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Chromosomal characteristics of Ph-positive chronic myelogenous leukemia in transformation
Chromosomal Characteristics of Ph-positive Chronic Myelogenous Leukemia in Transformation A Study of 23 Chinese Patients in Taiwan Hwei-Fang Tien, Sou-Ming Chuang, Chiu-Hwa Wang, Fen-Yu Lee, Su-Hui Chien, Yao-Chang Chen, Ming-Ching Shen, and Chen-Hui Liu
ABSTRACT: Cytogenetic study was performed in the past 3 years on 23 Chinese patients with Philadelphia chromosome (Phi positive chranic myelogenaus leukemia (CMI,) in transformation; seven ,.','ere in accelerated phase and 16 in acute blast crisis. ('hramosomal abnormalities in addition to Ph were found in three (43%) of the patients at accelerated phase and 14 (88%) of the patients at blast crisis. The common nonrundam chrnmosarnal aberratians were double Ph, trisomy 8. trisomy 19, and trisomy 21, which occurred in 47%, 41%, 35%, and 29%. respectively, af the total patients with extra chromasomal abnormalities. Isochramosamc for the long arm of chromosome 17 was flmnd in only one patient. In patients with blast crisis, the type of blast cell was characterized through morphalogic, cytochemical, and immunacytochemicaJ studies. Eleven cases were classified as myelaid and five as lymphoid transfiJrmation. Trisomy 8, 19, and 21 were detected only in patients with rnyeloid blast crisis. This study also revealed a high incidence oJ trisamy 21 and a low incidence of i(t7q) in Chinese patients with tmnsfarnmtian af CML.
INTRODUCTION For t h e m a j o r i t y of p a t i e n t s w i t h P h i l a d e l p h i a c h r o m o s o m e (Ph) p o s i t i v e c h r o n i c m y e l o g e n o u s l e u k e m i a (CML), t h e d i s e a s e u s u a l l y u n d e r g o e s t r a n s f o r m a t i o n w i t h i n a m e a n of 3 y e a r s after d i a g n o s i s I1, 2]. In m o r e t h a n 75% of p a t i e n t s , t r a n s f o r m a t i o n is a c c o m p a n i e d by k a r y o t y p i c e v o l u t i o n w i t h d e v e l o p m e n t of e x t r a c h r o m o s o m a l a b n o r m a l i t i e s , m o s t c o m m o n l y t r i s o m y 8, i(17q), or d u p l i c a t i o n of t h e P h (3]. Mit e l m a n a n d L e v a n first n o t e d u n e v e n g e o g r a p h i c d i s t r i b u t i o n of n o n r a n d o m a n d s p e c i f i c c h r o m o s o m e a b e r r a t i o n s in m a l i g n a n t d i s o r d e r s 14]. In a m o r e e x t e n s i v e r e v i e w , M i t e l m a n f o u n d t h a t t h e i n c i d e n c e of + 8 , i(17q), a n d + P h in a c u t e blast
l-'rom the Department of Internal Medicine (H. F. T., C H. I..), Pathoh~gy (S. M. C.. F. Y. 1..) and Clinical Pathology (C. H. W.. S. It. C., Y. C. C.. M. C. S.), National Taiwan University Hospital, Taipei, Taiwan, Republic of China. Address reprint requests to: Hwei-Fang Tien, M.D., Department of Internal Medicine, National Taiwan University" ltospital, No. 1 Chang-Te St., Taipei, Taiwan, Republic of China. Received October 4, 1988; accepted December 26, /988.
89 ~J 1989 Elsevier Science Publishing Co., Inc. 655 Avenue of the Americas, New York, NY 10Ill0
Cancer Genet Cytogenet 39:89-97 (1989) 0165-4608:89/503.50
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tl. F. Tien et al.
crisis of CML varies greatly in different areas ]5]. In blast transformation, the blasts can be either lymphoid or myeloid [6, 7]. The nature of the additional chromosomal abnormalities was also found to be different for these two phenotypes of blasts involved in the transformation [8, 9]. There have been few reports concerning karyutypes of Chinese patients with transformation of CML in the past. This report presents results of chromosome study for 23 Chinese patients with CML in transformation. Blast phenotypes were also characterized by morphology, cytachemistry, and immunocytochemistry in 16 patients with acute blast crisis who had more than 30% blasts in the bone marrow or more than 20% blasts in the peripheral blood and were correlated with the chromosomal abnormalities. MATERIALS AND METHODS
Cytogenetic study was performed on 23 unselected Chinese patients with Ph-positive CML in transformation at National Taiwan University Hospital from November 1985 to September 1988. Sixteen patients were in acute blast crisis and seven in accelerated phase. The criteria for diagnosis of blast crisis was the presence of more than 30% blasts in the bone marrow or more than 20% blasts in peripheral blood. Those patients in accelerated phase had clinical and hematologic deterioration suggestive of transformation but did not manifest the criteria for blast crisis [10]. Bone marrow (BM) ()r peripheral blood (PB) cells were harvested directly or after 1-3 days of unstimulated culture. The metaphase chromosomes were banded by the conventional trypsin-Giemsa banding technique [11] and then karyotyped according to the ISCN [12]. In patients with acute blast crisis, the phenotype of blasts was characterized through morphologic, cytochemical, and immunocytochemical studies. Cytochemical staining, which included myeloperoxidase, Sudan black B, periodic acid-Schiff. u-naphthyl butyrate esterase, and chloroacetate esterase staining, were performed on air-dried smears of BM aspirates or PB as previously described [13]. Immunocytochemical studies were carried out on blast cells isolated from BM or PB by FicollHypaque gradient centrifugation by indirect alkaline phosphatase method [14, 15] with a panel of monoclonal antibodies (Table 1). The enzyme terminal deoxynucleotidyl transferase (TdT) was shown by a peroxidase antiperoxidase method [16] on cytocentrifuge smears. The blastic transformation was considered myeloblastic when blasts were stained positively with peroxidase, Sudan black B or monoclonal antibodies My-7, My-9, or OKM-1; megakaryoblastic when they were positive for Table 1
Specificity of monoclonal antibodies
Antibody HI,A-DR (la) B4 J5 (CALLA) B1
Leu-9 Tll Leu-1 OKM-I My-7 My-9 HPI-ID
Specificity B lymphocytes, activated T lymphocytes, monocytes, non-T-ALL and AML blasts B lymphocytes Non T-ALL, Burkitt lymphoma, follicular lymphoma, some T-ALL B lymphocytes Pan-T lymphocytes Pan-T Iymphocytes (Sheep erythrocyte receptor) Pan-T lymphocytes Granulocytic, monocytic series and large granular lymphocytes Monocytes and Pan-myeloid cells Monocytes and myeloid precursors Megakaryocytes. platelets
Chromosomes of CML in Transformation
91
H P I - I D : and l y m p h o i d when the.,,, were peroxidase negative, TdT positive, and reacted with l y m p h o i d markers such as J5, B4, B1, or Leu-9.
