ara-C, and prednisone; ADOAP, doxorubicin +OAP.
*Achieved CR with subsequent therapy with methotrexate + asparaginase (patient 8) or with mitoxantrone and HDAC (patients 9, 11, 19, and 27).
tFew metaphases.
was defined as a morphologically normal marrow with the absence
of abnormal cells and the presence of normal peripheral counts
and differential, more than lo-' granulocytes/kL, and more than lo5
platelets/kL, lasting for at least 4 weeks. A partial remission (PR)
was as above, but with 6% to 25% abnormal marrow blastsz6
Survivaland remission duration curves were plotted by the KaplanMeier method,n and differences among curves were evaluated by
the log-rank test.=
RESULTS
RNA Splice Junction and bcr Rearrangement
HPA data were available on all 32 patients, and Southern
blotting of amplified product on 20 patients. In all 20 cases
in whom both studies were performed, HPA and Southem
blotting of amplified cDNA gave the same results. All
Ph-positive samples had HPA counts greater than 50,000
RLUs with the involved positive junction probes; counts
were usually less than 1,000 RLUs with the negative
junction probes. Figure 1 shows the results in 17 of these
patients in whom Southem blotting of amplified products
and HPA of amplified products were performed.
Immune complex kinase assay could be performed on
samples from 10 patients. Six patients had p190 and all of
these had the el-a2 junction. The four patients with p210
had either the b2-a2 or b3-a2 junction. Thus, HPA and
immune complex kinase assay were concordant in all 10
patients who were examined by both assays.
DNA analysis for bcr rearrangement was performed in 21
of the 32 patients. All eight patients with p210 had bcr
rearrangement, while none of the 12 patients with p190 did.
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KANTARJIAN ET AL
2414
Fig 1. Southemblattingand hybridization of PCR-amplified mmples from 17 patients. R o b e to b3-02 junction was used In (A) and to the 01-02
junction In (el.Each ampllfled product was also subjected to HPA analyals with acridinium ester-labeled probes. Results of HPA using the b3-a2
probe were: lane 1,425236 RLU; lane 2,93,366 RLU; lane 3,55,222 RLU; lane 4,751 RLU; lane 5,992 RLU; lane 6,52,143 RLU; lane 7,223 RLU; lane
8.910 RLU. Results of HPA urlng the el-a2 probe were: lane 9,556 RLU; lane 10,937 RLU; lane 11,567,789 RLU; lane 12,631,231 RLU; lane 13,
124,789 RLU; lane 14,223,431 RLU; lane 15,53,210 RLU; lane 16,789 RLU; lane 17,816 RLU.
Patient 30 with p210 and p190 had bcr rearrangement. The
results of HPA data and Southem blotting of DNA were
thus concordant in all 21 patients tested with both studies.
Among six patients in whom the bcr breakpoint was
localized, three had a 5’ breakpoint and three had a 3’
breakpoint.
Study Population and Treatment Outcome
A total of 32 adults with Ph-positive acute leukemia were
investigated. Thirty adults were untreated and two had
failed prior induction therapy (Table 1). Their median age
Table 2 Comlation Between Molecular Abnormalities and Other
Variables In N W D l a g n d Patients
No. of Patients ( W )
Characteristic
Age (VI
Splenomegaly
Hepatomegaly
Hemoglobin (g/dL)
WBC (XlO’lpL)
Platelet count
( x 1ovpL)
Marrow blasts
(%I
FAB morphology
Karyotypic
abnormality
Ph-positive
metaphases
Albumin (g%)
Lactic
dehydrogenase
(UIL)
Response to
therapy
Median remission
duration (wk)
Median survival
(wk)
Category
p210
(N = 14)
P1W
IN = 15)
P
Value
was 53 years (range, 24 to 71 years). Twenty-four patients
(75%) had ALL, four (12.5%) had AML, and four (12.5%)
had AUL. P190 was detected in 16of the 32 patients (SO%),
including 12 of 24 patients with ALL (50%), two of four
patients with AML (50%), and two of four patients with
AUL (50%). P210 was found in 15 patients: 11 of 24 ALL
(46%), two of four AML (50%), and two of four AUL
(50%). One patient with ALL (patient 30) had both p190
and p210.
With induction therapy, 17 of the 30 (57%) newly
diagnosed patients achieved CR, and three (10%) had PR.
CR was observed in 13 of 23 patients with ALL (57%), two
of three patients with AML (66%), and two of four patients
with AUL (50%). Five additional patients with ALL obtained CR after methotrexate-asparaginase (patient 8) or
mitoxantrone and high-dose ara-C (patients 9, 11, 19, and
27), yielding an overall response rate of 78% in ALL (18 of
23 patients).