RESULTS Seventeen (74%) of the 23 patients showed chromosomal abnormalities in addition to the Ph. Among them, the c o m m o n nonranctom chromosomal aberrations were +Ph, +8, +19, and +21 with an incidence of 47%, 41%, 35%, and 29%, respec'tively. Twelve (71%) of the 17 patients had either one of t 8 , +Ph, ~-19, or - 21, or most frequently two or more in combination, lsochromosome for the long arm of chromosome 17 was found in only one patient (case 19). Cytogenetic: study was also done at chronic phase for 13 patients including four or the five with trisomy 21: all showed simple Ph without other abnormalities. The detailed chromosomal changes at transformation are shown in Tables 2 and 3. Of the 16 patients in blast crisis, 11 had myeloid transformation (including myeloblastic: and megakaryoblastic) and five had l y m p h o i d transformation. All the patients with myeloid blast crisis had extrachromosomal abnormalities. Eight patients had - P h , six had - 8 , six had - 1 9 , and five had ~21 (Table 2). In ten, a h y p e r d i p l o i d clone was present, and none had a h y p o d i p l o i d clone. Three of the five patients with l y m p h o i d blast crisis showed additional chromosomal changes. One patient had +Ph but none had +8, -' 21, or +19 (Table 2). A h y p o d i p l o i d clone was present in one patient and a h y p e r d i p l o i d clone in another one. Isochromosome of 17q was not found in any patient in acute blast crisis of CML although other kinds of abnormalities involving chromosome 17 were present in three patients: two resulted in m o n o s o m y 17p (Figs. 1 and 2) and another in trisomy 17 (case 2). The mean survival after diagnosis of acute blast crisis was 11 months for patients with lymphold transformation and 3 months for those with myeloid transformation. Three of the seven patients in accelerated phase had extra chromosomal abnormalities. One showed i(17q), one - 1 7 , ~-r(4): and the remaining one t(3;7)(q26;q21) (Fig. 3), who had marked thrombocytosis with platelet c:ount of 2.1 × 10~//.d.
DISCUSSION Transformation of Ph-positive CML is usually accompanied by karyotypic evolution [3]. The most frequent chromosomal aberrations are +8, ,-Ph, and i(17q) [3]. The cells derived from the new clone have a growth advantage over both normal cells and the chronic-phase Ph-positive clones [10]. Our data revealect that 74% of the patients with CML in transformation (43% of those in accelerated phase and 88% of those in acute blast crisis) had chromosomal abnormalities in addition to the Ph. This incidence is comparable to that reported from other centers [3, 17]. However, some different characteristics of the c o m m o n extra chromosomal aberrations were found in this study. Among the 23 patients in transformation, eight had -~Ph, seven had ~ 8, six had --19, and five had +21, respec:tively. The occurrence of +21 was unexpectedly high (22')/0 of the total patients and 29% of the patients with additional chromosomal abnormalities] as compared with that in other areas [8, 9, 18-20]. Clinically, all five patients showed normal appearance and mentality without evidence of Down syndrome, and the karyotype of the four patients who had also been studied cytogenetically in chronic phase revealed a simple Ph without +21 at that time. On the other hand, the incidence of i(17ct), which is usually around 25% in other centers [3, 20, 21], was rather low in our series (7°/,,). Mitelman and Levan [4] first noted uneven geographic: distribution of n o n r a n d o m
92 Table 2
C y t o g e n e t i c r e s u l t s i n 16 p a t i e n t s w i t h C M L in a c u t e b l a s t c r i s i s
(;as(: number
Age/Sex
Blast p h e n o t y p e
3 '~
46;M
Myeloblastic
4
45,'F
Myeloblastic
9°
34/M
Myeloblastic
10
35/M
Myeloblastic
13
50/M
Myeloblastic
14 ~'
47/F
Myeloblastic
15"
42;M
Myeloblastic
7°
40/F
Megakaryoblastic
1"
43/M
12"
26,M
Myeloblastic -:megakaryoblastic Myeloblastic megakaryoblastic
16
26/F
2"
32/M
Myeloblastic lmegakaryoblastic I.ymphoid
5 6"
36/M 12/M
Lymphoid l.ymphoid
8
54/F
Lymphoid
11"
26/M
Lymphoid
N u m b e r of cells:Karyotype 17/46.XY.t{9;22}[q34:ql )1 l : 5 0 . X Y . - 4 . ~-13.41,q. ~ 21. + I ) . l q - . 7 p - , t [ 9 : 2 2 ) . - Ph 2."46,XX.t[9:22)Iq34;q 11 ) 22.'47,XX. - 8,t[9:22) 11."46,XY,t(9:22](q34;q I 1 ) 6/48,XY,*8,t(9:22h • Ph 3;46,XY,'8, 10,t(9;22) 2;48,XY, - 8 , - 19.t[9:22) 1"47.XY,+ 8 , - 10,t(9:22),.- Ph 2"46,XY,t(9;22)[q34;q 111 3,"46.XY.2q ÷ .t[9;22/ 4,"50,XY, ÷ 8,+ 10,+ 12,t19:22),- Ph 1;46,XY,t19;221{q34;q 11 ) :);47,XY, * 8,t(9;22) 1,'47,XY,t(9:221, + Ph 14,'49,XY. ~-8, 4 19.* 21,t(9;22) 6,'50,XY. + 8,4- 19, ~-21 ,t(9;22),- Ph 18."46,XX,t(9;22)(q34:q 11 } 7.;49,XX, + 8, + 19,4- 21 I(9:221 18/46,XY,t(9;22}(q34.q 11} 1,'47,XY,t(9:22), + Ph 2.'48, 4. 6,t(9;22). ~-Ph 23/46,XX,t(9;22][q34;q11 ) 2,'47,XX. 7 , ' 8 , + 2 1 . t [ 9 ; 2 2 ) 26."47,XY. * 21 ,t(9;22 )(q34;q 11 ) 8,'46,XY,t(9:22J(q34;q I 1 ] I 0/47,XY, + Ph,t(9;22J 1 ' 4 9 , X Y , - 1 , 4 2 , - 7 , - 8,+8, - 1 0 , - 1 2 , - 1 3 , 17. 2 0 . - 21 ,del[20)(q 11 ),t(9:221,~- der( 17J,t( 7;?: 17J ( 7 q t e r - . 7 q 11 ::?:: 17 p 11--~ 17qte.r)..'- Ph.-*- Ph 1/54,XY,+ 1 , - 2 , + 6 , - 7 . + 8 , + 8 , - 10, + 12,+ 19,t(9;22), + der(17),+ P h , + Ph 6/46.XX.del(7J(p 15),t(9:22}(q34;q 111 9i48,XX,+ 19.del[7J,t(9:22J,+ Ph 19i46,XY,t(9;22)(q34;ql 1 ),t(10;16)(q22;ql 3J,t[14:22) (q23;q13) 7;49,XY, + 15, + 17,t(9;22J,t(10;16),t(14:22), ~-Ph 26/46,XY,I(9;221(q34;q 11 J 15/46,XY,t(9;22}[34:q 11 ) 3/46.XY,-- 17,t(9;22), ~-der(l )t(1:17)(pl 1 ;ql 1 ) 15/46,XX,t(9;22)(q34;q11 ) 6/45,XX, 7,t(9;22) 26146,XY,t(9:22)(q34:ql 1 )
" The karyotypes of these~ patients had been studied in (:hronic phase and all showed simph: Ph :vithout additional change.
Chromosomes
o f C M L in T r a n s f o r m a t i o n
Table 3
93
C y t o g e n e t i c r e s u l t s in s e v e n p a t i e n t s w i t h C M L in a c c e l e r a t e d p h a s e
Case. n u m b e r
Age/Sex
Number of cells/karyotype
17
58:M
18"
35/M
2,'46,XY,t(9;22)((134;q 11 ) 13,,'46,XY.t(3;7)lq26:q211,t(9;221 3:45.X,-. Y,t(3:7],t(9:22) 1/46,XY 1/46.XY,t(9;22)(q34;ql 1 ) 22,'46,XY, - 17.4- r(4),t(9:22)
19" 2(1 21" 22 23
35,'F 65iF 27iM 38/M 12/M
9/46,XX,i(17q),t(9;22}(q34:q11) 26/46,XX,t(9:22}(q34:q 11 ] 7/46,XY,t(9:22)(q34:ql 1 ) 20/46.XY,t(9;22)(q34:q 11 ) 12/46.XY,t(9;22)(q34:q111
° The karyotypes of these patients had been studied in chronic phase and all showed simple Ph without additional change.
F i g u r e 1 Karyotype fronl one bone marrow (:ell of case 12, s h o w i n g 49,XY. ; 1. ~ 2, 7.+8, ÷ 8. -10,+12,-13, 17. 2l~de~l(2~((|~.t~9~22~((]34~q~)`(ier(~7~(7:`~7}I7qte`r~7q~?::~7p~7~ter + Ph, + l'h.
=
:.
@
D,
""
\
""
i¢
"ll
3
4
5
2 ~
ii
Jv
6
7~
• 8 \
I
9
~(
•
\ 10
11
12 ~
k
!
J~
\
13
15
14
16
/e 17
18 4
|
W(
•
19
2O
~1~,
22
X Y
],
94
tt. F. 'l'ien e,t al.