The median remission duration of the 17patients achieving CR was 22 weeks. The median overall survival was 48
weeks.
7(47)
1(7)
2(13)
lO(67)
4(13)
7(47)
6(40)
NS
NS
NS
NS
20-50
>50
9(64)
4(29)
2(14)
8(57)
5 (36)
3(21)
6(43)
NS
Correlationsof Molecular Abnormalities Wth Patient
Characteristics and Prognosis
<50
7 (50)
7(47)
NS
>50
ALL
AML
AUL
12(86)
11 (79)
l(7)
2(14)
15(100)
11 (73)
2(13)
2(13)
NS
There were no statistically significant differences in age,
degree of splenomegaly or hepatomegaly, Occurrence of
anemia leukocytosisthrombocytopeniaor blastosis, or albu-
Ph
Phi -7
Ph + other
3 (21)
4 (29)
7 (50)
4(27)
4 (27)
7 (47)
NS
< 100%
7(50)
6(43)
6(40)
6(40)
NS
NS
>50
Any
Any
< 10
<20
NS
Pk-P&r.
Tola R&PW
---- *
?
<3.5
-8
.-I
80-
-
lO(71)
8(53)
NS
7(50)
9(60)
NS
.
PZlO
6
PlW
I Wan-f.Lr.
I
60
-
I2
p = .08
0
0
5u
40-
2
>600
6
I
a
5a
Or-
n
20
-
L,,,--I
Complete
Abbreviation: NS, not significant.
I
I
0
21
29
.08
0
12
24
36
48
60
Remlsrlon Duratlon (Weeks)
40
55
.18
Fig 2 Remission duration in patients with Ph-po~ltiveacute
leukemia by molecular abnormality.
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2415
PH-POSITIVEACUTE LEUKEMIA; MOLECULAR STUDIES
cytes, 14% metamyelocytes, 3% bands, 71% polys) 1month
after achieving CR and for 1 month before recurrence of
acute leukemia. Her BM at that time showed 80% cellularity and only 1% blasts with a CML morphologic pattern.
Cytogenetic studies showed 100% Ph-positive metaphases.
None of the nine patients with p190 had a similar course
after achievement of CR.
CorrelationsBetween Cytogenetic and Molecular Studies and
Other Variables
Fig 3. Survival of patients with Ph-positive acute leukemia by
molecular abnormality.
min and lactic dehydrogenase levels between the two
groups of newly diagnosed patients with p190 or p210
(Table 2).
CR rates were also similar among patients with p190 or
p210 disease (60% v 50%; P not significant). Although the
remission duration was possibly longer in p190 acute
leukemia (P = .08, Fig 2), any differences were minor
(median CR duration 29 v 20 weeks) and were not paralleled by differences in survival rate (Fig 3).
Among patients achieving CR, one of seven patients
(14%) with p210 reverted to a chronic phase after remission
induction. This patient (patient 4) showed leukocytosis up
to a highest W C of 65.9 x 1@/pLwith a CML differential
1month after achieving CR (2% promyelocytes, 7% myelo-
Nine of our 32 patients (28%) had the Ph chromosome as
the only cytogenetic abnormality, eight (25%) had the Ph
chromosome and monosomy 7, and 15 (47%) had the Ph
chromosome and other additional abnormalities. These
occurred with equal frequency with p210 or p190 (Table 2).
Fourteen of the 29 newly diagnosed patients with p210 or
p190 had Ph and other abnormalities: a double Ph was
found in four patients, trisomy 8 in one, and both in one.
Four of the 14 patients (29%) with p210 had such abnormalities, compared with 2 of 15 (13%) with p190 (P not
significant). One of the seven patients with AML or AUL
had residual normal metaphases compared with 12 of 21
evaluable patients with ALL (14% v 57%; P = .05).
Ten patients had cytogenetic studies in remission, four
(40%) showing only normal metaphases. These included
two of six patients with p210 and two of four patients with
p190. The incidences of lymphoid markers were similar
with p190 and p210 (Tables 3 and 4). Myeloid markers were
present in 7 of 10 patients (70%) with p190 and in three of
eight (37%) with p210. There was no difference in response
rates within the p190 and p210 subgroups by the presence
or absence of myeloid markers.