1
17
9
22
d*d~),t(l:~ 7) F i g u r e 2 Partial karyotypes from two bone marrow c:ells of (:ase 6, showing -17,+der[1) t(l;17)(pllzqll), in addition to Ph chromosome.
F i g u r e 3 Partial karyotypes from two bone marrow (:ells of case 17 who had thrombo(:ytosis, showing t(3:7)(q26;q21) and Ph chromosome.
3
7
9
22
tlL~"
- I1 Jm
t(3:7)
-
t(S~2)
Chromosomes of CML in Transformation
95
and specific (:hromosome aberrations in malignant disorders. At the time of blast crisis of Ph-positive CML, Milehnan [5] foun(t that the incidence of the three most common additional abnormalities varies in different geographic areas: r8 from 21% in Sweden and New York to 46% in Japan, • Ph from 7% in the t;.S.S.R, to 54% in France, and i(17q) from 7% ira Sweden to 28% in the U.S.S.R. Trisomv 21 was not included in the study. As a whole, the incidence in lapan was most similar to ours: 46% for - 8 , 41.8% for - P h , and 15.2% for i{17q) in ]apan and 41%, 47%, 6%. respectively, in our series. The reason for this apparent geographic heterogeneity in the incidence of n o n r a n d o m chromosome aberrations is still u n k n o w n . Mitehnan 151 suggested differences ira the distribution of etiologic factors or genetic:ethnic hackground might be the possible explanation. More extensive studies in various t:ountries and areas are necessary to clarify, this issue. A total of five patients had abnormalities involving chromosome 17, which resuited in monosomy 17p in four of the five (cases 6, 12, 18, and 19). These, were i(17q), t{1:17}{p11:q11), a complex translocation involving #7, #17, and an u n k n o w n chromosome, and monosomy 17. Borgstrom et al. [221 suggested monosomv 17p might represent a crucial event in the terminal phase of CML. The aberration t{1;17)(pll;q11) was the same as that reported by Kerman el al. [231. They reviewed six such cases, two from their laboratory and four from the literature and concluded that this marker might be specific for the accelerating and blast phase of CMI,. In acute blast crisis of CML, blast cells can be either mw',loid or lymphoid [6.71. Rowley [24} noted that no patient with Ph-positive lymphoid dise, ase was reported to haw.' ~ 8, i(17ql, or +19. while most patients with n)yeloid disease had one or more of these abnormalities. 'rwo later reports [25, 26] did not support her findings. Blast phenotype (:an no,,',' be characterized more act:uratelv by (:vtochemical and (:omprehensive immunologic marker studies [6]. A correlation between extra chromosomal abnormalities and the type of blast crisis has emerged more (:learlv. Parreira et al. 181 found that +8, i{17q), anti - 1 9 were seen only in patients with myeloid but not lymphoid blast crisis of CMI,. Diez-Martin et aI. [9] reported similar findings. They also noted an association between abnormalities of chromosome 7 anti lymphoid blast crisis. Our data showed that 82% of the patients with myeloid blast crisis had ÷8, + 19 or both, while none of those with lymphoid transformation (lid. lsochromosome of 17q was seen in neither of them. It is of interest that • 21 ,,','as also present only in patients with myeloid crisis. Of the 18 cases in lymphoid crisis of CML ret)orted by Yao 127], six from his institute and 12 from the literature, none had ~ 21. It was also absent in the eight cases with lymphoid transformation reported by DiezMartin el al. [9]. However Parreira el a]. [8] found that one of their six patients with lymphoid blast crisis had ~21, though it is not known whether - 2 1 in this patient reflects constitutional change or not. It may be suggested that acquired +21 is a rare marker if not totally absent in lymphoid transformation of CMI,. Abnormalities of chromosome 7 in our study were present in both myeloid and lymphoid transformation in contrast to the previous reports [8, 9]. No definite (:nrrelation was found between the subtype of myeloid transformation and chromosomal abnormalities [9]. In one study [8} +19 was seen only in [)last crisis involving megakaryoblasts, which was not the case in the other report [91. In our series, ,19 was present not only in transformation involving megakaryoblasts but also in myeloblastic transformation. More studies are needed to clarify this issue. The authors thank Dr. 1. Whang-Peng for he,r kindness in checking the karyotypes. Dr. I,T Yam and Dr. CY Li for instruction in immunocytochemical staining, and Dr. Wl. Nichols for sending us monoclonal antibody HP1-1D. Supported in part by a grant from the National Science Council of the Republic of China, NSC 75-0412-B01}2-107 and 76-0412-B0112-121.
96
H. 1.'. Tien et al.
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23. Kerman SL. Miller RB, Heritage DW 11986]: Translocation (1:17) in accelerating and blast phases of chronic myelogenous leukemia, Cancer Genet Cytogenet 20:269-277. 24. Rowley JD (1980): Ph-positive leukemia including chronic myelogenotls leukemia. Clin Haematol 9:55-86. 25. Alimena G. Dallapiccola B, (_;astaldi R. Mai~delli t:. Brandt I.. Mitelman t", Nilsson P{; (1982): Chromosomal, morphological and {:linical (:orrelations in blast crisis of chronic mveloid leukemia-, a study of 69 cases. Stand I Haematol 28:103-117. 26..Muehh.~ck Si'}, M(:Kenna RW. Arthur DC. Parkin ]I., Brunning RD (1984): Transformation of chroni{: myelogenous leukemia: (21ini(:al. Inorphological. and t:ytogenetic features. Am ] Clin Pathol 8 2 : 1 - 1 4 . 27. Yao E (1986): Chromosomal abnormalities in ]ymt}hoid crisis of chrmlic myelogenous leukemia. Cancer J Cytogenet 16:235-243.
ABSTRACT: Cytogenetic study was performed in the past 3 years on 23 Chinese patients with Philadelphia chromosome (Phi positive chranic myelogenaus leukemia (CMI,) in transformation; seven ,.','ere in accelerated phase and 16 in acute blast crisis. ('hramosomal abnormalities in addition to Ph were found in three (43%) of the patients at accelerated phase and 14 (88%) of the patients at blast crisis. The common nonrundam chrnmosarnal aberratians were double Ph, trisomy 8. trisomy 19, and trisomy 21, which occurred in 47%, 41%, 35%, and 29%. respectively, af the total patients with extra chromasomal abnormalities. Isochramosamc for the long arm of chromosome 17 was flmnd in only one patient. In patients with blast crisis, the type of blast cell was characterized through morphalogic, cytochemical, and immunacytochemicaJ studies. Eleven cases were classified as myelaid and five as lymphoid transfiJrmation. Trisomy 8, 19, and 21 were detected only in patients with rnyeloid blast crisis. This study also revealed a high incidence oJ trisamy 21 and a low incidence of i(t7q) in Chinese patients with tmnsfarnmtian af CML.
INTRODUCTION For t h e m a j o r i t y of p a t i e n t s w i t h P h i l a d e l p h i a c h r o m o s o m e (Ph) p o s i t i v e c h r o n i c m y e l o g e n o u s l e u k e m i a (CML), t h e d i s e a s e u s u a l l y u n d e r g o e s t r a n s f o r m a t i o n w i t h i n a m e a n of 3 y e a r s after d i a g n o s i s I1, 2]. In m o r e t h a n 75% of p a t i e n t s , t r a n s f o r m a t i o n is a c c o m p a n i e d by k a r y o t y p i c e v o l u t i o n w i t h d e v e l o p m e n t of e x t r a c h r o m o s o m a l a b n o r m a l i t i e s , m o s t c o m m o n l y t r i s o m y 8, i(17q), or d u p l i c a t i o n of t h e P h (3]. Mit e l m a n a n d L e v a n first n o t e d u n e v e n g e o g r a p h i c d i s t r i b u t i o n of n o n r a n d o m a n d s p e c i f i c c h r o m o s o m e a b e r r a t i o n s in m a l i g n a n t d i s o r d e r s 14]. In a m o r e e x t e n s i v e r e v i e w , M i t e l m a n f o u n d t h a t t h e i n c i d e n c e of + 8 , i(17q), a n d + P h in a c u t e blast
l-'rom the Department of Internal Medicine (H. F. T., C H. I..), Pathoh~gy (S. M. C.. F. Y. 1..) and Clinical Pathology (C. H. W.. S. It. C., Y. C. C.. M. C. S.), National Taiwan University Hospital, Taipei, Taiwan, Republic of China. Address reprint requests to: Hwei-Fang Tien, M.D., Department of Internal Medicine, National Taiwan University" ltospital, No. 1 Chang-Te St., Taipei, Taiwan, Republic of China. Received October 4, 1988; accepted December 26, /988.
89 ~J 1989 Elsevier Science Publishing Co., Inc. 655 Avenue of the Americas, New York, NY 10Ill0
Cancer Genet Cytogenet 39:89-97 (1989) 0165-4608:89/503.50
90
tl. F. Tien et al.
crisis of CML varies greatly in different areas ]5]. In blast transformation, the blasts can be either lymphoid or myeloid [6, 7]. The nature of the additional chromosomal abnormalities was also found to be different for these two phenotypes of blasts involved in the transformation [8, 9]. There have been few reports concerning karyutypes of Chinese patients with transformation of CML in the past. This report presents results of chromosome study for 23 Chinese patients with CML in transformation. Blast phenotypes were also characterized by morphology, cytachemistry, and immunocytochemistry in 16 patients with acute blast crisis who had more than 30% blasts in the bone marrow or more than 20% blasts in the peripheral blood and were correlated with the chromosomal abnormalities. MATERIALS AND METHODS
Cytogenetic study was performed on 23 unselected Chinese patients with Ph-positive CML in transformation at National Taiwan University Hospital from November 1985 to September 1988. Sixteen patients were in acute blast crisis and seven in accelerated phase. The criteria for diagnosis of blast crisis was the presence of more than 30% blasts in the bone marrow or more than 20% blasts in peripheral blood. Those patients in accelerated phase had clinical and hematologic deterioration suggestive of transformation but did not manifest the criteria for blast crisis [10]. Bone marrow (BM) ()r peripheral blood (PB) cells were harvested directly or after 1-3 days of unstimulated culture. The metaphase chromosomes were banded by the conventional trypsin-Giemsa banding technique [11] and then karyotyped according to the ISCN [12]. In patients with acute blast crisis, the phenotype of blasts was characterized through morphologic, cytochemical, and immunocytochemical studies. Cytochemical staining, which included myeloperoxidase, Sudan black B, periodic acid-Schiff. u-naphthyl butyrate esterase, and chloroacetate esterase staining, were performed on air-dried smears of BM aspirates or PB as previously described [13]. Immunocytochemical studies were carried out on blast cells isolated from BM or PB by FicollHypaque gradient centrifugation by indirect alkaline phosphatase method [14, 15] with a panel of monoclonal antibodies (Table 1). The enzyme terminal deoxynucleotidyl transferase (TdT) was shown by a peroxidase antiperoxidase method [16] on cytocentrifuge smears. The blastic transformation was considered myeloblastic when blasts were stained positively with peroxidase, Sudan black B or monoclonal antibodies My-7, My-9, or OKM-1; megakaryoblastic when they were positive for Table 1
Specificity of monoclonal antibodies
Antibody HI,A-DR (la) B4 J5 (CALLA) B1
Leu-9 Tll Leu-1 OKM-I My-7 My-9 HPI-ID
Specificity B lymphocytes, activated T lymphocytes, monocytes, non-T-ALL and AML blasts B lymphocytes Non T-ALL, Burkitt lymphoma, follicular lymphoma, some T-ALL B lymphocytes Pan-T lymphocytes Pan-T Iymphocytes (Sheep erythrocyte receptor) Pan-T lymphocytes Granulocytic, monocytic series and large granular lymphocytes Monocytes and Pan-myeloid cells Monocytes and myeloid precursors Megakaryocytes. platelets
Chromosomes of CML in Transformation
91
H P I - I D : and l y m p h o i d when the.,,, were peroxidase negative, TdT positive, and reacted with l y m p h o i d markers such as J5, B4, B1, or Leu-9.