Table 3. CorrelationBetween Immunophenotype, Ph Molecular Abnormality, and Response in ALL
Percent of Cells Expressing Marker
Patient
Molecular
Abnormality
Response
1
2
p210
p210
CR
CR
3
4
p210
p210
5
6
7
8
9
10
11
12
p210
p210
p210
p210
p210
p210
p210
p190
13
14
15
p190
p190
p190
CR
CR
CR
16
17
18
19
20
21
p190
p190
p190
p190
p190
p190
CR
22
PI90
No.
Myeloid
Markers
CD14
(MY4)
CD13
(MY71
CD33
(MY91
CD34
(MY10)
CDlO
(CALLA)
CD19
CD20
-
-
39
84
83
91
83
83
31
CR
CR
+
31
46
91
81
93
CR
CR
CR
PR
PR
Res
+
-k
-
-
-
-
-
ND
ND
ND
ND
ND
ND
Res
-
CR
-
-
26
-
-
-
-
-
-
-
-
21
-
-
-
+
-
-
-
-
73
48
ND
ND
ND
49
ND
ND
ND
-
-
-
36
-
ND
ND
ND
24
92
89
90
-
-
47
90
-
49
Res
Res
+
+
+
+
+
-+
32
23
36
97
47
69
93
86
90
ND
Res
ND
ND
ND
ND
ND
PR
Res
Res
Abbreviations: Res, resistant: ND, not done: -, negative.
-
-
-
-
56
98
ND
ND
ND
88
93
99
97
98
ND
53
ND
87
93
99
93
90
98
95
73
90
93
89
91
98
95
85
95
95
55
ND
ND
ND
-
88
ND
40
ND
64
22
56
90
23
-
-
ND
From www.bloodjournal.org by guest on February 6, 2015. For personal use only.
2416
KANTARJIAN ET AL
Table 4. Correlations Between Immunophenotype and Molecular
Abnormality in ALL
No. PositivelNo. Tested (%)
lmmunophenotype Positivity
p210
PI90
CDlO
CD19
CD20
CD34
CD33
CD13
CD14
Positive myeloid markers
719 (78)
718 (87)
619 (66)
518 (62)
318 (37)
218 (25)
018 (0)
318 (37)
l o l l 0 (100)
9/10 (90)
3/10 (30)
819 (89)
3/10 (30)
5/10 (50)
0110 (0)
7/ 10 (70)
DISCUSSION
The distinction between de novo Ph-positive acute leukemia and Ph-positive CML in blastic phase presentation is
difficult to determine. Most patients with CML develop a
blastic phase, indistinguishable morphologically from acute
leukemia, and some of them could conceivably present in
blastic phase after a clinically silent or undiagnosed chronic
phase. This finding may in part explain the seemingly high
incidence of Ph-positive "ALL" in adults compared with
children, because CML is more common in the adult
population. It was hoped that the availability of Ph-related
molecular studies would increase our understanding of the
biology of Ph-positive acute leukemia, solve the above
diagnostic dilemma (CML blastic phase v de novo Phpositive acute leukemia), and provide prognostic correlations among patients with p210 or p190 disease. That p190
and p210 may discriminate between different acute leukemia subsets is suggested by (1) in vitro studies showing p190
to be a more active tyrosine kinase than ~ 2 1 0 , '(2)
~ the
consistent association of CML with p210 only, and (3) our
present ignorance of whether p190 and p210 activate the
same substrate. It was also thought that p210 might be a
specific marker for CML blastic phase presentation, identifying patients who are likely to have more frequently
observed features in CML (1) 100% Ph-positive metaphases
on cytogenetic analysis; (2) additional chromosomal abnormalities characteristic of CML blastic phase (double Ph,
isochromosome 17, trisomy 8); (3) persistence of Phpositive cells in remission; (4) development of a second
chronic phase after remission induction; and (5) possibly a
worse prognosis.
This is the first study to investigate systematically these
questions in a large number of patients with Ph-positive
acute leukemia, treated uniformly for the particular morphology, in a single institution. The incidence of p190
disease in our adult population was SO%, similar to that
reported by others, but lower than the 80% incidence
reported in children with Ph-positive acute leukemia3043
(Table 5). In contrast, p210 disease is more frequent in
adults with Ph-positive acute leukemia. In Ph-positive
AML or AUL, with the small number of patients studied,
the incidence of p190 disease is 50%, as it was for adult
ALL (Table 5).