RESULTS Seventeen (74%) of the 23 patients showed chromosomal abnormalities in addition to the Ph. Among them, the c o m m o n nonranctom chromosomal aberrations were +Ph, +8, +19, and +21 with an incidence of 47%, 41%, 35%, and 29%, respec'tively. Twelve (71%) of the 17 patients had either one of t 8 , +Ph, ~-19, or - 21, or most frequently two or more in combination, lsochromosome for the long arm of chromosome 17 was found in only one patient (case 19). Cytogenetic: study was also done at chronic phase for 13 patients including four or the five with trisomy 21: all showed simple Ph without other abnormalities. The detailed chromosomal changes at transformation are shown in Tables 2 and 3. Of the 16 patients in blast crisis, 11 had myeloid transformation (including myeloblastic: and megakaryoblastic) and five had l y m p h o i d transformation. All the patients with myeloid blast crisis had extrachromosomal abnormalities. Eight patients had - P h , six had - 8 , six had - 1 9 , and five had ~21 (Table 2). In ten, a h y p e r d i p l o i d clone was present, and none had a h y p o d i p l o i d clone. Three of the five patients with l y m p h o i d blast crisis showed additional chromosomal changes. One patient had +Ph but none had +8, -' 21, or +19 (Table 2). A h y p o d i p l o i d clone was present in one patient and a h y p e r d i p l o i d clone in another one. Isochromosome of 17q was not found in any patient in acute blast crisis of CML although other kinds of abnormalities involving chromosome 17 were present in three patients: two resulted in m o n o s o m y 17p (Figs. 1 and 2) and another in trisomy 17 (case 2). The mean survival after diagnosis of acute blast crisis was 11 months for patients with lymphold transformation and 3 months for those with myeloid transformation. Three of the seven patients in accelerated phase had extra chromosomal abnormalities. One showed i(17q), one - 1 7 , ~-r(4): and the remaining one t(3;7)(q26;q21) (Fig. 3), who had marked thrombocytosis with platelet c:ount of 2.1 × 10~//.d.
DISCUSSION Transformation of Ph-positive CML is usually accompanied by karyotypic evolution [3]. The most frequent chromosomal aberrations are +8, ,-Ph, and i(17q) [3]. The cells derived from the new clone have a growth advantage over both normal cells and the chronic-phase Ph-positive clones [10]. Our data revealect that 74% of the patients with CML in transformation (43% of those in accelerated phase and 88% of those in acute blast crisis) had chromosomal abnormalities in addition to the Ph. This incidence is comparable to that reported from other centers [3, 17]. However, some different characteristics of the c o m m o n extra chromosomal aberrations were found in this study. Among the 23 patients in transformation, eight had -~Ph, seven had ~ 8, six had --19, and five had +21, respec:tively. The occurrence of +21 was unexpectedly high (22')/0 of the total patients and 29% of the patients with additional chromosomal abnormalities] as compared with that in other areas [8, 9, 18-20]. Clinically, all five patients showed normal appearance and mentality without evidence of Down syndrome, and the karyotype of the four patients who had also been studied cytogenetically in chronic phase revealed a simple Ph without +21 at that time. On the other hand, the incidence of i(17ct), which is usually around 25% in other centers [3, 20, 21], was rather low in our series (7°/,,). Mitelman and Levan [4] first noted uneven geographic: distribution of n o n r a n d o m
92 Table 2
C y t o g e n e t i c r e s u l t s i n 16 p a t i e n t s w i t h C M L in a c u t e b l a s t c r i s i s
(;as(: number
Age/Sex
Blast p h e n o t y p e
3 '~
46;M
Myeloblastic
4
45,'F
Myeloblastic
9°
34/M
Myeloblastic
10
35/M
Myeloblastic
13
50/M
Myeloblastic
14 ~'
47/F
Myeloblastic
15"
42;M
Myeloblastic
7°
40/F
Megakaryoblastic
1"
43/M
12"
26,M
Myeloblastic -:megakaryoblastic Myeloblastic megakaryoblastic
16
26/F
2"
32/M
Myeloblastic lmegakaryoblastic I.ymphoid
5 6"
36/M 12/M
Lymphoid l.ymphoid
8
54/F
Lymphoid
11"
26/M
Lymphoid
N u m b e r of cells:Karyotype 17/46.XY.t{9;22}[q34:ql )1 l : 5 0 . X Y . - 4 . ~-13.41,q. ~ 21. + I ) . l q - . 7 p - , t [ 9 : 2 2 ) . - Ph 2."46,XX.t[9:22)Iq34;q 11 ) 22.'47,XX. - 8,t[9:22) 11."46,XY,t(9:22](q34;q I 1 ) 6/48,XY,*8,t(9:22h • Ph 3;46,XY,'8, 10,t(9;22) 2;48,XY, - 8 , - 19.t[9:22) 1"47.XY,+ 8 , - 10,t(9:22),.- Ph 2"46,XY,t(9;22)[q34;q 111 3,"46.XY.2q ÷ .t[9;22/ 4,"50,XY, ÷ 8,+ 10,+ 12,t19:22),- Ph 1;46,XY,t19;221{q34;q 11 ) :);47,XY, * 8,t(9;22) 1,'47,XY,t(9:221, + Ph 14,'49,XY. ~-8, 4 19.* 21,t(9;22) 6,'50,XY. + 8,4- 19, ~-21 ,t(9;22),- Ph 18."46,XX,t(9;22)(q34:q 11 } 7.;49,XX, + 8, + 19,4- 21 I(9:221 18/46,XY,t(9;22}(q34.q 11} 1,'47,XY,t(9:22), + Ph 2.'48, 4. 6,t(9;22). ~-Ph 23/46,XX,t(9;22][q34;q11 ) 2,'47,XX. 7 , ' 8 , + 2 1 . t [ 9 ; 2 2 ) 26."47,XY. * 21 ,t(9;22 )(q34;q 11 ) 8,'46,XY,t(9:22J(q34;q I 1 ] I 0/47,XY, + Ph,t(9;22J 1 ' 4 9 , X Y , - 1 , 4 2 , - 7 , - 8,+8, - 1 0 , - 1 2 , - 1 3 , 17. 2 0 . - 21 ,del[20)(q 11 ),t(9:221,~- der( 17J,t( 7;?: 17J ( 7 q t e r - . 7 q 11 ::?:: 17 p 11--~ 17qte.r)..'- Ph.-*- Ph 1/54,XY,+ 1 , - 2 , + 6 , - 7 . + 8 , + 8 , - 10, + 12,+ 19,t(9;22), + der(17),+ P h , + Ph 6/46.XX.del(7J(p 15),t(9:22}(q34;q 111 9i48,XX,+ 19.del[7J,t(9:22J,+ Ph 19i46,XY,t(9;22)(q34;ql 1 ),t(10;16)(q22;ql 3J,t[14:22) (q23;q13) 7;49,XY, + 15, + 17,t(9;22J,t(10;16),t(14:22), ~-Ph 26/46,XY,I(9;221(q34;q 11 J 15/46,XY,t(9;22}[34:q 11 ) 3/46.XY,-- 17,t(9;22), ~-der(l )t(1:17)(pl 1 ;ql 1 ) 15/46,XX,t(9;22)(q34;q11 ) 6/45,XX, 7,t(9;22) 26146,XY,t(9:22)(q34:ql 1 )
" The karyotypes of these~ patients had been studied in (:hronic phase and all showed simph: Ph :vithout additional change.
Chromosomes
o f C M L in T r a n s f o r m a t i o n
Table 3
93
C y t o g e n e t i c r e s u l t s in s e v e n p a t i e n t s w i t h C M L in a c c e l e r a t e d p h a s e
Case. n u m b e r
Age/Sex
Number of cells/karyotype
17
58:M
18"
35/M
2,'46,XY,t(9;22)((134;q 11 ) 13,,'46,XY.t(3;7)lq26:q211,t(9;221 3:45.X,-. Y,t(3:7],t(9:22) 1/46,XY 1/46.XY,t(9;22)(q34;ql 1 ) 22,'46,XY, - 17.4- r(4),t(9:22)
19" 2(1 21" 22 23
35,'F 65iF 27iM 38/M 12/M
9/46,XX,i(17q),t(9;22}(q34:q11) 26/46,XX,t(9:22}(q34:q 11 ] 7/46,XY,t(9:22)(q34:ql 1 ) 20/46.XY,t(9;22)(q34:q 11 ) 12/46.XY,t(9;22)(q34:q111
° The karyotypes of these patients had been studied in chronic phase and all showed simple Ph without additional change.
F i g u r e 1 Karyotype fronl one bone marrow (:ell of case 12, s h o w i n g 49,XY. ; 1. ~ 2, 7.+8, ÷ 8. -10,+12,-13, 17. 2l~de~l(2~((|~.t~9~22~((]34~q~)`(ier(~7~(7:`~7}I7qte`r~7q~?::~7p~7~ter + Ph, + l'h.
=
:.
@
D,
""
\
""
i¢
"ll
3
4
5
2 ~
ii
Jv
6
7~
• 8 \
I
9
~(
•
\ 10
11
12 ~
k
!
J~
\
13
15
14
16
/e 17
18 4
|
W(
•
19
2O
~1~,
22
X Y
],
94
tt. F. 'l'ien e,t al.
1
17
9
22
d*d~),t(l:~ 7) F i g u r e 2 Partial karyotypes from two bone marrow c:ells of (:ase 6, showing -17,+der[1) t(l;17)(pllzqll), in addition to Ph chromosome.