There were no significant clinical, laboratory, or karyotypic differences in patients with p210 versus p190 (Table
2). If patients with p210 had CML in blastic phase presenta-
tion rather than de novo Ph-positive acute leukemia, we
would have expected to observe in them a higher frequency
of the features associated with CML as listed above. The
molecular studies were also not helpful in distinguishing,
Table 5. Summary of the Molecular Studies in Ph-Positive Acute
Leukemia
No. of Cases'
Study
ALL
Adult
Bartram"
Chen et aV7
Hermans et al"
Erickson et ,Iu
Clark et a17
Denny et al"
Schaefer-Rego et al"
Dreazen et a P
Secker-Walker et ala
Kurzrock et aI4l
Kurzrock et aI6
Present study
Total
Pediatric
Bartram et alp
Chen et a13'
Clark et al'
Hermans et al"
Rodenhuis et aIJ2
Erikson et alu
Denny et al"
Dreazen et all9
Secker-Walker et at"
Total
Age unspecified
Hooberman et aIs
Saglio et al'O
Total
AML
Kurzrock et aP2
Bartram et al"
Hermans et aI3l
Erikson et a133
Chen et a137
Najfeld et al"
Saglio et al"
Present study
Total
No. of
Patients
17
4
5
3
1
p190
Disease
4
2
4
1
1
3
4
p210
Disease
13
2
1
2
-
3
9
2
10
4
2t
24$
82
It
It
12
39 (48)
11
42 (51)
9
3
2
4
2
2
7
3
2
34
7
2
2
4
2
2
7
1
1
28 (82)
2
1
2
1
6 (18)
9
8
17
5
4
9 (53)
4
4
8 (47)
It
3
1
1
6
1
2
4
18
6
2
5
2
4
2
-
It
-
2
1
1
1
3
3
1
1
2
9 (50)
1
2
9 (50)
AUL
Erikson et alu
Najfeld et at36
Present study
Total
1
1
4
-
1
1
2
-
6
3 (50)
3 (50)
2
Percentagesgiven in parentheses.
*Patients categorized based on either DNA, RNA, or protein studies.
tPatients are included in present study and are not counted in the
total number.
*One patient expressed both p190 and p210.
From www.bloodjournal.org by guest on February 6, 2015. For personal use only.
2417
PH-POSITIVEACUTE LEUKEMIA; MOLECULAR STUDIES
within Ph-positive acute leukemia, patients with different
presentations or prognoses. Patients with p190 and p210
disease had similar incidences of older age, splenomegaly,
hepatomegaly, anemia, leukocytosis, thrombocytopenia, and
blastosis (Table 2). The incidence of residual diploid cells
at diagnosis and in remission, and of karyotypic abnormalities such as monosomy 7 or double Ph/trisomy 8, were also
similar in the two subgroups. Prognostically, the CR rates to
induction therapy were not different with p190 and p210.
While patients with p190 disease had a trend for longer
remission durations, this was not statistically significant
(P= .08) and did not translate into significant survival
improvement. This finding suggests that the disease process
in Ph-positive acute leukemia is not influenced by the
different molecular abnormalities (p190 v p210), and that,
having produced disease evolution into the acute phase,
they are of little relevance to subsequent outcome.
In a small series of 10 adult patients with Ph-positive
acute leukemia, Secker-Walker et alQ also did not identify
significant associations between the molecular abnormalities and patient age, leukocyte count, or FAB type. Although the specific treatments were not detailed, no obvi-
ous differences in response to therapy or in prognosis were
seen. They also found a similar occurrence of normal
metaphases and of additional karyotypic abnormalities in
patients with or without bcr rearrangement. A mixture of
Ph-positive and diploid metaphases were noted in two of
five patients with bcr rearrangement and in five of seven
patients without bcr rearrangement (40% v 71%); a double
Ph or trisomy 8 were found in one patient and in two
patients in the two groups, respectively (20% v 29%). In
contrast to their study in which myeloid markers were
present more often in patients with bcr rearrangement, the
incidence of myeloid marker positivity was similar among
our patients with p190 or p210.
In summary, the present investigation of the different
messages in Ph-positive acute leukemia fails to show any
significant correlations with patient clinical characteristics
or laboratory features. Furthermore, the presence of p190
versus p210 does not appear to be prognostically relevant
within the context of our presently available treatment
modalities. In the future, improvement in the therapy (eg,
BM transplantation) may provide a differential benefit in
these two molecularly distinct entities.
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1991 78: 2411-2418
Significance of the P210 versus P190 molecular abnormalities in
adults with Philadelphia chromosome-positive acute leukemia
HM Kantarjian, M Talpaz, K Dhingra, E Estey, MJ Keating, S Ku, J Trujillo, Y Huh, S Stass and R
Kurzrock
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