F i g u r e 3 Partial karyotypes from two bone marrow (:ells of case 17 who had thrombo(:ytosis, showing t(3:7)(q26;q21) and Ph chromosome.
3
7
9
22
tlL~"
- I1 Jm
t(3:7)
-
t(S~2)
Chromosomes of CML in Transformation
95
and specific (:hromosome aberrations in malignant disorders. At the time of blast crisis of Ph-positive CML, Milehnan [5] foun(t that the incidence of the three most common additional abnormalities varies in different geographic areas: r8 from 21% in Sweden and New York to 46% in Japan, • Ph from 7% in the t;.S.S.R, to 54% in France, and i(17q) from 7% ira Sweden to 28% in the U.S.S.R. Trisomv 21 was not included in the study. As a whole, the incidence in lapan was most similar to ours: 46% for - 8 , 41.8% for - P h , and 15.2% for i{17q) in ]apan and 41%, 47%, 6%. respectively, in our series. The reason for this apparent geographic heterogeneity in the incidence of n o n r a n d o m chromosome aberrations is still u n k n o w n . Mitehnan 151 suggested differences ira the distribution of etiologic factors or genetic:ethnic hackground might be the possible explanation. More extensive studies in various t:ountries and areas are necessary to clarify, this issue. A total of five patients had abnormalities involving chromosome 17, which resuited in monosomy 17p in four of the five (cases 6, 12, 18, and 19). These, were i(17q), t{1:17}{p11:q11), a complex translocation involving #7, #17, and an u n k n o w n chromosome, and monosomy 17. Borgstrom et al. [221 suggested monosomv 17p might represent a crucial event in the terminal phase of CML. The aberration t{1;17)(pll;q11) was the same as that reported by Kerman el al. [231. They reviewed six such cases, two from their laboratory and four from the literature and concluded that this marker might be specific for the accelerating and blast phase of CMI,. In acute blast crisis of CML, blast cells can be either mw',loid or lymphoid [6.71. Rowley [24} noted that no patient with Ph-positive lymphoid dise, ase was reported to haw.' ~ 8, i(17ql, or +19. while most patients with n)yeloid disease had one or more of these abnormalities. 'rwo later reports [25, 26] did not support her findings. Blast phenotype (:an no,,',' be characterized more act:uratelv by (:vtochemical and (:omprehensive immunologic marker studies [6]. A correlation between extra chromosomal abnormalities and the type of blast crisis has emerged more (:learlv. Parreira et al. 181 found that +8, i{17q), anti - 1 9 were seen only in patients with myeloid but not lymphoid blast crisis of CMI,. Diez-Martin et aI. [9] reported similar findings. They also noted an association between abnormalities of chromosome 7 anti lymphoid blast crisis. Our data showed that 82% of the patients with myeloid blast crisis had ÷8, + 19 or both, while none of those with lymphoid transformation (lid. lsochromosome of 17q was seen in neither of them. It is of interest that • 21 ,,','as also present only in patients with myeloid crisis. Of the 18 cases in lymphoid crisis of CML ret)orted by Yao 127], six from his institute and 12 from the literature, none had ~ 21. It was also absent in the eight cases with lymphoid transformation reported by DiezMartin el al. [9]. However Parreira el a]. [8] found that one of their six patients with lymphoid blast crisis had ~21, though it is not known whether - 2 1 in this patient reflects constitutional change or not. It may be suggested that acquired +21 is a rare marker if not totally absent in lymphoid transformation of CMI,. Abnormalities of chromosome 7 in our study were present in both myeloid and lymphoid transformation in contrast to the previous reports [8, 9]. No definite (:nrrelation was found between the subtype of myeloid transformation and chromosomal abnormalities [9]. In one study [8} +19 was seen only in [)last crisis involving megakaryoblasts, which was not the case in the other report [91. In our series, ,19 was present not only in transformation involving megakaryoblasts but also in myeloblastic transformation. More studies are needed to clarify this issue. The authors thank Dr. 1. Whang-Peng for he,r kindness in checking the karyotypes. Dr. I,T Yam and Dr. CY Li for instruction in immunocytochemical staining, and Dr. Wl. Nichols for sending us monoclonal antibody HP1-1D. Supported in part by a grant from the National Science Council of the Republic of China, NSC 75-0412-B01}2-107 and 76-0412-B0112-121.
96
H. 1.'. Tien et al.
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C h r o m o s o m e s of C M L in T r a n s f o r m a t i o n
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23. Kerman SL. Miller RB, Heritage DW 11986]: Translocation (1:17) in accelerating and blast phases of chronic myelogenous leukemia, Cancer Genet Cytogenet 20:269-277. 24. Rowley JD (1980): Ph-positive leukemia including chronic myelogenotls leukemia. Clin Haematol 9:55-86. 25. Alimena G. Dallapiccola B, (_;astaldi R. Mai~delli t:. Brandt I.. Mitelman t", Nilsson P{; (1982): Chromosomal, morphological and {:linical (:orrelations in blast crisis of chronic mveloid leukemia-, a study of 69 cases. Stand I Haematol 28:103-117. 26..Muehh.~ck Si'}, M(:Kenna RW. Arthur DC. Parkin ]I., Brunning RD (1984): Transformation of chroni{: myelogenous leukemia: (21ini(:al. Inorphological. and t:ytogenetic features. Am ] Clin Pathol 8 2 : 1 - 1 4 . 27. Yao E (1986): Chromosomal abnormalities in ]ymt}hoid crisis of chrmlic myelogenous leukemia. Cancer J Cytogenet 16:235-243.